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.

Identification of microbial communities and multi-species biofilms contamination in seafood processing environments with different hygiene conditions.

Biofilms formed by spoilage and pathogenic bacteria increase microbial persistence, causing an adverse influence on the quality of seafood. The mono-species biofilms are widely reported, however, the contamination of multi-species biofilms and their matrix in food environments are still not fully understood. Here, we assessed the contamination of multi-species biofilms in three seafood processing environments with different hygiene levels by detecting bacterial number and three biofilm matrix components (carbohydrates, extracellular DNA (eDNA), and proteins). Samples comprising seven food matrix surfaces and eight food processing equipment surfaces were collected from two seafood processing plants (XY and XC) and one seafood market (CC). The results showed that the bacterial counts ranged from 1.89 to 4.91 CFU/cm2 and 5.68 to 9.15 BCE/cm2 in these surfaces by cultivation and real-time PCR, respectively. Six biofilm hotspots were identified, including four in CC and two in XY. Among the three processing environments, the amplicon sequence variants (ASVs) of Proteobacteria, Bacteroidetes, and Actinobacteria decreased with improved processing hygiene, while Firmicutes showed a decrease in the four most abundant phyla. The most prevalent bacteria belonged to genera Psychrobacter, Acinetobacter, and Pseudomonas, demonstrating the significant differences and alteration in bacterial community composition during different environments. From the biofilm hotspots, 15 isolates with strong biofilm forming ability were identified, including 7 Pseudomonas, 7 Acinetobacter, and 1 Psychrobacter. The Pseudomonas isolates exhibited the highest production of EPS components and three strong motilities, whose characteristics were positively correlated. Thus, this study verified the presence of multi-species biofilms in seafood processing environments, offering preliminary insights into the diversity of microbial communities during processing. It highlights potential contamination sources and emphasizes the importance of understanding biofilms composition to control biofilms formation in seafood processing environments.

A One-Step RPA-CRISPR Assay Using crRNA Based on Suboptimal Protospacer Adjacent Motif for Vibrio vulnificus Detection.

Vibrio vulnificus is a hazardous foodborne pathogen responsible for approximately 95% of seafood-related deaths. This highlights the urgent requirement for specialized detection tools to be developed and used by food enterprises and food safety authorities. The DETECTR (DNA endonuclease targeted CRISPR trans reporter) system that combines CRISPR/Cas and recombinase polymerase amplification (RPA) has been utilized to develop a molecular detection assay for V. vulnificus. However, because the incompatibility between RPA and Cas12a cleavage has not been addressed, it is a two-step assay that lacks convenience and presents contamination risk. Here, we developed a one-step RPA-CRISPR assay for V. vulnificus using a special crRNA targeting a sequence with a suboptimal protospacer adjacent motif (PAM). The entire assay, conducted at 37°C, takes only 40-60 min, yields results visualized under blue light, and exhibits exceptional specificity and sensitivity (detecting 4 pathogen genome copies per reaction). This study offers a valuable tool for detecting V. vulnificus, aiding in foodborne infection prevention, and exemplifies one-step RPA-CRISPR assays managing Cas-cleavage activity through PAM adjustments.

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.

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.

Inhibitory effect of chlorogenic acid-grafted chitosan on seafood isolates Pseudomonas fluorescens and its biofilm.

This study aimed to examine the inhibition of chlorogenic acid-grafted chitosan (CS-g-CA) on Pseudomonas fluorescens (P. fluorescens) and its biofilm. The minimum inhibitory concentration (MIC) of CS-g-CA against P. fluorescens was 1.25 mg/mL. Alkaline phosphatase (AKPase) leakage assay and scanning electron microscopy (SEM) observation showed that CS-g-CA causes structural damage to cell walls and membranes, resulting in the loss of function. In addition, CS-g-CA was able to disrupt the antioxidant system of P. fluorescens, interfere with energy metabolism, and interact with genomic DNA, affecting the normal physiological function of bacteria. It was also found that CS-g-CA inhibited the flagellar motility of P. fluorescens, which may be responsible for the inhibition of its biofilm formation. CS-g-CA at 2MIC was able to remove 71.64% of the mature biofilm and reduce the production of extracellular polysaccharides (EPS) by 60.72%. This was further confirmed by confocal laser scanning microscopy (CLSM), which showed a significant reduction in the amount of biofilm. In summary, CS-g-CA has strong antibacterial and anti-biofilm activities against P. fluorescens, and it can be applied as a potential seafood bacteriostatic agent.

