EXISTENCE OF LISTERIA SPECIES IN FRESHWATER FISH AND ITS CONTROL USING SANITIZED ICE

EXISTENCE OF LISTERIA SPECIES IN FRESHWATER FISH AND ITS CONTROL USING SANITIZED ICE

M.A.M. AMMAR^*; GHADA M. MOHAMED^* and NAHED M. ABD ELAZIZ^** 

^* Animal Health Research Institute, Assiut Regional Lab.

^** Fac. Vet. Med. Sohag, South-Valley University.

Email: mohmed2011eg@yahoo.coom    

Introduction

Fish is an excellent protein source (Jannat        et al., 2010) and provides many health benefits. One such benefit is its high level of omega 3 (n-3) fatty acids, which are known to reduce cholesterol level and the incidence of stroke, heart disease and preterm delivery, Willett (2005). Also fish is one of the most highly perishable food products, Ashie and Simpson (1996).

Normally fish muscle is sterile as its immune system prevents bacteria to proliferate easily whereas after death the fish's immune system collapses allowing easily penetration of microorganisms into the flesh, Huss (1995). This penetration increase in case of fish caught from polluted area where there are high densities of bacteria, Howgate (1985). Microbial contamination can reduce the quality of fresh fish, cause economic loss and health hazards, Gram and Huss (1996).

Listeria species could be isolated from a wide range of fishery products, including freshwater and seawater fish as well as frozen and processed fish (Farber and Peterkin 1991, Gudbjornsdottir et al., 2004 and Alves et al., 2005). One particular Listeria species (Listeria monocytogenes) has been recognized as a food borne pathogen since 1981. The bacterium is ubiquitous, Gram positive, facultative anaerobic, non-spore-forming, rod-shaped )Jacquet et al., 1992 and Jay 2000) and can grow over the temperature range of about 1 ºC to 45 ºC and pH range 4.1 to around 9.6. It may be expected to survive in foods for long periods of time, Ryser et al. (1985).

Contamination of fish by L.monocytogenes (LM) may take place during harvest, processing and distribution owing to improper handling and storage of fish, Kim et al. (1999). The ingestion of L M in food can pose a significant health risk with reported mortality rate. Most healthy humans are not significantly affected by the intake of small numbers of L M in foods. However, elderly people, pregnant women and unborn infants are the risk groups that are affected by the pathogen, FAO (1999). In human. The pathogen causes listeriosis which may range from mild to severe sickness, Adzitey and Huda (2010). The severe form of human Listeriosis is present as meningoenceophalitis followed by septic infections, Demetrios et al. (1996). Although an average of five to nine exposures to LM occur per person per year, Grif et al. (2003) listeriosis is a rather rare disease, Gerner-Smidt et al. (2005) but it is associated with a mortality rate of approximately 20% to 40%, Farber and Peterkin (1991), De Valk et al. (2005).

It has been reported that contamination of aquatic products with LMcannot be avoided totally, hence in order to inhibit the growth and development of this pathogen in the products and to ensure safety, antimicrobial additives are needed Nykanen et al. (2000). Antimicrobial activity of any preservative depends on its hydrophilic and hydrophobic properties i.e. solubility in water and fat, distribution in the model system, fat content pH and temperature, Glass and Doyle (1989). The antimicrobial additives must function the suppression of bacterial growth during storage with minor effects on the quality of the products, Zhu et al. (2005).

Trisodium phosphate (TSP) is generally recognized as safe by the US Food and Drug Administration and has been approved by the US Department of Agriculture-Food Safety and Inspection Service (USDA-FSIS) at levels of 8 - 12% as an antimicrobial agent. Trisodium phosphate has been evaluated by several investigators for its efficacy against pathogens and has been used as an effective compound in controlling the growth of Gram -negative pathogens, Ahmed and Abd El-Atti, (2012). Also, the efficacy of sodium acetate (SA) against LM has been investigated, Ghomi et al. (2011).

Research works have raised concerns about using solutions of TSP (Chang et al., 1995; Kim and Marshall, 2002) or SA (Sallam, 2007a and Sallam, 2007b) to reduce the load of LMon fish. There is scarce in research papers concerning the use of TSP or SA to sanitize ice used in fish storage.  

In the retail markets, fish is regularly displayed on ice to prevent spoilage and growth of pathogens. However if ice is made with sanitized water is used to store fish, it has potential to be bactericidal to the organisms, Felicano et al. (2010). Sanitized ice prepared with (TSP) or (SA) could help in achieving the goal. The aim of this study is to document the existence of listeria spp. in two economically, important freshwater fish species in Assiut markets (O. niloticus and Clarias gariepinus). A further goal is to evaluate the effectiveness of ices sanitized using TSP or SA in reducing populations of LM (on fish and in water from melted ice), their effect on the sensory quality and pH of stored fish as well as stability of ice.

