Improving the microbiological safety and quality of aquatic products using nonthermal processing.

Spoilage and deterioration of aquatic products during storage are inevitable, posing significant challenges to their suitability for consumption and the sustainability of the aquatic products supply chain. Research on the nonthermal processing of fruit juices, probiotics, dairy products, and meat has demonstrated positive outcomes in preserving quality. This review examines specific spoilage bacteria species and mechanisms for various aquatic products and discusses the principles, characteristics, and applications of six nonthermal processing methods for bacterial inhibition to maintain microbiological safety and physicochemical quality. The primary spoilage bacteria groups differ among fish, crustaceans, and shellfish based on storage conditions and durations. Four metabolic pathways utilized by spoilage microorganisms-peptides and amino acids, nitrogen compounds, nucleotides, and carbohydrates-are crucial in explaining spoilage. Nonthermal processing techniques, such as ultrahigh pressure, irradiation, magnetic/electric fields, plasma, and ultrasound, can inactivate microorganisms, thereby enhancing microbiological safety, physicochemical quality, and shelf life. Future research may integrate nonthermal processing with other technologies (e.g., modified atmosphere packaging and omics) to elucidate mechanisms of spoilage and improve the storage quality of aquatic products.

Recent advances in understanding the fitness and survival mechanisms of Vibrio parahaemolyticus.

The presence of Vibrio parahaemolyticus (Vp) in different production stages of seafood has generated negative impacts on both public health and the sustainability of the industry. To further better investigate the fitness of Vp at the phenotypical level, a great number of studies have been conducted in recent years using plate counting methods. In the meantime, with the increasing accessibility of the next generation sequencing and the advances in analytical chemistry techniques, omics-oriented biotechnologies have further advanced our knowledge in the survival and virulence mechanisms of Vp at various molecular levels. These observations provide insights to guide the development of novel prevention and control strategies and benefit the monitoring and mitigation of food safety risks associated with Vp contamination. To timely capture these recent advances, this review firstly summarizes the most recent phenotypical level studies and provide insights about the survival of Vp under important in vitro stresses and on aquatic products. After that, molecular survival mechanisms of Vp at transcriptomic and proteomic levels are summarized and discussed. Looking forward, other newer omics-biotechnology such as metabolomics and secretomics show great potential to be used for confirming the cellular responses of Vp. Powerful data mining tools from the field of machine learning and artificial intelligence, that can better utilize the omics data and solve complex problems in the processing, analysis, and interpretation of omics data, will further improve our mechanistic understanding of Vp.

Dissemination of IncC plasmids in Salmonella enterica serovar Thompson recovered from seafood and human diarrheic patients in China.

Salmonella Thompson is a prevalent foodborne pathogen and a major threat to food safety and public health. This study aims to reveal the dissemination mechanism of S. Thompson with co-resistance to ceftriaxone and ciprofloxacin. In this study, 181 S. Thompson isolates were obtained from a retrospective screening on 2118 serotyped Salmonella isolates from foods and patients, which were disseminated in 12 of 16 districts in Shanghai, China. A total of 10 (5.5 %) S. Thompson isolates exhibited resistance to ceftriaxone (MIC ranging from 8 to 32 µg/mL) and ciprofloxacin (MIC ranging from 2 to 8 µg/mL). The AmpC ß-lactamase gene blaCMY-2 and plasmid-mediated quinolone resistance (PMQR) genes of qnrS and qepA were identified in the 9 isolates. Conjugation results showed that the co-transfer of blaCMY-2, qnrS, and qepA occurred on the IncC plasmids with sizes of ∼150 (n = 8) or ∼138 (n = 1) kbp. Three typical modules of ISEcp1-blaCMY-2-blc-sugE, IS26-IS15DIV-qnrS-ISKpn19, and ISCR3-qepA-intl1 were identified in an ST3 IncC plasmid pSH11G0791. Phylogenetic analysis indicated that IncC plasmids evolved into Lineages 1, 2, and 3. IncC plasmids from China including pSH11G0791 in this study fell into Lineage 1 with those from the USA, suggesting their close genotype relationship. In conclusion, to our knowledge, it is the first report of the co-existence of blaCMY-2, qnrS, and qepA in IncC plasmids, and the conjugational transfer contributed to their dissemination in S. Thompson. These findings underline further challenges for the prevention and treatment of Enterobacteriaceae infections posed by IncC plasmids bearing blaCMY-2, qnrS, and qepA.

