Vibrio Parahaemolyticus In Seafood Research Articles

Rapid and accurate sensing platform based on lanthanide metal-organic framework and aptamer for Vibrio parahaemolyticus diagnosis

In this study, a rapid visual analysis strategy for pathogens was constructed based on lanthanide metal–organic frameworks (LnMOFs) and FAM-aptamer. The dual fluorescence signal changes were realized by the competitive binding effect of the target and LnMOFs on the FAM-aptamer. The nitrocellulose membrane (NC membrane) is used as the signal output platform, and the G/R value in Red, Green, Blue (RGB) of negative and positive signals is collected to output the detection results. Using Vibrio parahaemolyticus in seafood as model target, the detection performance of dual fluorescence sensing strategy was verified. Under the optimal conditions, the G/R difference between the positive and negative signals and the concentration of Vibrio parahaemolyticus was linear in the range of 102-105 colony forming units (CFU)/mL, and the detection limit was 102 CFU/mL. The method is simple to operate and the detection time can be controlled within 20 min. It can be developed as a portable detection kit, providing a new technical means for the safety and quality control of food supply chain.

Incidence, virulence genes and antimicrobial resistance of Vibrio parahaemolyticus isolated from seafood

The objective of the study was to establish the incidence, pathogenic factors and antimicrobial resistance of Vibrio parahaemolyticus in seafood from retail shops in Bulgaria. A hundred and eighty samples of sea fish, mussels, oysters, veined rapa whelks, shrimps and squids were included in the study. PCR methods were used to identify V. parahaemolyticus and prove tdh and trh genes. Antimicrobial resistance was established by disc diffusion method, and MAR index was calculated. The results proved the presence of V. parahaemolyticus in 24% (44/180) of the seafood samples. tdh-positive V. parahaemolyticus was not found, while the trh gene was detected in one veined rapa whelk isolate. All isolates were susceptible to Sulfamethoxazole/trimethoprim, Tetracycline, Gentamycin, Amoxicillin-clavulanic acid, Amikacin, Ciprofloxacin, and Levofloxacin. Intermediate resistance was found to Ampicillin (25%; 11/44), Cefepime (16%; 7/44), and Ceftazidime (2%; 1/44). The results showed that 16% (7/44) of the isolates were resistant to Cefepime, 9% (4/44) to Ampicillin, and 5% (2/44) to Ceftazidime. MAR-index values ranged from 0.10 to 0.30. The incidence of pathogenic and multidrug-resistant V. parahaemolyticus strains in seafood offered on the market poses a risk to consumer health.

Vibrio parahaemolyticus in sea food in north Chennai coastal retail shops

Vibrio parahaemolyticus in sea food in north Chennai coastal retail shops

Microfluidic thread-based electrochemical aptasensor for rapid detection of Vibrio parahaemolyticus

Vibrio parahaemolyticus is one of the foodborne bacteria that widely present in seafood as well as the leading cause of seafood-associated bacterial gastroenteritis. Traditional identification of such pathogens mainly relies on culturing methods, ELISA or PCR. These methods are usually laborious, time-consuming with poor diagnosis competences, or require costly and bulky equipment though of high sensitivity. In this study, a thread-based microfluidic electrochemical aptasensor was designed, fabricated and tested by using label-free aptamer immunosensing technology for rapid and highly sensitive detection of Vibrio parahaemolyticus in seafood. Both the microfluidic channels and electrodes were simply fabricated on threads. Molybdenum disulfide (MoS2) nanosheets were used to obtain enhanced sensitivity of the electrochemical measurement. When used in detecting Vibrio parahaemolyticus, the proposed aptasensor has a dynamic detection range of 10–106 CFU mL−1 with a detection limit of 5.74 CFU mL−1. Compared with traditional plate counting method, the proposed aptasensor has higher detection sensitivity and less assay time (30 min), while high specificity and accuracy are kept. The proposed microfluidic thread-based electrochemical aptasensor grabs the potential to detect other pathogens by simply functionalizing the threaded electrodes with aptamers for targeted biological substances.

