Isolation of Histamine-Producing Bacteria from Fish-Processing Facilities and FishingVessels

Abstract

The presence of histamine-forming bacteria in fishprocessing facilities was studied. Environmental sampling was conducted in the Hampton Roads area of Virginia in fishprocessing facilities that regularly handle scombroid fish or other fish which are known to potentially accumulate histamine levels greater than 50 ppm. Surfaces that come into contact with the fish were swabbed and the histamine-forming isolates from these areas were identified. One isolate each of Klebsiella ozaenaeand Vibrio alginolyticus, and two isolates of Aeromonassp. were found in the processing facilities. The study concluded that histamine-forming bacteria do not comprise a large part of the microflor a associated with fishprocessing facilities. Fishing vessels were also sampled and no histamine-forming bacteria were identified.     

Effect of Vacuum Tumbling with Direct Salting or Brining on Smoked Trout Fillets

Abstract

The following five brining and direct salting treatments were applied with and without vacuum tumbling to rainbow trout, Oncorhynchus mykiss, butterfly fillets: (1) brining, B; (2) direct salting, DS; (3) tumble followed by brining, TB; (4) tumble followed by direct salting, TDS; and (5) tumble with brine, TWB. The water-phase salt content of DS and TDS fillets was the highest while the lowest water-phase salt was found in TWB trout. DS and TDS treatments yielded product closest to the regulatory target of 3.5% water-phase salt. Tumbling followed by brining (TB) produced the softest texture. Tumbling performed before brining did not affect the water-phase salt content of the product.     

Effect of Trisodium Phosphate or Water Dip on the Survival of Salmonella and Listeria monocytogenes Inoculated Catfish Before and After Freezing

Salmonella and Listeria monocytogenes (LM) are occasional contaminants on raw fish. Catfish fillets were artificially contaminated with LM and Salmonella, dipped in a 1.5 % (30 min) trisodium phosphate solution (TSP), and cryogenically frozen. After 3 months frozen storage, Salmonella (2 log), but not LM, was inactivated on the fillets treated with the dip. This indicates that TSP dip followed by cryogenic freezing can be used to control Salmonella, but not LM, on catfish fillets.     

An overview of aquaculture in Turkey: With emphasis on sea bass and sea bream

Abstract

Aquaculture is a relatively recent industry in Turkey, enjoying great potential for development. Both freshwater and marine aquaculture are being practised in over 800 operational units. Rainbow trout (Oncorhynchus mykiss), Sea bream (Sparus aurata), Sea bass (Dicen‐trarchus labrax), comprise 90% of the total production, together with Atlantic salmon (Salmo salar), Common carp (Cyprinus carpio), Shrimp (Penaeidae spp.) and Mussel (Mytilus gal‐loprovincialis). Total aquaculture production reached 45 450 t in 1998, constituting 9% of total national fisheries production. Sea bass and sea bream farms are located along Aegean Sea and Mediterranean coasts and comprised 16% of the total number of aquaculture enterprises and 30% of total aquaculture production in 1997. Supplies of these two species have been steadily increasing in Turkey and other producing countries since 1990 resulting in market saturation and declining prices both locally and internationally. Mediterranean species including Epinephelus spp., Puntazzo puntazzo and Pagrus pagrus, are considered to be new candidates offering good prospects for mariculture.     

Comparative ecologies of domestic and naturalised rainbow trout in northern Lake Huron