MATERIALS and METHODS

1- Microbiological analysis of fish samples 

Fifty samples of freshwater fish namely Oreochromis niloticus and Clarias gariepinus (25 each) were collected from fish markets with different sanitation levels in Assiut city. All the collected samples were transferred to the laboratory in an ice box without delay where they were prepared and examined.

Sampling

Samples were taken from the left hand side of each fish in the anterior dorsal region. 25 gram muscle and its covering skin, was aseptically transferred to sterile stomacher bag.(Scott et al., 1992):

Isolation and identification of Listeria spp.: Tassou et al. (2004).

Two hundred and twenty five mL of sterile Listeria Selective Enrichment broth with antibioitic supplement (Difico Laboratories) was  aseptically added to 25 g prepared sample and pummeled in a stomacher (Seward BA7021, England) then transferred to sterile flask and incubated at 30ºC for 48h. After incubation the a loobfull of each broth was cultured onto Palcam Agar plates with antibioitic supplement (Difico Laboratories). After 48h of incubation at 35ºC the colonies morphologically suspected to be Listeria (black colonies with black sunken) were sub- cultured onto Tryptic Soy agar supplemented with 0.6% of yeast extract, incubated at 37 °C for 24 h and subjected to confirmatory tests.

All isolates were subjected to Gram's stain, motility and standard biochemical tests including, catalase, acid production from glucose, manitol, rhamnose, zylose, α-methyl-Dmanoside, nitrate reduction, methyl red /Voges-Proskauer, ß-hemolytic activity, and CAMP test, (Aygun and Pehlivanlar 2006).

2- Studying effect of ices formulated using TSP or SA on ice stability, L. monocytogenes load, sensory quality and pH of fish:

Preparation of ices: Feliciano et al. (2010).

Three concentrations, 2.5%, 5% and 10% (w/v) of TSP and one concentration (2.5%) of SA were prepared by reconstitution of target chemical in tab water. After stirring, the solutions were transferred to separate domestic ice cube trays and stored at -20 ºC for a week. Ices were weight into 700 g batches in polyethylene bags and stored at -20 ºC until used. Tab water ice cubes was also made as a control and stored under the same conditions.

Ice stability:

For each of ice formulations, the speed of melting was determined by placing the ice cubes (of the same dimensions) in a tray that was left uncovered at room temperature (25 ºC). At 30 min intervals the volume of water from melting ice was measured until the ice completely melted, Feliciano et al. (2010). All ices were tested at the same time and the test was repeated 3 times.

Preparation of bacterial inoculum: Ahmed and Abd El-Atti (2010).

One strain of L. monocytogenes (LM) previously isolated from the examined fish samples was used. The strain was maintained on 10 mL Tryptic Soya agar supplemented with yeast extract (Biolife). Tryptic soya broths (Difico Laboratories) were inoculated and incubated at 30 ºC for 24h to achieve viable cell populations approximately of 10⁹ cells/mL. Following incubation LM pool was prepared by diluting 10mL of the suspension with 90mL of sterile peptone water to yield a final inoculum approximately 10⁸cells/mL.

Fish used in the experiment:

Freshly caught O. niloticus (about 150 g each) were used. On arrival at the laboratory fish were gutted, washed with tab water, divided into two groups (GA = 20 fish and GB = 80 fish) and stored in the refrigerator until used (within 30 min).

Loading L. monocytogenes to fish:

The first group of fish (GA) was dipped for 60s in the LM pool (10⁸ cells /mL). After dipping, fish left to drain for 10 min in sterile tray fixed in an insulated cabinet and immediately used in the experiment.

Effect of ice formulation on L. monocytogenes load in fish and in water from melted ice:

 Five ice treatments were used for this experiment. Tab water ice (control), three concentrations (2.5%, 5% and 10%) TSP ices and 2.5% SA ice. Four fish were stored in each respective ice in a perforated sterile tray stand on sterile tray for collecting melted water. The fish were placed between two layers of crushed ice and left uncovered at 25 °C (ice: fish=1:1w/w and still constant until end of the experiment).

For each treatment a fish was sampled every hour and its load of LM was tested. Where the fish was grinded in a sterile mortar and a 25 gram portion was a aseptically sampled to sterile stomacher bag containing 225mL of 0.1 peptone water and pummeled for 1 min in a stomacher. Decimal dilutions were prepared and LM count was done following the technique recommended byTassou     et al. (2004). The counts were transferred to log10 CFU/g. At the end of storage period (4h), melted water from each treatment was randomly sampled and its LM load was determined.

Effect of ice formulation on sensory quality and pH of fish:

The 2^nd group of fish was subdivided into 5 subgroups (16 fish each). Each subgroup was stored between two layers of corresponding crushed ice treatment at 25°C. For each treatment, four fish were sampled every hour (3fish to be cooked for sensory evaluation and one for pH evaluation).