Development and comparison of immunochromatographic strips with four nanomaterial labels: Colloidal gold, new colloidal gold, multi-branched gold nanoflowers and Luminol-reduced Au nanoparticles for visual detection of Vibrio parahaemolyticus in seafood

In this study, the four nanomaterials: Traditional colloidal gold (AuNP), New colloidal gold (N-AuNP), Multi-branched gold nanoflowers (AuNF) and Luminol-reduced Au nanoparticles (L-AuNP) were respectively labeled with the monoclonal antibody against Vibrio parahaemolyticus (V. parahaemolyticus). We aimed to develop lateral flow immunoassays (LFIAs) with these nanomaterial labels and determine their performance in visual detection of V. parahaemolyticus. The results of four colloidal gold particle-labeled LFIAs and polymerase chain reaction (PCR) simultaneously detected in actual samples were to be highly consistent, indicating that strips have high accuracy. The four strips were also found to be stable up to 14 weeks under laboratory conditions. In terms of sensitivity, the N-AuNP-based strip was slightly better than the other three. For the N-AuNP-based strips, the difference between the results obtained for different batches was high consistency, and the stability was much better than that of the AuNP-, AuNF-and L-AuNP-based ones. Our results indicate that the N-AuNP can be better used as labels in immunochromatographic tests. It can also reduce the based antibodies, improve the detection sensitivity and reduce the production cost, thereby expanding the scale of production. The use of LFIA labeled with N-AuNP can be more effective, rapid and inexpensive methods for V. parahaemolyticus assay in on-site applications.

Determination of Vibrio parahaemolyticus in seafoods using direct plate counting, quantitative loop-mediated isothermal amplification and propidium monoazide-qLAMP

One of the most challenging aspects in culture independent methods for foodborne pathogens’ detection is discrimination of dead and live microorganisms. This study aimed to determine the Vibrio parahaemolyticus in seafoods via direct plate counting (DPC) and toxR-based quantitative loop-mediated isothermal amplification (qLAMP) and to discriminate dead and live cells using propidium monoazide (PMA)-qLAMP. A total of 200 samples including finfishes (n= 100) and shrimps (n= 100), representing the Mediterranean, Black and Aegean sea were collected from supermarkets and fish markets of Konya-Turkey. qLAMP was performed in a Real-Time Turbidimeter and the time threshold (tt) values were yielded in 60 minutes. On DPC, the colonies grown on TCBS Agar was further confirmed by conventional PCR based from gyrB1 gene of Vibrio spp. and toxR gene of V. parahaemolyticus. Virulence property of the isolates were determined by tdh based qLAMP. The detection limit of the qLAMP was 1.2×104 CFU/g in artificially contaminated seafoods. DPC, qLAMP and PMA-qLAMP detected V. parahaemolyticus in 8 (4%), 12 (6%) and 12 (6%) samples, respectively. The CFUs of V. parahaemolyticus detected in qLAMP (5.96±0.10 log10 CFU/ml) and PMA-qLAMP (4.71±0.13 log10 CFU/ml) were higher than those of DPC (1.99±0.44 log10 CFU/ml) (P<0.05). The mean tt reduction in PMA treated samples was 1.25±0.38 log10 CFU/sample. The tdh gene was not detected in any of the isolates. In conclusion, the toxR-based PMA-qLAMP method could be an alternative to be used more widely and effective assay for the quantification of live V. parahaemolyticus in seafoods.

Development and evaluation of a rapid and sensitive RPA assay for specific detection of Vibrio parahaemolyticus in seafood