Northern Lake Huron (Ontario, Canada) supports the largest concentration of freshwater salmonid cage culture farms in Canada and receives inputs of domestic‐strain rainbow trout (Oncorhynchus mykiss) through escapement. We assessed the potential effects of these domestic fish on the naturalised rainbow trout of this ecosystem by comparing their respective ecologies. Mature adults were sampled from spawning tributaries, primarily in spring, and to a lesser extent in fall. Fish of domestic origin comprised ~80% of rainbow trout sampled from sites near cage culture facilities but <20% of rainbow trout sampled from more distant sites. Domestic adults in spawning condition (gametes free‐flowing) were present in the spawning tributaries in both spring and fall sampling seasons, whereas naturalised fish in spawning condition were only observed in the spring. Domestic adults were younger and smaller (in length), appeared to have shorter lifespan and had a higher male to female ratio compared with their naturalised counterparts. Growth rates (change in length with age) of naturalised and domestic females in the wild were similar, but domestic males grew more slowly than naturalised males. Domestic females also produced smaller eggs than naturalised females. Food web positions (inferred from δ¹³C and δ¹⁵N) of domestic and naturalised fish were very similar but varied more strongly with body size in the former. Domestic‐strain rainbow trout of cage culture origin can survive, grow and attempt to spawn in northern Lake Huron and have the potential to compete for food, mates and spawning habitat with naturalised rainbow trout.     

Selection of lactic acid bacteria as candidate probiotics for Vibrio parahaemolyticus depuration in pacific oysters (Crassostrea gigas)

The application of probiotics in food is now widespread and is widely accepted by consumers. Lactic Acid Bacteria (LAB) were isolated from traditional salted fish and then characterized by its ability to inhibit Vibrio parahaemolyticus growth using minimal inhibitory concentration tests. Five out of these strains were identified as Lactobacillus delbrueckii, Lactobacillus casei and three Lactobacillus gasseri by PCR using 16S and 23S rRNA genes. Antibiofilm activity of Lactobacillus spp. extracts were also tested in 96 polystyrene plates. A potential antibiofilm effect was demonstrated as most LAB. Although most LAB extract were able to eradicate pre‐formed biofilm, results demonstrated that five Lactobacillus spp. exhibited a stronger inhibitory effect against V. parahaemolyticus in infected oysters. Vibrio parahaemolyticus viable cells number declined from 10⁶ UFC to 10⁴ UFC after 3 days of incubation with Lactobacillus spp. Probiotic applications, in biological control of seafood associated pathogens can be an alternative solution, providing consumer with a product of good quality owing to the use of non‐toxic compounds. Based on our results, LAB could be used as a bioprotective culture in oyster's depuration to prevent V. parahaemolyticus growth.     

Occurrence of Vibrio cholerae and Vibrio parahaemolyticus among milkfish farms in Zanzibar

ABSTRACT

Fishing is among the main economic activities of the people of Zanzibar. Few fish dealers are transforming this sector into mariculture. Among the farmed fish is milkfish. Diseases are among the limiting factors in the development of the mariculture industry. Among other zoonotic diseases, vibriosis is caused by bacteria from the genus Vibrio. This study aimed to establish the occurrence of Vibrio cholerae and Vibrio parahaemolyticus among milkfish farms in Zanzibar. A total of 380 milkfish were sampled. Swabs were collected from gills, intestine, and kidney of each sampled milkfish. Preliminary identification of V. cholerae and V. parahaemolyticus was done by biochemical tests. PCR was run on 16S rRNA, outer membrane protein W, and collagenase genes to confirm Vibrio species, V. cholerae, and V. parahaemolyticus respectively. Almost one-third (32.1%) of all sampled milkfish were found to contain targeted Vibrio; 18% and 29.5% of the sampled milkfish were positive for V. cholerae and V. parahaemolyticus respectively.     

Effects of fish species composition on Diphyllobothrium spp. infections in brown trout – is three‐spined stickleback a key species?