Epidemiological and Genomic analysis of Vibrio parahaemolyticus isolated from imported travelers at the port of Shanghai, China (2017-2019).

BACKGROUND: Vibrio parahaemolyticus is the predominant etiological agent of seafood-associated foodborne illnesses on a global scale. It is essential to elucidate the mechanisms by which this pathogen disseminates. Given the existing research predominantly concentrates on localized outbreaks, there is a pressing necessity for a comprehensive investigation to capture strains of V. parahaemolyticus cross borders. RESULTS: This study examined the frequency and genetic attributes of imported V. parahaemolyticus strains among travelers entering Shanghai Port, China, between 2017 and 2019.Through the collection of 21 strains from diverse countries and regions, Southeast Asia was pinpointed as a significant source for the emergence of V. parahaemolyticus. Phylogenetic analysis revealed clear delineation between strains originating from human and environmental sources, emphasizing that underlying genome data of foodborne pathogens is essential for environmental monitoring, food safety and early diagnosis of diseases. Furthermore, our study identified the presence of virulence genes (tdh and tlh) and approximately 120 antibiotic resistance-related genes in the majority of isolates, highlighting their crucial involvement in the pathogenesis of V. parahaemolyticus. CONCLUSIONS: This research enhanced our comprehension of the worldwide transmission of V. parahaemolyticus and its antimicrobial resistance patterns. The findings have important implications for public health interventions and antimicrobial stewardship strategies, underscoring the necessity for epidemiological surveillance of pathogen at international travel hubs.

Comprehensive shotgun proteomic characterization and virulence factors of seafood spoilage bacteria.

This article summarizes the characterization, by shotgun proteomics, of 11 bacterial strains identified as responsible for seafood spoilage. A total of 4455 peptide spectrum matches, corresponding to 4299 peptides and 3817 proteins were identified. Analyses of data determined the functional pathways they are involved in. The proteins identified were integrated into a protein-protein network that involves 371 nodes and 3016 edges. Those proteins are implicated in energy pathways, peptidoglycan biosynthesis, spermidine/putrescine metabolism. An additional 773 peptides were characterized as virulence factors, that participates in bacterial pathogenesis; while 14 peptides were defined as biomarkers, as they can be used to differentiate the bacterial species present. This report represents the most extensive proteomic repository available in the field of seafood spoilage bacteria; the data substantially advances the understanding of seafood decay, as well as provides fundamental bases for the recognition of the bacteria existent in seafood that cause spoilage during food processing/storage.

Microbiological safety of dry-cured fish from the raw material to the end of processing.

The commercialization of processed fish products is rising in restaurants and small to medium enterprises. However, there is a lack of data related to the microbiological safety of such products. In this study total aerobic colony count and Enterobacteriaceae, as proxy of process hygiene criteria, and detection of Listeria monocytogenes and concentration of histamine, as food safety criteria, were investigated in Salmo salar (salmon), Xiphias gladius (swordfish) and Thunnus albacares (yellowfin tuna), before, during, and at the end of a dry-curing process, performed in a dedicated cabinet, at controlled temperature, relative humidity and ventilation, up to 240 h. The microbiological parameters were investigated in the tested fish products by culture methods and shotgun metagenomic, while the presence of histamine, and other biogenic amines, was quantified by High Performance Liquid Chromatography. In the raw material, and up to the end of the dry curing process, the concentration of Enterobacteriaceae was always lower than 10 CFU/g, while total aerobic colony counts ranged between 3.9 and 5.4 Log CFU/g in salmon; 5.5 and 5.9 Log CFU/g in swordfish; 4.4 and 4.8 Log CFU/g in tuna. The pH values were significantly different between fish species, in the raw materials and during processing except for T4, occurring 70 h after the start of the process for salmon and after 114 h for swordfish and tuna. Water activity was different at specific sampling points and at the end of processing. Overall, 79 % of the sequences identified in the tested fish samples were assigned to y bacteria. The most abundant phyla were Pseudomonadota, Bacillota and Mycoplasmatota. The microbial populations identified by shotgun metagenomic in the tested fish species clustered well separated one from the other. Moreover, the microbial richness was significantly higher in salmon and tuna in comparison to swordfish. Listeria monocytogenes was not detected in the raw material by using the reference cultural method and very few reads (relative abundance <0.007) were detected in swordfish and tuna by shotgun metagenomic. Histamine producing bacteria, belonging to the genera Vibrio, Morganella, Photobacterium and Klebsiella, were identified primarily in swordfish. However, histamine and other biogenic amines were not detected in any sample. To the best of our knowledge this is the first paper reporting time point determinations of microbiological quality and safety parameters in salmon, swordfish and tuna, before, during and at the end of a dry-curing process. The data collected in this paper can help to predict the risk profile of ready to eat dry-cured fish products during storage before consumption.

