Prevalence of ِAeromonas Species in fresh and marine water fish from the seafood market of Zagazig, Egypt with a Trial to improve their shelf life at refrigeration
Eldaly, E.A.; Saleh, E.A. and Abd ElEl- Hafeez, M.M. Food Control Dept., Faculty of Veterinary Medicine, Zagazig University, Zagazig, Egypt.
Abstract
A total number of 120 fishes samples of Tilapia nilotica, Cat and Carp fish which represent the fresh water fish (20 of each) and Blue spot fish, Saurus and Pagrus fish which represent the marine water fishes (20 of each) were collected from different seafood markets in Zagazig city. Collected samples were bacteriologically investigated to explore the Prevalence of ِAeromonas Species in fresh and marine water fish with a Trial to improve their shelf life at chill temperature. Organoleptic examinations declared that all the investigated fresh and marine fish samples were considered fresh and fit for human consumption. Meanwhile, the bacteriological examination revealed that the incidence of aeromonas species was 65% , 100% and100% in the examined samples of Tilapia nilotica , Cat and Carp fish with a mean value of 21.1x104 ±7.7x104 CFU/g, 36x106 ± 6.5x106 and 7.7x106 ± 1.2x106 CFU/g respectively and 90%, 40% and 75% in the examined samples of Blue spot fish, Saurus and Saurus with a mean value of 36.9x104 ± 0.95x105 CFU/g, 13x105 ± 6.5x105 CFU/g and 13.7x105 ± 2.7x105 CFU/g respectively. A. hydrophila, A. sobria and A. caviae was isolated and identified from the examined fish samples with varying percentages. The obtained results from the present study indicated that the treatment of fish samples by lemon extract, thyme oil and acetic acid were the best as increasing the shelf life as well as the tenderness and enhancing the sensory evaluation which include appearance, odor, and color and over all acceptability. Introduction
Fish is a rich source of easily digestible protein that also provides polyunsaturated fatty acids, vitamins and minerals for human nutrition. Although some species of fish are used industrially for fish meal manufacture, a need for their preservation and utilization for human consumption has been recognized in order to prevent post-harvest fishery losses (Venugopal and Shahidi, 1995 and Koffi-Nevry et al., 2011). Aeromonas species are widely spread in waters, water habitants, and many food products (seafood, shellfish, and raw foods of animal origin like poultry, ground meat, raw milk, and raw vegetables). (Hristo, 2006). The high environmental prevalence of these bacteria should be regarded as an important threat to public health, since Aeromonas infections are generally acquired through consumption of water and food (Borrell et al., 1998) According to Adams and Moss (2000) and Kirov (2001) Aeromonas (principally A. hydrophila) currently has the status of a foodborne pathogen of emerging importance. It has attracted attention primarily because of its ability to grow at cold temperatures. Aeromonas spp. were first considered as possible causative agents of human gastroenteritis more than 30 years ago (Lautrop, 1961). Genus Aeromonas has emerged as an important human pathogen because of suspected food-borne outbreaks (Altwegg et al., 1991; Kirov et al., 1993) and the increased incidence of its isolation from patients with traveller’s diarrhea (Hanninen et al., 1995; Yamada et al., 1997). Among the 14 species of Aeromonas known to date (Carnahan 1
and Altwegg, 1996), A. hydrophila, A. caviae, and A. veronii biotype sobria have most commonly been involved in human infections and have been found to produce a variety of virulence factors such as hemolysins, cytotoxins, enterotoxins, proteases, leukocidin, phospholipases, endotoxins, outer membrane proteins, and fimbriae (Chopra and Houston, 1999 ). According to Kirov (1993, 2003), Kirov and Sanderson (1995), and Isonhood and Drake (2002), Aeromonas species have been recognized as pathogens which can cause a number of serious extraintestinal infection including bacteraemia , meningitis, pulmonary and wound