Subarctic populations of brown trout (Salmo trutta) are often heavily infected with cestodes of the genus Diphyllobothrium, assumedly because of their piscivorous behaviour. This study explores possible associations between availability of fish prey and Diphyllobothrium spp. infections in lacustrine trout populations. Trout in (i) allopatry (group T); (ii) sympatry with Arctic charr (Salvelinus alpinus) (group TC); and (iii) sympatry with charr and three‐spined stickleback (Gasterosteus aculeatus) (group TCS) were contrasted. Mean abundance and intensity of Diphyllobothrium spp. were higher in group TCS compared to groups TC and T. Prevalence, however, was similarly higher in groups TCS and TC compared to group T. Zero‐altered negative binomial modelling identified the lowest probability of infection in group T and similar probabilities of infection in groups TC and TCS, whereas the highest intensity was predicted in group TCS. The most infected trout were from the group co‐occurring with stickleback (TCS), possibly due to a higher availability of fish prey. In conclusion, our study demonstrates elevated Diphyllobothrium spp. infections in lacustrine trout populations where fish prey are available and suggests that highly available and easily caught stickleback prey may play a key role in the transmission of Diphyllobothrium spp. parasite larvae.     

Effects of the Depuration Process on Microbiological Quality and Nutritional Composition of Mediterranean Mussel (Mytilus galloprovincialis Lamarck, 1819)

In this study, the effects of the depuration period on Mediterranean mussel (Mytilus galloprovincialis L., 1819), collected from Band?rma Harbor Area by the Sea of Marmara, were investigated. Distinctive changes were found to exist in the percentage of moisture, protein, fat, and ash content. The process of depuration was determined to have an effect on the number of total aerobic mesophilic/psychrophilic bacteria. Salmonella spp., Listeria monocytogenes, Vibrio parahaemolyticus, Vibrio cholera, Escherichia coli, and Staphylococcus aureus could not be identified during the study. The substances of aluminum, iron, zinc, copper, manganese, nickel, cadmium, chromium, lead, and potassium were determined to decrease in the Mediterranean mussel from the baseline to the end of the depuration. It was found that the process of depuration has a positive effect on alanine, phenylalanine, lysine, tyrosine, valine, leucine, and isoleucine; a negative effect on aspartic acid, glutamic acid, threonine, proline, and arginine and no effect on methionine, histidine, glycine, and serine. The total saturated fatty acids, total monounsaturated fatty acids, and total polyunsaturated fatty acids at the end of depuration were found to be 29.29%, 12.51%, and 45.57%, respectively, and the change in the amount of polyunsaturated fatty acids was found to be significant (p < 0.05).     

Chitosan Coatings Containing Mentha spicata Essential Oil and Zinc Oxide Nanoparticle for Shelf Life Extension of Rainbow Trout Fillets

ABSTRACT

The objective of the current study was to determine the effects of chitosan (CH) coatings containing Mentha spicata essential oil (MSO; 0.2 and 0.4%) and zinc oxide nanoparticle (ZnO; 0.2 and 0.4%) on chemical (total volatile basic nitrogen (TVB-N), 2-thiobarbituric acid reactive substances (TBARS) and peroxide value (PV)), microbial (total viable count, psychrotrophic count, Pseudomonas spp., and Enterobacteriaceae) and sensory (odor, color, and overall acceptability) properties of raw rainbow trout fillets during 14 days of refrigerated storage. Final microbial population of CH + MSO 0.4% + ZnO 0.4% and CH + MSO 0.4% + ZnO 0.2% retarded approximately 3–5 log CFU/g as compared to control groups (P < 0.05). At the end of storage time in control samples, TVB-N, TBARS, and PV reached 49.3 mg N/100 g, 3.31 mg malonaldehyde/kg, and 2.04 meq peroxide/kg lipid, while the values for the treated samples with MSO (0.2 and 0.4%) in combination with ZnO (0.2 and 0.4%) remained lower than 25 mg N/100 g, 2 mg malonaldehyde/kg, and 1.1 meq peroxide/kg lipid, respectively. The results indicate a potential use of CH coating containing MSO and ZnO as an effective type of antimicrobial packaging to maintain quality of rainbow trout fillets.     

