Manual on Plant Health Clinic

2020

Manual on

PLANT HEALTH CLINIC Compiled by

Uma Shankar Ranbir Singh Neetu Sharma Ajay Pratap Rai Hafeez Ahmad D.P. Abrol

Faculty of Agriculture Sher-e-Kashmir University of Agricultural Sciences & Technology of Jammu Main Campus Chatha, Jammu-180 009 (J&K), India

Manual on

PLANT HEALTH CLINIC

Compiled by: Uma Shankar Ranbir Singh Neetu Sharma Ajay Pratap Rai Hafeez Ahmad D.P. Abrol

Faculty of Agriculture Sher-e-Kashmir University of Agricultural Sciences & Technology of Jammu Main Campus Chatha, Jammu-180 009 (J&K), India

Preface A variety of symptoms are produced in plants in response attack of different pests and diseases.

The situation becomes complex when similar symptoms are produced on the same plant by completely different factors such as disease, weeds competitions and nutrients deficiency. Therefore, accurate diagnosis of pest/disease including weeds and nutrients deficiency assumes utmost significance.

The reliable estimation of crop losses in relation to different pest and diseases is of paramount importance aimed at increased understanding of the factors responsible for plant damage at different stages of crop growth and development. To arrive at logical conclusions, one must understand the mechanics of insect-plant, diseases, weeds and nutrients interactions and how they affect crop yield.

Efficient pest management depends on an accurate diagnosis of the pest problem so that curative action is initiated well in time. Knowledge of the feeding mechanism of insects, intensity and infestation of disease, weeds competition and nutrients can play a role in arriving at right conclusions for timely management.

I express deep sense of gratitude to Prof. D. P. Abrol, Dean, Faculty of Agriculture, for his enormous help, guidance and interest in preparing this manaual. I owe my sincere thanks to the team of scientists associated with Plant Heath Clinic SKUAST-Jammu, for their timely help and cooperation to formulate this manual.

Dr. Uma Shankar Manager Plant Health Clinic

Contents Chapter

Title

Page No(s).

1.

About the Plant Health Clinic

1

2.

Introduction

3

3.

Advanced Techniques in Plant diagnostics

5

4.

Crop Loss assessment

12

5.

Field diagnosis of damage caused by different types of insect pests

21

6.

Procedure for collecting plant and insect Samples for problem diagnosis

34

7.

Plant Disease Diagnostic Techniques

42

8.

Acquaintance with Plant Pathology Laboratory and Equipments

47

9.

Seed Treatment

52

10.

Sterilization

55

11.

Isolation of Fungal and Bacterial Pathogens

57

12.

Preparations of Culture Media: Potato Dextrose Agar (PDA) for Fungi and Nutrient Agar (Na) for Bacteria

60

13.

Methods of Application of Fungicides: Soil Application

63

14.

Methods of application of fungicides: Seed and foliar application

65

15.

Major diseases of crops in Jammu region

68

16.

Identification of weeds in crops

71

17.

Methods of Herbicide Application

79

18.

Nutrients Deficiency Management

83

Plant Clinic-SKUAST-J Faculties

Chapter-1

About the Plant Clinic-SKUAST-Jammu

P

lant Clinic is the integration of four major disciplines of agriculture (Entomology, Plant Pathology, Agronomy and Soil science) which provides a common platform to the students, line

department officers and Farmers to train them in a better way to diagnose the insect-pests, pathogens, weeds, nutrient deficiency and their symptoms produced on different crops such as cereals, pulses, oilseeds, vegetable crops, fruit crops and ornamentals including medicinal plants and in various cropping ecosystem. Further, PC-SKUAST-J is crafted in such a way to suggest the Best Management Practices to up-gradation of skills of students, field functionaries and extension officers, and farming communities to impart the gap between the know-how and do-how of latest technologies and their proper dissemination.

Four Pillars of Plant Clinic Setting up Plant Clinic (Diagnostic Laboratory) The plant clinic acts as the students and farmer interface; the place where the individual questions are answered and needs are met with the use of audio-visuals and samples/specimens. It provides expert support, capacity building, training and diagnostics. The team works alongside local partners to train the local people to become ‘Plant Doctors’. Then share the knowledge in surveillance and diagnostic techniques, integrated pest management, technology development, judicious use of ecofriendly pesticide and reducing the pesticide pressure on ecosystem, markets and government policy.

How the plant clinic works?

When the farmer (people) has a problem with a crop, he/she can bring a sample along to the plant clinic. At the clinic, trained ‘plant doctor’ listens to the peoples’ problem, examines the sample, diagnoses the problem and offers a suggested treatment. Treatment suggestions are affordable for farmers with the use locally available resources. The correct chemicals are recommended only when necessary as the last resort. With access to these services, students/extension officers/ farmers can tackle Insect-pests, diseases, weeds and nutrients problem and produce healthy crops on their farm in a sustainable manner.

1

Table 1: Plant health clinics around the world Country No. Bangladesh 25 Bolivia 7 early DR Congo 8 India 2 Indonesia 2 Nicaragua 14 Uganda 4 Vietnam 2

Started Managed by 2004 RDA Bogra, AAS, and Shushilan 2004 CIAT Santa Cruz, PROINPA, and UMSS March, 2006 UniversitĂŠ Catholique du Graben, Butembo August, 2006 GB Pant University of Agriculture and Technology, Pantnagar October, 2007 University of North Sumatera (USU) March, 2005 Farmer organisations, NGOs, INTA, and others.

Supported by PASA II (danida) and other donors July, 2006 Socadido, SG2000, Caritas and MAAIF June, 2007 SOFRI

With India planning to introduce Plant clinics in all 40 states, the stage is set for providing poor farmers with better advice that helps them grow healthy crops with reduced risk and lower costs.

Role of Plant Health Clinics for Advisory Services: Experiences from Eastern Africa Negussie et al. (2011) highlighted that changing policies, declining public funding, new thinking and approaches, climate change and other environmental factors pose major challenges to public sector extension services. Pre-packaged, crop and region-biased extension approaches often failed to help remote and resource poor farmers to cope with rapidly changing realities. This has necessitated the search for alternative approaches. One such alternative, plant health clinics, has been developed and tested by the CAB International led Global Plant Clinic Alliance in many countries since 2001.

Generally, Plant health Clinic is operated by staff of local organizations in locations that are easily accessible to farmers and accept any crop problem. They provide regular, relevant and practical advisory services on plant health management. Clinic records generate useful information on priority problems and changing status of pests and diseases.

The plant health clinic approach has been widely introduced in Africa under a CAB International initiative, Plant wise. In Uganda, a pilot scheme began in 2005 and has expanded to 19 regularly operating plant clinics. Kenya established 24 plant clinics between mid-2010 and September 2011, while Rwanda started with five clinics in 2011. The clinics are run by a range of organizations, each adapting the basic model according to local circumstances but consistent with guidelines and principles established by CAB International. The approach demonstrates a number of strengths and benefits but is not without challenges.

2

Chapter-2

Introduction

A

large number of diverse forms of life have been scattered in nature like gems wherein, only plants are autotrophs which can be able to manufacture their own food. Besides plants all living

forms like animals and human being are directly or indirectly depend on plants for their survival.

Since antiquity, human being has transformed from hunter gatherer to food producer and this process is still continues.

Plant health is an important aspect which has been affected by a large magnitude of living and non-living factors. Living factors include insect-pests, mites, weeds, disease causing agents like fungi, bacteria, viruses, nematodes, rodents, birds, mammalian pest etc. while the non-living factors include environmental variables like temperature, relative humidity, rainfall, hailstorms, winds, drought, sunshine etc. Besides all these, soil acts like a living substrate to anchor, nourish and sustain the plants life forms and their diversity. Sometimes, the plant may adversely affected by the above mentioned factors which require a careful observation to come on any conclusion. Careful observation enables us to diagnosis the constraints accurately for initiation of right action.

Insects may harm the plants in different ways such as damage caused due to feeding, oviposition or disease transmission. However, diagnosis becomes easier if the insect causing damage is also observed and identified. Accurate insect-pest management depends on an accurate diagnosis of the problem. For this, the pre-requisite to determine whether an insect observed on a crop plant is a pest or not. The type of mouthparts of an insect species and the manner of feeding on the host plant are both important determinants of its pest status.

Typically, diagnostics is a process to come up with the best possible explanation of why a good plant has gone wrong. An incorrect diagnosis will lead to an incorrect treatment. A plant may be suffering from multiple problems, and the most obvious may not be most significant. For arriving at a logical conclusion, one has to observe the problem from different perspectives: 1.

Knowing the plants One must have to understand the difference between a normal and an abnormal plant, which

could provide a great early perspective in the diagnosis process. All plants have their own set of diseases and insect problems. Knowing the plants, their family and genus and their bio-ecology is a great starting point for diagnostics.

2.

Looking for abnormalities in the plants Plant abnormalities are categorized in terms of signs and symptoms. Signs are the actual causal

agents. Symptoms result from interaction between the plant and pests, pathogens, or environmental elements (e.g. high soil pH, nutrient deficiencies). However, some symptoms may be produced by multiple causes. Pattern of damage is characteristics of some particular insect species which can be very helpful in diagnosis.

3

Examples: Twisted leaves may be caused by sucking pests like leafhopper, thrips, or leafminer or exposure to plant growth regulator herbicides. Similarly, tiny leaf spots can be caused by a leaf spotting fungus or bacterium, or lace bugs and mites. Yellow leaves may be caused by sucking pests or by nutrient deficiencies in the soil or by a soil pH that makes the nutrients unavailable to the plants.

Some of the major symptoms produced in plants includea)

Chewed leaves or blossoms, e.g. defoliation, shot holes, margins notched, skeletonization; Discoloured leaves or blossoms eg. Stippling, streaking, mining, yellowing;

b)

Distorted leaves, branches, or trunks e.g. leaf cupping, leaf or twig galling, bark cracking; Dieback of shoots, twigs and branches e.g. shoot die-back, branch dieback;

c)

Products of insects e.g. honeydew and sooty mold, fecal spots, silk, protective coverings, fluffy white wax, soft or hard white, brown, gray or black covers, etc.

Therefore, examine all plant parts closely and carefully.

3.

Knowledge about plant site conditions and environmental history Conditions under which a plant is growing also affect plant growth and development. This may

not be confused with the stunted growth caused by sap sucking insects or nematodes infestation.

Poorly drained soil with poor internal aeration may result in death of plants. Acid loving plants often develop yellowing between the veins (interveinal chlorosis) if growing in alkaline soils (pH above 7) due to iron deficiency.

Plant stress may also produce a progression of symptoms. There are two types of plant stress: acute and chronic. a)

Acute stress is caused by an immediate event, such as feeding, drought, leaf defoliation, etc.

Symptoms are usually immediately visible and easy to diagnose.

b)

Chronic stress is caused by more subtle conditions such as site problem or agronomical problem. How harsh have been the winters or summers need to be known.

4.

Knowledge of available diagnostic tools Useful tools for diagnosis can be high tech, ranging from elaborate microscope and enzyme

linked immune-sorbent assay tests for virus and fungi in diagnostic labs to simpler tools e.g. soil probe (to check soil pH), hand lens (10X or 20X magnification), cutting tools (e.g. good sharp hand pruners and knife), digging tools (to check girdling roots e.g. spade), recording tools (e.g. a field notebook), a digital camera, a hand-held recorder, sampling equipment like specimen tubes, plastic bags, etc.

5.

Reporting of diagnosis and recommendations After diagnosis, it is important to put the suggested problem into proper perspective relative to

overall plant health. While recommending treatment for the problem, remember that sometimes “doing nothing� is the best recommendation when the problem is of minor importance.

Knowledge of pest life cycle and thereby crucial timing of initiating a control measure is very important. The recommendations should be made within a range of proper expectations.

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Chapter-3

Advanced Techniques in Plant diagnostics 1.

Use of computer based expert systems in pest diagnosis

A. Expert systems have developed from a branch of computer science known as Artificial Intelligence (A.I). Artificial intelligence is primarily concerned with knowledge representation, problem solving, learning, robotics, and the development of computers that can speak and understand human like languages. Thus, expert systems are computer programme designed to mimic the thought and reasoning processes of human expert.

B. Expert system can be developed for many kinds of applications involving diagnosis, prediction, consultation, information retrieval, control, planning, interpretation and instruction. In USA, computer based diagnostic systems for diseases, insect-pests and physiological disorders are available.

Examples: In citrus and selected tropical fruit crops, the TFRUIT.Xpert and CIT.Xpert computer based diagnostic programmes can quickly assist commercial producers, extension agents and homeowners in the diagnosis of diseases, insect-pest problems and physiological disorders. Users can also refer to summary documents and retrieve management information from the University of Florida’s Institute of Food and Agricultural Sciences extension publications through hypertext links. The programme is available separately on CD-ROM and each contains over 150 digital colour images of symptoms.

2.

Computer Simulation models Computer models can provide some theoretical explanations of the effect of injurious or

competitive organisms on crops. In general, computer models depend on a few known variables that influence plant growth, development, and production. However, in reality plants respond to damage or changes in the environment in a very complex manner. Thus far, such complexity cannot be incorporated into the models to simulate an actual situation.

However, good simulations or computer models can improve the theoretical understanding of the major effects of injuries or damages of pests on plants and their yield.

3.

Imaging technique New technologies and improvements to existing technologies are constantly changing

the way we view objects. When photographs or image recordings from a tower, balloon, plane, or satellite are available, they can give a useful indication of the area and intensity of dead or wilting plants or leaves and differences in crop yield caused by pest attack. Remote sensing techniques such as radar can automatically monitor the height, horizontal speed,

5

direction, orientation, body mass and the shape of arthropods intercepting the radar beam. It can provide information of aerial migration of pests and natural enemies. It can be particularly useful for monitoring locust swarms.

Radar entomology was first used in 1968 and since then comprehensive and intensive studies have been conducted in the UK, USA, Australia and China and it was predicted that fully automatic, season long and real time monitoring will be feasible with the verticallooking radar. Remote sensing technique relies on changes in the absorbance or reflectance of plants in response to pest attack. An instrument sensitive to specific wave lengths of radiation is used to detect such changes. Remote sensing in conjunction with ‘3S’ technique can help in achieving three-dimensional real time visualization of insect pest populations.

Imagery provided by remote sensing satellites could be utilized in identifying pest affected areas and intensity of pest damage. This could be particularly useful for pests which produce visible symptoms of crop damage over large area e.g. hopper burn symptoms in paddy, blacking of cotton leaves caused by sooty mould growing on honey dew secreted by aphid and whitefly, etc. Similarly, satellite data have also been used to identify areas of vegetation capable of supporting desert locusts. Further, such data can also find application in studying the effect of environmental changes on build-up, long distance migration and flight behaviour of air-borne pests.

The Australian Centre for Remote Sensing (ACRES) has introduced a new service to provide satellite data for real time application. The STAR (Speedy Transmission After Reception) service provides access to digital satellite data on various aspects which includes monitoring of pest infestations. The difficulty, apart from clouds, is to be able to relate pest and crop events on the ground to the pictures obtained.

The Distance Diagnostics through Digital Imaging project enhances the ability of the University of Georgia Cooperative Extension Service to evaluate and propose solutions for agricultural problems, including plant diseases and pests, through the use of digital imaging and the World Wide Web. Imaging stations consisting of computers, digital cameras, microscopes and image-capture devices have been deployed in 94 county offices and in 3 diagnostic labs.

4.

Acoustic and other tools Sensors which can detect the sounds of hidden insects like wood borers, termites,

stored grains pests, etc are finding applications in the advanced countries. Similarly, portable X-Ray machines are being employed for detection of insects attacking forest trees.

6

5.

Electronic nose In Oregan (USA), electronic devices programmed for detecting particular odour or

smell are being evaluated. One of these devices, Cyranose 3201, a portable electronic nose, has shown good promise in determining stink bug damage by external properties. The volatile compounds given off by sink bugs were identified and E-nose was trained to identify stink bugs’ (presence) smell prints. There was a strong correlation (R2 = 0.95) between the number of stink bugs in a sample and the Cyranose sensor’s response.

6.

Drones, or Unmanned Aerial Vehicles (UAVs) Technology Recent advances in remote sensed imagery and geospatial image processing using

Drones, or unmanned aerial vehicles (UAVs) have enabled the rapid and ongoing development of monitoring tools for crop management and the detection/surveillance of insect pests. Drones technology has enabled the agricultural farmers to see their fields from the sky. This bird's-eye view can reveal many issues such as irrigation problems, soil variation, and pest and fungal infestations. By identifying

the

farmer can attempt

trouble to

spots, improve

the crop

management and production.

Vanegas et al. (2018) described a (UAV) remote sensing-based methodology to increase the efficiency of existing surveillance practices (human inspectors and insect traps) for detecting pest infestations (e.g., grape Phylloxera in vineyards). The methodology uses a UAV integrated with advanced digital hyperspectral, multispectral, and RGB sensors. Further, they implemented the methodology for the development of a predictive model for Phylloxera detection. The drone, which is delivering a whole new perspective of crop disease and pest early warning system to farmers, has brought new hope to many. According to experts, drone’s precision at detecting pests and diseases is over 10 times accurate compared to the human eye. The drones are capable of collecting very high resolution images with very numerous details, thus, they are useful in mapping and surveying farms. Green means the crop has no stress, yellow the crop has scanty stress while red indicates that the crop has too much stress. The drones are mounted with a near-infrared sensor (NIR) capable of detecting pest and diseases, 10 days before it becomes visible to an expert or an extension worker. The gadgets ability to diagnose stress levels in plants at early

7

stages, according to experts, helps in minimising insecticide use. Drones, or unmanned aerial vehicles (UAVs), are playing the role of Plant doctor and are now able to diagnose the stress levels of plants, which could helps in minimising insecticide use. 

A professional UAV such as multispectral eBee SQ Agriculture Drone provides a holistic view on crop's growth, identify issues, and better target their field scouting.



UAV-IQ® improves traditional biocontrol applications by using drones to release beneficial insects exactly when and where they’re needed to suppress pests.



From Sri Lanka to Uganda, UAVs with near-infrared sensors are monitoring plants for pests and disease, with implications for agricultural policy worldwide



For governments and development agencies, drones can provide more accurate, up-todate information on what is being grown where.



For individual farmers, this kind of information could be the difference between a failed crop and a bumper harvest.

Case study In September, the International Water Management Institute (IWMI) carried out trials in Sri Lanka using an eBee drone equipped with a near-infrared sensor; the trials showed how this can give farmers early warning of problems anywhere in their fields. “When a plant goes into stress, it’s either due to a water or fertiliser shortage, or because it’s being attacked by a pest. Photosynthetic activity decreases and that affect the chlorophyll. That’s what the nearinfrared sensor can detect, but our human eye can’t see it until it’s more advanced.” That 10day warning could prevent large-scale crop losses. “If a crop is being attacked by insects, the whole area can be affected, not just one farmer. With UAVs, if you can figure this out before it spreads, you can save the whole area.”

A drone equipped with a camera hovers over potato crops in Peru (Source: International Potato Center)

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Drones in agriculture: a tool for early pest detection The FAO of the United Nations projects that by 2050, human population will likely to be grown to nearly 10 billion people. Farmers will need to produce more with less, while preserving our environment for future generations. And society has a duty to help them achieve this. Although agriculture is perceived as a traditional economic sector, precision agriculture technologies have already boosted crop yields significantly in the last decades.

What drones have to offer to agriculture?

Early pest detection is a major application of drones in agriculture. Depending on the crop, agricultural producers survey their land several times per week. For example, a potato producer is going in the field at least 3 to 4 times per week during the growing stage of the crop. No surprise here, as pests like the Colorado potato beetle, could spread extremely fast and destroy hundreds of hectares per day.

AgroHelper, a Bulgarian startup is working on developing a web-based solution that helps farmers process drone captured images and detect, in real time, zones with potential crop health issues. The platform is powered by a cloud infrastructure and does not require any specific hardware to be present on the farmer’s local machine (or a fast internet connection). Currently, most state-of-the-art software solutions use a process called “stitching” to create an orthophoto map from hundreds of individual overlapping aerial photos. Each individual photo captured by the drone camera contains different terrain features like crop rows, tractor trails or buildings. As the photos overlap, each individual feature is captured by the drone camera multiple times from different angles and perspectives.

AgroHelper’s Health Map feature indicates zones with potential issues Stitching, as the name suggest, is a mathematical process that matches the photos to solve the puzzle and create one high-resolution map. This is the most precise solution available today. However, the process takes from 5 to 10 hours, requires heavy computing power (which translates in high cost) and fails fairly often. The AgroHelper team aims to solve this problem by offering a real-time tool and eliminate stitching from the process. The results are less precise than stitching in terms of resolution but allow farmers to pinpoint an area that requires attention in real time. As the process is optimized, the cost is contained, and AgroHelper provides farmers the opportunity to process 3 maps per month for free.

Prediction of pest attacks and disease outbreaks using UAV/Drone based Hyperspectral Remote Sensing Technology

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Bharat Rohan is involved in empowering farmers by reducing losses through prediction of pest attacks and disease outbreaks in the crops. We are a team of aeronautical engineers and hyperspectral imaging experts implementing affordable UAVs/Drone, Hyperspectral Imaging, data analytics and machine learning technologies to meet local farm challenges.