Assessing biofilm formation and resistance of vibrio parahaemolyticus on UV-aged microplastics in aquatic environments.

UV degradation of marine microplastics (MPs) could increase their vector potential for pathogenic bacteria and threaten human health. However, little is known about how the degree of UV aging affects interactions between MPs and pathogens and how various types of MPs differ in their impact on seafood safety. This study investigated five types of UV-aged MPs and their impact on Vibrio parahaemolyticus, a seafood pathogen. MPs exposed to UV for 60 days showed similar physicochemical changes such as surface cracking and hydrophobicity reduction. Regardless of the type, longer UV exposure of MPs resulted in more biofilm formation on the surface under the same conditions. V. parahaemolyticus types that formed biofilms on the MP surface showed 1.4- to 5.0-fold upregulation of virulence-related genes compared to those that did not form biofilms, independently of UV exposure. However, longer UV exposure increased resistance of V. parahaemolyticus on MPs to chlorine, heat, and human gastrointestinal environment. This study implies that the more UV degradation occurs on MPs, the more microbial biofilm formation is induced, which can significantly increase virulence and environmental resistance of bacteria regardless of the type of MP.

High-Resolution Comparative and Quantitative Proteomics of Biogenic-Amine-Producing Bacteria and Virulence Factors Present in Seafood.

The presence of biogenic amines (histamine, tyramine, putrescine, and cadaverine) in seafood is a significant concern for food safety. This review describes for the first time a shotgun quantitative proteomics strategy to evaluate and compare foodborne strains of bacteria that produce biogenic amines in seafoods. This approach recognized 35,621 peptide spectrum matches, belonging to 20,792 peptides, and 4621 proteins. It allowed the determination of functional pathways and the classification of the strains into hierarchical clusters. The study identified a protein-protein interaction network involving 1160 nodes/10,318 edges. Proteins were related to energy pathways, spermidine biosynthesis, and putrescine metabolism. Label-free quantitative proteomics allowed the identification of differentially regulated proteins in specific strains such as putrescine aminotransferase, arginine decarboxylase, and l-histidine-binding protein. Additionally, 123 peptides were characterized as virulence factors and 299 peptide biomarkers were selected to identify bacterial species in fish products. This study presents the most extensive proteomic repository and progress in the science of food biogenic bacteria and could be applied in the food industry for the detection of bacterial contamination that produces histamine and other biogenic amines during food processing/storage.

Preservation effects of photodynamic inactivation-mediated antibacterial film on storage quality of salmon fillets: Insights into protein quality.

The preservation effects of a photodynamic inactivation (PDI)-mediated polylactic acid/5-aminolevulinic acid (PLA/ALA) film on the storage quality of salmon fillets were investigated. Results showed that the PDI-mediated PLA/ALA film could continuously generate reactive oxygen species by consuming oxygen to inactivate native pathogens and spoilage bacteria on salmon fillets. Meanwhile, the film maintained the content of muscle proteins and their secondary and tertiary structures, as well as the integrity of myosin by keeping the activity of Ca2+-ATPase, all of which protected the muscle proteins from degradation. Furthermore, the film retained the activity of total superoxide dismutase (T-SOD), suppressed the accumulation of lipid peroxides (e.g., MDA), which greatly inhibited four main types of protein oxidations. As a result, the content of flavor amino acids and essential amino acids in salmon fillets was preserved. Therefore, the PDI-mediated antimicrobial packaging film greatly preserves the storage quality of aquatic products by preserving the protein quality.

Photocatalytic inactivation mechanism of nano-BiPO4 against Vibrio parahaemolyticus and its application in abalone.