Effect of ice formulation on the sensory quality of cooked fish:

Treated fish were cooked separately for 15 min to an internal temperature of 75°C in a preheated conventional microwave oven adjusted to 180°C. One representative fish sample of the different treatments was individually presented in covered small porcelain dishes to each panelist. Panelists were asked to evaluate the overall acceptability with regard to appearance, odor intensity, flavor and aftertaste, tenderness, juiciness, off-odor and off-flavor according the score recommended by Sallam (2007b). Samples receiving overall scores of more than 4 were considered acceptable, while a score between 3 and 4 was considered the borderline of acceptability.

pH measurement:

Ten grams of each ice treated samples was srperately blended with 20ml distilled water in a blender for 30 s and the pH of fish homogenate was measured by a digital pH-meter (Gallenhamp No.101284) standardized at pH 4 and 7, Sllam, (2007b).

All experiments were repeated 3 times and the packaged SPSS program for windows version 12.0.1 was used for statistical analysis according to SPSS (2007). Data were expressed as mean ± standard error (SE). Differences between groups were determined by means of a Student "t"-test. Significance level was set at P < 0.05.

RESULTS

Table 1: Incidence of Listeria spp. in examined fish samples.

Table 2: Effect of ice formulation on the sensory quality of stored fish

Table 3: Effect of ice formulation on the pH of stored fish

*= significant (p<0.o5)

DISCUSSION

From the summarized results given in Table (1) it is evident that 10 % of examined fish samples contained Listeria spp. (5 out of 50 samples of O. niloticus and C. gariepinus fish). The percentages of Listeria sp. in each type of examined fish was 16 and 4%  respectively, where 4 and 6% of analyzed fish contained LMand L. Innocua, respectively. LM was found in a percentage of 4% in each of O. niloticus and C. gariepinus. While L.innocua could be detected in 12% of O. niloticus but failed to be recovered from C.garepinus.

The percentage of Listeria spp.in examined fish samples in our study is higher than that recorded by Rodas (2006) and Nikolaos (2007) where their findings were (4.5%) and (4.2%) respectively. Our records was lower than that reported by Joanne et al. (2004) Panda and Garg (2003) Jallewer et al. (2007), Hussein et al. (2011)  Siavash et al. (2011) and Mohamed (2012), where their findings were (23.3%), (20%), (17.1%( (18%), )26.7%), (16.7%), respectively. Some authors confirmed our results, they recorded nearly similar percentage of Listeria spp. as Adesiyum (1993) Modaresi  et al. (2011)  and  Shole  et al. (2013), where they recorded the presence of listeria spp. in 14.8%, 12.4% and 10.5% of examined raw fish samples, respectively.

The findings outlined in the same Table declared that the incidence of LM in raw fish  samples was (4%.(  This trend was higher than that obtained by Panda and Garg (2003), Nikolaos (2007) and Rahimi et al. (2012) which were (1.9%,1.7%  and 0.8%), respectvely. Our findings was lower than that reported by Rodas (2006), Jallewer et al. (2007), Modaresi et al. (2011), Mohamed (2012) and Shole   et al. (2013), where their records were (22.7%), (%10), )67%), (21%) and (7.3%), respectively. On the other hand, the incidence of  this organism was nearly agreed with that results obtained by Siavash    et al. (2011) Joanne et al. (2004) and Hussein et al. (2011) who recorded 4 %, 3.8% and 3.2 % respectively. Other studies have found that the prevalence of LM in raw fish is quite low, ranging from (0 to 1% (Autio    et al. (1999) and Johansson et al. (1999). Jemmi and Keusch (1994) and Hartemink and Georgesson (1991) stated that in Iceland 56% of fresh fish on sale were contaminated with LM and other listeria spp. However many investigators as karunasagar et al. (1992), Kamat and Nair (1994) and Papadopoulos et al. (2010) could not isolated LM from any of fresh water fish samples. Many results suggest that the absence of LM in tropical fish is due to using inadequate methodology. Reliable and accurate isolation and detection techniques are important in the surveillance of LM, Shole (2013).

L. innocua as presented in Table (1) was recovered from the fish in a percentage of 6% which was higher than that reported by Shole et al. (2013) who recorded 0.9% while Panda and Garg (2003) and Siavash et al. (2011) recorded higher percentages of L. innocua than in our study (17% and11%(, respectively. On the other side the percentage of L. innocua was somewhat agreed with the result detected by Hussein et al. (2011) and Rahimi et al. (2013) which were (8.4% and 5.7%, respectively). Our study showed that L.innocua was predominant among Listeria spp. and this agreed with the result obtained by Adzitey and Huda (2010 (Since both L M and L. innocua share the ecological niches, and the isolation of both bacteria is not surprising. where isolation of L.innocua in fishery products is considered as an indicator of possible contamination with L M.

The differences in prevalence of Listeria spp. may be due to the facts that: type of samples, methods of sampling, isolation techniques, geographical area and even climate of area which samples were collected, Hassan and Shole et al. (2013). In addition, contact with intestinal contents is risk factor for prevalence of Listeria spp. in seafood samples, Ertas and Seker (2005).