BackgroundVibrio parahaemolyticus (V. parahaemolyticus) is a leading cause of food poisoning and is of great importance to public health due to the frequency and seriousness of the diseases. The simple, timely and efficient detection of this pathogen is a major concern worldwide. In this study, we established a simple and rapid method based on recombinase polymerase amplification (RPA) for the determination of V. parahaemolyticus. According to the gyrB gene sequences of V. parahaemolyticus available in GenBank, specific primers and an exo probe were designed for establishing real-time recombinase polymerase amplification (real-time RPA).ResultsThe real-time RPA reaction was performed successfully at 38 °C, and results were obtained within 20 min. The method only detected V. parahaemolyticus and did not show cross-reaction with other bacteria, exhibiting a high level of specificity. The study showed that the detection limit (LOD) of real-time RPA was 1.02 × 102 copies/reaction. For artificially contaminated samples with different bacteria concentrations, V. parahaemolyticus could be detected within 5–12 min by real-time RPA in oyster sauce, codfish and sleeve-fish at concentrations as low as 4 CFU/25 g, 1 CFU/25 g and 7 CFU/25 g, respectively, after enrichment for 6 h, but were detected in a minimum of 35 min by real-time PCR (Ct values between 27 and 32).ConclusionThis study describes a simple, rapid, and reliable method for the detection of V. parahaemolyticus, which could potentially be applied in the research laboratory and disease diagnosis.

Point-of-care rapid detection of Vibrio parahaemolyticus in seafood using loop-mediated isothermal amplification and graphene-based screen-printed electrochemical sensor

Vibrio parahaemolyticus is one of the most important foodborne pathogens that cause various life-threatening diseases in human and animals. Here, we present a rapid detection platform for V. parahaemolyticus by combining loop-mediated isothermal amplification (LAMP) and disposable electrochemical sensors based on screen-printed graphene electrodes (SPGEs). The LAMP reactions using primers targeting V. parahaemolyticus toxR gene were optimized at an isothermal temperature of 65 °C, providing specific detection of V. parahaemolyticus within 45 min at the detection limit of 0.3 CFU per 25 g of raw seafood. The LAMP amplicons can be effectively detected using unmodified SPGEs, redox active molecules namely Hoechst-33258 and a portable potentiostat. Therefore, the proposed system is particularly suitable as a point-of-care device for on-site detection of foodborne pathogens.

Antimicrobial resistance, virulence and genetic relationship of Vibrio parahaemolyticus in seafood from coasts of Bohai Sea and Yellow Sea, China

Vibrio parahaemolyticus is an important foodborne pathogen which commonly inhabits estuarine and marine environments and seafood. In the present study, 90 V. parahaemolyticus isolates from the main seafoods from three coastal provinces surrounding Bohai Sea and Yellow Sea, China were analyzed to elucidate their antimicrobial resistance, virulence and genetic relationship by multilocus sequence typing (MLST). The results showed that the virulence genes tdh and trh were detected in one isolate and five isolates respectively. Most of isolates showed resistance to ampicillin (86/90) and cephazolin (75/90). Some isolates were resistant to amikacin (27/90), cefuroxime sodium (18/90), tetracycline (16/90), sulphamethoxazole/trimethoprim (16/90) and streptomycin (13/90). Forty isolates (44.4%) possessed multiple antimicrobial resistance to at least three antimicrobials. The V. parahaemolyticus population was composed of 68 sequence types, of which 41 were novel to the pubMLST database, displaying a high level of genetic diversity. The phylogenetic relatedness of V. parahaemolyticus isolates was irrelevant to the collection sources. Moreover, there were no associations of antimicrobial resistance and trh positive virulence with genetic population of V. parahaemolyticus isolates. These results indicated that the diversity of antimicrobial-resistant or pathogenic V. parahaemolyticus isolates from coasts of Bohai Sea and Yellow Sea, China could pose a potential risk to human health.

Prevalence of Vibrio parahaemolyticus in seafood and water environment in the Mekong Delta, Vietnam.

A total of 449 samples including 385 seafood and 64 water samples in the Mekong Delta of Vietnam collected in 2015 and 2016 were examined. Of 385 seafood samples, 332 (86.2%) samples were contaminated with Vibrio parahaemolyticus and 25 (6.5%) samples were pathogenic V. parahaemolyticus carrying tdh and/or trh genes. The tdh gene positive V. parahaemolyticus strains were detected in 22 (5.7%) samples and trh gene positive V. parahaemolyticus strains were found in 5 (1.3%) samples. Of 25 pathogenic V. parahaemolyticus strains, two strains harbored both tdh and trh genes and the other 23 strains carried either tdh or trh gene. Of 64 water samples at aquaculture farms, 50 (78.1%) samples were contaminated with V. parahaemolyticus. No tdh gene positive V. parahaemolyticus strains were detected; meanwhile, trh gene positive V. parahaemolyticus strain was detected in 1 (1.6%) sample. Twenty-six pathogenic V. parahaemolyticus strains isolated were classified into 6 types of O antigen, in which the serotype O3:K6 was detected in 4 strains. All pathogenic strains were group-specific PCR negative except for 4 O3:K6 strains. The result of antimicrobial susceptibility test indicated that pathogenic strains showed high resistance rates to streptomycin (84.6%), ampicillin (57.7%) and sulfisoxazole (57.7%). These findings can be used for understanding microbiological risk of seafood in the Mekong Delta, Vietnam.