Inhibitory Effect of Two Closely Related Phages on Vibrio parahaemolyticus.

Vibrio parahaemolyticus is a foodborne pathogenic bacterium commonly found in seafood. The emergence of drug-resistant strains poses a threat to human public health and economic development. Therefore, there are increasing needs to develop new technologies in controlling multidrug-resistant V. parahaemolyticus strains and to evaluate their practical efficiency in seafood or mariculture. In this study, we screened two genetically related V. parahaemolyticus phages, F23s2 and H256D1, which belonged to the siphoviridae family and podoviridae family, respectively. They showed 97.13% and 96.13% identity with Vibrio phage vB_Vpap_MGD1, respectively. Both phages were stable at pH 4-11 and displayed temperature tolerance (<70°C). Meanwhile they showed a broad host spectrum for multidrug-resistant V. parahaemolyticus, and Phage F23s2 lysed 16 of all 23 V. parahaemolyticus strains, while phage H256D1 lysed 10 strains. Phage F23s2 and H256D1 had a good inhibitory effect on V. parahaemolyticus in shrimp meat. Compared with the negative group, the bacterial amount of experimental group with phage F23s2 decreased by 1.60 log colony-forming unit (CFU)/mL at 12 h. For phage H256D1, the bacterial concentration of shrimp meat contaminated with V. parahaemolyticus H256 increased to 5.65 log CFU/mL at 72 h, while the concentration of the experimental group in presence of phage H256D1 was 3.58 log CFU/mL. All live clams infected with V. parahaemolyticus died after 96 h in the absence of phage, whereas clams with phage F23s2 and H256D1 still had a survival rate of 12% and 4%, respectively. Understanding the gene function and biology of phages facilitates its application for control of V. parahaemolyticus contamination worldwide.

A scoping review of the distribution and frequency of extended-spectrum ß-lactamase (ESBL)-producing Enterobacteriaceae in shrimp and salmon.

Antimicrobial-resistant (AMR) bacteria are a threat to public health as they can resist treatment and pass along genetic material that allows other bacteria to become drug-resistant. To assess foodborne AMR risk, the Codex Guidelines for Risk Analysis of Foodborne AMR provide a framework for risk profiles and risk assessments. Several elements of a risk profile may benefit from a scoping review (ScR). To contribute to a larger risk profile structured according to the Codex Guidelines, our objective was to conduct a ScR of the current state of knowledge on the distribution, frequency and concentrations of extended-spectrum ß-lactamase (ESBL)-producing Enterobacteriaceae in salmon and shrimp. Articles were identified via a comprehensive search of five bibliographic databases. Two reviewers screened titles and abstracts for relevance and characterised full-text articles with screening forms developed a priori. Sixteen relevant studies were identified. This review found that there is a lack of Canadian data regarding ESBL-producing Enterobacteriaceae in salmon and shrimp. However, ESBL- producing Escherichia coli, Klebsiella pneumoniae and other Enterobacteriaceae have been isolated in multiple regions with a history of exporting seafood to Canada. The literature described herein will support future decision-making on this issue as research/surveillance and subsequent assessments are currently lacking.

Circulating trimethylamine N-oxide levels following fish or seafood consumption.

PURPOSE: Some species of fish and seafood are high in trimethylamine N-oxide (TMAO), which accumulates in muscle where it protects against pressure and cold. Trimethylamine (TMA), the metabolic precursor to TMAO, is formed in fish during bacterial spoilage. Fish intake is promoted for its potential cardioprotective effects. However, numerous studies show TMAO has pro-atherothrombotic properties. Here, we determined the effects of fish or seafood consumption on circulating TMAO levels in participants with normal renal function. METHODS: TMAO and omega-3 fatty acid content were quantified across multiple different fish or seafood species by mass spectrometry. Healthy volunteers (n = 50) were recruited for three studies. Participants in the first study consented to 5 consecutive weekly blood draws and provided dietary recall for the 24 h preceding each draw. In the second study, TMAO levels were determined following defined low and high TMAO diets. Finally, participants consumed test meals containing shrimp, tuna, fish sticks, salmon or cod. TMAO levels were quantified by mass spectrometry in blood collected before and after dietary challenge. RESULTS: TMAO + TMA content varied widely across fish and seafood species. Consumption of fish sticks, cod, and to a lesser extent salmon led to significant increases in circulating TMAO levels. Within 1 day, circulating TMAO concentrations in all participants returned to baseline levels. CONCLUSIONS: We conclude that some fish and seafood contain high levels of TMAO, and may induce a transient elevation in TMAO levels in some individuals. Selection of low TMAO content fish is prudent for subjects with elevated TMAO, cardiovascular disease or impaired renal function.