Effect of ice formulation onice stability, L. monocytogenes load (in fish and melted water), sensory quality and pH of fish:

Ice stability:

Results from the ice stability in showed that, tap water ice appeared to melt faster than other ices (Figure 1). For example, after 60 min storage time, the volume of water collected from tap water ice was 265 mL whereas 255, 180, 186 and 207 mL were collected from (SA) 2.5%(TSP) 2.5%, (TSP) 5% and (TSP) 10%ices, respectively. However, for most storage times after that, the volume collected from all treatments appeared to be relatively similar, except around the 3-2.5 h storage period. In general, formulated ices are more stable (700 g ice melted completely in 5 h versus 4h to tab water ice).

Efficacy of ice formulations in reduction of Listeria monocytogenesload in fish and in water from melted ice:

To evaluate the effectiveness of antimicrobial ices, viable LM on agar surface were enumerated following treatments. As shown in Figure 2, the initial LM count was 5.5 log10 CUF/g. The initial count was reduced by 1.4, 0.2,1.4,1.6 and 0.6 log10 CFU/g for (SA) 2.5%_, (TSP) 2.5%, (TSP) 5%, (TSP) 10% and tape water ices at one hour sampling period, respectively. At two hours storage, the treatments were effective in achieving desired log10- unit reduction except TSP 2.5% and tap water ices. A reduction level of log₁₀-unit using (TSP) 2.5%ice was achieved after 4 h of storage. Whereas (SA) 2.5%ice was more effective at same concentration and storage time.

The rate of reduction was dependent on the concentration for TSP ices. For example, reductions in LM cells were 1.2,2.0 and 2.5 log10 CFU /g for (TSP) 2.5% _, TSP 5% and TSP 10%ices at the 4th sampling period, respectively. The results in Figure 2 also shows that (SA) 2.5%and TSP 10% ices were the most effective compared with the control (1.6  log10 CFU/g additional reduction to control) followed by (TSP) 5% (0.6log10 CFU/g reduction additional to control) at the 4^th hour of the storage period. Statistically, there were significant difference (data not shown) in the reductions between the ice treatments prepared from tap water and ices formulated using (SA) 2.5%or (TSP) 10%. Sodium acetate 2.5%or (TSP) 10% ices was efficient in reduction of LM in treated fish compared with control ice (P <0.05).

Figure 3 shows the amount of LM cells recovered in the waters collected from the ice treatments after 4h storage. The mean LM viable cells recovered were 3.7,3.8,3.5,3.0 and 4.0 log10 CFU / ml of melted ice for (SA) 2.5%, (TSP) 2.5%, (TSP) 5%, (TSP) 10%  and tap water ices, respectively. Statistically, (TSP) 10%ice was the most efficient in reduction of LM population in the water from melted ice (P<0.001) compared with tab water ice while only a significant difference (P<0.05) for other formulated ices compared with tab water ice.

This study, confirmed that formulated ices especially (SA) 2.5%or (TSP) 10%ice can help to overcome the LM burden in the water when it used to store the fish. This thus reduced the potential for cross–contamination to contact surfaces and even to the environment of the market.

To evaluate sanitizers some factors should be taken into consideration before making any assumption. For example, animal products (fish and meat) are foods rich in proteins and/or lipids. It has been reported that even low amounts of proteins or fats are capable for reducing  the  antimicrobial  efficacy  of   sanitizers) Vandekindern et al. (2009). In addition, it is documented that temperature also influences the efficacy of the sanitizer where Venkitanarayana et al. (1999)found that LM was more rapidly inactivated by sanitizers at 35 °C and 45 °C than at 4 °C or 23 °C. LM is also tolerant and survives in extreme conditions like a wide PH range (4.1-9.6), Adzitey and Huda (2010). Regarding TSP, Taormina and Beuchat (2013) revealed that LM survived at least 6 days when they were suspended in TSP at pH 9.0 and stored at 4 °C for 21 days. Capita et al. (2001) found that the rate of LM reduction was increased by increasing TSP concentration. The behavior of LM was significantly influenced by the origin of the strain and salt concentration in their experiment.

There is lake of researched papers about using of SA or TSP in ice formulations but the potential of using their solutions to reduce bacterial populations in fresh fishery products was explored. Kim and Marshall (2002) recorded a reduction of 0.6 to less than 1.8 log10 CUF of LM on fish skin when treated for 10 min using TSP 2-6%. Whereas, Mu et al. (1997) mentioned that 10 or 20% solution treatments of TSP didn't significantly reduce Listeria populations in fish fillets after 6 days of storage.

Sodium acetate is widely available, economical and generally "recognized-as-safe" Sallam (2007 a). The available literatures focused on using its solutions for dipping of fish. Golden (1995) explored that Listeria inactivation was directly related to the concentration and incubation temperature and inversely related to pH. Kouassi and Shelf (1996) found that SA was effective antilisterial at concentration of 3% at 37 °C. Sallam (2007 a) noticed that aqueous solution of (SA) 2.5% was efficient against proliferation of various categories of spoilage microorganisms of fish under refrigerated storage.

