www.ijcbs.org IJCBS RESEARCH PAPER VOL. 1 [ISSUE 1] April, 2014
ISSN:- 2349–2724
Antagonistic Activity of Trichoderma Spp, (A BioControl Agent) Against Isolated and Identified Plant Pathogens 1*Meenakshi
Kushwaha (Pursuing PhD) Biological Sciences Department National Botanical Research Institute Lucknow, India Email: meena.abc7@gmail.com
2Ajay
Kumar Verma (M.Tech.) Civil Engineering Department Madan Mohan Malveeya Engineering College Gorakhpur, India Email: ajaycrazy100@gmail.com
ABSTRACT: There is a great problem of farmers to protect their food crops from pathogenic microbes. Some fungi play a major role to have an effect on the crop’s yield and survival. On visiting farmer’s field in bakshi ka talab lucknow, some diseased crop plant and their fruits have been found. After screening the diseased area of sampled crop plants causal fungi get isolated and identified morphologically in the laboratory. The isolated and purified pathogenic fungi are Chollatotrichum from red rot of sugarcane, Alternaria from brown-black discoloration of cauliflower, early blight of tomato and black round spot of apple, Paracercospora from leaf spot disease of banana and Fusarium from wilt root of pegion pea and black margine of curled potato leaf. Some species of Trichoderma like T. hargianum, T. viride, T. hamantum have a potential to inhibit the growth of pathogenic fungi. Trichoderma had shown significant antagonistic activity against isolated pathogenic fungi Chollatotrichum, Alternaria, Paracercospora and Fusarium species. From this result we have found the trichoderma as a potential biocontrol agent against pathogenic fungi. Keywords: Trichoderma; antagonistic activity; fungal diseases of plants; Biological control; horticulture crop diseases; bio-control agent.
1. INTRODUCTION India with diverse soil and climate comprising several agro-ecological regions provides ample opportunity to grow a variety of horticulture crops. Cultivation of these crops is labor intensive and as such they generate lot of employment opportunities for the rural population. Fruits and vegetables are also rich source of vitamins, minerals, proteins, and carbohydrates etc. which are essential in human nutrition. Hence, these are referred to as protective foods and assumed great importance as nutritional security of the people. India is the second largest vegetable producer in the world, next only to China with an annual production of 81 million tons from 5.1 million hectares of land. However, the major factors responsible for low production of fruits, cucurbitaceous, brassicaceous and solanaceous vegetables are the diseases caused by Pathogenic fungi. The disease development is so fast that whole crop is lost in a few days. Therefore, the problem deserves immediate and effective measures of control (Neeraj, et.al.2010). Fruit plant diseases often are the worst natural hazards in horticulture. New diseases and new biotypes of existing disease-producing organisms appear from time to time in a more virulent form. With
the liberalization of import and export of large quantities of horticultural planting materials, the risk of spread of diseases into new areas has also increased. The most startling aspect of fruit plant diseases is that their control cost us a huge sum every year with serious environmental consequenes. Therefore, integrated disease management practices need to be adopted to reduce the losses Fruits and vegetables are highly perishable products, especially during the postharvest phase, when considerable. Losses due to microbiological diseases, disorders, transpiration and senescence can occur. A number of microorganisms, which effectively control postharvest pathogens, have been identified for post-harvest control (Wilson and Wisniewski, 1989). Biological control of plant pathogens by microorganisms has been considered a more natural and environmentally acceptable alternative to the existing chemical treatment methods (Baker and Paulitz, 1996). Trichoderma spp. is now the most common fungal biological control agents that have been comprehensively researched and deployed throughout the world. Several fungal cell wall
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IJCBS RESEARCH PAPER VOL. 1 [ISSUE 1] April, 2014
degrading enzymes, amongst them chitinase and glucanase, which seem to play an important role in the antagonistic action of Trichoderma against a wide range of fungal plant pathogens (Kucuk and Kivanc, 2008). T. harzianum shows antagonistic effects against, Alternaria spp. Colletotrichum spp.,Paracercospora spp., Fusarium spp. and, causative fungi of stem end rot, anthracnose and brown spot of rambutan respectively and also retained the overall quality and color of the fruits (Siva Kumar et al.,2000). The present study aimed to find out the efficiency of Trichoderma spp. against some post-harvest pathogens. MATERIALS AND METHODS
2.1 Survey and collection of variable diseased crop samples We have collected the following infected horticulture crop samples during the survey of farmer’s field at Bakshi Ka Talab, Lucknow and after collection we observed their symptom. In marigold leaf red & dry leaves, black margins on potato leaves, in pigeon pea root wilt root with black xylem, cauliflower with brown-black discoloration, in tomato fruit velvety green mat which is also known as early Blight of tomato, in banana leaves brown spots with yellow margin, black streaks & gray spot with black margin on different leaves of banana, leaf red thick streaks on sugarcane, which is also called red rot of sugarcane, apple fruit (post-harvest, from market) several black round spot with brown rot on side of tip fruit we have observed with naked eyes. These infected plant samples kept in sterile zipper lock plastic bags and carried to the laboratory where we have stored those samples at 4°C.