Improving the Quality of Silver Carp Fish Fillets by Gamma Irradiation and Coatings Containing Rosemary Oil

The objective of this work was to study the combined effect of gamma irradiation and coatings containing 0.5% rosemary (Rosmarinus officinalis) essential oil (RO) on the chemical, microbiological, and sensorial qualities of silver carp (Hypophthalmichthys molitrix) fish fillets (SFF) during cold storage (4°C). SFF were divided into three groups: uncoated (control), coated with edible coating (without additives), and γ-irradiated (0, 1, 3, and 5 kGy) coated with coating containing 0.5% rosemary. Gamma irradiation at 1, 3, and 5 kGy with coating reduced the initial total bacterial count, psychrophilic bacteria, and lactic acid bacteria and prolonged shelf life of the samples. Coated samples irradiated at 1 kGy reduced the counts of Enterobacteriaceae, Staphylococcus aureus, and Bacillus cereus, as well as eliminating Vibrio spp. and Salmonella spp., while coated samples irradiated at 3 and 5 kGy completely eliminated these bacteria. Combined treatment showed a slight increase in thiobarbituric acid-reactive substances postirradiation during cold storage but had no effect on the total volatile basic nitrogen and trimethylamine contents, while a gradual increase in these chemical quality indices was observed during cold storage. Combined treatment had no adverse effects on the sensory properties of SFF. The increase in the bacterial inhibitory effect is caused by both rosemary oil and irradiation.     

Microbial Growth Models in Gilthead Sea Bream (Sparus aurata) Stored in Ice

ABSTRACT

This study analyzes microbiological changes in whole, ungutted farmed gilthead sea bream (Sparus aurata) stored for an 18-day period in ice using traditional methods for mesophilic aerobic bacteria, psychrotrophic, Pseudomonas spp., Aeromonas spp., Shewanella putrefaciens, Enterobacteriaceae, sulphide-reducing Clostridium (Clostridia), and Photobacterium phosphoreum in muscle, skin, and gills, evaluating their seasonal differentiation. Two different statistical models were used to analyze microbiological growth. Simultaneously, physicochemical parameters such as the temperature, pH, biological oxygen demand (BOD), total dissolved solids, salinity, ammonia nitrogen, and total phosphorus content of growing waters were analyzed. The results showed that by the end of the storage time, specific spoilage bacteria (SSB) such as Pseudomonas spp., Aeromonas spp., and S. putrefaciens as H₂S-producing bacteria were dominant in sea bream harvested in temperate water in the Canary Islands. Muscle tissue had the least contamination, followed by skin and gills. The values of the analyzed seawater parameters were constant during the four seasons, except that the temperature showed a small difference between winter and summer. Seasonal effects were observed among the fish analyzed, suggesting that the lower levels of contamination detected in winter may have been due to the slight difference observed in water temperature in that season.     

Exploring the microbial diversity of the distal intestinal lumen and mucosa of farmed rainbow trout Oncorhynchus mykiss (Walbaum) using next generation sequencing (NGS)

In this study, next generation sequencing (NGS) was used to survey the 16S rRNA ribotypes of the distal intestinal lumen and mucosal epithelium of farmed rainbow trout. This approach yielded a library consisting of 2 979 715 quality filtered paired sequences, assigned to genus level of taxonomy using the Ribosomal Database Project (RDP). A high level of diversity was observed in both regions. A total of 90 bacterial genera were identified in the lumen of all fish sampled, compared with 159 in the mucosa. The allochthonous microflora was dominated by sequences belonging to the γ Proteobacteria (mean sequence abundance 54.3%), in particular the Enterobacteriaceae, with Yersinia, Serratia, Hafnia and Obesumbacterium the most abundant genera. Fewer γ Proteobacteria (mean sequence abundance 37%) were present in the mucosa, and autochthonous communities consisted of a more even split among the bacterial classes, with increases in sequences assigned to members of the β Proteobacteria (mean sequence abundance 18.4%) and Bacilli (mean sequence abundance 16.8%). The principal bacterial genera recorded in the mucosa were Cetobacterium, Yersinia, Ralstonia, Hafnia and Carnobacterium. The results of this study demonstrate that the luminal and mucosal bacterial communities may be different in their respective structures, and that the mucosal microflora of rainbow trout may be more diverse than previous research has suggested. This research also demonstrated a degree of conservation of bacterial genera between individual fish sampled, and is to the author's knowledge the first time the MiSeq^® NGS platform has been used to explore the rainbow trout intestinal microflora.     