Occurrence and antibiotic resistance of Vibrio parahaemolyticus isolated from the Tunisian coastal seawater.

Vibrio parahaemolyticus is a gram-negative bacterium ubiquitous in seawater or estuarine water throughout the world. It is a major cause of seafood gastroenteritis complications. In this study, the presence of V. parahaemolyticus was investigated in 66 seawater samples collected during 2018 from 15 stations spread along the Tunisian coast using selective media including CHROMagar Vibrio media. The results show that only eight samples contained V. parahaemolyticus. However, while Vibrio alginolyticus was detected in all samples; both Vibrio cholerae and Vibrio vulnificus were not found. Nine of the presumed V. parahaemolyticus colonies were purified on tryptic soy agar from eight positive samples then identified by the API 20E biochemical test and confirmed by the presence of a specific target toxR gene. The detection of virulence genes, thermostable direct haemolysin (tdh) and thermostable-related haemolysin (trh), by the polymerase chain reaction (PCR) showed the presence of only two trh-positive isolates. The assessment of antibiotic susceptibility of the V. parahaemolyticus isolated revealed a complete resistance to colistin, amikacin, penicillin and cefotaxime and a total sensitivity to chloramphenicol, nitrofurantoin and sulfamethoxazole-trimethoprim with a multiple antibiotic resistance index (MAR) ranging from 0.4 to 0.5.

Detection of trh+ Vibrio parahaemolyticus in seafood by lac dye coloration-based label-free lateral flow immunoassay strip.

Vibrio parahaemolyticus (V. parahaemolyticus) is an important foodborne pathogen known to cause severe enteric disease. Thus, timely detection of V. parahaemolyticus in seafood is crucial to prevent food poisoning and reduce economic losses. Traditional lateral flow immunoassay strips (LFIS) required good labelling materials and pairing of two antibodies, which made them costly and difficult to manufacture. In this study, a label-free and lac dye coloration-based LFIS (LD-LFIS) to detect trh+ V. parahaemolyticus was developed for the first time. Lac dye was used to stain V. parahaemolyticus, and LFIS was used to detect stained bacteria. Dimethyl sulphoxide (DMSO) and simultaneous mordanting were chosen as the best solvent and the best staining method for lac dye. In addition, three mordants [KAl(SO4 )2 ·12H2 O, NH4 Fe(SO4 )2 ·12H2 O, and AlCl3 ·6H2 O] were selected to improve dyeing efficiency. The detection limit of LD-LFIS was 3.9 × 105 CFU/ml when NH4 Fe(SO4 )2 ·12H2 O was used as mordant. Feasibility of the LD-LFIS method was verified by detecting trh+ V. parahaemolyticus in true and spiked seafood samples. The method developed in this study is expected to reduce restrictions on labelling materials and pairing of two antibodies on LFIS, and proposes a novel idea for the rapid detection of V. parahaemolyticus in seafood.

Depuration of live oysters to reduce Vibrio parahaemolyticus and Vibrio vulnificus: A review of ecology and processing parameters.

Consumption of raw oysters, whether wild-caught or aquacultured, may increase health risks for humans. Vibrio vulnificus and Vibrio parahaemolyticus are two potentially pathogenic bacteria that can be concentrated in oysters during filter feeding. As Vibrio abundance increases in coastal waters worldwide, ingesting raw oysters contaminated with V. vulnificus and V. parahaemolyticus can possibly result in human illness and death in susceptible individuals. Depuration is a postharvest processing method that maintains oyster viability while they filter clean salt water that either continuously flows through a holding tank or is recirculated and replenished periodically. This process can reduce endogenous bacteria, including coliforms, thus providing a safer, live oyster product for human consumption; however, depuration of Vibrios has presented challenges. When considering the difficulty of removing endogenous Vibrios in oysters, a more standardized framework of effective depuration parameters is needed. Understanding Vibrio ecology and its relation to certain depuration parameters could help optimize the process for the reduction of Vibrio. In the past, researchers have manipulated key depuration parameters like depuration processing time, water salinity, water temperature, and water flow rate and explored the use of processing additives to enhance disinfection in oysters. In summation, depuration processing from 4 to 6 days, low temperature, high salinity, and flowing water effectively reduced V. vulnificus and V. parahaemolyticus in live oysters. This review aims to emphasize trends among the results of these past works and provide suggestions for future oyster depuration studies.