Vibrio parahaemolyticus (V. parahaemolyticus) is the main pathogenic bacteria in seafood that can cause serious food-borne illness. The annual incidence of V. parahaemolyticus infection in the United States exceeds 45,000 cases, indicating there are potential shortcomings in seafood sterilization techniques. Meanwhile, the ongoing emergence of antibiotic-resistant strains highlights the urgent need for novel bacteriostatic strategies to eliminate V. parahaemolyticus. Nano-BiPO4 is a semiconductor with high H2O2 production efficiency and has potential for photocatalytic bacterial inactivation. But the effectiveness and mechanism of BiPO4 photocatalytic inactivation of V. parahaemolyticus has not been reported. In this study, nano-BiPO4 synthesized in pure water (P1) was found to exhibit optimal H2O2 production efficiency (1203 µmol h-1g-1) and antibacterial activity (in 0.8 g/L). Under UV light irradiation, P1 induced alterations in bacterial cell morphology, elevation in intracellular levels of ROS, H2O2, O2-, GSSG and MDA, and reduction in GSH level. Meanwhile, metabolomic analysis revealed that P1 stimulates the arginine biosynthesis, TCA cycle and alanine, aspartate and glutamate metabolism. These abnormal changes in the oxidative stress indicators and metabolic pathways proved that the bacterial damage was related to the H2O2 produced by nano-BiPO4 photocatalysis. Moreover, sliced abalone and hemolysis assay were used to demonstrate the applicability and biosafety of P1. This study provides theoretical support for exploring nano-BiPO4 as a bacterial inhibitor against V. parahaemolyticus.

Antibacterial properties of hexanal-chitosan nanoemulsion against Vibrio parahaemolyticus and its application in shelled shrimp preservation at 4 °C.

Vibrio parahaemolyticus has been considered as the leading pathogen associated with seafood-borne disease. Hexanal possesses antibacterial property but the hydrophobicity and volatility limit its application. The purpose of this study was to prepare hexanal-chitosan nanoemulsion (HCN), investigate its antibacterial ability against V. parahaemolyticus, and examine the combination of HCN with sodium alginate coating on the quality attributes of shrimp during cold storage. The mean droplet size of HCN fabricated by ultrasonic emulsification was 91.28 nm. HCN showed regular spherical shape and exhibited good centrifugation stability and storage stability at 4 °C. HCN exerted anti-V. parahaemolyticus effect with the minimum inhibitory concentration and minimal bactericidal concentration of both 5 mg/mL. Furthermore, HCN induced morphological changes and destroyed bacterial membrane, resulting in cell death. The results of preservation test showed that HCN alone and its combination with sodium alginate coating effectively retarded the quality deterioration and microbial spoilage of shelled shrimps during refrigerated storage. Comparatively, the combination treatment exhibited better preservation effect. The present study suggested that HCN prepared by ultrasonic emulsification is an effective alternative to control V. parahaemolyticus contamination in seafood and also shows great application potential in the quality maintaining of seafood during cold storage.

A novel phagomagnetic separation-ATP bioluminescence (PhMS-BL) for rapid and sensitive detection of viable Vibrio parahaemolyticus in aquatic product.

Detection of viable Vibrio parahaemolyticus (V. parahaemolyticus) is a major challenge due to its significant risk to food safety and human health. Herein, we developed a phagomagnetic separation-ATP bioluminescence (PhMS-BL) assay based on phage VPHZ6 for rapid and sensitive detection of viable V. parahaemolyticus. Phage as a recognition element was coupled to magnetic beads to capture and enrich V. parahaemolyticus, shortening detection time and improving method sensitivity. The intracellular ATP released by chemical lysis using CTAB was quantified using firefly fluorescein-adenosine triphosphate bioluminescence system to detect viable bacteria. So, PhMS-BL method was able to detect V. parahaemolyticus in a linear range of 2.3 × 102 to 1.3 × 107 CFU mL-1, with a detection limit of 78 CFU mL-1 within 15 min. It is successfully applied to detect V. parahaemolyticus in spiked lake water, lobster tail meat, and clam meat. The developed detection strategy can rapidly and sensitively detect viable V. parahaemolyticus in food matrixes.

Multi-omics analysis reveals the microbial interactions of S. cerevisiae and L. plantarum on Suanyu, Chinese traditional fermented fish.

