Quorum quenching Bacillus spp.: an alternative biocontrol agent for Vibrio harveyi infection in aquaculture.

Quorum sensing (QS) is a type of cell to cell communication in bacteria that can also regulate the virulence potential in pathogenic strains. Hence, QS disruption, i.e. the quorum quenching (QQ) mechanism, is presently being explored as a novel bio-control strategy to counter bacterial infections. In the present study, we characterized the QQ ability of Bacillus spp. strains to reduce the expression of some virulence factors of a shrimp pathogen, Vibrio harveyi. We screened a total of 118 spore-forming bacterial isolates from aquaculture ponds and mangrove soil for their ability to degrade the synthetic N-acyl-homoserine lactones (AHLs) C4-HSL, C6-HSL, C8-HSL, and C10-HSL. We then selected the top 17 isolates with high AHL-degradation ability for further study. Among them, B. subtilis MFB10, B. lentus MFB2, and B. firmus MFB7 had the highest ability for degradation. These 3 isolates suppressed the expression of virulence genes encoding protease, lipase, phospholipase, caseinase, chitinase, and gelatinase, and potentially inhibited the biofilm formation of V. harveyi MFB32. The reduction in expression of virulence genes like those coding for metalloprotease, serine protease, and haemolysin were confirmed by real-time PCR analysis. Moreover, in an in vivo challenge experiment, these Bacillus spp. protected Penaeus monodon post-larvae against V. harveyi MFB3 infection. Our results demonstrate the potential application of AHL-degrading Bacillus spp. as an alternative to antibiotics in shrimp hatcheries to control luminescent vibriosis. This novel bio-therapeutic method is a promising approach towards disease control in shrimp aquaculture.

Characterization of V. cholerae O1 biotype El Tor serotype Ogawa possessing the ctxB gene of the classical biotype isolated from well water associated with the cholera outbreak in Kerala, South India.

We investigated 22 water samples (17 well water and five pipe water - both chlorinated) and six soil samples from the surroundings of wells of the households of suspected patients from Palakkad district, Kerala (India), from where a cholera outbreak was reported during June-July 2016. A total of 25 Vibrio cholerae isolates were collected from three well water samples during a recent cholera outbreak. Biochemical and serological studies revealed that all of the isolates belonged to serogroup O1, biotype El Tor, serotype Ogawa. PCR assays confirmed the occurrence of ctxB, ctxA, hlyA, tcpA El Tor,VPI, ace, zot, ompW, rfbO1 and toxR genes in all isolates. The presence of the ctxB gene of the classical biotype in all of the El Tor isolates suggests that it is a new variant of El Tor biotype. Antibiogram profile of all V. cholerae O1 isolates revealed resistance towards five classes of antibiotics island and indicates that they were multidrug resistant. ERIC-PCR and PFGE finger prints showed the clonal relationship among the V. cholerae O1 isolates. The results of this study revealed the emergence of a new variant of El Tor biotype in the water samples from Palakkad district, from where a cholera outbreak was reported.

Vibrio harveyi virulence gene expression in vitro and in vivo during infection in black tiger shrimp Penaeus monodon.

Luminescent Vibrio harveyi is common in sea and estuarine waters. It produces several virulence factors and negatively affects larval penaeid shrimp in hatcheries, resulting in severe economic losses to shrimp aquaculture. Although V. harveyi is an important pathogen of shrimp, its pathogenicity mechanisms have yet to be completely elucidated. In the present study, isolates of V. harveyi were isolated and characterized from diseased Penaeus monodon postlarvae from hatcheries in Kerala, India, from September to December 2016. All 23 tested isolates were positive for lipase, phospholipase, caseinase, gelatinase and chitinase activity, and 3 of the isolates (MFB32, MFB71 and MFB68) showed potential for significant biofilm formation. Based on the presence of virulence genes, the isolates of V. harveyi were grouped into 6 genotypes, predominated by vhpA+ flaB+ ser+ vhh1- luxR+ vopD- vcrD+ vscN-. One isolate from each genotype was randomly selected for in vivo virulence experiments, and the LD50 ranged from 1.7 ± 0.5 × 103 to 4.1 ± 0.1 × 105 CFU ml-1. The expression of genes during the infection in postlarvae was high in 2 of the isolates (MFB12 and MFB32), consistent with the result of the challenge test. However, in MFB19, even though all genes tested were present, their expression level was very low and likely contributed to its lack of virulence. Because of the significant variation in gene expression, the presence of virulence genes alone cannot be used as a marker for pathogenicity of V. harveyi.