Characterization of Latilactobacillus curvatus MS2 isolated from Korean traditional fermented seafood and cholesterol reduction effect as synbiotics with isomalto-oligosaccharide in BALB/c mice.

This study investigated the properties of Latilactobacillus curvatus MS2 isolated from Korean traditional fermented seafood as probiotics and the effect of reducing cholesterol as a synbiotic with isomalto-oligosaccharide (IMO) in BALB/c mice. The isolated strain showed high resistance to acids and bile acids and exhibited a high DPPH scavenging capacity of 72.27 ± 0.38 %. In the intestinal adhesion test using HT-29 cells, the adhesion rate of MS2 was 17.10 ± 1.78 %, which was higher than the adhesion rate of the other investigated probiotics. MS2 showed good antimicrobial activity against food-borne pathogens, especially Staphylococcus aureus, S. epidermidis, Escherichia coli, and Vibrio vulnificus. This strain had high availability for IMO among the prebiotics of fructo-oligosaccharide, inulin and IMO. Oral administration of MS2 and IMO to BALB/c mice for 5 weeks resulted in a significant reduction in blood cholesterol levels by regulating liver lipid metabolism. These results suggest that the combination of MS2 and IMO has potential for application in functional foods.

Phylogenetic characterization and multidrug resistance of bacteria isolated from seafood cocktails.

The continual increase in resistance to antibacterial drugs has become a major public health problem, and their indiscriminate use in agriculture, aquaculture, and the treatment of human and animal diseases has severely contributed to the occurrence and spread of multidrug resistance genes. This study phylogenetically characterized multidrug-resistant bacteria isolated from seafood cocktails. Seafood cocktail dishes from 20 establishments on public roads were sampled. Samples were grown on TCBS agar and blood agar. Forty colonies with different macro- and microscopic characteristics were isolated. The 16S rRNA gene V4 and V6 hypervariable regions were amplified, sequenced and phylogenetically analyzed. Antibacterial drug resistance was determined by disk diffusion assay. Isolated bacteria were identical to species of the genera Enterococcus, Proteus, Vibrio, Staphylococcus, Lactococcus, Vagococcus, Micrococcus, Acinetobacter, Enterobacter, and Brevibacterium, with 75-100% presenting resistance or intermediate resistance to dicloxacillin, ampicillin, and penicillin; 50-70% to cephalosporins; 30-67.5% to amikacin, netilmicin and gentamicin; 40% to nitrofurantoin and other antibacterial drugs; 25% to chloramphenicol; and 2.5% to trimethoprim with sulfamethoxazole. In general, 80% of the bacteria showed resistance to multiple antibiotics. The high degree of bacterial resistance to antibacterial drugs indicates that their use in producing raw material for marine foods requires established guidelines and the implementation of good practices.

Halobacillus fulvus sp. nov., a moderately halophilic bacterium isolated from shrimp paste (Ka-pi) in Thailand.

An aerobic, Gram-stain-positive, endospore-forming, rod-shaped and moderately halophilic strain SKP4-6T, was isolated from shrimp paste (Ka-pi) collected from Samut Sakhon Province, Thailand. Phylogenetic analysis revealed that strain SKP4-6T belonged to the genus Halobacillus and was most closely related to Halobacillus salinus JCM 11546T (98.6 %), Halobacillus locisalis KCTC 3788T (98.6 %) and Halobacillus yeomjeoni KCTC 3957T (98.6 %) based on 16S rRNA gene sequence similarity. The digital DNA-DNA hybridization (dDDH) and average nucleotide identity (ANI) values between strain SKP4-6T and its related species were 18.2-19.3 % and 69.84-84.51 %, respectively, which were lower than the threshold recommended for species delineation. The strain grew optimally at 30-40 °C, at pH 7.0 and with 10-15 % (w/v) NaCl. It contained l-Orn-d-Asp in the cell wall peptidoglycan. The DNA G+C content was 44.8 mol%. The major fatty acids were iso-C15 : 0, anteiso-C15 : 0 and anteiso-C17 : 0. The predominant isoprenoid quinone was MK-7. Phosphatidylglycerol and diphosphatidylglycerol were present as major polar lipids. Based on this polyphasic approach, digital DNA-DNA relatedness and ANI values, strain SKP4-6T represents a novel species of the genus Halobacillus, for which the name Halobacillus fulvus sp. nov. is proposed. The type strain is SKP4-6T (=JCM 32624T=TISTR 2595T).