Work Mechanisms The BharatRohan technology is based on Hyperspectral Imaging which determines minuscule colour changes occurring in the plants due to physiological and phenological changes. For example: When a disease like bacterial blight infects a pomegranate plant, it is caused by a pathogen called Xanthomonas axonopodis pv. punicae. This pathogen releases a number of effector proteins including TAL effectors into the plant through their secretion system. This effector protein causes some biochemical change in the plants and leaves.

With human eyes, these changes become visible only when the water absorbing red-brown spots become visible on leaves. But with Hyperspectral imaging, we are able to identify the colour changes occurring in the leaves due to these biochemical changes even at just the onset of the infestation which helps us in providing early predictions and forecasts to the growers so that losses can be prevented.

They diagnose the problems of the farmer field with a non-destructive method of Hyperspectral Imaging and create prescription maps. These prescription maps are then converted into recommendations reports in vernacular language of the farmers. The recommendations are duly validated by the agronomists/pathologist/entomologists whom BharatRohan has access to being a portfolio company of a-IDEA, TBI of ICAR-National Academy of Agriculture Research Management, Hyderabad.

Current projects Presently Bharat Rohan is working on Mentha cropping system which is a very significant revenue source for the farmers in Uttar Pradesh where 90% of India's Mentha is cultivated and exported. The cropping system involves Mentha, paddy, mustard and potato crop. BharatRohan is building spectral libraries in association with CSIR-Central institute of Medicinal and Aromatic Plants, Lucknow. CSIR-CIMAP handles the microscopic inoculation of pathogens in controlled conditions. The phenological/biochemical changes are then used to generate a comprehensive spectral library which are then correlated with and applied on Drone/UAV based Hyperspectral Data acquired from farmers field using various proprietary unmixing models.

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The major objectives of this project are: -

Provide unprecedented advisory services through a proprietary ICT Platform to around 5000 acres of area in 6 tehsils of Barabanki District

-

Diagnose mineral deficiencies (NPK, Zinc and Iron) in Mentha Cropping System and suggest control methods.

-

Diagnose pest attacks, disease outbreaks in Mentha cropping system and suggest control methods.

-

Characterise weed infestations and suggest control methods.

-

Reduce irrigation frequency

-

Promote use of sustainable precision agriculture

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Chapter-4

Crop Lo oss Asse essmentt Backgroound: The looss suffered by a crop is a function off the pest poppulation, behhavior of thee pest and the bioloogical charaacteristics off the crop plant. Loss is caused by feeding f or dduring the prrocess of oviposittion. It may be b in the forrm of loss off production capacity, loss of producctive capacity y, loss of stand, diirect damagee, product coontaminationn and loss in storage etc. The type of loss by insect pest is influencced by severaal factors inccluding the organ o of the pest used too cause damaage, part of the plant attackedd, amount of destruction per p unit timee and damagee in relation to t insect num mbers.

Sinnce the beginnnings of agrriculture abouut 8-10,000 years y ago, huuman beingss had confron nted with the harm mful organism ms called pests which coomprises anim mal pests (innsects, mites,, nematodes,, rodents, slugs annd snails, birdds), plant paathogens (virruses, bacteriia, fungi) andd weeds (i.e.., competitiv ve plants) for cropp products grown g for huuman use annd consumpttion. These organisms o m may be contrrolled by applyingg cultural, physical, p mecchanical, Hoost Plant Reesistance, bioological andd chemical measures. m Crop prootection has been develooped for the prevention and a control of o crop lossees due to pessts in the field (prre-harvest losses) and duuring storagee (post-harveest losses). Biotic B stressoors have the potential to reduce the crop prroduction subbstantially.

Abiotic and biotiic factors ca ausing crop losses e of watter in the groowth season,, extreme Appart from biottic stress, abbiotic stress like lack or excess temperattures, high or o low irradiaance (factorss which can be controlleed only withiin narrow lim mits) and nutrient supply also play an impoortant role inn crop producction.

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Every year, a large proportion of crop yields are lost due to the attack of insect- pests, diseases, weeds and other pests like rodents, etc. Such losses are particularly high in the developing countries.

To determine what factors damaged the plants require investigative approach combined with careful observation and ability to put all the pieces together to reconstruct the event(s) that caused the damage. Accurate diagnosis must be made before undertaking corrective action.

In diagnosing plant damage, a series of deductive steps can be followed to gather information and clues from the complex and general situation down to specific, individual plant or plant part.

Thus, through the systematic diagnostic process of deduction and elimination, the most probable cause of the plant damage can be determined. Pest management decisions taken on the basis of incorrect identification of the causal agent of the damage could result in pest control failures and economic loss.

Pest infestations often have adverse effect on yield. Therefore, it becomes essential to accurately estimate the potential role of each agent in reducing yields so that based on their incidence the potential losses could be predicted. The understanding of the mechanisms which are involved in quantitative and qualitative crop losses could help in formulating appropriate strategies to minimize them. It would help in identifying the economic status of different pests.

Historical perspective: Zadoks (1981) identified three periods in the history of concern about crop loss assessment: exploratory, emergency, and implementation.

Zadoks and Koster (1976) reported that German Korn in 1880 was the first to stress the importance of using crop loss assessments for scientific and managerial purpose. Later on different countries like Sweden, Netherland and Prussia began to assess losses. The world’s first plant protection service started its work in the Netherland in 1899.

The exploratory period came to an end with the 1914 International Phytopathological Conference in Rome.

The periods of the two World Wars was the emergency period in which international exchange of commodities was hampered. Such situation coupled with droughts and famine caused food shortages resulting in loss of human life.

The implementation period was first initiated by the phytopathologist E.C. Large (1950) in the United Kingdom. However, international interest on this aspect was stimulated by Food and Agriculture Organization (FAO) symposium on crop losses held in 1967 in Rome, which was organized by L. Chiarappa and J. Vallega (FAO, 1967).

Work on crop loss methodology was strengthened by two publications produced under the aegis of FAO (Chiarappa, 1971, 1981).

Pest infestations often have adverse effect on yield. Therefore, it becomes essential to accurately estimate the potential role of each agent in reducing yields so that the potential losses based on their

13

incidence could be predicted. The understanding of the mechanisms which are involved in quantitative and qualitative crop losses could help in formulating appropriate strategies to minimize them.

Basic crop loss terminology (after Zadoks, 1985) Yield: A crop’s measurable economic production.

Injury: Any visible and measurable symptom caused by a harmful agent. The damage function translates injury into damage.

Damage: Any reduction in quantity and/or quality of yield. The loss function translates damage into loss.

Loss: The reduction in financial return per unit area due to harmful agents.

Therefore, the assessment of crop losses due to insect pests is of important from the following points of view: 1.

For proper planning of research. For example, if the mechanisms of crop yield are known, research can be directed toward increasing yields by reducing the effect of pests on yield and yield quality, increasing crop resistance to pests, reducing pest attack by forecasting pest outbreaks.

2.

For defining economic status of a pest species so that relative importance of different pests can be ascertained.

3.

For establishing economic threshold and economic injury levels.

4.

For evaluating crop varieties for resistance to insect-pests.

Oerke (2006) highlighted that the productivity of crops grown for human consumption is at risk

due to the incidence of pests, especially weeds, pathogens and animal pests and they may be prevented, or reduced, by crop protection measures. Estimates on potential and actual losses despite the current crop protection practices are given for wheat, rice, maize, potatoes, soybeans, and cotton for the period 2001–03 on a regional basis (19 regions) as well as for the global total. Among crops, the total global potential loss due to pests varied from about 50% in wheat to more than 80% in cotton production. The responses are estimated as losses of 26–29% for soybean, wheat and cotton, and 31, 37 and 40% for maize, rice and potatoes, respectively. Overall, weeds produced the highest potential loss (34%), with animal pests and pathogens being less important (losses of 18 and 16%). The efficacy of crop protection was higher in cash crops than in food crops. Weed control can be managed mechanically or chemically, therefore worldwide efficacy was considerably higher than for the control of animal pests or diseases, which rely heavily on synthetic chemicals.

Despite a clear increase in pesticide use, crop losses have not significantly decreased during the last 40 years. However, pesticide use has enabled farmers to modify production systems and to increase crop productivity without sustaining the higher losses likely to occur from an increased susceptibility to the damaging effect of pests. The concept of integrated pest/crop management

14

includes a threshold concept for the application of pest control measures and reduction in the amount/frequency of pesticides applied to an economically and ecologically acceptable level. Often minor crop losses are economically acceptable; however, an increase in crop productivity without adequate crop protection does not make sense, because an increase in attainable yields is often associated with an increased vulnerability to damage inflicted by pests.

Dhaliwal et al. (2010) had stressed upon that insect pest problems in agriculture have shown a considerable shift during first decade of twenty-first century due to ecosystem and technological changes. While there has been an overall decline in the severity of Helicoverpa armigera (Hubner), the incidence of several other insect pests like mealy bugs, particularly Phenacoccus solenopsis Tinsley on cotton; sugarcane woolly aphid, Ceratovacuna lanigera Zehntner on sugarcane; and tobacco caterpillar, Spodoptera litura (Fabricius), on several crops, has shown an increasing trend.

The diamondback moth, Plutella xylostella (Linnaeus), has consistently remained the most destructive pest of cruciferous vegetables. The global losses due to insect pests have declined from 13.6 per cent in post-green revolution era to 10.8 per cent towards the beginning of this century. In India, the crop losses have declined from 23.3 per cent in post-green revolution era to 17.5 per cent at present. In terms of monetary value, the Indian agriculture currently suffers an annual loss of about Rs 8,63,884 million due to insect pests.

A systematic approach to diagnosing plant damage Precise diagnosis must be made before corrective action can be taken. Probability of correct diagnosis based on only one or two clues or symptoms is low. Similarities of symptoms produced on the same plant by completely different factors frequently make the use of symptoms alone inadequate.

Factors causing plant damage can be grouped into two major categories: Living factors: Living organisms such as pathogens (fungi, bacteria, viruses, nematodes) and pests (insects, mites, molluscs, rodents..etc.).

Non-living factors: Mechanical factors (i.e. breakage, abrasions, etc); physical, environmental factors (extremes of temperature, light, moisture, oxygen, lightning); and, chemical factors (chemical phytotoxicities, nutritional disorders, etc). i.

Define the problem

A.

Plant identification and characteristics Establish what the “normal” plant would look like at this time of year. Describe the

“abnormality”: Symptoms & Signs.

B.

Examine the entire plant and its community Determine the primary problem and part of the plant where initial damage occurred.

ii.

Look for patterns: On more than one plant? On more than one plant species?

A.

Non-uniform damage pattern (Scattered damage on one or only a few plant species) is indicative of living factors (pathogens, insects, etc.).

15

B.

Uniform damage pattern over a large area (i.e. damage patterns on several plant species) and uniform pattern on the individual plant and plant parts indicates nonliving factors (mechanical, physical, or chemical factors).

iii. Delineate time-development of damage pattern A.

Progressive spread of the damage on a plant, onto other plants, or over an area with time indicates damage caused by living organisms.

B.

Damage occurs, does not spread to other plants or parts’’ of the affected plant.’

Clear line of demarcation between damaged and undamaged tissues. These clues indicate

nonliving damaging factors. iv.

Determine causes of the plant damage

A.

Distinguish among living factors

B.

1.

Symptoms and signs of Pathogens

2.

Symptoms and signs of insects, mites, and other animals

Distinguish among non-living factors 1.

Mechanical Factors

2.

Physical Factors

3.

a.

Temperature Extremes

b.

Light Extremes

c.

Oxygen and Moisture Extremes

Chemical Factors a.

Analyze damage patterns in fields and other plantings.

b.

Injury patterns on individual plants.

c.

Pesticide-pollutant phytotoxicities – damage patterns.

d.

Nutritional disorders -key to nutritional disorders.

If we suspect that it is a living damaging factor, we will look for signs and symptoms to distinguish between pathogens and insects. If the accumulated evidence suggests that it is a pathogen, we will seek evidence to distinguish among fungal, bacterial, viral pathogens and nematodes. If the evidence indicates the damaging factor is an insect or other animal, we will seek further evidence to distinguish between sucking and chewing types.

If evidence indicates that the damage is being caused by a non-living factor, we will seek further evidence as to whether the initial damage is occurring in the root or aerial environment. We will then attempt to determine if the damage results from Mechanical factors, from extremes in physical factors (i.e. environmental factors such as extremes of temperature, light, moisture, oxygen), or from chemical factors (i.e. phytotoxic chemicals or nutritional disorders). Once we have identified the plant and limited the range of probable causes of the damage, we can obtain further information to confirm our diagnosis from reference books, specialists such as plant pathologists, entomologists, horticulturists, and/or laboratory analyses.

16

Crop Losses In simple term it is a measure of reduction in either quantity and/or quality of yield. It may be taken as the difference between the actual Yield and the attainable yield.

Definition of loss due to pests Crop loss implies yield reduction which may be expressed as the percentage of reduction in potential yield in the absence of pests ‘m’. If yield in the presence of pests is ‘y’, then (m - y) × 100 m Yield loss (w ) =................................. m Estimation of crop losses caused by insects to economic crops are exceedingly difficult because, 1.

They are variable in nature of damage and, 2. Insect population fluctuates both in time and space.

The nature of damage caused by insect pests of crop plants is a function of pest population. So it is mostly insect capacity to increase in number rather than the nature of damage.

There are four general types of insect damage which contribute to yield loss: 1.

Indirect Damage- Pest feeds on non-marketable portion of plant, causing yield loss. (i.e. aphids on stems and leaves, root and seed maggot)

2.

Direct Damage- Pest feeds on marketable portion of plant, causing primarily quality loss. (i.e. earworm in sweet corn ears, cabbage worms)

3.

Damage by Vector Diseases- Insect transmits organism that causes plant disease, causing yield and quality losses. (i.e. cucumber beetles/bacterial wilt, flea beetles/Stewart’s wilt)

4.

Damage caused by Contamination - Presence of insects, insect parts, or insect products makes the product less marketable, and therefore less valuable, or completely unmarketable. (i.e. heads of cauliflower covered in aphids)

The following four points should be kept in view to estimate the losses.

1.

Any insect which cause some kind of the damage to crop can become pest when its population increase above a critical level. The critical level depends upon the nature of the damage caused by the insect. Example: In case of leaf feeders, the leaf eaten is near index of the losses caused by caterpillars while in case of insect vectors of virus of disease a very small population of infective individuals can spread the disease to whole crop.

2.

The losses caused vary both in time and space from 0 to 100 per cent. The estimation is fairly easy at these two extremes, but there are large numbers of factors which tend to invalidate any estimation in between these extreme limits.

3.

The loss may be either quantitative or qualitative. In case of quantitative loss, reduced yield is observed, whereas, in qualitative loss, quality may be affected. Example: In case of wheat bug (Eurygastor integriceps) is known to affect adversely the baking quality of wheat.

4.

Insect losses in terms of money are also objected. The selling price of the commodity would be reduced, if insect infestation will be of greater extent.

17

Methods of estimation of losses: The measures generally followed for estimating the losses caused by insect pests are based on either growing a crop, free from insect infestation as possible and then comparing its yield with that of check crop in which insect activity has been normal, or by making use of differential infestation and comparing the yield. The above ones are used in the following methods for estimating the crop losses.

The methods are as follows-

1.

1.

Mechanical protection of crop from insect pest damage

2.

Chemical protection of the crop

3.

Comparison of yields in different fields having different degrees of pest infestation

4.

Comparison of average yield of healthy plant with that of infested plants

5.

The average amount of damage caused by individual insect

6.

Manipulation of natural enemies

7.

Simulated damage

Mechanical protection of crop from insect pest damage: The crop is grown under the enclosures of wire gauze or cotton cloth. These enclosures keep the pest away from the crop.

Then, the yield of crop under such enclosures is compared with the yield obtained from the infested crop under similar conditions. This technique has been used with that various modifications for estimating the losses caused by leafhoppers and whitefly to cotton.

Flaw: a.

The limitation in the case of enclosures is that the plants generally become pale and weak due to changes in micro environment.

b.

This technique cannot be adopted on an extensive scale because it is very time consuming and impracticable on a field scale.

2.

Chemical protection of the crop / Paired Treatment Experiments: The crop is protected from pest damage by best scheduled chemical recommendation of pesticides. Then, the yield of treated crop is compared with that subjected to normal insect infestation. This technique has been very widely used and it can be adopted on a large scale in cultivator’s field.

Flaw: The crop treated with the chemical protection can also be physiologically affected for better or worse because of the effect of the chemical protectant.

3.

Comparison of yields in different fields having different degrees of pest infestation: The yield is determined per unit area in different fields having different degrees of pest infestation.

The correlation between the yield and degrees of infestation is worked out to estimate the loss in yield.

Flaw: The yield in different fields can also be influenced by the soil heterogeneity

18

4.

Comparison of the average yield of healthy plants with that of attacked plants: In this process individual plants from the same field are examined for the pest incidence and their yield is determined individually. The loss in yield is estimated by comparing the average yield of healthy plants with that of plants showing different degrees of infestation. The same data can also used for working out the correlation between the yield and infestation on the basis of infested individual plants. Advantage: The advantage of this technique over the above method is soil heterogeneity factor is considerably reduced

Flaw: a.

The different plants showing various degrees of infestation in itself is a proof that plants differ from one another in some unknown factors due to which they carried different degrees of infestation. This factor may be genetic or physiological or it may be mere soil heterogeneity in the same field.

b.

It is a time consuming and involves lot of labor.

5.

The average amount of damage caused by individual insect: For this method, the preliminary

information is obtained from studies on biology of the pest species. The details regarding the amount of damage caused by different stages or stages of the insect, and the exact nature and amount of loss caused are then worked out. Example: It has been estimated in the case of grasshopper. It consumes on average 42 grams of green leaves of maize during its life time. It was estimated that this insect would cause 18% loss in yield of maize at a population level of 10 grasshoppers per square yard.

Flaw: It is very difficult to use this technique over large area and it is time consuming.

6.

Manipulation of Natural Enemies: The manipulation of natural enemies of a pest species offers a means of evaluating plant damage. This technique has not been widely used. The pest is controlled by introducing predators or parasites into the field and the yield of such crop is compared that on which no such pest control measures have undertaken. This method is feasible only in small plots and is not practicable on field.

7.

Simulated damage: Many investigators have attempted to simulate pest injury by removing or injuring leaves or other parts of the plant. The simulated damage may not always be equivalent to the damage caused by an insect. Insects may persist over a period of time or inject long acting toxins rather than producing their injury. Feeding on margins of leaf may not be equivalent to tissue removal from the centre of the leaves. Insect feeding is usually extended over a period of time and is difficult to incorporate the concept of rate of injury.

Procedure: Loss estimation for the pod borer In order to assess the losses caused by insect pests of green gram the paired plot experiment is to be adopted. The method involves growing the crop in 26 plots, each measuring preferably 6m Ă— 3m.

Each plot should be separated by a buffer strip of one meter all around. One set of plots has to be kept protected from insect infestation by regular need-based application of recommended insecticides. The other set of plots has to be exposed to natural infestation and thus called unprotected. The unprotected

19

plots were allowed for a natural infestation of pests. About 5-10 randomly selected plants from each variety were taken for the pod damage per replication and pod damage percentage for each variety was calculated from protected and unprotected plots. Data of yield per plot were recorded and converted into yield per hectare basis from protected and unprotected plots and the avoidable losses was calculated. The avoidable yield loss due to pest was worked out by using following formula of Pradhan: X1 -X 2 Loss in yield (%) = ----------------------- Ă—100 X1 Where, X1 = Yield in treated plot =21.14 qt./ha X2 = Yield in untreated plot = 16.88 qt/ha 21.14 -16.88 The avoidable loss in yield =................................. x 100 =20.15 % 21.14

20

Chapter-5

Field diagnosis of damage caused by different types of insect pests Background: 

Insect and non-insect pests cause a particular type of damage to the plant parts often characteristic to particular pests



The pest mostly insect not present on the site of damage makes it difficult to know the casual organism.



Sometimes, the symptoms of damage caused by insects may closely resemble to those resulted due to pathogens or due to nutritional disorders.



So, practical experience makes familiar about the correct diagnosis on the basis of visual symptoms of damage to take appropriate control measures

Steps in Insect pest diagnosis in the field 1:

Insect- pests

A.

ROOT AND TUBER DAMAGE

(a) Root Damage 1.

2.

Wilting and drying of plants in patches: 

Leaves of affected plants turn pale, droop down and ultimately wither off.



Cut end of affected stem of collapsed plant swells, a characteristic diagnostic symptom.



Examples: Root grub, Holotrichia spp, Anomala spp.

Tillering is poor, affected plant turns yellow and stunted 

3.

4.

5.

Tunnels on pseudo-stem and plants break down at tunneled portion 

Plants bear few fruits and suckers



Circular holes with black rotten tissue of rhizome plugged with excreta



Example: Banana Rhizome weevil

Wilting and death of plants in Sorghum 

Damaged tubers in potato



Example: Termites

Wilting and Drying of plants and presence of large number of ants at the base of ragi tillers 

6.

Example: Rice root weevil

Example: Ragi root aphid

Root knots 

The tomato roots show knots



Example: Root knot nematode, Meloidogyne incognita

21

(b) Tuber Damage 1.

Potato tubers: with holes and tunnels inside Example: Potato tuber moth, Pthormia operculella and poatato white grubs

2.

Sweet potato tuber: with holes Example: Sweet potato weevil, Cylas formicarius

B.

Tree Damage: 

Yellowing of trees, withering of leaves, drying of twigs or complete drying of tree.