Effect of ice formulation on sensory quality of treated fish:

The mean values for odor and flavor intensity, juiciness, tenderness, appearance and off odor and flavor of cooked fish are illustrated in table (2). The analysis of these values showed non-significant difference between treated and control fish (data not shown). All treated fish were judged acceptable by the sensory panel. No off-odor or off-flavor could be detected in any of treated fish. The results regarding sensory evaluation of treated fish are in accordance with the finding of other researchers who treated fish with SA or TSP. Kilinc et al. (2009) found that dipping of fish in (TSP) 5% for 10 min did not affect the texture of fish fillets. Vyncke (1978) revealed that dipping  of  fish  in  a  solution  of  sodium tripolyphosphate for 5 min improved the appearance of treated fish. Sodium salt of acetic acid has been used to improve sensory quality attributes and extend the shelf life of fish. Sallam (2007 b) noticed no difference in appearance, Juiciness and tenderness in fish dipped in 2.5% aqueous solution of SA compared with control. Manjua et al. (2007) found that SA treated fish (2% Sol.) still acceptable during refrigerator storage for 15 day. Kim et al. (1995) summarized that fish fillets treated with (SA) 1% had an appearance and odor scores similar to fresh controls up to 3 days of refrigerator storage. We can summarized that treating fresh fish with the studied SA or TSP ice formulations can maintain the sensory quality of the treated fish when stored at room temperature (25 °C).

Effect of ice formulation on pH of treated fish:

Values regarding the effect of ices on the pH of fish muscles during storage at room temperature (25 °C) are shown in table3. The initial pH value of fresh control fish (6.03) was significantly higher (P< 05.0) than those treated with (SA) 2.5%or (TSP) 5% ices but not tap water ice at the end of 1^st hour of storage on ice. A significant decrease in the pH value of fish treated with (SA) 2.5%ice compared with the control was also observed at the 2^nd, 3^rd and 4^th hour of storage (P< 05.0). At the end of storage time (4h) a significance reduction of pH was observed in samples treated with (SA) 2.5% (6.1), (TSP) 2.5% (6.1) and (TSP) 5% (6.5)compared with the control (6.8). Trisodium phosphate 10%ice resulted in a significance (p<05.0) rising of pH of treated fish muscles at the 2^nd and 4^th sampling periods compared with the control, but still near the acceptable pH range of fish (6.6-6.8) recommended by Ross et al. (2000) The reductions of pH of fish muscles after treatment with SA in our experiment are consistent with those of Sallam (2007b) and Kim et al. (1995). In respect for TSP, Mu et al. (1997) recorded the rising in the pH values of fish fillets treated with TSP (10% solution) compared with the control.

We can summarized that SA ice treated fish had a significantly (p<05.0) low pH values compared with the control along the storage period (4h). The reduction of pH is the main function of acetic acid and it is salts when added to foods. That reduction did not affect sensory attributes of fish as revealed in the sensory evaluation. The relative high pH values in fish treated with (TSP) 10%ice compared with control may due to high skin pH value which recorded by Mu et al. (1997). The relatively high pH value (0.2 more than control) did not reflected in the sensory attribute tested by panels.

CONCLUSION

Chilling to about 0 °C is the most important means of fresh marketing in Egypt. The most common chilling media is ice. The finding during this study confirmed that Bolti (O. niloticus) and C. gariepinus from Assiut markets were contaminated by listeria (16% and 4%, respectively). Consequently raw fish and water from melting ice in contact can become a source of cross contamination if not dealt with properly. This potential could be significantly (p <05.0) reduced by the use of (TSP) 10%or (SA) 2.5%ice. These ice formulations did not affect any of sensory quality parameters or the overall acceptability of fish compared with control (p>05.0) and can be utilized as safe preservatives for fish under room temperature storage thus improving fish safety and protect public health.

REFERENCES

Adesiyum, A.A. (1993): Prevalence of Listeria spp., Campylobacter spp., Salmonella spp., Yersinia spp. and toxigenic E. coli on meat and seafood in Trinidad. Food Micribiol.10: 395-403.

Adzitey, F. and Huda, N. (2010): Listeria monocytogenes in foods. Incidences and possible control. Afri. J. of Microbial. Research 4: 2848-2852.

Ahmed, A.M. and Abd El-Atti, Nashwa, A. (2012): Existence of Listeria species in broiler carcasses with an attempt to control Listeria monocytogenes using trisodium phosphate. Afri. J. of Food Sci.4 (2), 046-051.

Alves, V.F.; De Martinis, E.C.; Destro, M.T.; Vogel, B.F. and Gram, L. (2005): Anti Listeria activity of Carnobacterium piscicola isolated from Brazilian smoked fish (surubim pseudoplatystoma spp.) and its activity against a persistent strain of Listeria monocytogenes isolated from surubim. J. Food Prot. 68: 2068-2077.

Ashie, I.N.A. and Simpson, B.K. (1996): Application of high hydrostatic pressure to control enzyme related fresh sea food texture deterioration. Food Research International. 29 (5-6) 569-575.