Strain variability and pathogenic potential of tdhcarrying Vibrio parahaemolyticus isolated from diseased Penaeus monodon.

A disease outbreak in 42-d-old black tiger shrimp Penaeus monodon juveniles from a commercial aquaculture farm in Kerala, India, was investigated. The cause of the disease outbreak was confirmed as Vibrio parahaemolyticus by biochemical tests, PCR targeting the toxR gene and pathogenicity testing of the isolates. All of the isolates tested negative by PCR specific for V. parahaemolyticus associated with acute hepatopancreatic necrosis disease (AHPND), implicating vibriosis unrelated to AHPND as the cause of mortality. Among the 19 isolates obtained, 2 possessed the tdh gene (coding for thermo-stable hemolysin), whereas none of the isolates possessed trh. The LD50 value of 8 isolates of V. parahaemolyticus from diseased and apparently healthy shrimp ranged from 2.7 × 104 to 4.9 × 105 CFU ml-1 by immersion challenge of P. monodon postlarvae. BOX-PCR and dendrogram analysis of the bacterial isolates revealed that the isolates from moribund and apparently healthy shrimp formed separate clusters, indicating that these isolates originate from separate clones. The isolates from moribund shrimp including tdh-positive V. parahaemolyticus clustered together. The present study represents the first report of tdh-positive V. parahaemolyticus causing disease in a shrimp farm.

Prevalence, molecular characterization, genetic heterogeneity and antimicrobial resistance of Listeria monocytogenes associated with fish and fishery environment in Kerala, India.

The prevalence of Listeria monocytogenes in the retail fish markets of the Kerala, India was investigated by screening 227 samples comprising of marine finfish (n = 97) shellfish (n = 19), ready-to-cook fish products (n = 47), ready-to-eat fish products (n = 10), dried fish (n = 11) and retail ice (n = 43). The prevalence of L. monocytogenes and L. innocua was 2·7% and 17·2% respectively. Sample category wise, prevalence of L. monocytogenes was higher in marine finfish (1·8%) and retail ice (0·9%). All the L. monocytogenes isolates carried virulent genes namely inlA, inlC, inlJ, hlyA, iap, plcA, prfA genes and majority (82%) belonged to 1/2a, 3a serogroups. L. monocytogenes isolates were multidrug-resistant and showed resistance to ampicillin, penicillin, erythromycin, tetracycline and clindamycin. Enterobacterial repetitive intergenic consensus-polymerase chain reaction (ERIC-PCR) delineated 58% genetic heterogeneity among the L. monocytogenes strains. The study reports that genetic similarities of the isolates were interlinked to their serogroup and sample origin. SIGNIFICANCE AND IMPACT OF THE STUDY: Prevalence of Listeria monocytogenes, in the retail fish markets of Kerala, India was low but their relatively higher presence in marine finfish and retail ice and virulent nature of the isolates signifies food safety concerns. Moreover, multidrug-resistant nature of these isolates may potentially lead to spread of antimicrobial resistance. This study identified retail ice as a vehicle for entry of L. monocytogenes in retail fish and hence, there is a need to ensure quality of retail ice used for maintaining the cold-chain.

Critical Role of Monoclinic Polarization Rotation in High-Performance Perovskite Piezoelectric Materials.