infections. Aeromonas spp. may play a significant role in ‘‘summer diarrhoea’’, a worldwide problem particularly in children under five years old, the elderly, and travellers. The role of these bacteria in foodborne incidences is not firmly established, but Aeromonas spp. has the potential to emerge as significant foodborne Pathogens. A. hydrophila were reported by Janda and Duffey (1998) to predominate in cases of Aeromonas-associated gastroenteritis. Kirov (2003) noted that the disease spectrum of A. hydrophila included gastroenteritis, septicemia, traumatic and aquatic wound infections, and infections. Therefore, the current study aimed to detect, count and identify Aeromonas spp. in some fresh and marine water fishes as well as to impose a trial to improve and extened their shelf life, Material and methods
A total number of 120 fishes samples of Tilapia nilotica, Cat fish and Carp fish which is fresh water fishes (20 of each) and Blue spot fish, Saurus and Pagrus which is marine water fishes (20 of each) were collected from different fish markets in Zagazig city. The collected samples were well identified and packaged separately in a sterile plastic bag; then directly transferred with a minimum of delay to the post-graduate laboratory of food Control Dept. Faculty of Veterinary Medicine, Zagazig University under aseptic conditions, where they were subjected to organoleptic and bacteriological examination A-Organoleptic examinations: This examination was adopted according to the procedure recommended by (Braumuller, 1958). B- Bacteriological examination:
Muscle samples of fish under examination were prepared according to ICMSF, (1978), enumeration, Isolation and identification of Aeromonas spp. according to the method recommended by ICMSF, (1978); Roberts et al., (1995) and MacFaddin, (2000) using Glutamate starch phenol-red agar medium (GSP Agar) which is (Pseudomonas Aeromonas Selective Agar Base) proposed by ICMSF, (1978) C-Trails to improve the shelf-life of fish:Antibacterial agents used * Tap water.* Lemmon extracts (120 PPM.) * Thyme oil 0.2 % (V/W).* Acetic Acid 1 % (V/W). A. Sampling:
30 random samples were collected from fishes markets in Zagazig city. The collected samples were identified and packaged separately in a sterile plastic bag under aseptic conditions; then directly transferred with a minimum of delay to the laboratory where they were subjected for treatment and examination protocols. 2
B. Treatment protocols:
In the laboratory, fresh Tilapia nilotica samples were divided into (6) groups, each group contains 5 fishes: First group untreated, the Second one was dipped in the previously isolated and identified A. hydrophila broth. Meanwhile, the third one was dipped in A. hydrophila broth then treated by Tap water for 10 minutes. Concerning the Fourth group it was treated by dipping in lemon extract (120 ppm) for 10 minutes, while the Fifth group was treated by dipping in Thyme oil 0.2 % (V/W) for 10 minutes. On the other side the Sixth group was treated by dipping in acetic Acid 1 % (V/W) for 10 minutes after their contamination with isolated and identified A. hydrophila broth. The control as well as treated groups were subjected periodically to the following examinations at zero time and after storage in domestic refrigerator (4º C + 1º C) every 2days till appearance of spoilage signs: 1- Organoleptic examination. 2- Measurement of pH according to (EOS/ 2760-1:2006): 3- Bacteriological examination which was carried out according to the method recommended in part I. Results and Discussion Table (1): Organoleptic examination of examined fresh and marine water fishes samples (No. = 20 of each) Fish Fishes Min Max Mean ± SEM Tilapia nilotica 61.00% 89.00% 72.00 ± 2.00% Fresh water fishes Cat fish 57.00% 89.00% 72.00 ± 2.00% Carp fish 64.00% 89.00% 74.00 ± 2.00% Blue spot fish 64.00% 76.00% 70.00 ± 2.00% Marine water Saurus 51.00% 86.00% 68.50 ± 2.00% fishes Pagrus 50.00% 89.00% 69.50 ± 3.00% Min = Minimum SEM = Standard Error Mean Max = Maximum No = Number of each fishes sample. Table (2): Incidence of Aeromonas in examined fresh and marine water fish samples. (No. = 20 of each):-