Sometimes gummy material oozes from the affected portion on the tree trunk



Examples: Tree borers of mango, cashew, coconut red palm weevil

C.

Bark Damage



Galleries of frassy web on the stem and near bark/angles of branches, Silken ribbon plastered on stem



Examples: Bark eating caterpillars of citrus, mango, guava, casuarina and jack

D.

Stem Damage

1.

Damaged part is cut off from the main plant and affected part wilts, dries up and exhibits symptoms like dead heart/white ear/bunchy top. Examples: Stem borers of paddy, millets, sugarcane and brinjal etc.

2. A) Bored hole on stem at or just below the ground level, Mature and developing pods are filled with mud in Groundnut B)

Earthen sheeting at the base of plant, mud filled galleries in shoots, drying of shoots in Sugarcane

C)

Mud galleries on tree trunk, if earthen sheet is removed eaten bark of trees is visible, Death of young plants and dry up in Mango

D) Mud galleries on tree trunk, bark and stem are eaten below the mud galleries, Nursery and transplanted fields show wilting of central shoot and stunted growth in coconut Example: Termites, Odontotermes obesus 3.

Ringing and girdling of stem bark

Example: Grape vine stem girdler, Sthenias grisator Amaranthus stem weevil, Hypolixus truncatulus Coccinia gall fly, Neolasioptera cephalandrae 4.

Galls on stem:

Examples: Tobacco stem borer, cotton stem weevil E.

Shoot Damage:

1.

Wilting, drooping of terminal plant part which later dries up

Examples: Shoot borers of brinjal, bhendi, cotton, castor, shoot fly of sorghum and black gram stem fly.

22

F.

Leaf Damage

1.

Mines in leaf: Insect larval stages cause different types of mines infesting between epidermal layers

Serpentine mines: Thread like Example: American serpentine leaf miner, Liriomyza trifoli Linear or serpentine mines: Thread like Example: Citrus leaf miner, Phyllocnistis citrella Blotch mines: Blight Example: Patch like Blotch leaf miner, Acrocercops syngramma 2.

Punctures on leaf: Leaves show punctures made by insects for feeding or oviposition Example: Bean flies, Ophiomyia phaseoli

3.

Galls on leaf: Due to insect feeding on the leaves, the plant reaction results in gall formation like warts, etc. Example: Mango gallfly, Procantarina mattesiana

4.

Out growths on leaves: The damage makes the plant to produce felt lie out growths Example: Jasmine mite, Aceria jasmini

5.

Shelter feeding i.

Leaf rolling/folding: Single leaf is folded to create shelter for feeding or pupation Example: Rice case worm, Nymphula depunctalis

ii.

Leaf webbing: Few leaves are webbed together and the larva feeds in between the leaves by scraping Example: Pumpkin caterpillar, Diaphania indica

iii. Terminal leaf webbing: The terminal tender leaves are webbed together and larvae and pupal stages occur in the webbed leaves. Larvae feed by scraping the surface. Example: Sapota leaf webber, Nephopteryx eugraphilla 6.

Leaf feeding

i.

Scraping: Early instar larvae feed by scraping the surface of leaves leaving papery patches.

Example: Larvae of Spodoptera litura

Papery windows: Ladder like papery windows caused by the feeding of adult beetles. Example: Brinjal Epilachna beetle or Hadda beetle, Henosepilachna vigintioctopunctata ii.

Holes

Small round holes result due to the feeding of small adult beetles on leaves Example: White spotted flea beetle, Monolepta signata Medium round symmetrical holes on leaves by the larval stages Example: Peanut defoliator, Helicoverpa armigera Irregular bigger holes on the leaves caused by the larval stages of many lepidopteran insects Examples: Slug caterpillar, Latoia lepida, Hairy caterpillar, Euproctis fraterna iii. Skeleenization of leaves by completely feeding on leaf lamina, leaving only the veins by larval stages Example: Castor semilooper, Achoea janata iv.

Free feeding. Completely the lamina and veins eaten away by the voracious feeder or in severe cases by the larvae Example: Lemon butterfly, Papilio demoleus.

23

v.

Nibbling: The leaves from margins in to small U shaped cuttings is done by adult beetles Example: Grey weevil, Myllocerus viridanus

vi.

Cutting the leaves in to big semi circular shape is done by adult bees Example: Leaf cutting bee, Megachile anthracina

vii. Crinkled leaves are due to sucking of cell sap by nymphs and adult insects Example: Aphids viii. Cupping /curling of leaves with result due to sucking of insects Example: Thrips G.

Fruit Damage

1.

Spots on outer surface indicate the oviposition punctures of insects on fruits Example: Mango fruit fly, Bactocera dorsalis, Ber fruit fly, Carpomyia vesuviana and cucurbit fruit fly Bactocera cucurbitae

2.

Corky layer on fruit surface is caused by laceration of cells Example: Grape vine thrips, Rhipiphorothrips cruentatus

3.

Bore holes plugged with excreta on the fruits are done by larvae Example: Bhendi shoot and fruit borer, Earias vittella

4.

Bore holes without excreta and sometimes, larva is seen outside Example: Tomato fruit borer, Helicoverpa armigera

H.

SEED DAMAGE: (STORED GRAIN PESTS)

1.

Webbed food items/stored seeds Example: Rice weevil, red rust flour beetle, rice moth

I.

GRAIN DAMAGE:

1.

Shriveled or chaffy grains during reproductive stage of the crop Examples: Rice gundhi bug, sorghum ear head bug, sorghum midge

24

J.

DIAGNOSIS BY LOCATING SIGNS ON/NEAR PLANTS

1.

Diagnosis by locating signs: Black ants, Componotus compressus movement on plant parts

2.

Honey dew secretion on plant parts:

3.

Sooty mould development on plant parts: All the above indicate the presence of Homopteran insects Example: Mealy bug, Planococcus lilacenus; Aphid, Aphis craccivora; Whitefly, Bemisia tabaci

4.

Excreta of Insects:



Black spots are found due to drying of excreta on leaves by thrips



Presence of excreta indicate usually the lepidopteran larvae

5.

Scale covering: The stem, leaves etc are covered heavily by the scale covering

6.

Exuviae of insects: 

Molted skins of leafhoppers, aphids etc present usually on lower surface of leaves



BPH molted skins at base of plant and floating in water

The Process of diagnosis of Pest Problems The field diagnosis of damage caused by different insect pests can be done on two broad basis i.e., A) on the basis of mouth parts and B)

on the basis of damage symptoms produced on different plant parts.

A) On the basis of mouth parts 1.

Chewing and Biting type: Most of the caterpillars (order Lepidoptera) and beetles (order Coleoptera) cause damage to the plants with the help of their biting & chewing type mouthparts.

Other chewing insects that are damaging to plants include grasshoppers, katydids, (order Orthoptera) termites (order Isoptera), cockroaches, and their relatives (order Blattaria).

Damage symptoms caused by Insects with chewing mouthparts: 1.

Chewing insects feed by biting, ripping or tearing plant tissue. They may damage all or part of the plant including roots, stems, leaves, buds and open flowers. Chewing insects produce varied plant damage including:

a)

Irregular holes in foliage or stems (shot holes / borer holes)

b)

Complete defoliation

c)

Leaves with "windowpanes" or leaf skeletonized, i.e., showing bared veins (Spodoptera, DBM and cabbage butterfly)

d)

Irregular margins of leaves or rugged appearance (Grasshoppers)

e)

Leaf mining (leaf miners)

f)

Discolored areas on the surface or margins of leaves or petals (thrips, psylla)

g)

Severed stems, leaves or buds or wilting of stems or canes (limb girdling)

h)

Wilting of plants (from root damage by white grubs or other root feeders/borers)

i)

Circular to semicircular holes in leaves (e.g., leaf-cutting bees)

25

j)

Caterpillars use their chewing mouthparts to consume several times their own weight in plant tissue over the course of their development. Much fibrous tissue passes through the caterpillar gut undigested and forms a major component of the large fecal pellets caterpillars leave behind. These pellets are a characteristic sign of caterpillar damage.

k)

Beneficial insects with mandibulate mouthparts can often keep pace with pest populations, aids in maintaining a more natural balance in our ecosystem

2)

Piercing & Sucking type of mouth parts: Many insects feed on a liquid diet (i.e., plant fluids)and have a beak, referred to as a proboscis

that is modified to suck up liquids. Sucking insects insert their beak (proboscis) into the tissues of leaves, twigs, branches, flowers, or fruit and then feed on the plant’s juices. The proboscis has a groove down its front inside which contains several very slender, sharp, and needle-like stylets that can pierce plant tissue to extract plant fluids.

Some examples of sucking insects are aphids, mealy bugs, true bugs, scale insects, white flies and leafhoppers. Other insects such as mosquitoes and sucking lice also consist of piercing & sucking type of mouth parts.

Damage symptoms caused by Insects with Sucking / Piercing Mouthparts: 2.

Insects with Sucking/Piercing Mouthparts produce varied plant damage including: a)

Discoloration (yellow or brown), mottled or necrotic (dead) spots on leaves or petals

b)

Wilted appearance of plant or plant parts

c)

Drooping of wilted parts.

d)

Curled, malformed leaves and petals

e)

Leaf spots (stippling)

f)

Many of insects pests that feed and secrets honeydew which is favourable for the growth of a black or sooty mold. Sooty mould reduces the photosynthetic area of plants.

g)

Reduces the vitality and vigour in affected plant.

h)

Many of these insects have ability to transmit the plant viruses, and human diseases.

i)

The mandible and maxillae are modified to form slender bristle like stylets which rest in the grooved labium.

j)

The stylets are pierced inside the plant tissues and generally they suck plant sap from phloem vessels.

3)

Rasping & Sucking / Lacerating & Sucking: Insects with rasping/sucking mouthparts actually rasps or scrapes the surface of plant tissue (such

as leaves or petals) and sucks up the fluids that ooze from the damaged area of tissue. Examples of pests with rasping-sucking mouthparts include thrips.

Damage symptoms caused by Insects with Rasping & Sucking / Lacerating & Sucking mouthparts are3.

Thrips prefer to feed on succulent plant tissues. They may feed on fully expanded foliage, open flowers, and even pollen grains. Light-colored flowers (white, yellow or other pale colors)

26

are often preferred. Affected tissue dies, turns brown and tears easily, a situation especially noticeable on the edges of pastel-colored rose petals. Leaves that are attacked become bleached and dry. Skin of damaged fruit appears sanded and the underlying tissues may be off-flavored, hard and/or dry.

B)

On the basis of damage symptoms produced on different plant parts Insects which are phytophagous can cause injury and damage to plants by defoliating, or sucking

their sap, insects can retard plant growth. Almost all the parts of the plant are being affected by various insect pests, and in most cases, the species of insect damaging different parts are always unique to the plant species. Based on the insect species affecting parts of the plant, which is usually typical, and the knowledge of the symptoms appearing are very important to understand, as the pest may sometimes disappear from the site of damage. Hence, we need to understand the Symptomology, so as to identify the pest, assessing the pest damage and also to recommend the proper management practices. The following classification of insect pests based on the damage with special reference to the parts they damage, is important to understand: 1.

Root feeding insects: Insect larvae feed on roots, root nodules; nymphs and adults suck sap from roots, resulting in stunted growth, poor tillering, drying of plants in isolated patches etc. White grubs, grubs of rhinoceros beetles, termites, rice root weevil & ragi root aphid

2.

Stem borers: Larvae enter into the shoot of main stem, tillers and feed on the central growing point. As a result, nutrient supply from the main plant beyond the infested part is affected leading to withering, wilting and drying up exhibiting symptoms such as dead heart / white ear / over growths of bunchy top etc. Stem borers of paddy, millets, sugarcane, brinjal, bhendi, cotton etc.

3.

Shoot borers: Larvae attack tender shoots and bore inside during the vegetative stage of crop growth and cause wilting, dropping of terminal plant part which later dries up. Shoot fly of sorghum, early stem borer in sugarcane, stem fly in black gram/ French bean, soybean, shoot borers of brinjal, bhendi, cotton, castor etc.

4.

Tree borers: Larvae bore deep into the tree trunk, tunnels in zigzag manner and feed on inner tissues, affecting nutrient and translocation of sap to upper portions of branches / tree exhibiting symptoms such as withering of leaves, drying of twigs or complete dying of tree. Presence of fresh powdered material, ooze of gummy exudations etc. from the affected portion on the tree trunk is also seen in some cases. Tree borers of mango, cashew; coconut red palm weevil etc.

5.

Bark borers: Larvae enter into the bark and tunnel into the branches. The larvae remain hiding in the galleries formed from floss / fecal matter and silken saliva on the stem and continue to scrape the bark. Larval feeding results in drying of branches and breaking of affected portion with wind or gale. Bark eating caterpillars of citrus, mango, guava, casuarinas, jack etc.

6.

Gall formers: Larvae/nymphs feeding inside the stem/ tiller /leaf/ flower bud affect the tissue by nibbling the meristamitic tissues and secretion of auxins that results in excessive growth of cells at the affected portion leading to distorted growth and malformation of plant parts known as 'Gall'.

27

Examples-Paddy gall midge, chilly midge, gingelly (Sesamum) midge, cucurbit stem borer, mango malformations, tobacco stem borer, cotton stem weevil, mango inflorescence midge, chilli midge etc.

7.

Leaf folders: Larvae fasten the margins of individual leaves from margins / fold longitudinally or roll leaves into bell shape and feeds within by scrapping the chlorophyll. Rice/ maize/leaf folder, cotton leaf roller, red gram / black gram leaf folder.

8.

Leaf miners: Larvae fasten the leaves /leaflets by means of silken threads (derived from saliva) and scrape the chlorophyll content by remaining within the web. Fecal pellets / frass remain present in the web. Leaf Webbers on groundnut / gingelly, Webbers of mango / sapota /Cashew.

9.

Leaf Webbers: Larvae fasten the leaves /leaflets by means of silken threads (derived from saliva) and scrape the chlorophyll content by remaining within the web. Fecal pellets / frass remain present in the web. Leaf Webbers on groundnut / gingelly, Webbers of mango / sapota /Cashew.

10. Defoliators / Skeletonizers: Larvae feed on the leaves completely leaving only midrib / veins or scrape the chlorophyll content of leaves or cause numerous holes. Castor Semilooper, red hairy caterpillar, Bihar hairy caterpillar, snake gourd Semilooper, ash weevils, tobacco caterpillar, brinjal Epilachna beetle.

11. Pod / Capsule borers: During the reproductive stage of crop, larvae bore into the flowers, pods, capsules and fasten the adjacent plant parts with silken threads, fross and excreta and feed on the internal contents within the web. Spotted pod borer in legumes, capsule borers of castor / gingelly; pod borer complex in pulses, gram caterpillar, pink bollworm, tobacco caterpillar, chilly pod borer etc.

12. Fruit borers / Bollworms: Larvae enter into the tender fruits bolls and feed on internal content /pulp and plug the larval burrow with excreta. Fruit borer of brinjal / bhendi or okra /tomato, mango fruit borer, fruit fly, mango stone weevil, cashew apple and nut borer, anar/ guava fruit borer, Cotton bollworm, etc.

13. Seed feeding insects / stored grain pests: Larvae feed on stored seeds either as internal/ external feeders / by webbing the food particles. Rice weevil, lesser grain borer, red rust flour beetle, rice moth, cigarette beetle, saw toothed beetle etc.

14. Sap sucking insects / feeders: (a) From tender plant parts: Nymphs and adults suck sap from the base of the plant/leaves / tender terminal plant parts and affect the vigor and growth of the plants. Different insects exhibit different symptoms. Most of the sap suckers suck sap in excess of their requirement and excrete honey dew, which is rich in sugars, a source for sooty mold development. Aphids, leafhoppers (jassids), plant hoppers, white flies etc on important crops. (b) From grains: Nymphs and adults suck juice from developing ovaries/milky grains resulting in the formation of shriveled /chaffy grains. Rice gundhi bug, Sorghum ear-head bug, Sorghum midge.

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Damage caused by insects to plant parts: seedlings Crop(s)

Insect(s)

Sci. Name of Insect

Symptom(s)

Rice

Rice grasshopper

Hieroglyphus banian

Irregular cuttings at leaf margins

Rice

Army worm

Spodoptera mauritia Spodoptera exigua

Defoliation

Sorghum

Red hairy caterpillar

Amsacta albistriga

Defoliation. Complete eating of plants

Brinjal

Hadda Beetle

Henosepilachna dodecastigma

Skeletonized patches on leaves forming ladder like

Cutworms

Agrotis spp.

Cut the seedling roots

Diamond Backed Moth

Plutella xylostella

Leaf mines, skeletonization

Cabbage, cauliflower

of leaves

Damage caused by insects to plant parts: roots & tubers Crop(s)

Insect(s)

Sci. Name of Insect

Symptom(s)

Wheat

Ghujia weevil

Tanymecus indicus

Grubs feed on root.

Rice

Rice root weevil

Echinocnemus oryzae

Stunted growth, yellowing, poor tillering.

Sorghum

White grub

Holotrichia consanguinea

Plant become pale and wilted appearance and ultimately dry up

Ragi

Root aphid

Tetraneura nigriabdominalis

Plants turn yellow and setting of seeds is reduced

Groundnut

White grub

Holotrichia serrata

Effected plants can be easily pulled out.Clumps dry up

Sugarcane

Termites

Odontotermes obesus

Wilting and drying of all stages of crop

Banana

Rhizome weevil

Cosmopolites scrdidus

Dull yellow green and floppy foliage, tunnels on pseudo-stem, plant easily blown down to mild winds

Cucumber

Striped cucumber beetle

Acalymma vittatum

Adult attack tender stem & leaves. Larva attack roots Adult attack tender stem & leaves. Larva attack roots Adult attack tender stem & leaves. Adult attack roots.

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Damage caused by insects to plant parts: stem Crop(s) Rice

Insect(s) Yellow stem borer Sorghum stem borer

Sci. Name of Insect Scirpophaga incertulas

Symptom(s) Dead heart, white ears

Chilo partellus

Ragi pink stem borer Early sugarcane borer

Sesamia inferens

Sugarcane

top shoot borer

Scriphophaga nivella

Sugarcane

Inter-node borer

Cotton

Stem weevil

Chilo sacchariphagus indicus Pempherulus affinis

Mesta

Stem weevil

Alcidodes affaber

Jute

Stem weevil

Apion carchori

Red gram

Stem fly (or) Bean fly Shoot & capsule borer

Ophiomyia phaseoli

Leucinodes orbonalis

Mango Mango

Shoot and fruit borer Stem borer Shoot and fruit borer Stem borer Stem borer

Numerous shot holes on leaves, dead heart, entrance and exit holes at base of stem Oblong holes in unfolded leaves.

Dead heart Entrance hole at ground level.

Dead hearth with offensive smell, which can be pulled out easily.

Shot holes, bunchy top appearance of side shoots dead hearth which cannot pulled out Tissue turned red color, holes plugged with excreta Gall like swellings on stem, mortality of plants Gall like swelling on stem, fross at hole Shoots formed on stem, and branching takes place Swellings at ground level on stem and cracks open fross matter at bored shoots, webbed condition of seedcapsule covered with dark excreta Affected shoots wither and holes on stems Complete withering of plant Flaring up of shoots, fruits

Potato

Cutworm

Agrotis epsilon

Sorghum

Sorghum Sugarcane

Castor

Brinjal Brinjal Bhendi

Chilo infuscatellus

Dichocrosis punctiferalis

Euzophera perticella Earias vitella Bactrocera rufomaculata Chlumetia transversa

Branches collapse Atta c ked shoots clipped off Attacked shoot clipped

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Damage caused by insects to plant parts: leaves Crop(s) Rice Rice Sorghum

Insect(s) Rice grasshopper Army worm Red hairy caterpillar Tobacco caterpillar Hairy caterpillar

Sci. Name of Insect Hieroglyphus banian Spodoptera mauritia Amsacta albistriga

Euproctis similis Spilosoma obliqua Acherontia styx

Defoliation

Brinjal

Hairy caterpillar Bihar hairy caterpillar Hawk moth (or) sphinx moth Hadda Beetle

Symptom(s) Irregular cuttings at leaf margins Defoliation Defoliation. Complete eating of plants Irregular holes on leaves,bolls skeletonized leaves Defoliation of leaves,damage to pods Defoliation of leaves Defoliation

Brinjal

Leaf webber

Henosepilachna dodecastigma Psara bipunctalis

Cabbage Cabbage & Cauliflower Mango Groundnut

Leaf webber Diamond Backed Moth Leaf webber Leaf miner

Crocidolomia binotalis Plutella xylostella

Castor

Liriomyza tritolii

Citrus

Serpentine leaf miner Serpentine leaf miner Serpentine leaf miner Leaf mine

Cotton

Leaf roller

Sylepta derogata

Sesamum Brinjal Cucumber

Antigastra catalunalis Antoba olivalea Acalymma vittatum

Cotton

Leaf webber Leaf roller Striped Cucumber beetle Aphids

Skeletonized patches on leaves forming ladder like windows Caterpillars feed on epidermal tissues Webbed Leaves on flowers and pods Leaf mines, skeletonization of leaves Skeletonized leaves Presence of small brown blotches on leaves field later “ burnt� from distance Serpentine shaped, white mines on leaves.

Serpentine shaped, white mines on leaves.