High-performance piezoelectric materials constantly attract interest for both technological applications and fundamental research. The understanding of the origin of the high-performance piezoelectric property remains a challenge mainly due to the lack of direct experimental evidence. We perform in situ high-energy x-ray diffraction combined with 2D geometry scattering technology to reveal the underlying mechanism for the perovskite-type lead-based high-performance piezoelectric materials. The direct structural evidence reveals that the electric-field-driven continuous polarization rotation within the monoclinic plane plays a critical role to achieve the giant piezoelectric response. An intrinsic relationship between the crystal structure and piezoelectric performance in perovskite ferroelectrics has been established: A strong tendency of electric-field-driven polarization rotation generates peak piezoelectric performance and vice versa. Furthermore, the monoclinic M_{A} structure is the key feature to superior piezoelectric properties as compared to other structures such as monoclinic M_{B}, rhombohedral, and tetragonal. A high piezoelectric response originates from intrinsic lattice strain, but little from extrinsic domain switching. The present results will facilitate designing high-performance perovskite piezoelectric materials by enhancing the intrinsic lattice contribution with easy and continuous polarization rotation.

Multilocus Sequence Typing and Staphylococcal Protein A Typing Revealed Novel and Diverse Clones of Methicillin-Resistant Staphylococcus aureus in Seafood and the Aquatic Environment.

Methicillin-resistant Staphylococcus aureus (MRSA) has been a global health concern since the 1960s, and isolation of this pathogen from food-producing animals has been increasing. However, little information is available on the prevalence of MRSA and its clonal characteristics in seafood and the aquatic environment. In this study, 267 seafood and aquatic environment samples were collected from three districts of Kerala, India. Staphylococcal protein A (spa) typing and multilocus sequence typing (MLST) was performed for 65 MRSA strains isolated from 20 seafood and aquatic environment samples. The MRSA clonal profiles were t657-ST772, t002-ST5, t334-ST5, t311-ST5, t121-ST8, t186-ST88, t127-ST1, and two non-spa assignable strains. Whole spa gene sequence analysis along with MLST confirmed one strain as t711-ST6 and another as a novel MRSA clone identified for the first time in seafood and the aquatic environment with a t15669 spa type and a new MLST profile of ST420-256-236-66-82-411-477. The MRSA strains were clustered into five clonal complexes based on the goeBURST algorithm, indicating high diversity among MRSA strains in seafood and the aquatic environment. The novel clone formed a separate clonal complex with matches to three loci. This study recommends large-scale spa typing and MLST of MRSA isolates from seafood and the aquatic environment to determine the prevalence of new MRSA clones. This monitoring process can be useful for tracing local spread of MRSA isolates into the seafood production chain in a defined geographical area.

Effect of chitosan on shelf life of restructured fish products from pangasius (pangasianodon hypophthalmus) surimi during chilled storage.

In the present study, restructured products were prepared from pangasius surimi and their qualities were analysed under chilled storage. Pangasius surimi had 75.82 % moisture, 16.91 % protein, 2.76 % fat and 0.95 % ash. Restructured products were prepared in three different formulations by incorporating corn starch (10 %) and chitosan (0.75 %). Formulation containing only corn starch (10 %) was served as control. In all the formulations, mono unsaturated fatty acids were higher (45.14 %). The total volatile base nitrogen (TVB-N) showed an increasing trend and it was found to be higher in control (4.8 mg/100 g) on 10(th) day than the chitosan incorporated sample (3.5-4.2 mg/100 g) on 17(th) day during chill storage. Similarly, peroxide value (PV) was found to higher (8.85 milliequivalent of O2/kg) in control than the chitosan incorporated sample (4.5-6.8 milliequivalent of O2/kg) on 10(th) day. All the three formulations had an acceptable level of thiobarbituric acid (TBA) value that ranged between 0.023-0.098 mg of malanoldehyde/kg during chilled storage. Based on the sensory and microbiological analysis, products prepared without chitosan had a shelf life of 10 day whereas, products incorporated with chitosan had an extended shelf life of 17 day.