Fish
Fresh water fishes
Marine water fishes
No. of positive % samples
Fishes Tilapia nilotica Cat fish Carp fish Blue spot fish Saurus Pagrus
3
13 20 20 18 8 15
65 100 100 90 40 75
Table (3): Aeromonas count (CFU/g) in examined muscles of fresh and marine water fish samples (No = 20 of each):-
Fish Fresh water fishes
Marine water fishes
Fishes Tilapia nilotica Cat fish Carp fish Blue spot fish Saurus Pagrus
Min ≤100 6x106 2x106 ≤100 ≤100 ≤100
Max 12x105 104x106 19x106 17x105 11x106 4x106
Mean ± SEM 21.1x104 ± 7.7x104 36x106 ± 6.5x106 7.7x106 ± 1.2x106 36.9x104 ± 0.95x105 13x105 ± 6.5x105 13.7x105 ± 2.7x105
Table (4): Frequency distribution of isolated Aeromonas organisms recovered from examined fresh and marine water fish samples. No. of Aeromonas Fish Fishes Aeromonas No. % isolates spp. isolates A. hydrophila 8 61.5 Tilapia A. sobria 4 30.8 13 nilotica A. caivae 1 7.7 A. hydrophila 13 65 Fresh Cat fish 20 A. sobria 6 30 water A. caivae 1 5 fishes A. hydrophila 13 65 Carp fish 20 A. sobria 4 20 A. caivae 3 15 A. hydrophila 14 77.8 A. sobria 1 5.6 Blue spot 18 fish A. caivae 3 16.6 A. hydrophila 8 100 Marine water Saurus 8 A. sobria 0 0 fishes A. caivae 0 0 A. hydrophila 10 66.7 Pagrus 15 A. sobria 0 0 A. caivae 5 33.3
4
Table (5): Organoleptic examination of untreated samples and treated samples of Tilapia nilotica fishes:Day Control group 1 3 5 7 9 11 13 15
87.86±0.61% 61.43±1.32% 30.71±1.82% AD
In vitroTape water Lemon extract contaminated treated group treated group group 87.14±1.82% 87.14±1.82% 87.14±1.43% 30±1.82% 60±2.07% 73.57±1.43% AD 31.43±1.32% 62.14±1.82% AD 30.71±1.82% AD
Thyme oil treated group
Acetic acid treated group
86.43±1.32% 77.14±1.82% 65±1.75% 53.57±1.14% 31.43±1.32% AD
90.71±1.82% 83.57±1.82% 75±2.25% 67.14±2.07% 61.43±1.32% 52.86±1.32% 34.29±1.82% AD
AD = Apparently Decomposed Table (6): pH of examined untreated and treated samples of Tilapia nilotica fish:Day
Control group
1 3 5 7 9 11 13 15
6.82±0.04 6.24±0.07 6.34±0.17 *
In vitrocontaminated group 6.84±0.07 6.48±0.07 *
Tape water treated group 6.64±0.04 6.34±0.02 6.44±0.11 *
Lemon extract Thyme oil treated group treated group 6.14±0.18 6.08±0.21 6.22±0.09 6.52±0.16 *
Acetic acid treated group
6.54±0.09 5.54±0.06 5.82±0.06 6±0.09 6.42±0.13 *
5.09±0.07 5.38±0.11 5.64±0.07 5.92±0.05 6.1±0.04 6.32±0.05 6.44±0.07 *
* = pH not measured Table (7): Aeromonase hydrophila count (CFU/g) in the examined untreated and treated samples of Tilapia nilotica fish. Day 1 3 5 7
In vitroTape water Control group contaminated treated group Group 4 2.5x10 ± 6x106± 9.5x104± 4 6 0.2x10 1.1x10 0.6x104 5 6 2.2x10 ± 12x10 ± 7.2x105± 5 6 0.33x10 1.2x10 0.39x105 5 41.2x10 ± 45.2x105± * 5 6.2x10 4.2x105 *
Lemon extract treated group
Thyme oil treated group
Acetic acid treated group
2.2x104± 0.2x104 2.2x105± 0.33x105 9x105± 1.8x105 2.9x106± 0.43x106
8.4x102± 1.7x102 2.3x104± 0.29x104 22.4x103± 2.3x103 2.3x105± 0.3x105 17.8x105± 3.1x105
4x102± 0.7x102 2.4x103± 0.51x103 15x103± 1.2x103 32.4x103± 3.1x103 4.8x104± 0.8x104 22.8x104± 2.3x104 15.6x105± 4.8x105
*
9
*
11
*
13 15
*
* = Not counted 5
Discussion I- Organoleptic examination:
Consumers are the ultimate judges of quality and acceptance depends on satisfying their sensory requirements. Freshness declines as the storage life passes, until the product is no longer acceptable to the consumers. Therefore, Freshness implies acceptability; loss of freshness indicates loss of storage life when the product is unacceptable (Hall, 1992). It is evident from the results recorded in table (1) that the mean values of organoleptic examinations of the examined samples of Tilapia nilotica and cat fish (Clarias lazera) were 72.00 ± 2.00%, for each while in Carp fish it was 74.00 ± 2.00%.as fresh water fish. All the examined samples were considered fresh and fit for human consumption. Nearly similar results were recorded by Sallam (1990), Abo Samara (2001) and Morshdy et al., (2002). On the other hand, the mean values of organoleptic examinations of the examined samples of marine water fishes, Blue spot fish, Saurus and Pagrus were 70.00 ± 2.00%, 68.50 ± 2.00% and 69.50 ± 3.00% respectively (table 1). All the examined samples were considered fresh and fit for human consumption. The results showed that physical examination must be associated with bacteriological examination to give the accurate judgment. II-Bacteriological examination:
(A)- Incidence and count of Aeromonas species in the examined fresh water fishes samples: The obtained results in tables (2 &3) revealed that the incidence of Aeromonas species was 65% , 100% and100% in the examined samples of Tilapia nilotica , Cat and Carp fish with a mean value of 21.1x104 ±7.7x104 CFU/g, 36x106 ± 6.5x106 and 7.7x106 ± 1.2x106 CFU/g respectively. The obtained results were coincid with that reported by El-Kelish (1995), El-Atabany (1995) and Lamada (1999), higher findings were reported by Bastawarows and Mohamed (1999) ,on the other side, lower findings were reported by Naser (1991). Such variations may be attributed to presence of aeromonas spp. in mud and cat fish as a bottom feeder most contaminated species. Comparatively, the obtained results declared that the Cat fish had a higher count of Aeromonas spp. than that of Tilapia nilotica and Carp fish. This could be attributed to dirty habitat in which Cat fish live and prolonged exposure of such fishes to various types of contamination during marketing where the fishes live for a relatively long time after catching while other fish species dead shortly after harvesting and this result agrees with Abo Samara (2001). (B)- Incidence and count of Aeromonas species in the examined marine water fishes samples: It is evident from the results recorded in tables (2&3) that the incidence of Aeromonas spp. was 90%, 40% and 75% in the examined samples of Blue spot fish, Saurus and Saurus with a mean value of 36.9x104 ± 0.95x105 CFU/g, 13x105 ± 6.5x105 CFU/g and 13.7x105 ± 2.7x105 CFU/g respectively. Such results substantiate what has been found by Lamada (1999). C- Incidence of isolated Aeromonas organisms recovered from examined fishes samples: The identification of Aeromonas species (A. hydrophila, A. sobria and A. caviae) was demonstrated in table (4) which showed that these strains were isolated from the examined samples of Tilapia nilotica in percentages of 61.5, 30.8 and 7.7%, respectively. While in Cat 6
fish they were recovered from 65, 30 and 5%, samples respectively. On other hand, these strains were identified in 65, 20 and 15% of the examined Carp fish samples, respectively.In Egypt, Mousa, (1986) and Khalil et al., (1990) isolated aeromonas from the muscles of Tilapia nilotica and Mugil cephalus. The widespread occurrence of A. hydrophila has been proved by many investigators, who isolated this organism from human beings, fowls and sea-foods (Fehlhaber et al., 1985 and Palumbo et al., 1989). Its wide spread in the environment causes food related illness and becomes of concern to food microbiologist as has been isolated from a variety of foods (Palumbo et al., 1989). Data presented in table (4) revealed that the incidence of identified aeromonas spp. (A. hydrophila, A. sobria and A. caviae) isolated from examined marine water fishes samples was 77.8, 5.6 and 16.6% for Blue spot fish samples, 100, 0 and 0%, for Saurus fish samples and 66.7, 0 and 33.3% for Pagrus fish samples, respectively. From the aforementioned results it can be concluded that A. hydrophila was the most predominant identified Aeromonas spp. isolated from both fresh and marine water fish samples. Conclusions regarding the significance of A. hydrophila in foods should be made with circumspection. These findings substantiate what have been reported by García-López et al (2011) who was able to isolate five strains of Aeromonas species during a three-year period in the same geographic in Spain. IV. Trials for improve the shelf life of fresh water fishes: 1-Organoleptic examination:
The basic goal of all decontamination trails is to reduce the risk of pathogenic microorganism and decrease the microbial load as well as to prolong shelf life of fish. Advantages and disadvantages of the chemicals used in decontamination must be taken in to account on choosing the best one for commercial uses. The fitness of any article of food should be based on combined information obtained from organoleptic examinations which includes smell, appearance and texture, chemical and microbial evaluation. The pH value of live fish muscle is close to 7.0, however post mortem pH can vary from 6.0 to 7.0 depending on season, species and other factors (Simeonidou et al., 1998). Organoleptic examination of Tilapia nilotica over all the refrigeration of storage at 4+1º C is presented in table (5). The results revealed that Tilapia nilotica control group samples, in vitro-contaminated group samples, and treated groups with tape water, lemon extract, thyme oil and acetic acid were organoleptically rejected shown extreme discoloration and off odor after 7th, 5th, 7th, 9th, 11th and 15th days of refrigeration storage, respectively. Moreover, it is evident that the mean values of organoleptic examination at 1st day in control group , in vitro-contaminated group samples, tape water, lemon extract, thyme oil and acetic acid in Tilapia nilotica fish samples was 87.86±0.61%, 87.14±1.82%, 87.14±1.82%, 87.14±1.43%, 86.43±1.32% and 90.71±1.82% respectively. At the same time, the pH of the same examined control and treated groups were 6.82±0.04, 6.84±0.07, 6.64±0.04, 6.14±0.18, 6.54±0.09 and 5.09±0.07 respectively (Table 6). On the other hand, at the 3rd day of refrigeration storage at 4+1º C the mean values of organoleptic examination of Tilapia nilotica fishes was 61.43±1.32%, 30±1.82%, 60±2.07%, 73.57±1.43%, 77.14±1.82% and 83.57±1.82% in control group, in vitro-contaminated group, tape water, lemon, thyme oil and acetic acid treated samples respectively. The pH of the 7
same examined treated and untreated groups was 6.24±0.07, 6.48±0.07, 6.34±0.02, 6.08±0.21, 5.54±0.06 and 5.38±0.11 respectively (Table 6). Meanwhile, at the 5th day of refrigeration storage at 4+1º C of Tilapia fishes, the mean values of organoleptic examination for untreated control group samples and treated samples with tape water, lemon extract, thyme oil and acetic acid was 30.71±1.82%, 31.43±1.32%, 62.14±1.82%, 65±1.75% and 75±2.25% respectively. The pH of the same examined treated groups was 6.34±0.17, 6.44±0.11, 6.22±0.09, 5.82±0.06 and 5.64±0.07 respectively (Table 6). The in vitro-contaminated group samples show signs of decomposition. By 7th day of refrigeration storage at 4+1º C of Tilapia fishes the mean values of organoleptic examination for treated samples with lemon extract, thyme oil and acetic acid was 30.71±1.82%, 53.57±1.14% and 67.14±2.07% respectively. The pH of the same examined treated groups was 6.52±0.16, 6±0.09 and 5.92±0.05 respectively (Table 6). Also at 7th day of refrigeration storage at 4+1º C the untreated control group samples and treated samples with tape water of Tilapia fishes showed decomposition. Such results nearly similar to that of Abd Al-Rahman (2010) where the Tilapia nilotica control samples were still organolyptically acceptable till 10th day of refrigeration storage at 4+ 1º C. At 9th day of refrigeration storage at 4+1º C the mean values of organoleptic examination in treated samples of Tilapia nilotica fish with thyme oil and acetic acid was 31.43±1.32% and 61.43±1.32% respectively, beside that the mean values of pH for treated samples of Tilapia nilotica with thyme oil and acetic acid was 6.42±0.13 and 6.1±0.04 respectively (Table 6). The lemon extract treated fish samples show signs of decomposition. By 11th day thyme oil treated samples of Tilapia fishes were decomposed while the mean value of the organoleptic examination of refrigeration storage at 4+1º C for Tilapia fishes for treated samples with acetic acid was 52.86±1.32%and the pH was 6.32±0.05. At 13th day the mean value of organoleptic examination of acetic acid treated samples of Tilapia nilotica fishes was 34.29±1.82%, and the value of pH was 6.44±0.07. Finally at 15th day the acetic acid treated samples of Tilapia nilotica fishes were decomposed. Nearly similar findings were found by Sallam et al. (2007) and Abd AlRahman (2010) where the Tilapia nilotica acetic acid treated samples were still organolypticaly acceptable till 18th day. The use of 1% acetic acid could retard the microbial growth, delay chemical changes, and improve the sensory attributes and extend shelf life of fishes. 2- Bacteriological examination:-