Serpentine shaped, white mines on leaves Mines on young leaves on sever case, mines turns to silver colour Presence of large number of leaf rolls Presence of large number of leaf rolls Presence of large number of leaf rolls Rolled up leaves and flowers Rolled up leaves Retardation of plant

Cotton Sesamum

Thrips Leaf and pod

Thrips tabaci Antigastra catalaunate

Cotton Sun hemp Mesta Castor Sesamum

Tomato Pumpkin

Spodoptera litura Utethiecia pulchella

Orthaga exvinacea Aproaerena modicella

Liriomyza tritolii Liriomyza tritolii Phyllocnistis citrella

Aphis gossypii

Downward crushing of leaves sooty mould development Upward curling Rolled up leaves and

31

Pulses Mustard Bottle gourd Cotton

Cabbage

borer, blossom webber Leaf and pod borer, blossom webber Leaf aand pod borer, blossom webber G.Nut aphid/ Black fly Aphid Red pumpkin beetle Red spider mite

Cabbage borer or web worm

webbed flowers Rolled up leaves and webbed flowers

Aphis craccivora

Sooty mould development

Lipaphis erysimi Aulacophora foveicollis

Curling leaves, sooty mould Chewing large holes leaving only veins Large number of leaves turns red color, pin spots Large number of leaves turn red colour and pin spots Infested plant hillas (or) produce side shoots Infested plants produce side shoots

Tetranychus telaris

Hellula undalis

Damage caused by insects to plant parts: floral parts (flowers, buds): Crop(s) Insect(s) Sci. Name of Insect Symptom(s) Cotton Pink bollworm Rosette flowers. Premature Pectinophora gossypiella flower & bud drop Chili Blossom midge Maggots scrape and draw fluid Contarinia solani from flower buds Mango Inflorescence Small exit holes on the axis of Eroisomyla indica midge inflorescence, Drying and shedding of floral parts Sorghum Orange banded Presenting grain setting by Mylabris postulata blister beetle feeding on tender ear heads and flowers Presenting grain setting by feeding on tender ear head and flowers Bean, Soybean Green stink bug Premature abscission of flower Nezara viridula buds Sesamum Blossom webber Antigastra catalaunate Webbed flowers Moringa Moringa Shedding of flower buds Noorda morinage budworm Chilli Chilli blossom Discoloration and premature Contarinia solani midge blossom drop Citrus Black Aphid Feed on flower,Transmit tristeza Toxoptera citricida disease Rose Rose Thrips Rhipiphorothrips cruentalis Partial opening of flowers, brown discoloration of edges of petals

32

Rose Rose

Aphids Rose chaffer beetle Jasmine stink bug Jasmine bud borer Spotted bollworm American bollworm Pink bollworm Tobacco caterpillar Red cotton bug Gram pod borer

Macrosiphum rosae Oxcetonia versicolor

Plume moth spotted pod borer

Exelastis atomosa Maruca vibrate

Grams

Lentil pod bore

Etiella zinckenella

Sesamum Sunflower

Gall fly borer

Asphondylia sesame Heliothis armigera

Bhendi

Earias vitella

Bhendi Melons

Shoot and fruit bore Gram pod borer Melon fly

Onion Coriander

Head borer Coriander aphid

Heliothis arnigera Hydaphis coriandri

Pulses

Tobacco caterpillar Cutworm Shoot and fruit borer

Spodoptera litura

Jasmine Jasmine Cotton Cotton Cotton Cotton Cotton Red gram, bhendi Red gram Pulses

Pulses Brinjal

Hendecasis duplifascialsis

Dropping fading of flowers Irregular feeding marks on flowers Flowers damaged due to sap sucking Premature and drop

Earias vitella

Bolls are damaged

Heliothis armigera

Bolls are damaged

Pectinophora gossypiella Spodoptera litura

Stained lint irregular holes on bolls

Dysderlus cingulatus Heliothis armigera

Staining of lint by Bolls damaged,Round holes on pods Bud, flower and pod drop Webbed flower Webbed flower Dropping of flower buds and young pods Formation of galls on pods Circular large entry hole,white feeding hand inside rest body outside Flaring up of young fruits

Nezara vividula

Heliothis armigera Bactrocera cucurbitae

Spodoptera exigua Leucinodes orbonalis

Circular large entry hole Watery fluid oozing from punctured hole in fruits Cutting of pedicel of flower Devitalization of leaves and flower Irregular holes on leaves, pods, skeletonized leaves Holes on podes Affected shoots wither and droop, holes on fruits Affected shoots wither and droop, holes on fruits

33

Chapter-6

Procedure for collecting plant and insect Samples for problem diagnosis

M

ost plants are prone to attack by various insect pest and plant disease organisms. Pest outbreaks and diseases must be identified accurately to enable their efficient management.

Basic requirements for collecting, preserving and submitting plant, insect and disease samples (A) Collecting plant samples It is important to gather the best plant samples possible and to record all pertinent background information for the diagnostician. Following are general guidelines for collecting plant samples.

1.

Examine the entire plant for symptoms Plants may be affected by one or more pathogenic microorganisms or pests, although they may

also have an abiotic disease that does not involve a plant pathogen. Affected Plants often display a range of symptoms—visual signs of the infection or pest damage.

2.

Collect several plant specimens A single plant sample may not be enough to allow a correct diagnosis of the problem; several

plant samples showing the range of symptoms may be needed. If possible, select samples with various stages of disease development (early and late stages). but submitting excessive amounts of leaves or soil should be avoided.

How to collect data in the field?

Identifying the sampling sites (a) Tagging the site Mark sampling sites in the field whenever possible, even if you do not intend to return to the same site so that if a specimen or observation taken islost or destroyed, you would be able to revisit the site if needed. Remember to choose tags that will survive a variety of weather conditions, and use a pencil or ink that does not smear when wetted to label the tags. Options for marking the site include: 

Spray-painting a mark



Placing sticks with a bright tassel or tag, particularly where a pest has been completely Removed (such as weeds), but only when the stick or marker will not interfere with the



Management of the site, such as getting caught in harvesting equipment



Tying a tag or tassel to a plant stem or branch.

(b) Recording site details The location and unique identifying details of each site need to be recorded in a notebook. These details may be entered using a standard form that can be used for each site. Describing the sampling

34

site would include information such as a GPS reading, a unique number, distances from visual cues (e.g. 20 metres from roadside), number or nearest number of plant in a row (e.g. tenth tree in third row from the northeastern corner), or any distinguishing topographical features (e.g. edge of a ravine, in a ditch).

What data to record in the field The most important tool you will have with you in the field will be your notebook and notes. In your notes you would record any information that could otherwise be forgotten, such as the dates of surveying, the weather at the time, the site details, the names and contact details of the local people involved. Notebooks with carbon paper duplicate pages can be very useful when recording information to accompany a specimen taken. In this way, the details are written once only but you then have a permanent record in your notebook and a copy to be kept with the specimen.

Designing a form The simplest way to record data is to design a form that allows for recording all the information that you intend to collect. A simple way to save a lot of time is to work out ahead of the survey how the data will be stored and to design your form so that it is easy to transfer the information to the storage system. When designing a form, you could include the followings: 

Observer’s name



Field site number or name



Sampling site number or name



Targeted pest names—common and scientific



Time and date



Brief description of weather conditions



Locations, such as by GPS readings, of sampling sites



Description of habitat (e.g. aspect, vegetation, soil type)



Scale/population density categories that could be ticked



Symptoms of the pest or host



Pest life stage or state (e.g. larvae, pupae, adults for insects; anamorph/teleomorph state



for fungi; seedling, budding, senescent, first flush for plants)



Caste of colonial insects surveyed, such as of termites, ants and some wasps



Behavioural notes on possible vectors (e.g. ‘insect ovipositing on fruit’ or ‘insect restingon



plant leaf area or length of plot or transect assessed



Cross-reference to pest example in a pest photo library



Colour of identifying features, such as of flowers



Any quarantine measures applied at the field site, such as hygiene measures



Treatments applied to site



Additional comments

35

Units for data Data are normally reported in terms of a unit of measure, usually the number of pests per unit area. The number might be a direct count of the pests or could be a scale of intensity of the pest that is recorded. The area examined might be per tree, fruit, field, crop, kilometre, quadrat, sweep of a net, trap etc. For example: number of shoots attacked per plant, number of trees affected as compared with the total number of trees examined.

Use of scales and scores In some cases where the pest is numerous, or particularly for symptoms of plant pathogens, whole numbers of pests are not possible or useful. Instead, a scale of cover of the host or a standardised measure of the pest could be used. Scales are semi-quantitative as the scale intervals can be wide and may not be consistent in their range.

3.

Preserving plant samples After collecting the samples, do not expose them to direct sunlight. Keep them cool and do not

allow them to dry out or cook. Place samples in plastic bags in the shade or in a cooler until they are ready for delivery to the plant clinic. Leaves may be pressed between the pages of a book or magazine or wrapped in tissue.

(A) Collecting insect samples 

Collect whole insects in good condition.



Collect as many insect stages as possible: eggs, larvae, pupae, and adults.



Place the insects in 70 per cent isopropyl alcohol immediately. Keep moths and butterflies intact in small containers or wrapped with plastic or paper.



Spiders should be collected alive, dropped into hot (180 degrees F) water, and transferred to 70 per cent isopropyl alcohol after cooling.



If the insect was causing plant damage, include a plant specimen showing evidence of the plant injury.

Avoid touching insects with fingers Some insects can injure humans. Handling insects can also cause damage to their bodies that may prevent their identification.

Collect different life stages of the insect Sometimes insects cannot be properly identified unless a certain life stage is present. For example, adults may be needed for correct identification.

Collect multiple specimens Collect several specimens of the insect. Time of day matters. Many leaf-feeding insects (such as caterpillars) may hide from predators during daylight hours. It may be necessary to capture insects during twilight in the evening or early morning.

36

(B) Preserving insect specimens (i)

Most insects Termites, bugs, beetles, flies, wasps, ants, maggots, spiders, etc. should be immersed in isopropyl

(“rubbing�) alcohol, which kills and preserves them. (ii) Mites, scales, aphids, thrips Send these in alive on some of the affected foliage or stems, collected as you would a plant specimen. Place in a plastic bag when collected. refrigerate until sent. (iii) Butterflies and moths Kill the specimens by freezing, wrap lightly in tissue paper, and place in a crush-proof box.

Careful handling is required because the pattern of scale coloration is often used in identification. (iv) Caterpillars Send in alive on some of the host plant tissues in a plastic bag. Refrigerate until sent. (v) Grubs Send in alive in a pint or two of soil enclosed in a plastic bag. Refrigerate until sent.

(C) Packaging plant and insect samples It is important to package the samples properly to ensure they arrive in good condition at the plant clinic. Following are general guidelines for handling and packaging plant and insect samples.

Use plastic bags For most samples including leaves, stems and roots, use plastic bags to prevent plant samples from drying out during transport. However, fleshy fruits, vegetables, or tubers in stages of decay should be wrapped individually in dry newspaper.

Submit samples as soon as possible Decayed plant or insect samples are useless for an accurate disease diagnosis. Always plan to have samples arrive at the Centre within one or two days of their collection, if possible, or take steps to inhibit the deterioration or decay of samples (i.e., by refrigeration).

Representative, moderate symptoms Do not submit dead plants for diagnosis. Place roots and soil together in a Plastic bag and close it securely. Place several branches showing decline or dieback in a separate plastic bag. For smaller plants, submit an entire plant (confine the root ball in a plastic bag tied tightly to the stem). Place the entire plant in another plastic bag and close it securely. Be sure there is no water on the foliage surfaces (this causes deterioration during shipping).

General Packaging Guidelines 1.

Take your samples before applying pesticides; otherwise the ability to recover disease pathogens may be limited.

37

2.

Don’t add water or pack a sample that is wet or in wet paper

3.

After your samples are collected keep them refrigerated until submitted.

4.

Don’t mix samples in the same submission bag. Moisture from root samples will contribute to the decay of foliage if they are mixed together.

5.

Plant disease identification procedures do not utilize soil. Excess soil can be hand shaken from root systems.

6.

Please mark sample packages with a “Warning” if there are thorns or spines

7.

All samples must be accompanied with a completed “Plant Disease Diagnostic Form.”

8.

Note recent pesticide history on the form accompanying the sample

9.

Samples arriving from sites that are two days or less mailing time from a clinic can be sealed in plastic bags for shipping

10. Samples arriving from distances greater than two days mailing time from a clinic should be packed tightly in a box with dry paper.

11. Mail samples early in the week to avoid the weekend layover in the post office.

12. For emergency samples or anything you suspect might be a dangerous exotic, use overnight courier services or overnight mail.

Plant and Insect Sample Submissions 

Try to collect several specimens in different stages of development. Some identification keys we use are for adults, while other are for immature bugs.



Insects submitted whole are more useful than when submitted in segments.

Packing Insects 

Insects should be killed before shipping. Live caterpillars often pupate during shipment and beetles may eat their way out of the shipping container.



Send all mature and immature insects (except butterflies and moths) in a glass vial or bottle containing ethyl or isopropyl (rubbing) alcohol.



The vial or bottle must be properly padded in a mailing tube or other container to prevent breaking. Make sure that the cap for the vial is well secured so the alcohol doesn’t leak from within the vial during hipping.



Send butterflies or moths dry in pill boxes or a similar container with tissue paper to prevent the specimen from being broken.



It is often easier to identify an insect by seeing the damage it is doing to foliage, twig, fruit or other plant parts.



If foliage or tender twigs are sent, they should be placed in a plastic bag and sealed. During the summer months, add a paper towel with the plant material when mailing specimens in a plastic bag. It absorbs excess moisture and helps prevent the plants from decaying and molds forming en route.

38



Thuus, plant maaterial will remain moisst and will arrive in a condition thhat enables analysis.

Maailing leaves in paper envvelopes resullts in their drying d out so that insect ddamage is diifficult to dettermine.

39

40

41

SKUAST MAJOR INSECT PESTS OF FRUIT CROPS IN JAMMU JAMMU

Mango leaf hoppers

Mango mealy bugs

Mango fruitflies

Mango leaf webber

Citrus leaf miner

Citrus psylla

Citrus whiteflies

Citrus blackflies

Citrus caterpillar

Fruit sucking moth

Damaged fruit by moth

Peach leaf curl aphids

Peach Fruit fly

Anar Fruit borer

Aonla fruit borer

Bark eating caterpillar

Aonla twig Gall maker

Aonla leaf folder

Guava Fruit fly

Guava Fruit borer

Banana weevil

Walnut weevil

Apple wooly aphis

Parrot damaging pear

Parrot damaging litchi

Bulbul damaging guava

Blue throated Barbet

© Dr. Uma Shankar, Plant Clinic, Nov. 2018

Squirrel damaging ber

Chapter-7

Plant Disease Diagnostic Techniques Importance of the Plant Diseases Globally, enormous losses of the crops are caused by the plant diseases. The loss can occur from the time of seed sowing in the field to harvesting and storage. Important historical evidences of plant disease epidemics are Irish Famine due to late blight of potato (Ireland, 1845), Bengal famine due to brown spot of rice (India, 1942) and Coffee rust (Sri Lanka, 1967). Such epidemics had left their effect on the economy of the affected countries.

Objectives of Plant Pathology Plant Pathology (Phytopathology) deals with the cause, etiology, resulting losses and control or management of the plant diseases. The objectives of the Plant Pathology are the study on: 

the living entities that cause diseases in plants;



the non-living entities and the environmental conditions that cause disorders in plants;



the mechanisms by which the disease causing agents produce diseases;



the interactions between the disease causing agents and host plant in relation to overall environment; and



the method of preventing or management the diseases and reducing the losses/damages caused by diseases.

Causes of Plant Diseases 

Plant diseases are caused by pathogens. Hence a pathogen is always associated with a disease. In other way, disease is a symptom caused by the invasion of a pathogen that is able to survive, perpetuate and spread. Further, the word “pathogen” can be broadly defined as any agent or factor that incites 'pathos or disease in an organism or host. In strict sense, the causes of plant diseases are grouped under following categories:

1.

Animate or biotic causes: Pathogens of living nature are categorized into Fungi, algae, bacteria, phanerogams, phytoplasma, protozoa and Rickettsia-like organisms

2.

Mesobiotic causes: These disease incitants are neither living or non-living, e.g. viruses and viroides

3.

Inanimate or abiotic causes: In true sense these factors cause damages (any reduction in the quality or quantity of yield or loss of revenue) to the plants rather than causing disease. The causes are: (i) Deficiencies or excess of nutrients (e.g. ‘Khaira’ disease of rice due to Zn deficiency) (ii) Light (iii) Moisture (iv) Temperature (v) Air pollutants (e.g. black tip of mango) (vi) Lack of oxygen (e.g. hollow and black heart of potato) (vii) Toxicity of pesticides (viii) Improper cultural practices (ix) Abnormality in soil conditions (acidity, alkalinity)

42

Plant disease diagnosis is the identification of nature and cause of diseases based on signs and symptoms. Identification of symptoms and signs and comparative symptomologies of infectious and non infectious diseases are considered to be most essential for diagnosis of a unknown plant diseases.

The presence of the pathogens or various structures viz., mycelium, sclerotia, sporophores and spores produced on the surface of the host are called signs whereas symptoms refer to only to the appearance of infected plants or plant tissues. Diagnosis of a plant disease is one of the most important and useful techniques in plant pathology and familiarity with the basic classification of plant diseases, the characteristics of organisms that cause a particular diseases, the symptoms and signs associated with different types of disease is a pre-requisite to diagnose a plant disease. Majority of plant diseases can be diagnosed by a relatively straight-forward procedure involving an evaluation of background information and a macroscopic and often microscopic examination of diseases plant. However, some diseases can be diagnosed correctly through the use of electron microscope and serology. A majority of abiotic and biotic factors may cause similar disease symptoms and the best proof that a particular organism is the cause of disease is fulfillment of Koch’s postulates. Koch's postulates are performed infrequently, except when the disease agent is suspected to be new and previously unreported.Most of the plant disease diagnoses done today involve identification of plant diseases that have been previously described and named. Several techniques may be performed to determine the identity of diseases. Visual studies of symptoms and signs, microscopy, culture media studies and serology techniques are the most frequently used techniques in diagnostic clinics.

Identification of nature of a disease In determination of a plant disease the first step is to determine the infectious and non infectious nature of the disease.

Infectious diseases An infectious disease will spread to other plants in the field by various means and is characterized by the presence of pathogens on the surface of the plants or inside the plant. In diseases caused by pathogens viz., fungi, bacteria, nematodes, viruses, mollicutes, a few or large numbers of these pathogens may be present on the surface of the plants or inside the plants. The presence of such pathogens in an active state on the surface of a plant indicates that they are probably the cause of the diseases. Their detection and identification can be determined with the experienced naked eye or with a magnifying lens and if no such pathogens are present on the surface of a diseased plants then it will be necessary to look for additional symptoms, especially for pathogens inside the diseased plant. Such pathogens are usually at the margins of the affected tissues, in vascular tissues or at the base of the plant or roots. Certain infectious pathogens like viruses are neither seen nor can be grown on artificial media. They produce symptoms similar to those resulting from nutritional deficiencies.

Non infectious diseases These are the diseases with which no parasite is associated; hence they are called as abiotic

43

diseases. They remain non infectious and cannot be transmitted from diseased plant to healthy plant.

If no organism is found in association with the diseased part and if viral symptoms are not present, the diseases may be due to inanimate cause. If symptoms look like those of nutritional deficiencies the identification can be confirmed by spraying a solution of the possible element in its salt form and usually recovery will occur within a week and identification can be confirmed. These non parasitic, non infectious diseases are due to disturbances in the plant body caused by lack of proper environmental conditions of soil and air, low and very high temperatures, unfavourable oxygen relations, unfavourable soil moisture, pH, presence of toxic gases in the atmosphere, mineral excess and deficiencies in the soil etc., are the major causes of non-parasitic diseases.

The diagnostic laboratory Location of the laboratory The diagnostic laboratory should be in a building with walls protected from rain. In tropical regions fungi commonly grow on the inside of walls exposed to rain. Such fungal growth can produce spores which contaminate cultures. Ideally, the laboratory should be located on the second level of the building. This reduces problems with rats and other pests such as ants. It is recommended that the laboratory consist of two large rooms, a preparation room and a clean room. We also recommend that a room or covered area be used for initially examining field samples and removing soil from root samples by washing. In this area small plant samples should be selected for later isolation of fungal or bacterial pathogens in the clean room. This area can also be used to extract plant parasitic nematodes from soil.

Preparation room The preparation room is used for preparing media, including sterilizing items in the autoclave, sterilizing Petri dishes in an oven, washing glassware and storing glassware, chemicals and other basic items. This room should have an exhaust fan to remove hot air produced by the autoclave and the oven.

Clean room The clean room is used for isolating fungi and bacteria from cleaned subsamples of diseased plant tissue into pure cultures. It is also used for growing cultures under clean conditions. The microscopes are located in this room for examining cultures and fungal structures This room should be air-conditioned, if possible, to protect equipment and cultures. It should also be kept free from dust and insects. However, do not have an airtight clean room or humidity will be too high and fungus (mould) will develop on walls and equipment. A dehumidifier is useful in this room. No soil is allowed in the clean room as soil is a source of fungus-eating mites that can contaminate cultures.