Vibrio parahaemolyticus in seafood - associated outbreaks

Among the 30 species of the genus Vibrio, only 13 of them are pathogenic to humans. All pathogenic vibrios have been reported to cause foodborne diseases, although Κ parahaemolyticus is considered the most important pathogenic Vibrio. V parahaemolyticus is a halophilic bacterium that occurs naturally in aquatic environments worldwide. The pathogen caused sporadic diarrhoea mainly associated with the consumption of raw or undercooked seafood up to recent years. Since 1996, the incidence of V. parahaemolyticus infections has increased dramatically. V. parahaemolyticus is the leading cause of seafood associated bacterial gastroenteritis in the United States and of the half foodborne outbreaks in some Asian countries. This increase in incidence has been related to the emergence of the 03:K6 serovar. The pathogenic V. parahaemolyticus strains can produce a thermostable direct hemolysin or a thermostable direct hemolysin-related hemolysin, which arc encoded by the tdh and trh genes, respectively. Vibrio parahaemolyticus has not been included in the microbiological criteria of E.U. Food legislation, probably because the risk by this pathogen was considered rather low in liurope. However, climate changes favour the growth of the pathogen in seawater. Recent studies in Spain and France have shown that V. parahaemolyticus infections from seafood consumption have been increased. The emergence of the pathogen in liurope is of public health concern and emphasizes the importance of microbiological surveillance and control programs for V, parahaemolyticus.

Rapid real-time PCR for the detection of Vibrio parahaemolyticus in seafood

Rapid real-time PCR for the detection of Vibrio parahaemolyticus in seafood

Identification of Vibrio parahaemolyticus in Seafood by Multiplex PCR

ABSTRACTVibrio parahaemolyticus is a human pathogen frequently found in seafood. Once the seafood is contaminated by V. parahaemolyticus, it can become a vehicle for foodborne illness. The conventional culture methods for detection of V. parahaemolyticus are time-consuming and cannot differentiate pathogenic strains from nonpathogenic ones. In this study, a multiplex polymerase chain reaction (PCR) technique was investigated for detecting tdh, chiA, and toxR of V. parahaemolyticus. The sensitivity of the multiplex PCR was determined by testing 28 strains of V. parahaemolyticus, 15 non-V. parahaemolyticus strains, and fresh seafood spiked with cells of V. parahaemolyticus. All the strains were analyzed for production of thermostable direct hemolysin (TDH) and chitinase. This study showed that both the chiA and toxR are excellent markers for detecting V. parahaemolyticus strains, and a multiplex PCR targeting chiA and tdh genes can be applied to simultaneously detect environmental and pathogenic V. parahaemolyticus.

Molecular characterizations of Vibrio parahaemolyticus in seafood from the Black Sea, Turkey.

Vibrio parahaemolyticus is a marine bacterium that is considered as one of the major causes of bacterial food-borne outbreaks at a global scale. A total of 114 samples including mussel (n=42), seawater (n=22) and fish (n=50) samples were collected and subjected to investigation. Vibrio parahaemolyticus was detected in 45 (39%) of 114 samples with an occurrence in mussel, seawater and fish samples of 76, 40·9 and 8% respectively. A total of 96 isolates were positive for the species-specific genes toxR and tlh and confirmed as V.parahaemolyticus. Presence of the virulence marker gene tdh was not identified in any of the strains investigated; however, four of strains were positive for the trh gene. Serological analysis of eight randomly selected trh-negative isolates identified three different serotypes: O4:K untypeable (KUT), O2:KUT, O3:KUT. Conversely, all four trh-positive strains belonged to a single serotype (O1:K1) and share an undistinguishable genetic profile by PFGE analysis, suggesting the existence of a dominant clone for the trh-positive strains in the region. Vibrio parahaemolyticus is the most prevalent food-poisoning bacterium associated with seafood consumption. The number of infections is increasing worldwide and is being reported in areas with no previous incidence. This study provides the first instance of the occurrence ofV.parahaemolyticus strains with virulence traits in the Black Sea, contributing to gain a better understanding about potential risk associated with this pathogen in the region.