Occurrence of viral pathogens in Penaeus monodon post-larvae from aquaculture hatcheries.

Viral pathogens appear to exert the most significant constraints on the growth and survival of crustaceans under culture conditions. The prevalence of viral pathogens White Spot Syndrome Virus (WSSV), Hepatopancreatic Parvo Virus (HPV), Monodon Baculo Virus (MBV) and Infectious Hypodermal and Hematopoietic Necrosis Virus (IHHNV) in Penaeus monodon post-larvae was studied. Samples collected from different hatcheries and also samples submitted by farmers from Kerala were analyzed. Out of 104 samples collected, WSSV was detected in 12.5% of the post-larvae samples. Prevalence of concurrent infections by HPV, MBV and WSSV (either dual or triple infection) was present in 60.6% of the total post-larvae tested. Out of the 51 double positives, 98% showed either HPV or IHHNV infection. HPV or IHHNV was detected in 11 post-larval samples showing triple viral infection. This is the first report of IHHNV from India. Result of this study reveals the lack of efficient screening strategies to eradicate viruses in hatchery reared post-larvae.

Biochemical, textural, microbiological and sensory attributes of gutted and ungutted sutchi catfish (Pangasianodon hypophthalmus) stored in ice.

Pangasianodon hypophthalmus (sutchi catfish) is a fresh water catfish extensively being cultured in the South East Asian countries in the recent years. The present study provides the first report on the effects of gutting on the quality characteristics of aquacultured sutchi catfish stored in ice. pH of whole ungutted and gutted catfish didn't show significant difference (p > 0.05) during ice storage period. Total Volatile Base Nitrogen (TVB-N), Alpha Amino Nitrogen (AAN), Free Fatty Acids (FFA) and Thio Barbituric Acid Reactive Substance (TBARS) were lower in gutted fish compared to whole ungutted fish at any particular day during ice storage. However, gutted fish expressed higher rate of primary lipid oxidation than ungutted fish. Textural degradation of the fish muscle as indicated by hardness, cohesiveness, springiness and chewiness was lower in gutted fish. Results of sensory evaluation revealed that gutting has significantly improved the sensory quality of the fish. However, microbiological analysis revealed higher Total Plate Count (TPC) and Enterobactereaceae count in gutted fish. The shelf life of gutted and whole ungutted sutchi cat fish as determined by microbiological analysis was 16-18 days and 18-20 days respectively while storage in ice.

White spot syndrome virus infection: Threat to crustacean biodiversity in Vembanad Lake, India.

The Vembanad Lake located on the south-west coast of India, an ecological hotspot is the nursing ground of many economically important crustaceans. The prevalence of white spot syndrome virus (WSSV) among crustaceans from farmed, estuarine and marine environments surrounding the Vembanad Lake, India was detected using PCR. A total of 308 samples from aquaculture ponds consisting of six species of crustaceans collected from five different farms were tested for the presence of WSSV. Of these, 67% were found to carry the virus. A total of 258 samples of crustaceans from the Cochin backwater system that forms a part of the Vembanad lake viz., Metapenaeus dobsoni, Metapenaeus monoceros, Penaeus monodon and Penaeus indicus were found to contain WSSV in 62% of the samples. Fifteen species of crustaceans caught from the seas off Cochin were also screened for the presence of WSSV. Out of these, twelve species had WSSV incidence levels ranging from 6-23%. WSSV was not detected from three species of deep sea crustaceans tested. The black tiger shrimp, Penaeus monodon had the highest incidence of WSSV among the species screened in farmed, estuarine and marine environments.

Draft Genome Sequence of the Halophilic and Highly Halotolerant Gammaproteobacteria Strain MFB021.