A compound microscope fitted with ×10, ×20, ×40 and ×100 (oil immersion) objective lenses. A basic student-grade microscope is sufficient for most diagnostic work. If funds are available the microscope can be fitted with a ×20 metallurgical lens with a long working distance. This lens is ideal

44

for examining fungal structures in situ in cultures, as it has a long depth of field A dissecting microscope for examining diseased plant samples for fungal structures. This is especially important for many leaf infecting pathogens which cannot be grown in artificial media. It is also used for transferring germinated single spores or hyphal tips for purifying cultures, and for studies of plant pathogenic nematodes.

A sterile work chamber for pouring media and isolating fungi from plant tissues. The tropical climate of Vietnam means that there are many fungal spores in the air. These spores contaminate media while pouring it, plating tissue or performing culture transfers, unless a sterile work chamber is used.

A bench with overhead fluorescent lights for stimulating sporulation and pigment production of many fungal species, either in culture or on leaves in moist chambers. Ideally there should be one bench for clean cultures and one bench for plates with diseased tissues. A culture cupboard is useful for incubating cultures in the dark. This is necessary for cultures on media containing antibiotics that are affected by light (e.g. Phytophthora selective medium).

A refrigerator for storing media in bottles, Petri dishes with media (in plastic bags or foil to stop the media drying out), as well as antibiotics, cultures and small tissue samples.

An electronic balance with an accuracy of 0.001 g is recommended for weighing small amounts of antibiotics or chemicals.

Comfortable chairs for sitting at work benches.

Accession books, for recording details of each diagnosis and for recording a list of stored cultures. 

Small instruments for isolation and cultural work, including: –

Fine Forceps

–

Inoculating Needles

–

Surgical Scalpel Handles

–

Transfer Loops (For Bacterial Work)

–

Surgical Scalpel Blades

–

Marker Pens

–

Small Knives

–

Ethyl Alcohol

–

Transfer Needles (Flat Tip) Tissues

–

Cutting Boards

–

Microscope Slides and Cover Slips

–

Filter Paper

Check the walls and equipment regularly for fungal growth.

The floor of the clean room should be mopped regularly to remove any dust particles from the area. Fans should also be turned off and windows closed whilst culturing, to reduce the movement of air in the laboratory. Critical work should be carried out in a laminar flow cabinet which has been wiped down with 70% alcohol.

45

Essential items of equipment for the preparation room are listed below o

An oven for sterilizing glass Petri dishes, which should be wrapped in newspaper or in paper bags.

o

A small autoclave suitable for sterilising volumes of 1–2 litres of media or water in flasks or Schott bottles. The autoclave is also used to sterilise media or water in glass test tubes or McCartney bottles, pipettes and other glassware wrapped in paper or aluminium foil. 

A pressure cooker to sterilise small amounts of media and water. This can be purchased at most large markets.

o

A balance (0.1 g accuracy) for weighing chemicals, potatoes, carrots and so on for media preparation.

o

An electric hot plate for boiling potatoes and carrots for media.

o

A bench for media preparation.

o

A sink for washing Petri dishes and other glassware.

o

A storage cabinet.

46

Chapter-8

Acquaintance with Plant Pathology Laboratory and Equipments The student should to get acquainted with the chemicals glassware and equipments of the plant pathology laboratory listed below.

Instruments 1.

Microscope Microscope is a device, which can magnify a microbial cell or a group of microbial cells to

enable the human eye to study its structures, morphology etc.

A.

Simple microscope: Consists of a simple lens system

B.

Compound microscope: It consists of 2 or more lens systems- Depending on source of illumination, they are of two kinds:

a.

Light microscope Specimen is illuminated by visible light or U.V. rays with a max magnification of 1000 or more.

These are used for observing stained and unstained specimens and counting of microbes. They

include the bright field, dark field, U.V phase contrast and the fluorescent microscope. b.

Electron microscope Here the images are formed on a fluorescent screen by electron beam focused by magnets instead

of lens, with a magnification of 1, 00,000. These are used for observation of viruses and ultra structures of cells.

2.

Autoclave It is an apparatus in which saturated steam under pressure affects sterilization called autoclaving.

The pressure increases boiling point of water and produces steam with a high temperature. Cells are destroyed by high temp and not by the pressure. Most of the organisms are killed at 121 °C and 151b pressure per sq. inch in 15 min. It is more efficient and common instrument used to sterilize solids and liquid media for microbial culture. It is not recommended for oils, powders, heat sensitive fluids and plastics. Autoclave is a double walled cylindrical metallic vessel made of thick stainless steel copper, lid of which is opened to receive the material to be sterilized. The lid is provided with pressure gauge noting the pressure, steam clock for air exhaustion of the chamber. It is also provided with safety valve to avoid explosion. The materials to be sterilized are kept in a basket provided with holes all around for the free circulation of steam. Moist air has most penetrating power than dry heat and hence it is more efficient than dry- heat.

3.

Pressure cooker: It is a suitable alternative to an autoclave. Some labs will have a big size

pressure cookers implanted with a pressure gauge. In case of power failure materials are sterilized in pressure cooker.

47

4.

Hot air oven It is an electrically operated equipment with a thermostat (ambient Temp, to 300°C) used for

sterilizing glassware. An oven consists of an insulated cabinet, which is held at a constant temp, by means of an electric thermostat. Some ovens are also fitted with fan to keep hot air uniformly circulated at constant temperature. For proper circulation of hot air, the shelves are perforated. The scheduled temperature for sterilization with dry air is given in table.

5.

Incubator It is used for incubation (culturing of microbes) at a constant temp. It is similar to an oven in

construction and consists of an insulated cabinet fitted with a heating element at the bottom. The temp, of the incubation is maintained at desired level (ambient to 110°C) by an automatic device called thermostat. It is provided with double doors, made of glass so that the contents of incubator maybe viewed without admitting outside air. Most incubators can be supplied by placing a beaker of water in it to retard the dehydration of medium during growth of micro organisms. Some incubators are provided with fluorescent light that can be used to encourage sporulation.

Temperature and humidity control chamber: In this one can adjust both temperature and humidity.

6.

Colony counter It is an electronic apparatus used to count the number of colonies on a Petri plate. A Petridis fits

into the recess in the platform. The colonies on plates are counted on an illuminated screen, illuminated from beneath with a large magnifying lens which provides 1.5X magnification. Some instruments are also fitted with electronic micro switch with pen and counter. The counter bar is depressed and the number of colonies is instantly displayed on digital read out.

7.

Inoculation chamber Most of the aseptic transfers are made using inoculation chamber made of wood. Now-a-days

laminar airflow system is used as inoculation chamber. It is used for reducing danger of infection while working with infective microorganisms and for preventing contamination of sterile materials. It is a hood like structure having germicidal ultraviolet lamp and Bunsen burner. It consists of mid table as working place onto which sterile air is pumped at uniform velocity either in horizontal or vertical direction. It works on the principle of application of high efficiency particulate filters (HEPA)-or fibre glass filter which can retain all particles including bacteria whose diameter is more than 5 microns.

8.

Ultraviolet lamps U.V. rays with 200-300 nm wave length is germicidal. The lamp which produces U.V. rays of

near 200 300 nm wavelength kill or inactivate most of the virus and vegetative form of microorganism present in laboratory or on an inoculation chamber.

9.

pH meter It is used to determine the pH of solutions of unknown pH as well as for setting of pH of various

media, and testing biochemical activity of microorganisms. pH is expressed as a number from 0 to 14.

48

The number is an expression of the concentration of H ion in the solution. The optimum range of pH for bacteria is 6.5 to 7.5 and for fungi it is 4-6. The measure of pH with pH meter is done Electrometrically.

Measurement of pH depends upon the" development of membrane potential by a glass electrodes. As an alternate, pH papers are used to measure the pH of the medium.

10. Water bath It is an insulated metallic box fitted with an electric heating mechanism and a thermostat, which maintains the temperature at desired level. There is racks form holding test tubes. These are usually used for melting of media, testing enzymatic activities of various microorganisms, widal test etc.

11. Centrifuge It is an apparatus that rotates at high speed and separates substances as particles on the basis of mass and density by means of centrifugal force. The microbes are arrested from sediments settled at the bottom of the tube after centrifugation. The centrifugal force is noted in rpm of angular speed. A centrifuge consists of head which is rapidly revolving on upright motors. Generally four metal caps are attached to the head for holding rubes or other container of the material from which particulate matters to be separated. During centrifugation liquid containing particulate matter is kept in the tubes, run at a particular speed and when centrifugation is completed, the particulate matter gets settled at the bottom of the tubes. The commonly used centrifuges are of low speed, high speed and ultracentrifuge with highest speed limit of 5000 rpm, 18000 rpm, and 20,000 to 60,000 rpm, respectively. These are used for separation of virus particles, bacterial cells, and fungal spores, separation of mixtures of liquids varying in their density and concentrating microorganisms in various samples for enzymatic and other studies.

12. Balance Various media components for culture media preparation and samples etc. are weighed on an ordinary balance. Whenever precision is required an electronic monopan balance is recommended. As most of the media ingredients are highly hygroscopic, the balance should be cleaned immediately after use.

13. Spectrophotometer or colorimeter It is an electrically operated simple instrument used for estimating population of bacteria, based on the principle of turbidity determination. Turbidity is the cloudiness of the suspension. The more turbid a suspension, less light will be transmitted through it. In other words, the amount of light absorbed and is scattered is proportional to the mass of cell. As bacteria grow in a broth, the clear.broth becomes turbid. Since turbidity increases as the number of cells increases, this is used as an indicator of bacterial density in broth: The turbidity is expressed in unit of optical density (O.D.) which is expressed using Spectrophotometer.

49

14. Haemo-cytometer The number of microorganisms present in a given liquid sample can be counted and morphology of bacteria can be observed by direct cell count method using haemocytometer. It is a special glass slide with a depression (0.1 mm - 0.02 mm deep) at the centre covering an area of 1 mm, the area of 1 mm is further divided into 400 small squares. To get the number of cells per ml of sample the following formula is used. The number of cells per ml = average number of cells in a small square x 400 x 104 (factor).

15. Filters Heat sensitive materials like vitamin solutions are sterilized by filtration technique as they are destroyed by heating at temperature normally used for sterilization e.g. Seitz filter.

16. Refrigerator It is a basic requirement in the microbiological laboratory and used for storing stock cultures of microorganism at 4°C to save sub-culturing every few days. The stored cultures at low temperature are fairly inactive and will not suffer damage due to evaporation of medium. It is also used to store sterilized media to prevent dehydration and to serve as a repository for thermo-labile solutions, serums, antibiotics and biochemical reagents.

17. Bunsen Burner It is named after R.W. Bunsen. It is a type of gas burner with which a very hot particularly nonluminous flame is obtained by allowing air to enter at the base and mix with gas. In the absence of Bunsen burner, alcoholic lamp is used. They are used to sterilize inoculation needles / loops before they are inserted into culture. It is also used for flaming the mouth of test tubes, media containing flasks and other glass apparatus to avoid contamination by other microorganisms.

18. Hot plate stirrer It is useful to stir the chemicals in water without heat to make suspension. It is fined with the stirrer and heat control. Stirring is done by creating magnetic field, which causes the bar magnet kept in the container to spin resulting in the stirring of the medium.

1.

Inoculation loop or Inoculation needle Used for aseptic transfer of culture. It consists of an insulted handle provided with screw device

at the tip which holds a heat resistant nichrome or platinum wire approximately 3 inches long. The end of wire is bent to form a loop. Inoculation needle is similar to loop, but the holder contains a straight piece of wire instead of a loop. They are sterilized by flaming in the blue portion of burner flame until it is red. The loop is mainly used to transfer culture of microorganism growing on liquid cultures. Inoculation needles are used to transfer cultures of microorganisms growing on solid medium in form of colonies.

2.

Glass spreader It is bent T or L shaped glass rod used for spreading of liquid culture and sample on sterile agar

plate.

50

3.

Glass marking pen All the culture material is labelled with the use of Glass marking pen.

4.

Petri-dish Cans It is made of copper and used as container for keeping Petri-dishes. The Petri-dishes in cans are

sterilized in hot air oven at required temperature.

5.

Glass Ware

Test Tube: a) Test tubes of 18 x 150 mm size are used for preparation of broth, agar slants and agar stabs, b) 25 x 150 mm size are used for preparation of dilution blanks c) Screw caps tubes with round bottom of size 15 x 125 mm are used for maintenance of culture.

Petri-dishes: Petri-dishes, a pair of circular glass containers named after Petri, are used for the preparation of agar plates. The common size is 100 mm in diameter.

Pipettes, Flask and Beakers: Different sizes of pipette and conical flask are used for preparation of dilutions and plating. Generally pipette of 10 ml and 1 ml are used for sterile transfer of known volumes of liquid. For preparation of dilutions conical flasks of 250, 500 and 1000 ml are used for preparation of medium. The volume of media should not exceed 2/3 of the volume of flask. Beakers of size 250, 500 and 1000 ml are used for preparation of medium.

Durham Tubes: These are small tubes of 3 x 25 mm size used for melting of chemicals during preparation media in broth tubes for observing gas production in various fermentation tests.

Slides and cover slips: Rectangular slides of 75 x 25 mm size made of glass with polished edge are used for observation of microorganisms. Square or circular cover slips of size 18x18mm or 20 mm diameter are used for covering the specimen glass slide while observing under high power objective of a microscope. The thickness of cover slip shouldn't exceed 0.016 mm.

Cleaning of glass ware: All the glass ware before put to use for microbiological work should be cleaned with a detergent (chromic acid mixture) followed by thorough wash with clean water and distilled water. All the used glassware with cultures has to be autoclaved before subjecting to further cleaning with detergent. Glassware to be cleaned should be left in the chromic acid mixture over night and later repeatedly washed in running water to remove traces of detergent.

Exercise: Get acquainted with handling and use of instruments and apparatus which are commonly used in a plant pathology laboratory.

51

Chapter-9

Seed Treatment

S

eed treatment with bio control agent and / or fungicides will not only control seed borne diseases but also soil borne diseases. This is the cheap and most economical method of plant disease

control. Seed treatment can be done both in dry and wet states. The details pertaining to the fungicidal seed treatment are furnished in exercise no. 9 of this manual. The seed treatment with bio-control agents can be done in two ways i.e. wet and dry treatments.

Wet treatment 1.

Grow Trichoderma viride or T harzianum bio control fungi in Petri plates on PDA medium for 15 days for abundant conidial production.

2.

Harvest the conidia from well sporulated agar culture plates with the help of a cotton swab into a clean beaker.

3.

Prepare 1 ml of spore suspension of 106 to 108 load with the help of a Haemocytometer in a test tube.

4.

Weigh 4 grams of seed and add to it 1 ml of spore suspension having stickers like Carboxy methyl cellulose 1 per cent to smooth surfaced seeds for 30 minutes. Seeds with rough surface do not require a sticker and treat the seeds before sowing.

Dry treatment 

Weigh 4 grams of Trichoderma talc formulation.



Weigh 1 kilogram of seed to be treated.



Add 4 grams of Trichoderma formulation to 1 kg seed and thoroughly mix them and use before sowing.



This treatment can be combined with fungicidal seed treatment provided the biocontrol agent and fungicide are compatible. The fungicide such as metalaxyl, carbendazim and mancozeb are compatible with bio control agents like T viride, T harzianum etc.

Note: Before combining fungicides with bio-control agents it is mandatory to know the sensitivity of bio-control agents with test fungicides.

Advantages: Combined treatment of seeds have a synergistic effect in controlling seed and soil borne diseases of crops.Seed treatment with bio-control agents also promotes the seedling vigour, population stand under field conditions.

Preservation of living cultures Living cultures are stored for use as reference cultures, or for later use in pathogenicity tests or other experiments. Cultures are stored in national culture collections as part of reference materials that support a national database of plant pathogens.

Storage in sterile water—Pythium and Phytophthora

52

This is a low-cost, simple method that is particularly suitable for Pythium and Phytophthora. A sterile work chamber should be used for this procedure. Agar blocks 1 cm square are cut from the margin of a young, actively growing fungal colony. These are placed in sterile water in a McCartney bottle and the cap is screwed down. The bottles are stored under cool conditions. Do not store in a refrigerator as some species are killed at low temperatures. Cultures can be stored between 6 months to 2 years, depending on the species. Cultures are revived by removing a block of agar from the bottle and placing mycelium side down on fresh medium. It is essential to ensure that the water and agar blocks are not contaminated by bacteria—the presence of bacteria will lead to rapid death of the fungus.

Storage of sclerotia Sclerotia can be stored for long periods under cool dry conditions in a small screw cap glass bottle or ampoule. This is a suitable technique for storage of species such as Sclerotium rolfsii, Sclerotinia sclerotiorum, Rhizoctonia spp. (sclerotial forming isolates).

In tropical regions it is best to store sclerotia on sterile paper tissue over blue silica gel in a McCartney bottle (or similar screw cap bottle) to ensure very low humidity for storage.

Storage as colonized pieces of plant stem or leaves Cultures are grown on sterile WA containing pieces of sterile plant tissue or seeds. The colonised pieces are air dried and then stored in a small glass tube. Alternatively they can be stored on a sealed container on sterile paper above blue silica gel to ensure very dry storage conditions.

Lyophilisation by freeze drying Lyophilisation, or freeze-drying, is the method of choice for long-term preservation of many fungi and is used routinely in most major culture collections. Its major drawback is the requirement and expense of specialised equipment. It is best suited to fungi which grow and sporulate well in culture on sterile plant tissue such as green rice stem-pieces or carnation leaf-pieces. There are also many fungi which cannot be freeze-dried successfully, such as oomycetes, rusts and mildews.

Cultures are lyophilised by drying colonised stem or leaf pieces in small glass ampoules under high vacuum (10–1 to 10–2 Torr). The ampoules are prepared by inserting a small cotton wool plug and then autoclaving in a loosely covered beaker. Five stem or leaf pieces are taken from a culture (which is two weeks old and initiated from a single conidium), and aseptically transferred to the ampoule. The ampoule is replugged, labelled (with an internal label) then heated and drawn out to an hourglass shape using a gas torch. The ampoules are attached to the freeze dryer for 12–24 hours, then sealed under high vacuum and stored at room temperature or at 5 °C. Many species of Fusarium and other fungal genera have been successfully lyophilised using this technique and have retained viability for many years. Cultures can be revived by aseptically plating the dried stem or leaf pieces onto a suitable medium. The ampoule is first surface sterilised before it is shattered to release the leaf pieces.

53

Other preservation techniques for living cultures Cultures can also be stored as spore suspensions in glycerol in a –80°C freezer for long-term storage. Many species have also been stored successfully in liquid nitrogen. However, these are very expensive techniques.

Preservation of fungal cultures for herbarium records Cultures are initiated from single germinated conidia and grown under standard conditions of temperature and light for 2 to 3 weeks. Cultures are then killed by exposing the plates to formalin in a closed container for 3 days. Preservation of the culture is achieved using agar and glycerine. Three grams of agar are dissolved in 147 mL water, which is then dispensed as 6 mL aliquots into test tubes before autoclaving. The lid of the culture dish is inverted, 1.5–1.75 mL glycerine is added and then the 6 mL aliquot of hot agar is poured over the glycerine. The culture is aseptically lifted from the Petri dish and floated on the mixture in the lid. Cultures are then allowed to dry in a drawer for 3–5 days covered with a sheet of paper. When dried, the culture is rubbery and can be removed from the Petri dish for storage. This procedure was originally developed for use with Fusarium species at the Fusarium Research Centre, Pennsylvania State University. It is suitable for many fungi.

Preservation of fungi under mineral oil Many fungi can be stored in culture under sterile mineral (paraffin) oil for 4–5 years at 15–20°C. Cultures should be grown on PDA amended with 0.1% concentrated yeast extract (e.g. Vegemite®).

The mineral oil should be prepared as follows: 1.

Dispense 11 mL paraffin oil into 25 mL McCartney bottles without rubber seals.

2.

Replace lids loosely and autoclave at 121 °C for 20 minutes.

3.

Allow to cool completely in the autoclave.

4.

Remove any water from the oil by heating at 120 °C in an oven for 8 hours and leaving in the oven overnight to slowly cool to room temperature. Discard any bottles that contain cloudy oil or repeat the oven-heating process.

Cultures can be re-grown as follows: 1.

Aseptically remove a small piece of agar from the preserved culture.

2.

Blot the piece on sterile filter paper or blotting paper to remove the oil.

3.

Plate the piece of agar on appropriate medium.

Note: It is recommended that three cultures of each fungal isolate are preserved at any one time, and that mineral oil cultures are renewed every 4–5 years

54

Chapter-10

Sterilization

S

terilization is the process of killing all living organisms in a culture medium or on the surface of glassware used for sterile work, such as glass Petri dishes.

Heat sterilisation The temperature and time required for killing are inversely related. Table shows the minimum times required for effective sterilisation at the temperatures given for both moist and dry heat: These times do not guarantee sterility. They are times calculated from experience and are based on normal levels of contamination with heat resistant organisms. The species, strain and spore forming ability of a microbe greatly affects its susceptibility to heat. In moist heat the vegetative forms of most bacteria, yeasts and fungi and most animal viruses, are killed in 10 minutes by temperatures between 50 °C and 60 °C. However bacterial spores require 15 minutes at temperatures ranging from 100 °C to 121 °C. In dry heat bacterial spores require 1 hour at 160 °C The nature of the material in which the organisms are heated is also an important factor. A high content of organic substances generally tends to protect spores and vegetative organisms against the lethal action of heat. Proteins, gelatin, sugars, starch, nucleic acids, fats and oils all act in this way. The effect of fats and oils is greatest in moist heat as it prevents access of moisture to the microbes. The pH is also very important. The heat resistance of bacterial spores is greatest at neutral pH and decreases with increasing acidity or alkalinity.