Characteristics of Vibrio vulnificus isolates from clinical and environmental sources.

Researchers have developed multiple methods to characterize clinical and environmental strains of Vibrio vulnificus. The aim of our study was to use four assays to detect virulence factors in strains from infected patients and those from surface waters/sediments/oysters of South Carolina and the Gulf of Mexico. Vibrio vulnificus strains from clinical (n = 81) and environmental (n = 171) sources were tested using three real-time PCR methods designed to detect polymorphisms in the 16S rRNA, vcg and pilF genes and a phenotypic method, the ability to ferment D-mannitol. Although none of the tests correctly categorized all isolates, the differentiation between clinical and environmental isolates was similar for the pilF, vcgC/E and 16S rRNA assays, with sensitivities of 74.1-79.2% and specificities of 77.4-82.7%. The pilF and vcgC/E assays are comparable in efficacy to the widely used 16S rRNA method, while the D-mannitol fermentation test is less discriminatory (sensitivity = 77.8%, specificity = 61.4%). Overall percent agreement for the D-mannitol fermentation method was also lower (66.7%) than overall percent agreement for the 3 molecular assays (78.0%-80.2%). This study demonstrated, using a large, diverse group of Vibrio vulnificus isolates, that three assays could be used to distinguish most clinical vs environmental isolates; however, additional assays are needed to increase accuracy.

Coordination of primary metabolism and virulence factors expression mediates the virulence of Vibrio parahaemolyticus towards cultured shrimp (Penaeus vannamei).

AIMS: Acute hepatopancreatic necrosis disease (AHPND) caused by Vibrio parahaemolyticus has emerged as a severe bacterial disease of cultured shrimp. To identify the key virulence factors, two AHPND-causing V. parahaemolyticus (VpAHPND ) strains (123 and 137) and two non-VpAHPND strains (HZ56 and ATCC 17082) were selected. METHODS AND RESULTS: Challenge tests showed that the four strains exhibited different virulence towards shrimp with cumulative mortalities at 48 h postinfection (hpi) ranging from 10 to 92%. The expression of pirABVP in strain 123 and 137 was not significantly different. Genomic analysis revealed that the two VpAHPND strains contain a plasmid with the PirABVP toxins (pirABVP ) flanked by the insertion sequence (ISVal1) that has been identified in various locations of chromosomes in VpAHPND strains. The two VpAHPND strains possessed almost identical virulence factors, while ISVal1 disrupted three genes related to flagellar motility in strain 137. Phenotype assay showed that strain 123 possessed the highest growth rate and swimming motility, followed by strain 137, suggesting that the disruption of essential genes mediated by ISVal1 significantly affected the virulence level. Transcriptome analysis of two VpAHPND strains (123 and 137) further suggested that virulence genes related to the capsule, flagella and primary metabolism were highly expressed in strain 123. CONCLUSIONS: Here for the first time, it is demonstrated that the virulence of VpAHPND is not only determined by the expression of pirABVP , but also is mediated by ISVal1 which affects the genes involved in flagellar motility and primary metabolism. SIGNIFICANCE AND IMPACT OF THE STUDY: The genomic and transcriptomic analysis of VpAHPND strains provides valuable information on the virulence factors affecting the pathogenicity of VpAHPND.

Aeromonas spp. isolated from ready-to-eat seafood on the Norwegian market: prevalence, putative virulence factors and antimicrobial resistance.