ISSN:- 2349–2724
for a while so as to kill all the microbes present over its surface, it is then dipped in ethanol and again placed over the flame, but for a short while. It is held for a few seconds to cool it down. Then sample was taken and the loop was scrapped on the infected part of the samples so as to pick a good amount of inoculums, inoculation done properly with the help of loop. After that the loop is taken out and the PDA plate is covered. The whole process of inoculation was repeated with each sample. The inoculated PDA plates were incubated in BOD at 26º C for 72 hrs. with one PDA plate without inoculum taking as control. Pure colonies are isolated from inoculated petri plates separately in aseptic condition. Pure cultures are made with the help of sterilized inoculation loop on PDA plate. The isolated pure cultures on separated PDA plates are then kept on BOD incubator for incubation at 26º C for 72 hrs. Observation was taken daily to record the growth of microbes. 2.3 Microscopic identification The pure colonies were picked and identified under high quality microscope.
2.2 Isolation and pure culture of pathogens Fungal pathogen was separated from infected area of plant sample and pure cultured on PDA plates in the laminar air flow. For preparation of 300 ml Synthetic PDA medium 39 gm. of PDA powder (Himedia laboratories Pvt. Ltd.) is used for making 1 ltr. of PDA medium as recommended. After autoclave at 121 °C for 15– 20 minutes, cool the media up to tolerable temperature then pouring was done into sterile petri dishes under the flaming area in the laminar air flow. The PDA plates were successfully prepared. Taking small part of the infected part of each sample separately, surface washing of inoculums with sterile water was done. Now in the laminar air flow, the inoculation loop is taken and kept on the flame
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IJCBS RESEARCH PAPER VOL. 1 [ISSUE 1] April, 2014
ISSN:- 2349–2724
Table No. 2
2.4 Management of isolated disease causing fungus Antagonistic activity We have performed the antagonistic activity of Trichoderma against Colletotrichum, Alternaria, Paracercospora & Fusarium. Switch on the UV of laminar air flow. After switching off, switch on the air flow. Take 24 sterile petriplates and pour the PDA medium in each petriplate and keep it for solidification. Keep 4 pour plates as control (pure media) for checking the medium and inoculate 8 petriplates with Trichoderma and Alternaria, Trichoderma and Colletotrichum, Trichoderma and Paracercospora, Trichoderma and Fusarium respectively which worked as control 2. In other 12 petriplates inoculate Trichoderma and just opposite of this inoculation, inoculate Alternaria Colletotrichum, Paracercospora & Fusarium, Replicate it three times. We Kept all the petriplates in the BOD incubator at 27oC for 96 hours and taken records after every 24 hours.
Table No. 3
Table No. 1
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IJCBS RESEARCH PAPER VOL. 1 [ISSUE 1] April, 2014
ISSN:- 2349–2724
3. RESULT The samples that we were collected from farmer’s field at BKT have following diseases which are wilt root (black xylem) of pigeon pea root, brown spots with yellow margin of banana leaf, bacterial watery marks of cauliflower, red leaves of marigold leaf, red rot of sugarcane of sugarcane leaf, black margins of potato leaf, black round spot of apple fruit, early blight of tomato. Table No. 5: Isolated fungi from each infected plant samples
Table No. 4
Trichoderma show strong antagonistic activity against Colletotrichum causing red rot of sugarcane, Alternaria spp. causing Alternaria leaf spot disease, Paracercospora spp. causing leaf spot disease of banana, Fusarium spp. causing wilt disease in pigeonpea.
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CONCLUSION Several strains of Trichoderma have been developed as biocontrol agents against fungal diseases of plants. The various mechanisms include antibiosis, parasitism, inducing host plant resistance and competition. Most bio control agents are from the species T. harzianum , T. viride and T. hamatun. The biocontrol agent generally grows in its natural habitat on the root surface and so affects root disease in particular but can also be affected against foliar diseases. REFERENCES 1. Neeraj and Shilpi Verma,(2010). FGC,Alternaria diseases of Vegetable Crops and New Approaches for its Control. ASIAN J. EXP. BIOL. SCI., VOl 1 (3):681-692 2. Rajendiran, R.1, Jegadeeshkumar, D.1, Sureshkumar, B.T.,2 and Nisha, T.26 (2010).In vitro assessment of antagonistic activity of Trichoderma viride against post-harvest pathogens. Journal of Agricultural Technology Vol.6(1): 31-35 3. Singh A, Islam MN (2010) In vitro evaluation of Trichoderma spp against Phytophthora nicotianae. Int. J. Expt. Agric. 1(1), 20-25.
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