Dry heat sterilisation Dry heat kills microbes by oxidation. The dry heat process is the best method for the sterilisation of dry glassware such as test tubes, glass Petri dishes, flasks, pipettes, all glass syringes and instruments such as forceps, scalpels and scissors.

Moist heat sterilisation Moist heat kills microorganisms, probably by coagulating and denaturing their enzymes and structural proteins, a process in which water participates. All culture media therefore are sterilised by moist heat. An effort should be made to avoid sterilising large and small volumes of media in one load as time must be allowed for large volumes to reach the required holding temperature, and this will result in small volumes receiving too much heat.

Sterilisation of instruments Forceps, inoculating needles and other instruments must be sterilised before contact with a culture to avoid cross-contamination. Inoculating needles are best sterilised by heating to red heat in a flame. Forceps and scalpels are sterilised by dipping in alcohol. Before use, the alcohol is burnt off by passing the forceps through a flame to ignite it. Do not hold the instrument in the flame, since this will heat it up too much. Be very careful not to place hot or flaming instruments in or near alcohol, since this is a fire hazard.

55

Sterilisation of work surfaces Trays, benches and other surfaces may be sterilised with a liquid disinfectant. Alcohol is the most commonly used. Alcohol works best as a sterilant if it contains some water, and a solution of 70% ethyl alcohol is suitable. Methylated spirits is also suitable.

Health and safety In the field 

Take care to follow all recommended safety precautions when applying pesticides, particularly those used for insects (insecticides). Only use registered chemicals.



Wash hands carefully before eating meals, especially when soil has been handled.



Drink adequate water on hot days in the field.



Take care with machetes so that you do not cut yourself or other people.

In the laboratory 

Check the safety aspects of all chemicals before use. Such information can be found on the product packaging or on the internet. Major chemical companies supply links to the chemical Material Safety Data Sheets that correspond with their products.



Use gloves where appropriate.



Ethyl alcohol is highly flammable. Do not wipe bench



Keep a fire blanket in the laboratory to put out clothing fires.



Wear shoes in the laboratory to protect feet from sharp instruments dropped accidentally. Closed shoes also protect feet from broken glass and chemicals.



Do not open the autoclave until the internal air pressure reaches atmospheric pressure (reading 0 on the dial). Always use heavy duty material gloves when removing any material from the autoclave or oven.



Take care when opening the oven. High temperatures and steam can cause serious burns.

56

Chapter-11

Isolation of Fungal and Bacterial Pathogens

I

solation of the fungal pathogens from diseased material is made by surface sterilizing the diseased area with surface sterilizing agents, removing a small portion of the infected tissue (leaves, stems,

fruits etc.) with a sterile scalpel, and plating it in a plate containing a nutrient medium. The most common method, for isolating fungal pathogens from infected leaves as well as other plant parts involves cutting several small sections 5-10 mm-square from the margin of the infected lesion to contain both diseased and healthy looking tissue. These are placed in surface sterilizing agents solutions for about 15-30 seconds the sections are taken out aseptically and blotted dry on clean, sterile paper towels or washed in three changes of sterile water and are finally placed on the nutrient medium, usually three to five per dish. The pathogen will grow from the sections and the colonies of the pathogen are sub cultured aseptically for further study.

Materials required Infected young leaves,sterile Petri -dishes, PDA slants, sodium hypochlorite solution ( 1 % ), sterile water, razor blade, forceps, inoculation needle, burner/spirit lamp, spirit, incubator, PDA medium.

Procedure 1.

Select infected host tissue from the advancing margin of the lesions.

2.

Cut into small pieces (2-5 mm ) containing both the diseased and healthy tissue and keep in sterile Petri dishes

3.

Dip the pieces into 1 % sodium hypochlorite solution for about one minute.

4.

Transfer the pieces to Petri - dishes containing sterile distilled water and wash thoroughly in two changes of sterile water to free them from the chemicals if any.

5.

Wash hands with rectified spirit and wipe the table top of inoculation chamber-'with rectified spirit.

6.

Lit the burner

7.

Hold the flask containing sterile Luke warm PDA in the right hand and remove plug near the flame. Lift the lid of Petri dish gently with left hand and pour about 20 ml of medium. Close the mouth of the flask with plug near the flame

8.

After solidification of the medium, place four sterilized pieces at different distance in a single PDA plate.

9.

Incubate the Petri dishes in an inverted position at 25° C and examine for 3-5 days.

Observations and results Observe the incubated plates from the second day onwards for the growth of the fungus.

Aseptically transfer the bits of mycelia from the margin of the colonies on fresh PDA slants for

57

further study. Mycelia growth on the medium from the infected tissues, indicates that the disease may be due to a fungus.

Isolation of phyto-pathogenic bacteria from diseased plants Isolation and identification of bacteria associated with diseased plant is important to determine whether bacteria are involved in plant disease. The method normally used to isolate phyto-pathogenic bacteria differs from that used for fungi. A suspension of bacterium is prepared from the infected material and loopfuls of this are streaked onto nutrient agar plates. The aim is to produce single colony that can be sub-cultured pure. Pure cultures are absolutely essential for pathogenicity assays and characterizing the pathogen for identification. The serial dilution method is used for isolating bacteria from diseased tissues contaminated with other bacteria. After surface sterilization of sections of diseased tissues, the sections are ground in small volumes of sterile water and then part of this homogenate is diluted serially. Finally, plates containing nutrient agar are streaked with a loop dipped in each of the different serial dilutions and single colonies of the pathogenic bacterium are obtained from the higher dilutions that still contain bacteria.

Choice of material: Selection of the diseased tissue is important because pathogenic bacteria may occupy different locations in the plant. In isolation of bacteria, it is generally better to use newly collected material. The earliest stages of symptom development should be used. Old lesions and dead areas usually contain few pathogens and many saprophytes. Necrotic diseases usually start with tiny, dark greenish, spots, which are excellent for isolations. Cankers and soft rots should either be at an early stage, or if, older lesions only are available, the advancing edge must be used, where the disease is spreading into healthy tissue. When crown gall is suspected in a woody plant a search must be made for young galls on young green stems. With wilts and other vascular infections small pieces of infected stem are usually good for isolation.

Preparation of material: Clean leaves and stems, carefully chosen and "handled aseptically, can often be used without surface sterilization. Roots and parts contaminated with soil should be gently washed with clean water as soon as possible after collection.

Medium: Nutrient agar is suitable for the isolation of most plant pathogens. The medium used for isolations must have a dry surface. If water is present the bacteria move around and a carpet of mixed growth results instead of the required single colony. This exercise deals with the isolation of bacterium, Xanthomonas axonopodis pv. citri causal agent of citrus canker.

Materials Fresh citrus leaves infected by Xanthomonas axonopodis pv. citri, nutrient agar medium, surface sterilizing agents (1 % sodium hypochlorite), sterile razor blade, glass rod, sterile water, sterile test tubes and Petri-dishes, sterile pipettes (I ml), inoculation loop.

Procedure Put on the U.V lamp of inoculation chamber for 5 mts. Wipe the table top with rectified spirit

58

Wash hands with rectified spirit and air dry. Lit the burner or spirit lamp, arrange sterile Petri dishes near the burner.

1.

Select a diseased citrus leaf infected by canker and cut out a small portion of the diseased tissue from the advancing lesion using sterile razor blade in a drop of sterile water and after several minutes, examine under microscope. If bacterial ooze is seen, proceed for isolation.

2.

Surface disinfests the cut portions by dipping in sodium hypochlorite solution for 60 sec. and then immediately rinse three times with sterile water.

3.

Immerse the disinfested cut portions in I ml of sterile water taken in a clean sterilised test tube.

4.

Crush the cut portions of the leaf with a sterile glass rod. Allow it to stand for 5 minutes to allow the bacteria to diffuse out of the cut tissue and into the water.

5.

Gently lift the lid of a Petri dish with left hand and using inoculation loop transfer several loopfuls of the bacterial suspension to sterile Petri-dishes (three) containing 1 ml of sterile water and mix thoroughly.

6.

Hold flask filled with sterile Luke warm nutrient agar medium in the right hand and remove cotton plug near the flame and pour about 20 ml of medium into each dish and mix thoroughly by gentle rotation. Allow time for solidification of medium.

7.

Incubate the dishes in an inverted position at 25°C for 36 to 72 hours.

8.

Observation: Observe the dishes for appearance of desired bacterial colonies. If colonies appear, select consistently found and well isolated colonies of the pathogen, for sub-culturing and further studies.

9.

Select the isolated colonies and streak on the surface of a solidified medium in a zigzag manner and incubate the dishes at 25oC. Bacteria isolated from nature may be contaminated with saprophytic species; hence, re-streaking for isolation ensures a pure culture. Transfer some of the purified colonies to NA slants and grow them for further use.

59

Chapter-12

Preparations of Culture Media:

Potato Dextrose Agar (PDA) for Fungi and Nutrient Agar (Na) for Bacteria

M

edium (media pl.) is the substance which provides nutrients for the growth of microorganisms.

The nutrient preparation on which culture is grown in the laboratory is called culture medium

Microbes require different nutrients for their growth. There is no single medium which can support the growth of majority of microbes. Thus, different types of media and environmental condition are to be used for a given group of microbes. Many special purpose media are needed to facilitate, recognition, enumeration and isolation of certain microbes. Based on chemical composition, media can be classified into.

1)

Natural 2) Semi-synthetic 3) Synthetic.

1.

Natural medium: The exact chemical composition of this media isn't known properly. It includes ingredients of natural origin like yeast extract, beef, milk, tomato juice, blood etc.

Sometimes this medium is also referred to as complex medium or non-synthetic medium because medium is of complex type and contain various ingredients of unknown chemical composition.

This type of media is useful for cultivation of microbes whose specific growth factor requirement is not known.

Eg. Carrot slices, potato plugs, twigs, milk, meat extract, peptone etc.

2.

Semi-synthetic: The chemical composition of media is only partially known. Media, which contains Agar, is semi-synthetic medium.

Eg. Potato Dextrose Agar medium, Nutrient Agar media.

3.

Synthetic medium: The chemical composition of the medium is completely known.These media are very useful in studying the physiology, metabolic nature and nutritional requirements of microbes. Both autotrophs and heterotrophs can be grown in these media.

Eg. Mineral glucose medium, Richard's solution, Raulins medium etc.

Based on consistency the media are of three types as 1) Liquid 2) Semisolid 3) Solid medium

1.

Liquid medium: Nutrient broth is the common liquid medium used in a microbiological laboratory. Its drawback is that the morphology of bacterial colony cannot be studied. But it supports a high microbial population.

2.

Semi-solid medium: A semisolid medium is prepared with agar of concentration of 0.5% and is useful in the cultivation of micro aerophilic or studying bacterial motility.

3.

Solid medium: If agar is added to a nutrient broth, it becomes solid medium. It is used for isolating microbes and to determine characteristics of colonies. It remains solid on incubation and not destroyed by proteolytic bacteria. The addition of 15g of agar in 1 I of liquid culture will produce a gel that liquefy at 95°C and solidifies at 40-45°C into gel.

Based on application or function, media can be classified as follows.

1.

Selective media: Provide nutrients that enhance the growth and predominance of particular microbe and don't enhance or may inhibit other types of organisms that may be present. For

60

instance, isolation of bacterium Neisseria gonorrhoeae from a clinical specimen is facilitated by the use of media containing certain antibiotic. These antibiotics don't affect pathogenic but inhibit the growth of contaminating bacteria.

2.

Differential media: Certain reagents or supplements when incorporated into culture media may allow differentiation of kinds of bacteria. If a mixture of bacteria is inoculated on to blood agar media, some of bacteria destroy the RBC and others don't. Thus one can distinguish between haemolytic and non-haemolytic bacteria on the same medium.

3.

Assay media: Media of prescribed composition are used for the assay of vitamins, amino acids, antibiotics etc.

4.

Enumeration media: Specific kinds of media are used for determining the bacterial population in milk, water, soil and food etc.

5.

Maintenance media: It is used for satisfactory maintenance of viability and physiological characteristics of culture.

Preparation of basic liquid Medium (broth) for routine Cultivation of Bacteria Bacteria are often cultivated in liquid broth (media lacking agar) Materials: Peptone 5g, Beef extract 3 g, distilled water 1 I, 0.1 N HCl, 0.1 N NaOH, pressure cooker, 1 L beaker measuring cylinder, non-absorbent cotton, test tube and pH paper.

Procedure: Take the weighed amounts of peptone and beef extract and mix in 50 ml of distilled water and heat it is dissolve the contents. Add more distilled water to make it to 1 L. Adjust the pH to 7 using pH papers by adding either acid or alkali as the case may be. Take this into the test rube and apply cotton plug, sterilize at 15 Ibs pressure for 15 mts in pressure cooker. Allow the pressure cooker to cool, remove the nutrient broth tubes and store at room temp and cover with butter paper.

Preparation of Basic Solid Medium Liquid broth media containing nutrients are usually solidified by the addition of agar. Eg. Potato Dextrose agar medium, Nutrient agar medium.

A) Preparation of Potato Dextrose Agar Medium: Used in isolation and maintenance of common fungi.

Materials: Peeled potatoes - 200g, Dextrose - 20 g. Agar - 20 g, Distilled water 1 L, beaker 1L, 250 ml conical flasks, knife, muslin cloth, measuring cylinder, cotton nonabsorbent, pressure cooker.

Procedure 1.

Take 500 ml of distilled water in 1L beaker and add 200g of peeled and sliced potato boil the potatoes till they become soft.

2.

Filter the contents of the beaker through muslin cloth and squeeze out all liquid

3.

Add the dextrose dissolved in water to this extract.

4.

Adjust the pH of medium to 6 to 6.5 using 0.1 N HCl or 0.1N NaOH as the ease maybe

5.

Add the dissolved agar to dextrose-potato extract and make the volume to 1lt and dispense 200ml each to 5 conical flask and plug with non absorbent cotton. Sterilise the flasks at 15 Ibs pressure for 15 mts in a pressure cooker.

6.

Allow the pressure cooker to cool, "Remove the conical flask and store at room temperature.

61

Allow the flask to cool until the flask can be held by hand.

7.

Prepare agar plate by pouring the media into Petri-dish quickly. Using aseptic condition, allow the media in Petri-dish to solidify to produce the agar plate.

B.

Preparation of Nutrient Agar Medium: Used for the maintenance and isolation of bacteria.

Materials: peptone - 5g, beef extract - 3g, Agar - 20g, distilled water -1lt, Petri-dish, 1lt beaker, 250 ml conical flasks, measuring cylinder, non absorbent cotton, pressure cooker and hot plate.

Procedure 1.

Dissolve the weighed amounts of peptone and beef extract into 500 ml of water.

2.

Heat and dissolve the chemicals and adjust the pH of medium to 7 by adding 0.1N HCl or 0.IN NaOH.

3.

Weigh 20g agar and dissolve in 500 ml of distilled water in another beaker

4.

Mix the dissolved agar with chemical solution and make up the vol. to 1lt.

5.

Dispense 200 ml each into 5 conical flasks.

6.

Plug the flask with non absorbent cotton and sterilise at 15 Ibs pressure for 15 mts in a Pressure cooker.

7.

Allow the cooker to cool, remove the conical flask and store at room temp.

8.

Allow the flask to sufficiently cool and prepare agar plates by pouring media into

Petri-dish under aseptic condition; allow the media with Petri-dish to solidify.

Precautions 1.

Don't pour the media over 2/3 of flask capacity.

2.

Cotton plug must be loose whale autoclaving.

3.

Don't pour media to Petri-plate when the medium is too hot since it produce condensation of water on underside of Petri plate lid and thus can fall on to agar surface and may lead to contamination and spreading of colonies.

4.

Pour medium quickly to avoid contamination by air-pores and close lid down as soon as possible.

5.

Perform the pouring of medium in inoculation chamber fitted with U. V. lamp with filtered air.

6.

Pouring should be performed near the flame.

Observation 

After sterilization of medium observe the medium in conical flask and plate for solidification.



After incubation period of 24-48 hrs for nutrient agar medium and 7 days for PDA observe the growth of any microbe on the surface of the medium.

Exercise 1.

Prepare 500 ml nutrient agar medium.

2.

Prepare 500 ml potato dextrose agar medium

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Chapter-13

Methods of Application of Fungicides: Soil Application Purpose: To eradicate or reduce the inoculums density of soil-borne plant pathogens. Methods of soil treatments Soil drenching: In this method fungicides are mixed in water and about the same Concentration as for spraying and applied to the soil surface either before or after plants emerge. The required quantity of fungicide suspension is applied with a sprinkler or rose can per unit area so that the fungicide reaches a depth of at least 10-15 cm. This method is followed for controlling damping off. Root rots or infections at the ground level.

Quantity of fungicidal solution required for soil drenching a)

For Mini plots or shallow flats: 2.5L/m^2

b)

Pots up to 40 cm size and propagative beds up to 10 cm in depth: 3.0 1/m^2

c)

Pots and propagative beds greater than 10 cm in diameter or depth: 3.5 to 6.0 L/ m^2

Important fungicides used for soil drenching Fungicide Dosage (%) Pathogens controlled Bordeaux mixture 1 % Pythium, Phytophthora Cheshunt compound 0.3 % Pythium, Sclerotium.

Copper oxy chloride 0.2 to 0.3 % Pythium, Phytophthora Captan 0.2 to 0.3% Pythium, Phytophthora Captafol 0.3 % Macrophomina,Pythium, Rhizoctonia, Phytophthora Dexon 0. 1 to 0.2 % Pythium, Phytophthora Carbendazim 0. 1 5 % Fusarium, Rhizoctonia, Sclerotium Benomyl 0.15% Fusarium, Rhizoctonia, Sclerotium Fosetyl-AL 0.2% Phytophthora, Pythium Metalaxyl (Ridomil-MZ) 0.2 % Pythium, Phytophthora Vitavax 0.15% Fusarium, Rhizoctonia, Sclerotium 2.

Broadcast: In this method fungicides are mixed with soil or fertilisers, and scattered with hand as uniformly as possible over the-field and mixed with the soil with a suitable implement. This method involves greater quantity of fungicides than other methods.

For example: Benomyl @ 12-45 kg a.i. /ha for soil treatment by broadcast controls pea root rot and Verticillium wilt of potato.

3.

Furrow application Fungicides are applied either as dusts or mixed with water to the furrow at the time of planting

for control of diseases that occur at the base of the plant. This method 'requires’ much less quantity of fungicides per hectare than broadcast method.

63

For example Captan @ 25-30 kg/ha for furrow application for the control of onion smut Thiram @ 15-25kg/ha for furrow application for the control of smut and neck rot of onion.

4.

Fumigation: This method is usually restricted to small areas and high value crops. Fumigants are used to control soil-borne fungi and nematodes. These chemicals produce a gas that distributes itself through the soil. After application of chemicals to soil, it should be covered with athin polythene sheet for some time.

Fumigant Pathogens controlled Methyl bromide: Rhizoctonia, Nematodes, Pythium Vapam: Pythium and Nematodes Methyl isothiocyanate:Rhizoctonia Sodium azide,Potassium azide: Sclerotium rolfsii Nemagon Nematodes and Pythium Formaldehyd (37-40 % soln): Pythium and Rhizoctonia Chloropicrin: Pythium and Nematodes Precautions 1.

Soil should be in good planting conditions

2.

Soil should be moist enough to permit seed germination

3.

Manure, peat, compost and other humus material must be added before treatment, when using soil fumigants, fertilizers containing ammonia or ammonium salts should not be added to the soil at the time of treating.

4.

Planting should be done two to four weeks after the treatment.

64

Chapter-14

Methods of application of fungicides: Seed and foliar application a)

Seed treatment: Seeds, tubers, bulbs, setts and other propagating materials are given physical / chemical

treatment for eradication of pathogens present on them and for preventing their rot in the soil after planting.

Types of seed treatment: There are various types of seed treatment and the important ones are physical arid chemical methods.

Physical methods 1.

Mechanical separation: Healthy seed contaminated with ergot of rye, ergot of bajra, false smut of rice, ear cockle of wheat can be separated mechanically because the pathogen causes alteration in size and weight of the seed.

2.

Steeping the seed in Brine Solution: The diseased grains are removed by steeping the seed in the brine solution and the diseased grains float due to their lightweight. Examples: Ergot of rye, ear cockle of wheat.

3.

Hot water treatment: Jensen (1887) suggested this for control of loose smut of wheat. Hot water treatment aims at destroying the infection in the seed without harming the embryo.

Examples: Hot water treatment of seeds at 52°C for 10 min.: controls loose smut of wheat, Whip smut of sugarcane - hot water treatment of setts at 52°C for 30 minutes, Grassy shoot of sugarcane - hot water treatment of setts at 52°C for 30 minutes.

Diseases controlled by Hot Water Treatment Crop Disease Temperature 

Sesamum Bacterial leaf spot Treat seeds at 44°C / 10 mts.



Mustard Alternaria Treat seeds at 50°C / 30 mts.



Potato Virus infection Tuber treatment at 50-56°C for 15-20 mts(or) 50°Cfor 10-12 mts.