AIMS: We aim to investigate the prevalence, putative virulence factors and antimicrobial resistance of mesophilic Aeromonas isolated from ready-to-eat (RTE) seafood available on the Norwegian market, and to assess the potential risks by consuming RTE seafood to consumers. METHODS AND RESULTS: The prevalence of mesophilic Aeromonas in 148 RTE seafood was investigated and the highest prevalence was found in retail sushi (17%), followed by oysters (10%), fresh salmon loins (10%) and scallops (4%). Among 43 Aeromonas isolates, 75% of them were identified as A. media, 23% as A. salmonicida and 2% as A. bestiarum based on partial gryB gene sequencing. Aeromonas isolates were potentially pathogenic due to the presence of four virulence genes: alt (73%), hylA (22%), aerA (17%) and act (6%). In addition, all isolates were resistant to ampicillin and erythromycin. Most of the isolates (98%) were multidrug resistant. CONCLUSIONS: The occurrence of potentially pathogenic and multidrug-resistant Aeromonas strains in RTE seafood implies a potential risk to consumers. Our finding suggests that RTE seafood could be a potential vehicle for the transfer of virulent and multidrug-resistant Aeromonas. SIGNIFICANCE AND IMPACT OF THE STUDY: To our knowledge, this is the first study to report multiple antibiotic resistance in Aeromonas associated with RTE seafood in Norway.

Prevalence, antibiogram and virulence characterization of Vibrio isolates from fish and shellfish in Egypt: a possible zoonotic hazard to humans.

AIMS: Infection of seafood with pathogenic species of the genus Vibrio causes human food-borne illnesses. This study was executed to examine the antimicrobial resistance phenotypes, biofilm-forming capability and virulence-associated genes of Vibrio from fish and shellfishes. METHODS AND RESULTS: Three hundred fresh water and marine fish and shellfish samples were collected from wet markets and supermarkets in Mansoura, Egypt. Bacteriological examination and PCR amplification identified 92 Vibrio spp., including 42 Vibrio parahaemolyticus and 50 Vibrio alginolyticus isolates from the examined fish and shellfish (infection rate: 30·67%). However, V. vulnificus was not found in this study. Vibrio spp. exhibited variable frequencies of antimicrobial resistance with higher percentages to ampicillin and penicillin. Multidrug resistance (MDR) was detected in 69·04 and 38% of V. parahaemolyticus and V. alginolyticus respectively. PCR testing of virulence genes, tdh, trh and tlh revealed the presence of tlh and trh in 100 and 11·9% of V. parahaemolyticus isolates respectively and none of V. alginolyticus carried any of these genes. Biofilm-forming capability was displayed by 76% of V. parahaemolyticus and 73·8% of V. alginolyticus isolates. Both V. parahaemolyticus and V. alginolyticus showed nonsignificant weak positive correlations (r < 0·4) between antimicrobial pairs belonging to different classes; however, a significant positive correlation (P <0·05) between trh and resistance to erythromycin (r = 0·45) and imipenem (r = 0·38) was only identified in V. parahaemolyticus. CONCLUSIONS: This study reports the existence of MDR strains of V. parahaemolyticus and V. alginolyticus from the common types of fishes and shellfishes in Egypt. Furthermore, the presence of virulence genes in these isolates and the ability to produce a biofilm in vitro pose potential health hazards to consumers. SIGNIFICANCE AND IMPACT OF THE STUDY: Frequent monitoring of seafood for the presence of Vibrio spp. and their antimicrobial susceptibility, virulence determinants and biofilm-forming capability is important for assessing the risk posed by these organisms to the public and for improving food safety.

Role of RpoS in stress resistance, biofilm formation and quorum sensing of Shewanella baltica.

Shewanella baltica is one of the most important bacterial species contributing to spoilage of seafood. Principally, RpoS has been recognized as the central regulator of stress resistance in many bacterial species. However, little is known about the role of RpoS in S. baltica. In this study, an rpoS mutant of S. baltica was constructed and analysed for its functions. The results showed that the survival rate of rpoS mutant decreased when treated with heat, ethanol and H2 O2, while increased the resistance to NaCl. Moreover RpoS promoted the biofilm formation of S. baltica at 30°C, while declined at 4°C. Interestingly, the rpoS-deficient mutant showed increased swimming motility. Furthermore, the results revealed that the production of quorum-sensing (QS) signals such as cyclo-(l-Pro-l-Leu) and cyclo-(l-Pro-l-Phe) reduced in rpoS mutant. Mainly, rpoS positively regulated QS response regulators, as the expression of all luxR genes in rpoS mutant significantly decreased relative to wild type. This study reveals that RpoS is a major regulator involved in stress responses, biofilm formation and quorum sensing system in S. baltica. The present work provides significant information for the control of microbiological spoilage of seafood.