The quality of fresh fishes is a major concern to consumer. Fresh water fishes are extremely perishable food commodities. Deterioration of fishes mainly occurs as a result of bacteriological activity leading to loss of quality and subsequent spoilage. Bacteriological spoilage in refrigerated fishes under aerobic storage condition is caused by gram –ve psychotropic organisms such as A. hydrophila. A. hydrophila which they are capable to grow in refrigerate temperature and in anaerobic environments. They are gram –ve aerobic rods. Many are capable of breaking down proteins and /or fats and can digest materials such as gelatin, and hemoglobin. Most spoilage bacteria found in fishes are gram –ve and include aerobic and facultative anaerobic heterotrophic strains, although some gram +ve bacteria can also be found in high numbers. 8
From the results obtained in table (7), it is evident that the mean values of A. hydrophila count at the 1st day in control group , in vitro-contaminated group , tape water , lemon extract , thyme oil and acetic acid treated group samples in Tilapia fishes was 2.5x104 ± 0.2x104 CFU/g, 6x106 ± 1.1x106 CFU/g, 9.5x104 ±0.6x104 CFU/g, 2.2x104 ± 0.2x104CFU/g, 8.4x102 ±1.7x102 CFU/g and 4x102 ± 0.7x102 CFU/g respectively. Meanwhile, the 3rd day refrigeration storage at 4+1º C revealed that the mean values of A. hydrophila count of Tilapia fishes was 2.2x105± 0.33x105 CFU/g, 12x106±1.2x106 CFU/g, 7.2x105 ± 0.39x105 CFU/g, 2.2x105 ± 0.33x105 CFU/g, 2.3x104±0.29x104 CFU/g and 2.4x103±0.51x103 CFU/g in control, in vitro-contaminated group, tape water, lemon extract, thyme oil and acetic acid treated group samples respectively. On the other side, at the 5th day of refrigeration storage at 4+1º C for Tilapia fishes the mean values of A. hydrophila count for untreated control samples, treated samples with a tape water, lemon extract, thyme oil and acetic acid was 41.2x105 ± 6.2x105 CFU/g, 45.2x105 ± 5 5 3 3 3 3 4.2x105 CFU/g, 9x10 ±1.8x10 CFU/g, 22.4x10 ±2.3x10 CFU/g and 15x10 ±1.2x10 CFU/g respectively. The in vitro-contaminated group samples show signs of decomposition. By 7th day of refrigeration storage at 4+1º C for Tilapia fishes the mean values of A. hydrophila count for treated samples with lemon extract, thyme oil and acetic acid was 2.9x106± 0.43x106 CFU/g, 2.3x105±0.3x105 CFU/g and 32.4x103±3.1x103 CFU/g respectively. Also at the same day of refrigeration storage at 4+1º C, the untreated control and treated samples with a tape water of Tilapia fishes showed signs of decomposition. Concerning the 9th day of refrigeration storage at 4+1º C, the mean values of A. hydrophila count in treated samples of Tilapia fishes with thyme oil and acetic acid was 17.8x105±3.1x105 CFU/g and 4.8x104±0.8x104 CFU/g respectively. The lemon extracts treated fishes samples show signs of decomposition. Then at 11th day Thyme oil treated samples of Tilapia fishes were decomposed, while the mean value of the A. hydrophila count of refrigeration storage at 4+1º C for Tilapia fishes for treated samples with acetic acid was 22.8x104±2.3x104 CFU/g. Finally at 13th day the mean value of A. hydrophila count of acetic acid treated samples of Tilapia fishes was 15.6x105±4.8x105 CFU/g. Nearly similar results were obtained by Abd Al-Rahman (2010) who found that the mean value of psychrotrophic bacteria count was 73x105+ 1.4x105 CFU/g at 11th day of Tilapia nilotica samples collected from markets in Zagazig city. In view of the aforementioned