Aygun, O. and Pehlivanlar, S. (2006): Listeria spp. in the raw milk and dairy products in Antakya, Turkey, Food Control. 17: 676-679.

Autio, T.; Hielem, S.; Miettinen, M.; Sjoberg, A.M.; Aarnisalo, K. and Bjork roth, J. (1999): Sources of L. monocytogenes contamination in a cold –smoked rainbow trout processing plant detected by pulsed-field gel electrophoresis typing. Appl. and Environ. Microbiol. 65, 150-155.

Aygun, O. and Pehlivanlar, S. (2006): Listeria spp. in the raw milk and dairy products in Antakya, Turkey. Food Control. 17: 676-679.

Ben Embarek, P.K. (1994): Presence, detection and growth of Listeria monocytogenes in seafood. A review. Int. J. Food Microbiol 23:17-34.

Capita, R.; Alonso-Calleja, M.C. and Moreno, B. (2001): Influence of strain and trisodium phosphate concentration on growth parameters of Listeria monocytogenes in vitro. Lett. Appl. Microbiol. 32(6), 428-32.

Chang, K.R.; James, H.O.; Amy, V.P.; Charles, W.H.; Marshall, H.and Douglas, L. (1995): Extending shelf life of refrigerated cat fish fillets using sodium acetate and monopotassium phosphate. J. Food Prot. 6: 597-708.

Demetrios, K.; Bori, M. and Antonios, M. (1996): Growth of Listeria monocytogenes in the whey cheeses, Myzi theria, Anthotyros, and Manouri during storage at 5,12 and 22 C^о. Food Prot. 59: 1193–1199.

De Valk, H.; Jacquet, C.; Goylet, V.; Vaillant, V.; Perra, A.; Smon, F.; Desenclos, J.C. and Martin, P. (2005): Surveillance of Listeria infections in Europe. Eurosurveillance 10:251-255.

Ertas, H.B. and Seker, E. (2005): Isolation of Listeria monocytogenes from fish intestines and RAPD analysis, Turkish J. of Vet. and Animal Scie. Vol. 29: 1007-1011.

FAO (1999): Report of the FAO expert consultation on the trade impact of Listeria in fish products. FAO Fisheries Report No 64. FIIU/ESNS/R604, Amherst, MA, United States.

Farber, J.M. and Peterkin, P.I. (1991): Listeria monocytogenes, a foodborne pathogen. Microbiol. Rev. 55: 476-511.

Feliciano, L.; Lee, J.; Lopes, J. and Pascall, M.A. (2010): Efficacy of sanitized ice in reducing bacterial load on fish fillet and in the water collected from the melted ice. J. Food Sci. 75(4), 231-238.

Gerner-Smidt, P.; Ethelberg, W.; Schiellerup, P.; Christensen, J.; Engeberg, J.; Fussing, V.; Jensen, A.; Petersen, A.M. and Bruun, B.G. (2005): Invasive listeriosis in Denmark 1994-2003: a review of 299 cases with special emphasis on risk factors for mortality. Clinic. Microbiol. Infect, 11: 618-624.

Ghomi, M.R.; Nikoo, M.; Heshmatipour, Z.; Amir, J.A.; Ovissipour, M.; Hashemi, F.L.H.; Hasandoost, M. and Jadiddokhani, D. (2011): Effects of sodium acetate and nisin on microbial and chemical changes and fatty acid composition of grass carp Ctenopharyngodon idella during refrigeration storage. Food Saf.,Doi:10.1111/j.1745-4565.2010.00281.x.

Glass, K.A. and Doyle, M.P. (1989): Fate of Listeria in processed meat products during refrigerated storage. Appl. Environ. Microbiol. 55: 1565-1569.

Golden, M.H.; Buchanan, R.L. and Whiting, R.C. (1995): Effect of sodium acetate or sodium phosphate with EDTA and ascorbic acid on the inactivation  of L. monocytogenes Food Safety Vol.15,1:53-56   

Gram, L. and Huss, H.H. (1996): Microbiological spoilage of fish and fish products. Int. J. Food Microbiol. 33: 121-137.

Grif, K.; Patscheider, F. and Dierich, M.P. (2003): Incidence of fecal carriage of Listeria monocytogenes in three healthy volunteers: a one-year prospective stool survey. Eur. J. Clinic. Microbiol. Infect. Dis. 22: 16-20.

Gudbjornsdottir, B.; Suiko, M.L.; Gustavsson, P.; Thorkelsson, G.; Salo, S. and Sjoberg, A.M. (2004): The incidence of L. monocytogenes in meat, poultry and seafood plants in Nordic Countries. J. Food Microbiol. 21:217-225.

Hartemink, R. and Georgsson, F. (1991): Incidence of Listeria species in sea food and seafood salads. Int. J. of Food Microbiol. 12:189-195.