Cotton Bacterial blight(black arm) Seed treatment at 52-56°C/10 to 15 mts



Sugarcane Red rot Sett treatment at 52°C / 30 mts.



Whip smut Sett treatment at 52°C / 30 mts.



Grassy shoot Sett treatment at 50°C / 30 mts

Solar treatment: It is a safe and convenient method than hot water treatment. J.C. Luthra (1931) suggested this method for control of loose smut of wheat. Wheat seed is pre soaked in water for 4-5 h in the shade or in a room and then dried on ground/concrete floor in a thin layer in sun for 1 hr usually at noon.

65

Chemical Methods Fungicidal seed treatment: The treatment of seed with fungicides is commonly called seed dressing and the fungicide used is known as Seed Dressing Agent (or) Seed Dresser. Fungicidal seed treatment not only kills the pathogen present on / in the seed but also protects the germinating seeds from other soil-borne pathogens till they become established into young plants. Fungicidal seed treatment methods include wet seed dressing and dry seed dressing.

Wet seed dressing a)

Seed dip method: Dipping the seed or seed materials in fungicidal solution for 5-20 minutes and drying them in shade before sowing. This method is particularly useful to check externallyseed borne diseases.

Fungicides / Bactericides used: Captan, carbendazim, Agrimycin-100 and 500 Aureofungin.

Concentration: 0.1 to 0.5 %( 2g/kg seed / 2 1 of water) Diseases controlled by wet seed treatment Diseases controlled by wet seed treatment Crop Disease Procedure Sugarcane: Red rot, Whip smut, Pine apple rot; Dipping of setts in carbendazim (5 g/10 litres) solution for 15 minutes Turmeric Rhizome rot; Dipping of rhizomes in 0.3 % mancozeb solution for 30 minutes (3 g/lt.)

Ginger Rhizome rot; Dipping of rhizome in 0.2 % mancozeb solution for 2 hr (2 g/lt.)

Betelvine Stem rot, Leaf spot; Dipping of vines in 0.5 % Bordeaux mixture + 500 ppm streptocycline solution for 30 mts.

Chilli Virus infection; Dipping of seeds in Trisodium orthophosphate solution (90 g/lt.) for 15 mts.

Sesamum Bacterial leaf spot; Dipping of seeds in 200 ppm streptomycin sulphate or 250 ppm, agrimycin solution for 1 hour.

Beans Bacterial leaf spot; Dipping of seeds in 500 ppm streptomycin solution for 1 hour.

Cotton Black arm; Dipping of seeds in 1000 ppm streptomycin sulphate Rice Bacterial leaf blight; Dipping of seeds in 250 ppm agrimycin solution for12 Hours Cabbage and Cauliflower Black rot; Dipping of seeds in 100 ppm agrimycin or streptomycin solution for 30 minutes b)

Slurry treatment: The seed is mixed with a dust fungicide in a special treater (Slurry treater) in which small calibrated amounts of concentrated liquid (about 5-20 ml/kg seed) are added, thus forming a soap-like slurry to ensure coating without undue wetting. This treatment is common in almost all seed processing plants owned by Government as well as private producers.

Fungicides used: Thiram, Captan. c)

Sprinkle treatment: The seed is sprinkled with a fungicidal liquid, solution or suspension, left damp with this for a definite period of tune and then dried. This method is widely used in countries like USA, Europe and UK.

66

Dry seed dressing: This method is simple and economical. The method consists of adding the required fungicide (usually 0.3 % to the seeds and shaking them in a closed vessel or a 'rotary drum' for 5-15'minutes to facilitate even spreading of fungicides over the surface of all seeds.

Commonly used seed dressers Sulphur group Sulphur dust (5 g / kg seed) Organic fungicides Mancozeb, Thiride 75 SD, Captan 75 SD Systemic fungicides Vitavax, Plantvax, Benomyl, Carbendazim, Ridomil (Metalaxyl), Tricyclazole.

Diseases controlled by dry seed treatment Crop

Fungicides

Diseases

Rice

Tricyclazole,Thiram,Captan, Carbendazim

Brown spot and blast

Sorghum and Pearl millet Thiram,Captan

Grain moulds and smuts

Maize

Thiram

Grain moulds, wilt, stalk rots, smut

Groundnut

Captan

Afla root, stem rot

Castor

Captan

Seedling blight and wilt

Sunflower

Thiram, Mancozeb,Metalaxyl

Altemaria blight, Sclerotium wilt, Downy mildew

Tobacco

Thiram, Metalaxyl

Damping-off in nurseries

Chilli

Captan,Thiram

Damping-off, Die-back and fruit rot

Safflower

Thiram

Wilt and rust

67

Chapter-15

Major diseases of crops in Jammu region Followed with the Photographs‌

68

69

70

Chapter-16

Identification of weeds in crops Objective 1)

To appraise the students regarding common weeds of the locality.

2)

To impart the information regarding preparation of weed herbarium.

Materials required 1)

Knife/Scalpel

2)

Plant press

3)

Scrap book/ Herbarium sheets

4)

Glue/Gum/Tape

5)

Newspaper

Procedure Herbarium/ Weed Collection 1)

Collect the weed plant from the field

2)

Maintain all the plant parts intact (leaf, stem, flowers, fruits)

3)

Collect fresh part of the plant

Herbarium pressing and drying 1)

The weed plant should be pressed with wooden press board.

2)

Collected specimen should be placed in between the folds of newspaper for blotting. Ensure that plant is intact.

3)

Ensure that the specimen is spread properly in the newspaper folds.

4)

The large specimens are to be cut in parts according to convenience and the cut parts are to be arranged on the same sheet.

5)

If the foliage is very thick, it has to be pruned assuring that the portion of the cut parts are identified.

6)

The newspaper should be changed after 12 hours in first instance and there after 24 hours, 48 hours and 72 hours is done till the specimen is dried completely. This is known as natural drying.

7)

In artificial drying, after sweating period, specimens can be dried in hot air oven by maintaining 62oC temperature.

Herbarium mounting 1)

Thick herbarium sheets are used for mounting

2)

Keep the specimen in the centre and spread properly.

3)

Fix the specimen to the mounting sheet with glue/gum/tape

71

Herbarium labeling 1)

Label the specimen in the space provided on the lower side of the herbarium sheets.

2)

The label information should contain scientific name, local name and habitat.

Table 1: List of kharif season weeds Sl.No. Scientific Name

Local Name

Family

Habitat

1.

Ammania baccifera

Monarch red stem

Lythraceae

A

2.

Amaranthus viridis

Amaranthus

Amarthanceae

A

3.

Cleosia argentia

Cox comb

Asteraceae

A

4.

Commelina benghalensis

Day Flower

Commelinaceae

A

5.

Cynodon dactylon

Bermuda grass

Gramineae

P

6.

Cyperus rotundus

Purple nut sedge

Cyperaceae

P

7.

Cyperus difformis

Small flowered nut sedge

Cyperaceae

P

8.

Cyperus iria

Rice flat sedge

Cyperaceae

P

9.

Dactyloctenium aegyptium

Crow foot grass

Gramineae

A

10

Echinochloa crusgalli

Barnyard grass

Gramineae

A

12

Echinochloa colonum

Jungle rice

Gramineae

A

13

Eicchornia crassipes

Water hycainth

Ponderiaceae

P

14

Eleusine indica

Goose grass

Gramineae

A

15

Imperata cylindrical

Thatch grass

Gramineae

P

16

Panicm repens

Torpedo grass

Gramineae

P

17

Parthenium hysterophorus

Congress grass

Compositae

A

18

Physalis minima

Sunberry

Solanceae

A

19

Solnum nigrum

Black night shade

Solanceae

A

20

Sorghum halpense

Johnson grass

Gramineae

P

21

Setaria glauca

Cat tail millet

Gramineae

A

22

Tridax procumbens

Coat button

Compositae

A

23

Trianthema monogyna

Horse purslane

Aizoaceae

A

Common cattail

Typhaceae

A

24 Typha latifolia *A- Annual, P-Perennial Table 2: List of Rabi season weeds Sl.No.

Scientific Name

Local Name

Family

Habitat

1

Anagallis arvensis

Krishna neel

Primulaceae

A

2

Argemone maxicana

Satyanashi

Papavaraceae

A

3

Avena fatua

Jangli jayee

Gramineae

A

4

Asphodelus tenuifolius

Pyazi

Liliaceae

A

5

Chenopodium album

Bathua

Chenopodiaceae

A

6

Cirsium arvensis

Kateli

Compositae

P

7

Chichorium intybus

Kasani

Asteraceae

A

8

Convolvulus arvensis

Hiran Khuri

Convolvulaceae

A

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10

Cuscuta chinesis

Amar bel

Convolvulaceae

A

11

Fumari parviflora

Gajri

Papaveraceae

A

12

Lathyrus aphaca

Chatri matri

Leguminoceae

A

13

Medicago denticulata

Maina

Leguminoceae

A

14

Malwa parviflora

Sonchal

Malvaceae

A

15

Melilotus indica

Yellow Senji

Leguminoceae

A

16

Melilotus alba

White Senji

Leguminoceae

A

17

Phalaris minor

Gulli danda

Gramineae

A

18

Portulaca oleracea

Jangli Palak/Qualfa

Portulacaceae

A

19

Poa annua

Bueen

Gramineae

A

20

Pluchea lanceolata

Rasna

Asteraceae

A

21

Ranunculus repens

Chambl

Ranunculaceae

A

22

Rumex spinosus

Jangli palak

Polygonaceae

A

23

Saccharum spontaneum

Kans

Gramineae

P

Akari

Leguminoseae

A

24 Vicia hirsuta *A- Annual, P-Perennial, B-Biennial

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75

76

77

78

Chapter-17

Methods of Herbicide Application

H

erbicides are applied either to the soil (soil application) or to the foliage (foliar application) to kill the herbaceous plants. Depending on the properties of the herbicides, mode of action and

selectivity, different methods are adopted. Environmental factors, convenience and cost are other factors that influence the choice of correct method of application. An improper method of application can result in poor weed control and/or severe crop injury.

I)

Soil Application

a)

Soil Surface Application: Herbicides are sprayed on the soil surface to form a uniform herbicide layer. The herbicide thus sprayed, due to their low solubility may penetrate only few centimeters into the soil. Weeds germinating in the top layer are killed due to incidental absorption of herbicides. Most of the triazines, ureas and amides are applied by this method. The herbicides for surface soil application has to be soil active less soluble and less volatile. After application, the surface soil should not be disturbed.

b)

Soil Incorporation: Some of the herbicides belonging to aniline and carbamates groups are volatile. When they are applied to the soil surface, they are lost by volatilization. When applied to soil surface a third of flucholarlin is lost within three days. Volatile herbicides are incorporated into the soil to reduce losses. Generally, these herbicides are applied before planting as it is difficult to incorporate the herbicide after sowing.

c)

Sub-Surface Application: When perennial weds like Cyperus rotundus and Cynodon dactylon are to be controlled, the herbicides are applied to the lower layers of the soil. This is done by injecting herbicides into the soil at several points.

d)

Band Application: Herbicides are applied as narrow bands over or along the crop row. The weeds in between the rows can be controlled by intercultivation or by non-selective herbicide.

This method is useful where labour is expensive and intercultivation is possible. Weeds in maize can be controlled effectively by spraying atrazine on seed row at the time of sowing and nonselective herbicides in between the rows after weeds have emerged.

II) Foliar Application a)

Blanket Application: It is application of herbicides over the entire leaf area. Only selective herbicides are applied by this method.

b)

Directed Application: Application of herbicides in between the crop rows directing towards weeds is known as directed spray or directed application. Care is taken to avoid spray fluid falling on the crop.

c)

Spot Application: Herbicide solution is poured on weeds in cropped and uncropped fields infested with obnoxious weeds in isolated patches. This method of application of heavy doses of herbicides in isolated patches is called spot application

79

d)

Basal Application: Brush-wood and unwanted trees are treated with herbicides by different methods depending on the situation. Generally, foliar application is the most common method of treating brush. However, if the trees are taller say more than 2.5 m, it is not possible by ground sprays. Under such situation, basal bark application is resorted to. The bark of the trees at the base of the stem upto 30cm height is removed and drenching spray of herbicide is given to the base.

e)

Cut-Surface Application: The most difficult brush weeds or trees are cut to the ground surface, concentrated herbicides or paste of herbicides is applied to the stump or cut surface.

Computation of herbicide dose

Rate of application: It is the amount of active ingredient or acid equivalent of herbicide applied to a unit area of land or water body. It is usually given in terms of kg a.i. or a.e./ha Active ingredient (a.i.): A chemical or commercial product that is directly responsible for its herbicidal effect is called active ingredient. The ingredient may be as per cent by weight or volume.

For example Herbicide concentrate 500 w/v contains 500g of active ingredient per liter of the liquid product.

1)

For Crops

Recommended dose in kg a.i. or a.e./ha x 100 Dose of commercial product (kg/ha) = ...........................................................................

Percent concentration in the product Acid Equivalent: It refers to that part of a formulation that theoretically can be converted to the acid.

Some herbicide structures are active organic acids and some are not dissolved in water for example phenoxyalkonic acids. They are prepared in the form of their salts and esters for the ease of their field preparation. For instance 2,4-D in acid form is water insoluble then we have to use its sodium and amine salts and esters. The acid equivalent of sodium salt of 2,4-D is 92.5%. It indicates 2, 4-dichloro phenoxy acetic acid is 92.5% sodium salt of 2, 4-D. Liquid formulation may include both per cent active ingredient and acid equivalent as weight per liter. In such cases, the concentration in terms of acid equivalent may be considered. A commercial formulation of 2,4-D containing 580 g of diethanolamine salt per litre would have a concentration of a.i. 58 per cent, but the concentration will be 58 x Molecular weight of 2,4-D acid= 58 x 221 = 39.32 per cent Molecular weight of 2,4-diethanolamine 326 The acid equivalent of a concentrate is always less or more than its content of active ingredient.

2)

For aquatic weeds For controlling aquatic weeds, herbicides are applied on weight basis (of water body) Weight of water (in lbs) = Volume x 62.3 Volume= L x B x D

80

Where L=Length of water body in feet B=Breadth of water body in feet D=Average depth of water body in feet One cubic feet of water weight=62.3 lbs.

Weight of water (lbs) x Recommended dose of herbicide (ppm) lbs of herbicides in a.i./a.e./ha= .............................................................................................................

1000,000 Numerical Problems Problem 1: Calculate the amount of commercial product (Stomp 30 EC) of pre-emergence herbicide pendimethalin required for 6000 m2 area to control weeds in wheat. The recommended rate of application is 1 kg/ha.

Solution: Recommended dose x 100 Dose of commercial product (kg/ha)= ................................................ a.i.

1_ x 100=3.33 kg/ha Dose of commercial product (kg/ha) = ...........................................

30 1 hectare=10,000m2 10,000 m2 require pendimethalin= 3.33 3.33 1m2 require pendimethalin= .......................

10,000 6000m2 require pendimethalin= 3.33____ x 6000 = 1.99 kg 10,000 Hence, pendimethalin require for 6000m2 is 1.99 kg Problem 2: Calculate the amount of commercial product of (Satrix 10% SL) post-emergence herbicide imazathapyr required for 5000 m2 area to control weeds in chickpea. The recommended rate of application is 75g/ha.

Solution: Recommended dose x 100 Dose of commercial product (kg/ha) = ................................................ a.i.

75_ x 100=750g/ha Dose of commercial product (kg/ha) = ....................................

10 2 1 hectare=10,000m 10,000 m2 require imazethapyr = 750 1 m2 require imazethapyr = 750 10,000

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6000m2 require imazethapyr = 750 -------------- x 5000 = 375g 10,000 Hence, Imazethapyr require for 5000 m2 is 375g Conversion of Field measurement into different units 1. 1hectare = 10,000 square meter = 2.5 acres = 20 kanals = 400 marlas 2. 1 acre = 4000 square meter = 8kanals = 160 marlas 3. 1 kanal = 500 square meter =20 marlas 4. 1 marla =25.29 square meter=272.25 square feet (161/2ft x 161/2ft) General conversion factors Sl.No. 1 2 3 4 5 6 7 8 9 10 11 12 13 14 14 15 16 17 18 19 20 21 22

To convert Acre Acre Centimeter Foot Square foot,ft2 Cubic foot,ft3 Gram/ cubic centimeter Gram/cm3 Hectare Hectare Inch Inch2 Inch3 Kilogram Meter Meter Meter Meter2 Meter3 Meter2 Mile Quintals Tons

To Square meter, m2 Square Yard,yd2 Inch Milimeter Square meter, m2 Litres Pounds/feet3 Kg/litre Acre Meter2 Centimeter Cm2 Cm3 Gram Centimeter Foot Yard Foot2 Foot3 Yard2 Kilometer Kilogram Quintals

Multiply by 4047 4840 0.393 304.8 0.093 28.3 62.428 1.0 2.47 10,000 2.54 6.45 16.39 1000 100 3.28 1.09 10.76 35.31 1.31 1.61 100 10

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Chapter-18

Nutrieents Defficiency y Manag gement Nutrien nts deficiencyy

S

oil conditions greatly g influeence the avaailability of mineral m elem ments. Underr field condittions, the conncentration of o mineral nutrients, n orgganic matterr, lime, magnesia, phospphorus, soil reaction,

presencee of reducingg agents, andd oxidation-rreduction reaactions in thee soil determ mine the deficciency or responsees to these elements. e Deeficiency or unavailabilitty of mineraal elements iin soil condiitions are broadly categorized into two brooad and briefl fly described belowP M , Sulphur) A. Macrronutrients (Nitrogen, Phosphorus , Potassium, Calcium, Magnesium, B. Micrronutrients (Boron, ( Zincc, Copper, Iron, I Magneese, Molybdeenum)

Nitrogen Nittrogen deficiiency occurs on all classees of soil, altthough, of coourse, some ssoils are less prone to the deficciency than others. o Lightt sands lackinng organic matter m are perrhaps the pooorest of all as a regards nitrogenn. The nitrogeen supplies of o soils are greatly g affectted by croppiing and mannagement. So oils under grass annd leguminouus covers geet enriched with w nitrogen n, and whenn these are pploughed under, they generally yield a fluush of nitroggen to the suubsequent cro ops. The conntinuous croopping of araable soils with noon leguminouus crops greeatly depletees the nitrog gen supply, and even tthe continuo ous clean cultivatiion of a soill without croopping impooverishes thee soil of nitrrogen by stim mulating nitrrification processees and destrooying the orgganic matter. Under thesee, the nitrate, which is forrmed, is lost from the soil by leaching.

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Examplle: When fruuit trees, afteer growing under u clean cultivated coonditions forr a number of years, have eveentually sufffered severe nitrogen n starrvation due to o destructionn of or matterr.

Deficien ncy: Restriccted shoot growth g with yellowish green coolour of the foliage, thinn and short rooots, angle betweenn stem and leeaf is reduceed and so pllants looks erect, latteral shoots fail to develoop and lateraal bud may die or remain dorm mant. In accute deficienncy cases, leaves are a much sm maller and deevelop highlyy coloured tints of yellow, oraange or red; first appearr on older a proceedss towards thhe younger leeaves, preleaves and mature defoliation and supprressed lateraal growth, n defficiency always look flowerinng and fruitiing is greatlly reduced; plants suffeering from nitrogen conspicuuously thin and a poorly fuurnished.

Diseasess: In the t case of nitrogen n deficciency, plantts look so sicckly and consspicuously ppale that the condition c is calledd “general sttarvation�. This T is true foor all the speecies of crop plants.

Phosphoorus Phoosphorus is present p in sooils in a variety of formss mainly com mpounds of ccalcium, mag gnesium, in combination with organic mattter, compouunds of iron, aluminum annd phosphoruus present in n the pare rock. Onnly a small percentage p off the total phhosphorus preesent in the soil s is availabble to plants during a crop seeason. The most availlable soil phosphorus p compounds

are

t those

co-taaming

calccium

and

phosphoorus in the form of moonocalcium phosphate (water soluble) or dicalcium phosphate. The least availablee forms a thhe compoundds of phosphhorus with iron andd aluminum and a the comppounds of paarent rock.

Deficien ncy: Phosphoorus deficiennt plants in many m ways are simiilar to thosee caused byy nitro deficciency, but differ inn that the leaaves are gen nerally dulll bluish green in colour which usuually develop p purple tints ratther than yelllow or red or they mayy turn into dull d bronzingg with purple or brown spotting; scorchinng on leaf margins m mayy also appearr; retarded growth g of toop and rootss, smaller leaaves and prematuure defoliation occurs in the t same wayy as in the caase of nitrogeen deficiencyy.

Diseasess: In some speciees of plantss, phosphoruus deficiency y induces chhlorosis adjacent to maain veins with those arrising out followedd by leaf asyymmetry. Thiis condition is termed sicckle leaf andd resembles w due to ziinc deficienccy.

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Potassium Potassium is widely distributed in the earth's crust as a constituent of soil minerals which is slowly rendered available in water soluble form. Heavy soils are generally richer in potassium than light soils. Potassium is fairly well fixed in soils which is related to the Mount of colloidal matter present therein. Fixation of potassium is important factor as it controls rate of solution and consequent leaching and moreover, regulates the supply of available potassium to the plants. Potassium salts are more easily fixed than salts carrying nitrogen, but less so than the phosphates. Although the potassium salts dissolve readily in the soil moisture, they are soon taken out into less soluble forms as they unite with the colloidal complex and replace calcium or some other elements associated with the very finest soil particles. Potassium so held move slowly in the soil, at a rate depending on the amount and nature of the colloidal complex. In the lighter soils, the movement may eventually result in loss of potassium but in soil having a high clay and silt content, the movement is slow and often limited to the tipper few inches of soil.