S. cerevisiae and L. plantarum play important roles in Suanyu fermentation. This study investigated the interaction between S. cerevisiae and L. plantarum during fermentation and its impact on metabolic pathways. Co-culturing S. cerevisiae and L. plantarum increased pH to 5.72, reduced TVB-N to 9.47 mg/mL, and achieved high utilization rates of sugars (98.9%) and proteins (73.7%). During microbial interactions, S. cerevisiae and L. plantarum produced antibiotics, including phenyllactate and Gentamicin C1a, inhibiting the growth of each other. S. cerevisiae used S-adenosyl-l-methionine to counteract acid production of L. plantarum, establishing dominance in Suanyu fermentation. Microbial interactions influenced carbohydrate and energy metabolism pathways, such as nicotinate and nicotinamide metabolism and purine metabolism. S. cerevisiae significantly impacted gene expression in protein synthesis and cell growth pathways, including ribosome, SNARE interactions, basal transcription factors, and MAPK signaling. These findings offer insights into microbial interactions and metabolic processes during Suanyu fermentation.

Modeling naturally-occurring Vibrio parahaemolyticus in post-harvest raw shrimps.

There is little known about the growth and survival of naturally-occurring Vibrio parahaemolyticus in harvested raw shrimps. In this study, the fate of naturally-occurring V. parahaemolyticus in post-harvest raw shrimps was investigated from 4℃ to 30℃ using real-time PCR combined with propidium monoazide (PMA-qPCR). The Baranyi-model was used to fit the growth and survival data. A square root model and non-linear Arrhenius model was then used to quantify the parameters derived from the Baranyi-model. The results showed that naturally-occurring V. parahaemolyticus were slowly inactivated at 4℃ and 7℃ with deactivation rates of 0.019 Log CFU/g/h and 0.025 Log CFU/g/h. Conversely, at 15, 20, 25, and 30 °C, the average maximum growth rates (µmax) of naturally-occurring V. parahaemolyticus were determined to be 0.044, 0.105, 0.179 and 0.336 Log CFU/g/h, accompanied by the average lag phases (λ) of 15.5 h, 7.3 h, 4.4 h and 3.7 h. The validation metrics, Af and Bf, for both the square root model and non-linear, indicating that the model had a good ability to predict the growth behavior of naturally-occurring V. parahaemolyticus in post-harvest raw shrimps. Furthermore, a comparative exploration between the growth of artificially contaminated V. parahaemolyticus in cooked shrimps and naturally-occurring V. parahaemolyticus in post-harvest raw shrimps revealed intriguing insights. While no substantial distinction in deactivation rates emerged at 4 °C and 7 °C (P > 0.05), a discernible disparity in growth rates was observable at 15 °C, 20 °C, 25 °C, and 30 °C, with the former surpassing the latter. Which indicated the risk of V. parahaemolyticus using models derived from cooked shrimps may be biased. Our study also unveiled a discernible seasonal effect. The µmax and λ of V. parahaemolyticus in shrimps harvested in summer were similar to those harvested in autumn, while the initial and maximum bacterial concentration harvested in summer were higher than those harvested in autumn. This predictive microbiology model of naturally-occurring V. parahaemolyticus in raw shrimps provides relevance to modelling growth in situ.

Antimicrobial resistance and antagonistic features of bivalve-associated Vibrio parahaemolyticus from the south-west coast of India.

Vibrio parahaemolyticus, a potent human and aquatic pathogen, is usually found in estuaries and oceans. Human illness is associated with consuming uncooked/partially cooked contaminated seafood. The study on bivalve-associated V. parahaemolyticus revealed that the post-monsoon season had the highest bacterial abundance (9 ± 1.5 log cfu) compared to the monsoon season (8.03 ± 0.56 log cfu). Antimicrobial resistance (AMR) profiling was performed on 114 V. parahaemolyticus isolates obtained from bivalves. The highest AMR was observed against ampicillin (78%). Chloramphenicol was found to be effective against all the isolates. Multiple antibiotic resistance index values of 0.2 or higher were detected in 18% of the isolates. Molecular analysis of antimicrobial resistant genes (ARGs) revealed the high prevalence (100%) of the TEM-1 gene in the aquatic environment. After plasmid profiling and curing, 41.6% and 100% of the resistant isolates were found to be sensitive to ampicillin and cephalosporins, respectively, indicating the prevalence of plasmid-associated ARGs in the aquatic environment. A study to evaluate the antagonistic properties of Bacillus subtilis, Pseudomonas aeruginosa, and Bacillus amyloliquefaciens against V. parahaemolyticus isolates identified the potential of these bacteria to resist the growth of V. parahaemolyticus.

Isolation and characterization of bioprotective lactic acid bacteria from Moroccan fish and seafood.