Production Characteristics of Rainbow Trout,Oncorhynchus mykiss and Brook Trout,Salvelinus fontinalis Under Seasonal Pond Conditions

Abstract

The production characteristics of juvenile rainbow trout, Oncorhynchus mykiss and brook trout, Salvelinus fontinalis were compared under winter pond conditions. Juvenile rainbow trout (55.1 ±1.5 g) and brook trout (28.9 ±0.4 g) were stocked at a density of 8,750 fish/ha into six 0.04-ha ponds. After 163 days, survival, growth, and feed conversion were similar (P >0.05). The results of this study suggest that brook trout may attain growth rates similar to rainbow trout under winter pond conditions in temperate regions of North America.     

Pathogenicity of Vibrio harveyi to salmonids

Out of 19 Vibrio harveyi isolates obtained from a diversity of hosts and geographical locations, 14 were pathogenic to rainbow trout, Oncorhynchus mykiss (Walbaum), and Atlantic salmon, Salmo salar L., with mortalities of up to 100% following intraperitoneal injections of 106 cells fish?1. The extracellular products (ECPs) of only five pathogenic isolates were harmful to fish. Both pathogenic and non‐pathogenic cultures produced ECPs containing caseinase, gelatinase, phospholipase, lipase and haemolysins. Vibrio harveyi VIB 645, which was the most pathogenic isolate, produced ECPs with a maximal effect on salmonids from preparations obtained by using cellophane overlays on tryptone soya agar supplemented with 1% (w/v) sodium chloride with incubation at 28 °C for 24 h. This preparation contained the highest titre of haemolytic activity to Atlantic salmon (1:256) and rainbow trout (1:32) erythrocytes.     

Microbiological quality of Litopenaeus vannamei culture using conventional and biofloc systems

Shrimp farming is a fast‐expanding activity that has supported the growth in the supply of these crustaceans to consumers around the world. However, the end product is vulnerable to contamination at all stages of the process, including the rearing tanks, where current practices prioritize to raise stocking densities and the minimization of water renewal. It is thus important to evaluate the potential of these systems for the proliferation of undesirable microorganisms, which may render the product unfit for human consumption. In the present study, the presence of coagulase‐positive Staphylococcus, Salmonella spp., Vibrio spp., and total and thermotolerant coliforms was verified in biofloc tank and conventional pond systems used for the rearing of Litopenaeus vannamei in Pernambuco, Brazil, and the results were compared with the legislation regulating the marketing of fresh shrimp. Samples were collected from two biofloc tanks with a density of 375 shrimp m^?2, and two conventional ponds with 12 shrimp m^?2. None of the samples tested positive for either Salmonella spp. or coagulase‐positive Staphylococcus, which is consistent with the legislation. While no standards are defined legally for Vibrio spp. or coliforms, very low concentrations were recorded in both systems, in comparison with other studies. While some variation in bacterial contamination was observed over the rearing process, the end product of both systems was fit for human consumption. The results of the study indicate that, while the water is not renewed in the biofloc system, the development of undesirable microorganisms can be controlled, with no adverse effects for the end product.     