Hassan, M. and Shole, Y. (2013): Molecular characterization of Listeriamonocytogenes isolated from fresh seafood samples in Iran. http://www.diagnosticpathology.org/content/8/1/149.

Howgate, P. (1985):The self-life of fish products. J. Sci. Food Agric. 36 (2), 126-127.

Huss, H.H. (1995): Quality and quality changes in fresh fish. Food Agriculture Organization (FAO), Fisheries Technical Paper 348. Rome: FAO.

Hussein, A.; Othman, R.E.E.; Sayed, S.M.A.; Hassanein, R. and Abushahahba, F.N.M. (2011): Occurrence of Listeria monocytogenes in poultry, fish and their products as well as its public health hazard on women. Animal hygiene and sustainable livestock production. Proceedings of the XVth International Congress of the International Society for Animal Hygiene, Vienna, Austria, 3-7, Vol.2: 987-992.

Jacquet, C.; Aubert, S.; El Solh, N. and Rocourt, J. (1992): Use of rRNA gene restriction patterns for the identification of Listeria species. Systematic and Appl. Microbiol. 15: 42-46.

Jallewar, P.K.; Kalorey, D.R.; Kurkure, N.V.; Pande, V.V. and Barbuddhe, S.B. (2007): Genotypic characterization of Listeria spp. isolated from freshwater fish. Internat. J. Food Microbiol. 114: 120-123.

Jammi, T. and Keusch, A. (1994): Occurrence of L. monocytogenes in fresh water fish farms and fish smoking plants. Food Microbiol. 11:309-316.

Jannat, A.H.; Shabanpoor, B.; Shabani, A. and Sadeghi, A. (2010): Effects of cooking methods on physico-chemical and nutritional properties of Persian sturgeon Acipenser persicus fillet. Int. Aquat. Res., 2: 15-23.

Jay, J.M. (2000): Modern Food Microbiology. An Aspen Publication and Aspen publishers, Inc. Gaithersburg, Maryland.

Joanne, T.; Kendra, K.N.; Ken, G.; Virginia, N.S. and Martin, W. (2004): Tracking of Listeria monocytogenes in smoked fish processing plants. J. of Food Prot. Vol.67 (2), 328-341.

Johansson, T.; Rantala, L.; PaLMu, L. and Honkanen-Huzalski, T. (1999): Occurrence and typing of L. monocytogenes strains in retail vaccum-packed fish products and in a production plant. Internat. J. Food Microbiol. 47:111-119.

Kamat, A.A. and Nair, P.M. (1994): Incidence of Listeria species in India sea foods and meat. J. Food Safety 14: 117-130.

Karunasagar, I.; Segar, K.; Karunasagar, I. and Goebel, W. (1992): Incidence of Listeria spp. in tropical seafoods. in: Listeria. Abstract No. 155. Eleventh international Symposium on problems of Listeriosis (ISOPOL  XI), May 11^th -14^th, Copenhagen, Denmark.

Kilinc, B.; Cakli, S.; Candun, A. and Sen, B. (2009): Effects of phosphate dip treatments on chemical, microbiological, color, textural and sensory changes of Rainbow trout fillets during refrigerate storage. Aquatic Food Product Teechnology (24), 110-119.   

Kim and Marshall (2002): Influence of catfish skin mucus on trisodium phosphate inactivation of attached Salmonella typhimurium,Edwardsiella tarda, and Listeria monocytogenes. J.Food Prot. 65(7), 1146-1151.

Kim, C.R.; James, O.H; Vickery, A.P.; White, C.H. and Marshall, D.L. (1995): Sodium acetate and Bifidobacteria increase shelf life of refrigerated catfish fillet. Food Sci. 60 (1),    15-27.        

Kim, J.M.; Huang, T.S.; Marshall, M.R. and Wei, C.I. (1999): Chlorine dioxide treatment of seafood to reduce bacterial loads. Food Sci. 64:     1089-1090.

Kouassi, Y. and Shelf, L.A. (1996): Metabolic activities of L. monocytogenes in the presence of sodium propionate,acetate ,lactate and citrate,Appl.Bacteriol. 81(2), 147-153.

Modaresi, R.; Mardani, K.; Tukmechi, A. and Ownagh, A. (2011): Prevalence of Listeria spp. in fish obtained from Urmia fish markets. African J. Microbiol. 5(30), 5398–5401.

Manjua, S.; Ravishankarb, C.N. and Lalithac, K.V. (2007): Effects of sodium acetate dip treatment and vacuum- packaging on chemical, microbiological, textural and sensory changes of (Pearlspot(Etroplus saratensis during chill storage. Food Chemistry. 102(1) 27-37.

Mohamed, G.M. (2012): Comparative study between raw and cooked fish sold in Assiut city on the incidence of some foodborne pathogens. Assiut Vet. Med., J. Vol. 58, 133: 99-108.

Mu, D.; Huang, Y.; Gates, K. and WU, W. (1997): Effectof trisodium phosphate on L. monocytogenes attached to Ranibow trout and shrimp during refrigerated storage. Food Safety. 17(1) 37-46.