We report the 4.25-Mbp first draft sequence of Gammaproteobacteria strain MFB021, a moderate halophile isolated from petroleum-contaminated soil in Cochin, India. The genome of the strain MFB021 was sequenced to understand the mechanism of hydrocarbon degradation and the halophilicity of the bacterium.

First draft genome sequence of a member of the genus mangrovibacter, isolated from an aquaculture farm in India.

Mangrovibacter sp. MFB070, a Gram-negative, facultatively anaerobic, nitrogen-fixing bacterium, was isolated from an aquaculture farm in Cochin, India. Here, we report the first draft genome sequence of a member of the genus Mangrovibacter, which may help us to elucidate the evolutionary status of this genus. The draft genome sequence of the Mangrovibacter sp. consists of 5,361,682 bp, encoding 4,971 predicted coding sequences in 57 contigs.

Identification of Degradation Products and a Stability-Indicating RP-HPLC Method for the Determination of Flupirtine Maleate in Pharmaceutical Dosage Forms.

In this stability-indicating, reversed-phase high-performance liquid chromatographic method for flupiritine maleate, forced degradation has been employed and the formed degradants were separated on a C18 column with a 80:20% v/v mixture of methanol-water containing 0.2% (v/v) triethylamine; the pH was adjusted to 3.1. The flow rate was 1 mLmin(-1) and the photodiode array detection wavelength was 254 nm. Forced degradation of the drug was carried out under acidic, basic, thermal, photolytic, peroxide, and neutral conditions. Chromatographic peak purity data indicated no co-eluting peaks with the main peaks. This method resulted in the detection of seven degradation products (D1-D7). Among these, three major degradation products from acidic and basic hydrolysis were identified and characterized by (1)H-NMR, (13)C-NMR, and mass spectral data. The method was validated as per International Conference on Harmonization guidelines (Q2). The linearity of the method was in the concentration range of 20-120 µgmL(-1). The relative standard deviations for intra- and interday precision were below 1.5%. The specificity of the method is suitable for the stability-indicating assay.

Digoxigenin-labeled probe-based colony blotting assay for rapid quantification of Salmonella serovars in seafood and water.

A non-radio-labeled probe-based detection method was developed for rapid enumeration of Salmonella in seafood and water samples. A Salmonella-specific invA gene probe was developed using a digoxigenin-based non-radio labeling assay, which was evaluated with naturally contaminated seafood and water samples. The probe-based technique was further compared with the quantitative PCR assay. The method was specific for detection of different Salmonella serovars without any nonspecific hybridization with other Salmonella-related Enterobacteriaceae. The optimum labeling efficiency was determined for the labeled probe, and 10 pg/microL probe concentration was observed to be most efficient for detection of Salmonella colonies on nylon membrane. Quantification of Salmonella in naturally contaminated seafood and water samples (n = 21) was in the range 10-10(2) CFU/mL. The assay successfully quantified Salmonella in spiked seafood and water samples in the presence of background flora, and the entire assay was completed within 48 h. The probe-based assay was further evaluated with real-time PCR, and results showed that the assay was comparable to real-time PCR assay. Thus, this probe-based assay can be a rapid, useful, and alternative technique for quantitative detection of Salmonella in food, feed, and water samples.

Differential gene expression profile of the hepatopancreas of white spot syndrome virus infected Fenneropenaeus indicus by suppression subtractive hybridization.