Deficiency: Deficiency of this element is characterized by retarded growth, suppression of terminal shoot growth, and in acute cases, shoot may die back; leaves are dull bluish green with interveinal chlorotic areas; browning of the tips; marginal scorching with numerous brown spots near the margins. In broad leaved plants, deficiency results in upward or downward curling of leaves; scorched leaves often turn forward; shorter internode a and poor root system is the chief symptom of potassium deficiency.

Diseases: In beet, celery and carrot, bushy growth identical to "rosette" condition develops due to potassium deficiency. Such effects are also visible in pea, cereals and potato.

Acute deficiency of potassium causes loss of apical dominance and regeneration of many laterals and bushy growth. Prolonged deficiency results in the "die back" of laterals resulting from the loss of apical dominance.

Calcium Calcium is present in soils in a variety of combinations.

It is easily leached from most soils and especially from sand.

Clay soils often have only poor reserves of the element.

Calcium does not move freely from the older to the younger parts of plants; young tissues contain lower concentrations than older ones. Calcium deficiency occurs more frequently on acid soils than in neutral soils.

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Deficiency: Symptoms of calcium deficiency first appear in the young leaves, growing points of stem and in the roots. The young eaves are distorted, tips are hooked back and margins curled backward or forward, irregular and ragged leaf margins with brown scorching or spotting effects or cholorotic marginal bands. Poor root system which appears gelatinous.

Diseases: In cauliflower, beet and tobacco, calcium deficiency produces characteristic "hooking" of the leaf tip. This, condition arises because of differential growth in marginal and central regions of the leaf causing strain and ultimately resulting in the change of leaf shape.

Blossom end rot: This disease is very common in tomato. Symptoms include a depressed region near the distal end of the youngest fruit. The depressed portion is surrounded by dark green tissues and flesh is orange coloured.

Magnesium The mode of occurrence of magnesium in soils is fairly similar to that of calcium. It occurs as carbonate and in a variety of minerals, it is readily brought into the soil solution from the carbonate,

and

is

held

in

soils

as

an

exchangeable base. It is somewhat easily leached, and for this reason may become deficient in sandy soils during wet period. Large part of soil magnesium is found in the colloidal fraction and only 1 to 1/5 of this is in the exchangeable or soluble form. Magnesium is easily leached from soils, especially from the lighter soils, during wet periods.

Deficiency: Symptoms of magnesium deficiency are more pronounced on older leaves and proceed towards younger ones. Chlorosis of leaves having brilliant tints, veins remaining green and in the case of severe deficiency necrosis occurs. Premature shedder and withering of leaves is also common. Leaf mottling and defoliation may be extremely severe. The veins are bordered by a pale green region which encloses the pigmented spot. Later the leaves exhibit yellow-green interaveinal chlorosis and profuse necrotic spotting.

Diseases: Sand-drown: In tobacco, magnesium deficiency causes a disease designated as "sand drown". Lower leaves lose their colour at the tips and between the veins. The veins remain green, but in acute cases the leaf becomes nearly white. This in turn results in reduced elasticity leaves and irregular colouring which decreases the trade value.

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Sulphurr In soils, s sulphuur occurs in both b organic and inorganiic forms. A large l part of soil sulphur seems to be contaained in the organic o matteer while, the inorganic su ulphur is pressent chiefly aas sulphates.. The end product of oxidationn processes within w the sooil and the co ombination of o sulphuric acid with baases such m, and amm monium. Wheen sulphur is applied to thhe soil, it maay be rapidly oxidised as calciuum, potassium to sulphhuric acid. Thhis reaction is i commonlyy used to low wer the pH value v of alkaaline soils as sulphate element moves easily through the t soil. Thee sulphate iss not stronngly held byy the ordinarry clay of thee soil, but iss readily leached. l Deficien ncy: Sulphuur deficiencyy symptomss some way y resemblees those cauused by nitroogen deficienncy. Marked d decreasee in leaf sizee, general paaling with reed or purplee pigmenttation are the chief symptoms of sulphurr deprivedd plants. However, H symptoms of sulphurr deficiency appear on o younger leaves l first and a later on n older leeaves. In case c of sevvere sulphurr deficiency y necrosiss of young leeaf tips and margins, shorter interno odes, prematture developm ment of lateeral buds, multiplee branches with dead tips and shoot die back occu ur.

Diseasess: Tea yellow: Chloroosis starts froom leaf marrgins and sp preads inwardds. Under seevere deficieency, the entire pllant may turrn yellow leaading to prem mature leaf fall, f reducedd flowering aand fruiting. Number and size of leaves remains small..

Iron i may be due to the (i) lack of iro on, (ii) lime induced defi ficiency in caalcareous Deficiency of iron f other minerals, m and d (iv) deficieency from exxcess of pho osphorus, soils, (iiii) deficiencyy resulting from manganeese, zinc andd copper.

Lim me induced chlorosis c is associated a wiith high potaash and low calcium/pota c ash ratio or decreased d availablee iron. Carbbonate lime in soil is suuggested to react with w iron, foorming ferric carbonatee and thus reducingg the availabbility of iroon. Iron chloorosis may also be due to preseence of zinc in the nutriient media. a induce The preesence of biicarbonate irrons may also chlorosis. Bicarbonaate tends to decrease caalcium and increasee potassium content of chlorotic leeaves thus producinng the highh K:Ca ratioo familiar inn chlorotic foliage.

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Deficien ncy: The faillure of leavess to develop chlorphyll iss most charaacteristic effeect of iron deeficiency.

Youngesst leaf is affeected first whhich may beccome compleetely chlorottic. In most pplants initial chlorosis developss in interaveeinal tissues adjacent a to all a veins and produces a fine reticulate pattern. In n cereals, this connsists of brigght yellow-grreen or yelloow chlorosiss in longituddinal strips w which finally y become ivory in colour.

Diseasess: Lime in nduced chlorrosis: Excesss of lime toggether with higher h pH of soil causess general chllorosis of leaves reesulting from m iron deficiiency and is known as "lime inducedd chlorosis".. The younger leaves are moree severely affected than older ones, sometimes becoming b coompletely whhite or yello ow white.

Fruit treees, evergreens and decidduous are moore susceptib ble to iron deeficiency thaan the field crops, c but it may occur o in brasssicae, beet, sppinach and cereals. c

Mangan nese Tottal quantity of manganeese present in the soil iss of little siggnificance; qquantity abso orbed by plants iss more impoortant. The availability of manganeese to plantss from soils,, largely dep pends on nature of o salt soluttion, pH, tyype of reduccing agents, amount and nature of o higher oxiides of mangganese, lime G it and orgganic matterr present in the soil. Generally becomess unavailablle when the soil is richh in organic matter and a soil reacctions from weakly acidd to weakly alkaline. Applicatioon of calciium carbonnate greatly reduces its availabiility and abssorption. Waater soluble s sollubility of exxchangeable calcium in the soil supresses i deficient. manganeese. In humuus rich soils, manganese is

88

Deficiency: The younger leaves are principally affected, but in some crop older leaves display the initial symptoms of manganese deficiency. Leaves become chlorotic leaving midrib and principal veins. This is followed by profuse small dark brown necrotic spots. Leaflets of very young leaves become totally chlorotic. The interveinal chlorotic area finally show total bleaching associated with fine brown or purple tinted necrotic spots near major veins. In some crops, manganese deficiency cause chocolatee brown lesions, streaks or speck. These spots begin between veins near the base which collapses and form specks moving towards the tip.

Diseases: Grey speck: This disease is also called grey stripe, grey spot or dry spot and is caused by manganese deficiency. Oats, barley, wheat, rye and maize display symptoms of grey speck which is characterized by the appearance of greyish spot, small chlorotic areas, chiefly in the lower half of the leaf. These coalesce together and form elongated streaks which finally turn brown and occurs most y in third or fourth leaf.

Pahla blight of sugarcane: The disease is characterized by chlorotic areas forming a long white streak confined to leaf blade leaving the sheaths. The youngest leaves are most affected. With the death of the chlorotic cells, red spots coalesce resulting in continuous red streaks which may follow splitting of the leaf along the line of the streak.

Marsh spot of pea: The disease occurs on the seeds in the pod and is characterized by brown, black spots or cavities on the internal surface of the cotyledons. Sometimes plumule may also be affected.

Foliage symptoms may or may not appear. qI fight paling of leaves and small nectrotic areas may appear in acute deficiency.

Speckled yellows of sugarbeet: This disease resulting from manganese deficiency involves interveinal chlorosis in the leaves. The margins of the affected leaves curl upwards over the upper surfaces of the leaves.

Boron Soil types differ greatly in respect of boron deficiency. Highly leached soils, old soils or soils derived from igneous rocks are generally deficient in boron content. Medium-textured sandy soils have a lower optimum range of boron than clay. Continuous cropping, leaching and soil exhaustion resulting from intensive cultivation, application of organic manures all regulate boron response. Boron deficiency occurs at pH 7 and becomes more severe at high pH. Excessive limestone in sandy soil induces boron deficiency.

Deficiency: Death or abnormal growth of the stem apical meristem is the chief symptom of boron starvation, which results in regeneration of lateral buds producing a bushy habit. This is preceeded by

89

crinklingg of veins, leathery andd rudimentryy leaves wh hich are disttorted in shaape. Leaves become abnormaally turgid, brittle b and roolled. Stem and petioless also becom me brittle; pooor root deveelopment with shoort thick secoondary root growth g follow wed by crack king and deccay. Flowerinng is reduced d with increased sterility and poor bearingg are the chieef characterisstics.

Diseasess: Heart rot r of Sugarbeet and Mangold: M Thee disease is also a known as a heart rot, crown rot orr dry rot.

The most prominennt symptoms are the neccrosis of tissue of interioor root. The youngest leeaves are markedlly curled, veiins becomingg yellow andd the petioless are brittle. The main grrowing pointt dies and new shooots developiing in the axiils of the leavves are affeccted later in thhe same wayy as the main n shoot.

Canker and intern nal black sp pot of gardeen beet: The prominentt symptom oof this diseaase is the ment of inteernal hard black b necrottic masses, irregular in size and shhape which may be developm confinedd to the centrral region or scattered thee root. Micro oorganism laater attack thee root causin ng canker and renddering it unfiit for use.

Brownin ng of Cauliflower: Thee characteristtic symptom ms of this dissease is the aappearance of o "water soaked" areas on thee developingg curd whichh turn brown and becomee hard. The laves becomee thicker, brittle, curl c downwarrd and blisteering may occcur on the peetiole and aloong the midrrib.

Yellow top of Luceerne: Boron deficiency causes c yellow w top in luceerne. The paathological sy ymptoms y orr interveningg bronzing, shorter interrnodes follow wed by deatth of the include a uniform yellowing growingg points.

Top sick kness of tob bacco: The first f externall symptom of o this diseasse is the palee green colour of the terminall leaves withh bases afterr than the tiips. The bassal tissue off young leavves breaks down d and buddies.. The older leaves becoome thicker and brittle, midrib mayy break and upper laves tends.to droop.

Hard frruit of Citrus: Boron deficiency d inn citrus resu ults in the diie back of aapical growin ng point, reduced flowering annd shedding of fruits. Thhe fruits are badly shapedd, thick skinnned and imp pregnated wn as "hard fruit". irregularrly by m arouund the centrral axis, know Zinc Soiil reaction veery often altters the uptakke of zinc by plantts. Zinc defi ficiencies aree commonlyy observed on soilss with pH above 6.0 Effect E of pH H on the availabillity of zinc is associated with the abiility of the plant rooots to absorbb or to transpport to the other o plant organs the t absorbedd ions or varriations in thhe stability of solubble and insoluuble organicc complexes of zinc or a changee in the solubbility of antaagonistic ionss.

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Deficiency: Symptoms usually appear both on younger and older Ieaves. Leaf size is considerably reduced with yellow mottling and rosette formation. Leaf margins are distorted become twisted or wavy which later curl and Look 'sickle shape". Leaf colour become dull yellow, or ivory with interveinal mottling as the Chief Characteristics symptom of zinc deficiency. Interveinal tissues develop necrotic areas which are' irregularly arranged, eventually resulting in breakdown of the tissues. Seed production and fruit size is greatly reduced in zinc deficient plants.

Diseases: Khaira of paddy: The young seedlings of paddy first show the sign of mild chlorosis embeded with small brown spots on the younger leaves. These spots coalesce forming bigger ones and the entire looks brown in colour and ultimately die.

White Bud of Maize: The characteristic symptoms of this disease occuring due to dine deficiency is the appearance of Light yellow streaks between the veins of older leaves followed by the rapid development of white necrotic spots. Unfolding newer leaves are often pale yellow to white in colour.

Rosette of fruit trees: The disease is also known as "little leaf". The most characteristic symptom is the yellow mottling of the leaves and the reduction in leaf size with the format to of rosette like appearance. As these symptoms proceed, newer leaves become smaller, elongated, yellow mottled and de-shaped ultimately resulting in the die back of the affected branches.

Frenching of Citrus: The first symptom of this disease is the appearance of yellow areas between the veins; leaves are progressively smaller and develop chlorophyll only at the basal end of the midrib. In extreme cases die-back may follow.

Copper Copper occurs in the form of organic or inorganic ions of soils. Soils containing large quantity of organic matter are generally rich in copper content. Copper deficiency depends not only due to low copper content, but also on its availability. In general, when the percentage of mobile copper goes below fifty per cent of total copper content, deficiency symptoms set in. Humus compounds, reaction of top soil and underground drained laver, drainage and climatic conditions all determine the availability of copper in soil. Copper deficiency generally occurs in peaty soils, but it has also been noted on light sandy soils.

Deficiency: Symptoms of copper deficiency appear in young leaves which include necrosis and withering. Foliage is initially dark blue green in colour, but fading or chlorosis may follow. In severe cases of deficiency, leaves become limp and chlorotic which tightly curl. The tip of the younger emerging leaves is trapped within the rolled ones. Growing point is killed and produces excessive number of tillers or multiple bird formation is common in many plants.

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Diseases: Exanthema or die-back of fruit trees: This disease affects various fruit trees including citrus species, rosaceous trees, glum, apple and pear. The pathological symptoms include formation of strong water shoots bearing large leaves, gummous tissue or the bark and longitudinal breaks. Fruits are generally brownish, glossy and splitted. Affected shoots lose their leaves and die-back and lateral shoots produce typically bunchy appearance.

Reclamation disease: Pathological symptoms in cereals, oats, beet and leguminous crop plants due to deficiency of copper is known as reguminous disease or "white tip". In affected plants, the tips of the leaves become chlorotic followed by a failure of the plants to set seed.

Molybdenum In soils, molybdenum is present as soluble molybdate, components of soil organic matter and a fraction as exchangeable anion. Primary soils formed from such parent material having low molybdenum content are liable to cause deficiency. Mineral oils and coal ashes are rich source of molybdenum. Availability of molybdenum is greatly influenced by soil pH and unlike other elements it is more available at high pH than at lower pH. Deficiency of nitrogen gives poor response to molybdenum application, while iron and boron at too high or low level hinders molybdenum availability to plants. Lime and cobalt salts, on the other hand, enhances its availability.

Deficiency: The symptoms of molybdenum deficiency commences on older leaves and progresses towards the apex. Bright Yellow green or Zale orange interveinal mottling followed by marginal wilting and frequently cupping in broad leaved plants or various types of inrolling of lamina with water soaked appearance is the general characteristic of molybdenum deficiency. Severe deficiency results in the development of papery necrosis, mottling and withered condition. Flowering is suppressed, which sheds before opening.

Diseases: Whiptail of Brassica: Characteristic symptoms arising out due to molybdenum deficiency in brassies are grouped under the name "whiptail". The pathological symptoms commence as translucent oval areas near the midrib which become ivory tinted d necrotic. The leaf margins come ragged with upward curling. Leaf elongation and suppression of lamina tissue occur prior to death of the growing point causing a typical 'whiptail' condition.

Scald of legumes: In peas, lucerne, beans and allied species, molybdenum deficiency is reflected as leaf paling, wilting, marginal rolling and scorching. "Scald" disease of bean which is caused by molybdenum deficiency is greatly regulated by seed reserves of this element.

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Generalized Visual Leaf and Plant Nutrient Element Deficiency and Excess Symptoms Element/status Nitrogen (N) Deficiency

Excess

Ammonium toxicity

Phosphorus (P) Deficiency Excess

Potassium (K) Deficiency

Excess Calcium (Ca) Deficiency

Excess

Visual symptoms Light green leaf and plant color with the older leaves turning yellow, leaves that will eventually turn brown and die. Plant growth is slow, plants will be stunted, and will mature early.

Plants will be dark green in color and new growth will be succulent; susceptible if subjected to disease and insect infestation; and subjected to drought stress, plants will easily lodge. Blossom abortion and lack of fruit set will occur.

Plants fertilized with ammonium-nitrogen (NH4- N) may exhibit ammonium-toxicity symptoms, with carbohydrate depletion and reduced plant growth. Lesions may occur on plant stems, there may be a downward cupping of the leaves, and a decay of the conductive tissue at the base of the stem with wilting of the plants under moisture stress. Blossom-end rot of fruit will occur and Mg deficiency symptoms may also occur.

Plant growth will be slow and stunted, and the older leaves will have a purple coloration, particularly on the underside.

Phosphorus excess will not have a direct effect on the plant but may show visual deficiencies of Zn, Fe, and Mn. High P may also interfere with the normal Ca nutrition, with typical Ca deficiency symptoms occurring.

On the older leaves, the edges will look burned, a symptom known as scorch. Plants will easily lodge and be sensitive to disease infestation. Fruit and seed production will be impaired and of poor quality.

Plants will exhibit typical Mg, and possibly Ca deficiency symptoms due to a cation imbalance The growing tips of roots and leaves will turn brown and die. The edges of the leaves will look ragged as the edges of emerging leaves stick together. Fruit quality will be affected with the occurrence of blossom-end rot on fruits.

Plants may exhibit typical Mg deficiency symptoms, and when in high excess, K deficiency may also occur.

Magnesium (Mg) Older leaves will be yellow in color with interveinal chlorosis (yellowing between the Deficiency veins) symptoms. Plant growth will be slow and some plants may be easily infested by disease.

Results in a cation imbalance showing signs of either a Ca or K deficiency.

Excess Sulfur (S) A general overall light green color of the entire plant with the older leaves being light Deficiency green to yellow in color as the deficiency intensifies.

A premature senescence of leaves may occur.

Excess Boron (B) Abnormal development of the growing points (meristematic tissue) with the apical Deficiency growing points eventually becoming stunted and dying. Rowers and fruits will abort.

For some grain and fruit crops, yield and quality is significantly reduced.

Leaf tips and margins will turn brown and die.

Excess Chlorine (Cl) Younger leaves will be chlorotic and plants will easily wilt. For wheat, a plant disease Deficiency will infest the plant when Cl is deficient.

Premature yellowing of the lower leaves with burning of the leaf margins and tips. Leaf Excess abscission will occur and plants will easily wilt.

Copper (Cu) Plant growth will be slow and plants stunted with distortion of the young leaves and Deficiency death of the growing point.

An Fe deficiency may be induced with very slow growth. Roots may be stunted.

Excess Iron (Fe) Interveinal chlorosis will occur on the emerging and young leaves with eventual Deficiency bleaching of the new growth. When severe, the entire plant may be light green in color.

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Excess

A bronzing of leaves with tiny brown spots on the leaves, a typical symptom frequently occurring with rice.

Manganese (Mn) Interveinal chlorosis of young leaves while the leaves and plants remain generally green Deficiency in color. When severe, the plants will be stunted.

Older leaves will show brown spots surrounded by a chlorotic zone and circle.

Excess Molybdenum (Mo) Symptoms will frequently appear similar to N deficiency. Older and middle leaves Deficiency become chlorotic first, and In some instances, leaf margins are rolled and growth and flower formation are restricted.

Not of common occurrence.

Excess Zinc (Zn) Upper leaves will show interveinal chlorosis with an eventual whiting of the affected Deficiency leaves. Leaves may be small and distorted with a rosette form.

An Fe deficiency will develop.

Excess

Terminologies: 

Chlorosis is a physiological disorder that occurs to deficiency of mineral elements (eg; Mn, K, Zn, Fe, Mg, S and N). Leaves or plant parts become abnormally yellow.  Mottled is surface marked with coloured spots (anthocyanin develops) eg. Due to deficiency of N, Mg, P, S.  Necrosis refers to patch of dead tissues, due to the deficiency of Mg, K, Zn, Ca and Mo. (Source: TNAU-2016)

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95

Table: Nutrient deficiency of single elements that often occur because of an excess of other nutrients Visible deficiency

caused by an excess of

Potassium

Nitrogen, calcium, magnesium, sodium

Magnesium

Potassium and/or calcium

Nitrogen

Chloride

Sulfur

Chloride

Calcium

Magnesium, potassium

Boron

Calcium

Iron

Nitrogen, phosphorus, manganese, molybdenum, nickel, zinc, bicarbonate

Manganese

Phosphorus, iron

Copper

Phosphorus, molybdenum

Zinc

Phosphorus

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2020

Manual on

PLANT HEALTH CLINIC

© Dr. Uma Shankar 2017

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