Nikolaos, S.; Amin, A.; Konstantinon, P. and Vasikios, S. (2007): Incidence of Listeria spp in fish and environment of markets in Northern Greece. Food Control 18: 554-557.

Nykanen, A.; Weckman, K. and Lapvetalainen, A. (2000): Synergistic inhibition of listeria monocytogenes on cold-smoked Rainbow trout by nisin and sodium lactate. Internat. J. Food Microbiol. 61: 63-72.

Pand, A.K. and Garg, S.R. (2003): Prevalence of Listeria in foods of animal origin. Indian J. Anim. Sci. 73: 967-968.

Papadopoulos,bTh.;bAbrahim, A.; Sergelidis, D.; Kirkoudis, I. and Bitchava, K. (2010): Prevalence of Listeria spp. in freshwater fish (Oncorhynchus mykiss and Carassius gibelio) and the environment of fish markets in Northern Greece. J. Hellenic Vet. Med. Soc.61(1), 15-22.

Rahimi, E.; Shakerian, A. and Raissy, M. (2012): Prevalence of Listeria species in fresh and frozen fish and shrimp in Iran,” Annals of Microbiol. 62:37-40.

Rodas, 0.R.; Flores, J.F.; Betancourt, J.M.; Quinones, E.I. and Vazquez, C. (2006): Occurrence and antibiotic Sensitivity of Lisreria monocytogenes strains isolation from oysters,fish,and estuarine water. Appl, Environ. Microbiol. 72(11), 7410-7412.

Ross, T.; Dalgaard, P. and Tienungoon, S. (2000): Predective modeling of the growth and survival of Listeria in fishery products. Internat. J. Food Microbiol. 62: 231-245.

Ryser, E.T.; Marth, E.H. and Doyle, M.P. (1985): Survival of Listeria monocytogenes during manufacture and storage of cottage cheese. J. Food Prot. 48: 746–750.

Sallam, K.I. (2007 a): Chemical, sensory and shelf life evaluation of sliced salmon treated with salts of organic acids. Food Chem. 101 )2), 592-600.

Sallam, K.I. (2007b): Antimicrobial and antioxidant effects of sodium acetate, sodium lactate and sodium citrate in refrigerated sliced salmon. Food Control. 18(5), 566-575.

Scott, D.; Fletcher, G.C.; Charles, J.C. and Wang, R.J. (1992): Spoilage changes in the deep-water fish Smooth oreodory during storage in ice. Internat. J. Food Sci. Technol. 27: 577-587.

Shole, Y.; Hassan, M.M.D. and Elahe, T.B. (2013): Listeria monocytogenes serotypes in fresh fish, shrimp, crab and lobster in Isfahan and Shahrekord, Iran. Inte. J. of Advan. Biological and Biomedical Research,Volume 1, Issue 5,: 493-504.

Siavash, M.; Ali, F. and Samhra, E. (2011): Incidence of Listeria Species in farmed tropical fish in Khuzestan, Iran. World J. of Fish and Marine Scie. 3(3), 206-209.

SPSS (2007): Sample Power Statistic, SPSS, 12.01Syntax Reference Guide for SPSS Base. SPSS Inc, 233 South Wacker Drive, Chicago, IL.pp111-119.

Taormina, P.J. and Beuchat, L.R. (2003): Survival and heat resistance of Listeria monocytogenes after exposure to alkali and chlorine. Appl. Environ. Microbiol.79: 24-29.

Tassou, C.C.; Lambropoulou, K. and Nychas, G.J.E. (2004): Effect of prestorage treatment and storage condtion on the survival of Salmonella enteritidis PT4 and Listeria monocytogenes on fresh marine and freshwater aquaculture fish. Food Protect. 67(1), 193-198.

Vandekinderen, I.; Devliehgfrf, E.; Van Camp, J.; Kerhaert, B.; Cucu, T.; Ragaert, P.; De Bruyne, J. and De Meulemaer, B. (2009): Effects of food composition on the activation of foodborne microorganisms by chlorine dioxide. Internat. J. Food Microbiol. 131: 38-44.  

Venkitanarayanan, K.S.; Ezeike, G.O.; Hung, Y.C. and Doyle, M.P. (1999): Efficacy of electrolyzed oxdizing water for inactivation of Escherichia coli O157:H7, Salmonella enteritidis and Listeria monocytogenes. Appl. Environ. Microbiol. 65: 4276.

Vyncke, W. (1978): Infuence of sodium tripolyphosphate and citric acid on the shelf life of thornback ray (Raja clavata L.). Zlebensum Unters 28, 166(5)284-286.

Willett, W.C. (2005): Fish balancing health risks and benefits. Amer. J. Prev. Med. 29: 320-32 Zhu, M. Du. M.; Cordray, J. and Ahn, D.U. (2005): Control of Listeria monocytogenes Contamination in Ready-to-Eat Meat Products. Comprehensive Rev. Food Saf., 22: 34-42.