Incidence and prevalence of Vibrio parahaemolyticus in seafood: a systematic review and meta-analysis.

Vibrio parahaemolyticus is an important seafood borne human pathogen worldwide due to it occurrence, prevalence and ability to cause gastrointestinal infections. This current study aim at investigating the incidence and prevalence of V. parahaemolyticus in seafood using systematic review-meta-analysis by exploring heterogeneity among primary studies. A comprehensive systematic review and meta-analysis of peer reviewed primary studies reported between 2003 and 2015 for the occurrence and prevalence of V. parahaemolyticus in seafood was conducted using “isolation”, “detection”, “prevalence”, “incidence”, “occurrence” or “enumeration” and V. parahaemolyticus as search algorithms in Web of Science (Science Direct) and ProQuest of electronic bibliographic databases. Data extracted from the primary studies were then analyzed with fixed effect meta-analysis model for effect rate to explore heterogeneity between the primary studies. Publication bias was evaluated using funnel plot. A total of 10,819 articles were retrieved from the data bases of which 48 studies met inclusion criteria. V. parahaemolyticus could only be isolated from 2761 (47.5 %) samples of 5811 seafood investigated. The result of this study shows that incidence of V. parahaemolyticus was more prevalent in oysters with overall prevalence rate of 63.4 % (95 % CI 0.592–0.674) than other seafood. Overall prevalence rate of clams was 52.9 % (95 % CI 0.490–0.568); fish 51.0 % (95 % CI 0.476–0.544); shrimps 48.3 % (95 % CI 0.454–0.512) and mussels, scallop and periwinkle: 28.0 % (95 % CI 0.255–0.307). High heterogeneity (p value <0.001; I2 = 95.291) was observed mussel compared to oysters (I2 = 91.024). It could be observed from this study that oysters harbor V. parahaemolyticus based on the prevalence rate than other seafood investigated. The occurrence and prevalence of V. parahaemolyticus is of public health importance, hence, more studies involving seafood such as mussels need to be investigated.

Quantitative Analysis of Pathogenic and Nonpathogenic V ibrio parahaemolyticus in Shrimp Derived from Industrial Processing

Vibrio parahaemolyticus is an important seafood-borne pathogen. V. parahaemolyticus carrying either tdh or trh or both genes is considered a pathogenic strain. The objectives of this study were to examine the prevalence and concentration of V. parahaemolyticus in shrimp processed in a factory using most probable number (MPN) combined with either polymerase chain reaction (PCR) or loop-mediated isothermal amplification (LAMP) techniques. The pathogenic group (pathogenic Vp) is composed of strains carrying tdh (tdh+ Vp) and/or trh (trh+ Vp) and nonpathogenic group (total Vp) is consisted of both pathogenic and nonpathogenic strains. There was no difference in the total Vp detection by PCR and LAMP techniques. However, combined MPN with LAMP techniques, an increase in detection of pathogenic Vp in samples was identified and the concentrations of pathogenic Vp in shrimp from the first five processing steps were 3.2, 3.0, 3.1, 3.5 and 4.3 MPN/g and were below the detection level (<3 MPN/g) in shrimp from the last three steps. LAMP technique estimated significantly higher positive samples and MPN values for pathogenic V. parahaemolyticus than PCR technique. Practical Applications Loop-mediated isothermal amplification technique is more sensitive than polymerase chain reaction at determining low numbers of pathogenic Vibrio parahaemolyticus in seafood. Decreased temperature in the peeling step as well as frequent change in dipping water in the grazing step will reduce the total numbers of V. parahaemolyticus contaminated in shrimp.