The microbiota of aquatic animals is heavily influenced by their environment, offering a potential source for biotechnologically relevant microorganisms. In this investigation, bacterial strains from fish and fish products were investigated to determine their antimicrobial effects against fish and food pathogens. Twelve strains, including five Lactococcus, two Enterococcus hirae, two Enterococcus mundtii, and three Latilactobacillus sakei were selected as producing bacteriocin-like substances with antimicrobial properties that were active against a broad spectrum of bacteria, such as Listeria monocytogenes, Staphylococcus aureus, and Pseudomonas aeruginosa. Selected strains were identified via 16S rRNA sequencing. Most strains exhibited sensitivity to eight types of antibiotics (erythromycin, tetracycline, chloramphenicol, vancomycin, fosfomycin, gentamicin, ampicillin, and netilmicin), lacked hemolysin and gelatinase virulence factors, and did not produce histamine. These findings suggest that marine fish may be a promising source of lactic acid bacteria strains with antimicrobial potential for use as biopreservatives in the food industry.

Effect of alginate-gallic acid coating on freshness and flavor properties of Mackerel (Scomberomorus commerson) fillets under refrigerated storage.

This study investigates the effect of sodium alginate-gallic acid (ALG-GAL) coating on mackerel's flavor compounds and quality properties during cold storage at 4 °C for 12 days. To this end, freshness quality indicators, including biogenic amines (BAs), volatile organic compounds (VOCs), ATP-related compounds, K value, total viable counts (TVC), thiobarbituric acid (TBA), and sensory assessment, were measured. During storage, eight BAs, i.e., histamine (HIS), tyramine (TYR), putrescine (PUT), cadaverine (CAD), 2-phenylethylamine (2-PHE), agimation, spermine (SPM), and spermidine (SPD) were identified in control and treated samples. The biogenic amine index (BAI) for control samples was 56.25 at the time of sensory rejection (day 6). BAI for samples coated with ALG-GAL did not exceed 20 mg/100 g at the time of sensory rejection (day 12). The fillets treated with the ALG alone or incorporated with GAL possessed a different trend in the retardation of VOCs, including aldehydes, ketones, alcohols, and hydrocarbons. Seven key flavors VOCs, including 3-methylbutanal, phenylacetaldehyde, E-2-hexanal, 1-hexanol, 1-octen-3-ol, 2,3 pentanedione, and hydroxyl-2-butanone, were identified in control and coated samples. Samples coated with ALG and GAL were of significantly higher quality (p < 0.05) throughout storage, which could result in lower Inosine (HxR) concentrations and K values. The results of TVC showed that use ALG-GAL had lower bacterial counts compared to control (p < 0.05). The ALG-GAL-coated samples retarded the increase in the contents of TBA during storage. In addition, significant differences in sensory scores between ALG and ALG-GAL were observed (p < 0.05). In this study, aldehydes and hypoxanthine (Hx) were the main compounds in the formation of off-flavor. These results revealed that ALG coating combined with GAL improved the quality of refrigerated mackerel fillets by decreasing off-flavor compounds and TVC population.

ESBL-producing Vibrio vulnificus and V. alginolyticus harbour a plasmid encoding ISEc9 upstream of blaCTX-M-55 and qnrS2 isolated from imported seafood.

In recent years, trade liberalisation has led to the spread of antibiotic-resistant bacteria (ARB) in food products. Because ARB has reportedly been found in imported foods, the spread of plasmid-mediated ARB through food products is a concern. Here, we report the complete genome sequences of ESBL-producing Vibrio vulnificus and V. alginolyticus strains harbouring a plasmid isolated from imported seafood. First, V. vulnificus and V. alginolyticus were isolated from purchased frozen and thawed Litopenaeus vannamei shrimp, and genome extraction and sequencing were performed. Hybrid genome assemblies were performed using Unicycler and annotated using DFAST. Then genome analysis was performed using BRIG. Plasmid comparisons showed that the plasmids carried by both Vibrios are remarkably similar and encode the same antibiotic-resistance genes. The 270-310 kb region specific to both Vibrios were isolated in this study and encodes the antibiotic-resistance genes blaCTX-M and qnr. Furthermore, the mobile genetic factors ISEc9, ISVch4, and ISVpa4 are located upstream and downstream of these genes. This is the first report of ESBL-producing V. vulnificus and V. alginolyticus harbouring a common plasmid encoding ISEc9 upstream of blaCTX-M-55 and qnrS2 isolated from imported seafood.