results, it is clear that the untreated samples (control group) showed the expected increase in A. hydrophila count with time. On the other hand, the maximum decontaminating effect was achieved by dipping Tilapia nilotica samples in acetic acid 1 % for 10 minutes which led to increase shelf-life of Tilapia nilotica samples during refrigerated storage up to 13 days. The obtained results from the present study indicated that the treatment of fish samples by lemon extract, thyme oil and acetic acid were the best as increasing the tenderness and enhancing the sensory evaluation which include appearance, odor, and color and over all acceptability. References Abd Al-Rahman, R. A. (2010): Improving the sanitary status of some fresh fish, M.V.Sc. Thesis, Fac. Vet. Med, Zagazig Univ, Egypt. 9
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Kirov, S. M. (1993): The public health significance of Aeromonas spp.in foods. International Journal of Food Microbiology, 20, 179–198. Kirov, S. M. (2001): Aeromonas and PlesiomonasSpecies. In M. P.Doyle, L. R. Beuchat, & T. J. M ontville (Eds.), Food microbiology:fundamentals and frontiers (second ed., pp. 301– 328). ASM Press. Kirov, S. M. (2003): Aeromonas Species. In A. D. Hocking (Ed.), Foodborne microorganisms of public health significance (sixth ed.,pp. 553–575). AIFST Inc. (NSW Branch). Kirov, S. M., and Sanderson, K. (1995): Aeromonas: Recognising the enemy.Today’s Life Science, 11, 30–35. Kirov, S.M., Ardestani, E.K., and Hayward, L.J.,(1993): The growth and expression of virulence factors at refrigeration temperature by Aeromonas strains isolated from foods. Int. J. Food Microbiol. 20, 159–168. Koffi-Nevry, R. ; Ouina , T.S.T. ; Koussemon, M. and Brou, K.(2011): Chemical Composition and Lactic Microflora of Adjuevan,A Traditional Ivorian Fermented Fish Condiment. Pakistan Journal of Nutrition 10 (4): 332-337. Lamada, H. M. O. (1999): Prevalence of some potential pathogens in Nile fishes. M.V.Sc. Thesis Fac. Vet. Med., Moshtohor; Zagazig University, Bena Branch, Egypt. Lautrop, H. (1961): ‘‘Aeromonas hydrophila’’ isolated from human faeces and its possible pathological significance. Acta Pathologica Microbilogica Scandinavica, 51, 299–301. Macfaddin J. F. (2000): Biochemical tests for identification of medical bacteria 3rd Edi. Lippincoll Williams & wilkins Washington, hiladelphia, U.S.A. Morshdy, A. M.; Eldaly E. A. and Sallam, K. I. (2002): Sanitary evaluation of some marketed fishes in Sharkia Province. 6th Vet. Zag. Conference: 313. Mousa, M.M.I. (1986): Microbiology of some fish and shellfish in local markets and its relation to public health. Ph. D. Thesis, Fac. Vet. Med., Alex. Univ. Naser, G. N. (1991): Occurrence of some food-poisoning microorganisms in natural and farm fishes. M.V. Sc. Thesis. Fac. Vet. Med., Alexandria Univ.,Egypt. Palumbo, S. A.; Bincivengo, M. M.; CorralF, D.; William, A. C. and Buchanan, R. L. (1989): Characterization of A. hydrophila Group isolated from retail foods of animal origin. J. Clin. Microbiol.; 27(5): 854. Roberts, D.; Hooper W. and Greenwood, M. (1995): Practical Food Microbiology. 2nd edi. Public Health Laboratory Service, London, UK. Sallam, A. A. M. (1990): Studies on the sanitary condition of fishes marketed in Ismailia. M.V.Sc. Thesis, Fac. Vet. Med., Zagazig Univ. Sallam, K. I., Ahmed A. M., Eigazzar M. M. and Eldaly E. A. (2007): Chemical quality and sensory attributes of marinated pacific saury during vacuum-packages storage at 4°C. Food Chemistry, 102: 1061-1070. Simeonidou, S.; Govaris, A. and Vareltzis, K. (1998): Quality assessment of seven Mediterranean fish species during storage on ice. Food Res. Int. 30: 479–484. Venugopal, V. and F. Shahidi, (1995): Value-added products from underutilized fish species. Crit. Rev.Food Sci. Nutr., 35: 431-453. Yamada, S., Matsushita, S., Dejsirilet, S., and Kudoh, Y., (1997): Incidence and clinical symptoms of Aeromonas-associated travellers’ diarrhoea in Tokyo. Epidemiol. Infect. 119, 121–126.
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