Suppression Subtractive Hybridization was employed in order to identify the differentially expressed genes in the hepatopancreas of white spot syndrome virus infected Fenneropenaeus indicus. A forward subtracted cDNA library generated 356 clones following a white spot syndrome virus infection. A total of 345 clones with more than 100 nucleotides were selected for further analysis using bioinformatics tools after vector screening. Twenty-three contigs and 111 singletons were generated from a total of 134 consensuses. The consensuses, on a sequence homology search using BLASTX (NCBI), revealed that 74 (55%) of them had no significant match to reported sequences in the database, suggesting that they were found for the first time and are probably associated with shrimp immune function. Out of the remaining 60 (45%) consensuses, 43 had significant homology to known protein sequences in the database while 17 consensuses are homologous to unknown proteins in the database which are considered novel. The most abundant genes in the subtracted library were antimicrobial peptides accounting for 56 clones; among which one is a member of SNF2 family of proteins and another belonged to PfP1 family of proteins on analysis using Antimicrobial peptide predictor software. The other predicted genes in the subtracted library include signal transduction molecules (GTPase, Serine threonine kinase, Armadillo repeats etc), antioxidant enzymes (Cytochrome oxidase, Monomeric sarcosine oxidase and Catalase), active transporters (Nuclear Localization Signal [NLS], calcium ATPase, sodium glutamate symporter, Store-Operated Calcium Entry [SOCE] and ribonucleoprotein [RNP]) contributing to 19, 14 and 5 clones respectively. Three clones are homologous to reverse transcriptase; a first time report in shrimp and one each belong to cell adhesion molecule and Proteinase. InterProScan at EMBL, when used for an integrated search at PROSITE predicted; signal sequences and transmembrane regions for 13 clones. This is the first report on the differential gene expression in WSSV-infected F. indicus. The high expression of immune related genes in response to virus infection in shrimp will provide a new insight into the crustacean innate immunity. Further work on the functionality of the unknown genes in shrimps will give an overview on the role of the differentially expressed genes during viral infection and increase our understanding for developing antiviral measures by making use of the shrimp defense mechanism.

Effect of reduced oxygen atmosphere and sodium acetate treatment on the microbial quality changes of seer fish (Scomberomorus commerson) steaks stored in ice.

The effect of reduced oxygen atmosphere and sodium acetate treatment on the microbial quality of seer fish (Scomberomorus commerson) steaks was determined during chilled storage (1-2 degrees C). The O2 absorber reduced the oxygen content in the pack to less than 0.01% corresponding to 99.96% reduction within 24 h. The use of O2 absorber with sodium acetate dip treatment (2% w/v) extended the sensory shelf life up to 25 days compared to only 12 days for control air packs and 20 days for untreated samples with O2 absorber. A prominent lag phase was observed for many bacterium studied, particularly for the sodium acetate treated samples with O2 absorber. On the day of sensory rejection, both the total mesophilic and psychrotrophic counts reached 7.7-8.1 and 7.1-7.9 log cfu/g, respectively. The sodium acetate treatment and reduced O2 atmosphere affected the type of major spoilers. In air packed samples, H2S-producers predominated followed by Brochothrix thermosphacta, Pseudomonas spp., where as in the untreated samples with O2 absorber, H2S-producers predominated the microbial flora followed by Lactobacillus spp. For treated samples with O2 absorber, B. thermosphacta formed the major micro-flora followed by Lactobacillus spp. The use of O2 absorber inhibited the growth of Pseudomonas spp., and total Enterobacteriaceae.

Effect of packaging atmosphere on the microbial attributes of pearlspot (Etroplus suratensis Bloch) stored at 0-2 degrees C.