Mysterious syndrome causing high mortality in wild brown trout in Eastern Switzerland,pathology and search for a possible cause

Since 2016, annually occurring species-specific die-offs of brown trout (Salmo trutta fario) occurred in the Thur River, situated in the Eastern part of Switzerland. These events lead to drastically reduced population densities in the impacted river regions. Clinical signs in brown trout and mortality were restricted to few weeks in August/September. To characterize the syndrome and to find possible causes, from end of March to November 2018, one-year-old brown trout (Salmo trutta fario) and rainbow trout (Oncorhynchus mykiss) were exposed to water from Thur River, fish were sampled regularly and screened for infectious agents, including viral metagenomics, and pathology was described. Starting approximately four months post-exposure, brown trout showed severe lymphohistiocytic pancarditis and necrotizing and haemorrhagic hepatitis. These lesions were recorded until the end of the experiment in November. Rainbow trout were not affected at any point in time. No infectious agents could be identified so far as cause of disease, especially no viral aetiology. Even if pathogenesis and pathology point in the direction of an infectious agent, a causative relationship could not be confirmed and aetiology remains unclear.     

A simplified method to estimate Diphyllobothrium spp. infection in salmonids

Some fish parasites constitute severe management problems as they may cause mortality of their fish host or are important zoonoses of humans. Parasite assessments are therefore critical to keep track of infections. If conventional sampling techniques can be simplified, parasite assessments might be easier to obtain, less time‐consuming and more extensive. In this study, we compare the assessed number of Diphyllobothrium spp. cysts (CYST) with the counted number of Diphyllobothrium spp. plerocercoid larvae recovered using a conventional digestive technique (LARV). The aim was to determine the potential of using CYST as a simplified methodology for assessing Diphyllobothrium spp. infection in salmonids. In total, 365 brown trout and 424 Arctic charr were sampled from nine lakes in subarctic Norway. Strong correlation, significant linear relationship and large amount of explained variation were found between log₁₀CYST and log₁₀LARV in both fish species. The method had a slight, but not significant tendency to work better in charr compared to trout. In addition, absolute difference between CYST and LARV increased at parasite intensities >100 indicating that the method has reduced functionality when estimating parasite intensity in heavily infected salmonid populations. However, overall, using this simplified and less time‐consuming methodology, a good indication of Diphyllobothrium spp. intensity, abundance and prevalence was obtained. We suggest that this method provides a sound proxy of the Diphyllobothrium spp. burden and have the potential to be used in parasite assessment during fish monitoring and fisheries management surveys, particularly if the time and resources for detailed parasite studies are not available.     

Microbiological controls across the productive cycle of Dicentrarchus labrax L. and Sparus aurata L.: a study from the environment to the final product

The microbiological quality of water, sediments and seafood products of three Sicilian marine fish farms (Castellammare Gulf, Capo d'Orlando and Porto Palo) was investigated in order to draw a complete picture of the health conditions from the start (environment) to the end (seafood product) of the productive cycle. Before the beginning of fish farming, low concentrations of faecal contamination indicators (faecal coliforms, Escherichia coli, enterococci) were found in the water of each examined area. Due to the enhanced organic load released from cages, the set‐up of fish farming significantly altered the distribution of faecal indicators and sometimes that of halophilic vibrios in the pelagic compartment. Significant differences in the density of heterotrophic bacteria were sometimes recorded at the sediment level. Despite this increase in microbial abundance, the microbiological conditions remained acceptable for the productive process. Pathogens (Salmonella spp., Vibrio parahaemolyticus, Listeria monocytogenes, Staphylococcus aureus) were mostly absent in seafood products. The study underlines that the achievement of good quality levels in aquaculture strongly depends on the conformity of the rearing environment to qualitative microbiological standards, highlighting the importance of sanitary controls along the different steps of the productive cycle.     

Effects on Lipid Oxidation and Bioactive Properties of Rainbow Trout Fillets Fed with Barley

Barley concentrations ranging from 0% to 32% were incorporated into rainbow trout (Oncorhynchus mykiss) diets. The effect of barley concentration on lipid peroxidation and antioxidant activity of fish fillets were analyzed. Results showed that the inclusion of barley in rainbow trout diets had an inhibitory effect on lipid oxidation, probably associated with certain bioactive compounds in barley that could interact with scavenging and reducing metabolites involved in lipid oxidation. Concentrations up to 8% of barley produced fish fillets with high antioxidant activity and higher levels of alpha-tocopherol.