Validation of the Thermo Scientific™ SureTect™ Vibrio cholerae, Vibrio parahaemolyticus, and Vibrio vulnificus PCR Assay for the Detection of Vibrio cholerae, Vibrio parahaemolyticus, and Vibrio vulnificus in Seafood Matrixes: AOAC Performance Tested MethodsSM 022301.

BACKGROUND: The Thermo Scientific™ SureTect™ Vibrio cholerae, Vibrio parahaemolyticus, and Vibrio vulnificus PCR Assay method is a real-time PCR method for the multiplex detection of Vibrio cholerae, Vibrio parahaemolyticus, and Vibrio vulnificus in seafood. OBJECTIVE: The Thermo Scientific SureTect Vibrio cholerae, Vibrio parahaemolyticus, and Vibrio vulnificus Assay was evaluated for AOAC Performance Tested MethodsSM certification. METHOD: Inclusivity/exclusivity, matrix, product consistency/stability, and robustness studies were conducted to assess the method's performance. For the matrix study, the method was validated using the Applied Biosystems™ QuantStudio™ 5 Real-Time PCR Food Safety Instrument and the Applied Biosystems™ 7500 Fast Real-Time PCR Food Safety Instrument against the U.S. Food and Drug Administration Bacteriological Analytical Manual, Chapter 9 (2004), Vibrio and ISO 21872-1:2017 Microbiology of the food chain-Horizontal method for the determination of Vibrio spp.-Part 1: Detection of potentially enteropathogenic Vibrio parahaemolyticus, Vibrio cholerae, and Vibrio vulnificus reference methods. RESULTS: Matrix studies showed equivalent or superior performance of the candidate method compared to the reference method and, overall, no difference between presumptive and confirmed results, except for one matrix due to high background flora. The inclusivity/exclusivity study correctly identified/excluded all strains analyzed. Robustness testing showed no statistically significant differences in assay performance under varied test conditions. Product consistency and stability studies demonstrated no statistically significant differences between assay lots with different expiration dates. CONCLUSIONS: The data presented show that the assay constitutes a rapid and reliable workflow for the detection of V. cholerae, V. parahaemolyticus, and V. vulnificus in seafood matrixes. HIGHLIGHTS: The SureTect PCR Assay method allows for fast, reliable detection of stipulated strains in seafood matrixes with results obtained in as little as 80 min post-enrichment.

Characterization of ready-to-eat fish surface as a potential source of contamination of Vibrio parahaemolyticus biofilms.

The worldwide consumption of ready-to-eat seafood products has steadily increased due to a range of health benefits. However, depending on the handling or cutting process of raw fish, ready-to-eat sashimi can be exposed to microbiological risks that can lead to foodborne infection by marine pathogens. Since surface characteristics are key factors for microbial adhesion and biofilm formation, the present study aims to determine the correlation between raw fish skin properties and Vibrio parahaemolyticus biofilm formation. We analyzed V. parahaemolyticus biofilms (ATCC 17802; initially inoculated ca. 2 or 4 log CFU/cm2) on fish skin (gizzard shad, pomfret, red snapper, and mackerel; fish species served as sashimi without peeling the skin) formed under simulated marine environments (incubating in artificial seawater with rocking motion at 30 °C, the maximum temperature of seasonal seawater) for 24 h. The characteristics of fish skin were determined using confocal laser scanning microscopy/scanning electron microscopy. V. parahaemolyticus showed higher biofilm counts on fish skins than on stainless steel, which was used as a control (P < 0.05). In particular, V. parahaemolyticus formed biofilms with significantly higher levels of bacterial populations on gizzard shad and pomfret (ca. 5 log CFU/cm2; P < 0.05), highlighting the relationship between the biofilm formation level and the characteristics of gizzard shad and pomfret skins. The surface roughness of fish skins, including the main roughness parameters (Ra, Rq, and Rz), influenced the attachment of V. parahaemolyticus (P < 0.05). Additionally, images of V. parahaemolyticus biofilms suggested that different topographical profiles of fish species (e.g., mucus, unique structural features, etc.) could cause V. parahaemolyticus to exhibit different biofilm phenotypes, such as sticking to or entangling on the fish skin surface. The major findings of this study provide various phenotypic adhesions of V. parahaemolyticus to fish skin in considerations of the potential hazard for the consumption of ready-to-eat sashimi served with its skin.