Effect of packaging atmosphere (air and under different modified atmospheres (MAs), 40% CO2/60% O2, 50%/50% O2, 60% CO2/40% O2, 70% CO2/30% O2 and 40% CO2/30% O2/30% N2) on the microbial and biochemical attributes of fresh pearlspot (Etroplus suratensis Bloch) stored at 0-2 degrees C was investigated. Trimethylamine nitrogen (TMA-N) and thiobarbituric acid (TBA) values remained lower than the proposed acceptability limits throughout the storage period. Results demonstrated that storage of pearlspot under air and MA 40% CO2/30% O(2)/30% N(2) resulted in growth of Enterobacteriaceae, Aeromonas and H(2)S-producing bacteria including Shewanella putrefaciens, while all other packaging atmospheres did not allow multiplication of Enterobacteriaceae and Aeromonas within 3 weeks. Aeromonas spp. identified were Aeromonas hydrophila, Aeromonas veronii biovar sobria and A. veronii biovar veronii. Significant reduction (p<0.01) was noticed in Aeromonas population of pearlspot stored under MA 60% CO2/40% O2 and 70% CO2/30% O2. A delay of growth of Pseudomonas below 5.0log(10)cfug(-1) was observed during the 15th day of storage at 0-2 degrees C under modified atmosphere packaging (MAP) conditions. Growth of faecal streptococci was significantly inhibited in all the packaging atmospheres at 0-2 degrees C during the entire storage period. Survival of coagulase positive Staphylococci (<50cfug(-1)) in low numbers was noticed during storage in all the packaging atmospheres. Clostridium botulinum toxin was not detected. All the packaging atmospheres did not allow multiplication of sulphite-reducing clostridia at 0-2 degrees C during the entire storage period. Packaging in MA 60% CO2/40% O2 resulted in the inhibition of growth of Aeromonas and Enterobacteriaceae, and the slowest growth of psychrotrophic bacteria, H(2)S-producing bacteria, including Shewanella putrefaciens and Pseudomonas and extended microbiological shelf life to 9-10 days. This study confirms the survival of potentially pathogenic A. hydrophila, A. veronii biovar sobria and A. veronii biovar veronii capable of growth at low temperature in pearlspot stored under MA.

Microbiological and biochemical changes in pearl spot (Etroplus suratensis Bloch) stored under modified atmospheres.

AIMS: This study aimed to determine the effect of packaging [air, modified atmosphere (MA)] on microbial growth, sensory and chemical parameters and also on shelf life of fresh pearl spot (Etroplus suratensis Bloch) and on the selection of microbial association. METHODS AND RESULTS: Fresh pearl spot (whole, gutted) were packaged under both 100% air and MAs (40%CO(2)/60% O(2), 50%CO(2)/50%O(2), 60% CO(2)/40%O(2), 70% CO(2)/30% O(2) and 40% CO(2)/30% O(2)/30% N(2)) and stored at 0 degrees C. Microbial growth (counts of total aerobic bacteria, H(2)S-producing bacteria, Lactic acid bacteria, Brochothrix thermosphacta, yeast and mould), chemical spoilage indicators (pH, total volatile basic nitrogen) and sensory characteristics were monitored. Microbial changes in Pearl spot packed under 100% air and 40% CO(2)/30%O(2)/30% N(2) were similar. The total volatile basic nitrogen values increased, but the values never exceeded the acceptability limit of 25 mg 100 g(-1). CONCLUSIONS: MA 60% CO(2) : 40%O(2) was found to be better with a shelf life of 21 days whereas air stored samples had a shelf-life of 12-14 days only. SIGNIFICANCE AND IMPACT OF THE STUDY: Storage of pearl spot under MAs 60% CO(2) : 40%O(2) is a promising method to extend shelf-life. Longer shelf life expands the market potential of pearl spot and reduces waste during distribution and retail display.

Occurrence of Clostridium botulinum in fresh and cured fish in retail trade in Cochin (India).

The occurrence of Clostridium botulinum in fresh (67) and cured fish (278) samples in retail trade in Cochin was studied. An overall prevalence of 19% (13/67) was found in fresh retail fish and types A to D were detected in the positive samples. In pelagic fish, incidence of C. botulinum was 18% (7/39) whereas in demersal fish, 21% (5/24) of the samples harboured C. botulinum. Incidence of C. botulinum in shrimp was 25% (1/4) and type D was found in the positive sample. Of 257 cured fish in retail trade tested for the presence of C. botulinum, the overall contamination level was 10% (25/257). Among the C. botulinum types A to D prevalent in fresh fish, types C and D were the predominant types found in cured products. In salt-dried shrimp, of the 21 samples analysed, 10 samples (48%) harboured C. botulinum.