Molecular Detection Methods for Vibrio parahaemolyticus in Seafood

To detect Vibrio parahaemolyticus in seafood, we evaluated efficient combinations of molecular methods with DNA extraction methods using heat extraction and alkaline heat extraction, and PCR, real-time PCR and loop-mediated isothermal amplification (LAMP) assays were performed targeting V parahaemolyticus species-specific genes (tlh and rpoD) and pathogenic factors genes (tdh and trh). The species-specific genes were detected in all combinations of two strains (a tdh * trh1-positive strain and a trh2-positive strain), two kinds of shellfish (oyster and bloody clams) and molecular methods with tlh-real time PCR or rpoD-LAMP assays with DNA of alkaline heat extraction at 85-145cfu/test level. tdh was detected in both seafoods with real time PCR assay with DNA of heat extraction at 85cfu/test level, and detected with the LAMP and real time PCR assays with DNA of alkaline heat extraction at 85cfu/test level. Detection of both trh1 and trh2 with the PCR assay with DNA of alkaline heat extraction was comparatively high though trh2 was detected with the LAMP assay with DNA of alkaline heat extraction at 145cfu/test level. It, however, is necessary to investigate more sensitive trh-detection methods. In this study, the results indicated that tlh-real time PCR or rpoD-LAMP, tdh-real time PCR and tdh-LAMP assays with DNA of alkaline heat extraction are relatively-sensitive methods to detect V. parahaemolyticus in seafood.

제외국 식중독균 위해관리 정책 비교 연구

본 연구는 제외국의식중독균별저감화사례를조사, 비교하여 정보를 획득하고 식중독균 저감화 매뉴얼을 작성하며 저감화 정책방안을 제시하기 위한 기반 자료 확보를 목적으로 수행하였다. 선진 외국(미국, 덴마크, 일본)의 식중독 저감화 정책의 현황 및 사례조사에는 인터넷 자료, 관련 기관 전문가 인터뷰를 통하여 조사하였다. 미국의 신선농산물에서 미생물학적 위험요소 저감화(FDA), 미국 분쇄쇠고기에서 E. coli O157:H7의 저감화(USDA), 덴마크의 닭고기, 돼지고기, 계란에서 살모넬라 저감화, 일본의 어패류에서 장염비브리오 저감화 프로그램을 사례 연구로 제시하였으며 이들 국가의 저감 프로그램의 배경, 전략, 효과를 조사하고 장단점 분석을 실시하고 시사점을 도출하였다. 이들 제외국 정책 사례 연구와 국제기구 CODEX의 위해관리 체계를 결합하고 국내 현실을 반영하여 식중독 저감화 관리 매뉴얼로 저감 프로그램에 관한 일반 모델을 다음 순서로 제시하였다: 1) 위해관리 초기판단, 2) 식중독 저감화 프로그램 계획, 3) 대안 확인 및 선택, 4) 대안 실행(이해당사자별 역할 설정 및 대안 방법 적용), 5) 감시 활동, 6) 중간 재검토, 7) 목적 달성 때까지 대안 실행 지속적 실시 (만일 대안이 효과가 없으면 대안을 대체하거나 수정하여 식중독 저감화 목표 달성 시까지 실시), 8) 최종 평가에 보건에 미치는 영향을 확인하고 필요시 비용 편익 분석실시. 본 연구를 통해서 제시된 식중독 저감화 정책에 관한 시사점 및 식품안전의 이해당사자별 역할별로 도출된 식중독 저감화 정책 모델 및 매뉴얼은 미생물학적 위해관리를 수행하는데 향후 활용될 수 있다. This study was performed to acquire the information on the foodborne pathogen risk management programs in a couple of developed countries by the expert meeting and searching the information on the web. The backgrounds, strategies and effects related to microbial hazards of the foodborne pathogen reduction programs in fresh produce (US), Escherichia coli O157:H7 in ground beef (US), Salmonella in chicken, pork and eggs (Denmark), and Vibrio parahaemolyticus in seafood (Japan) were investigated for case study. A comparison among the pathogen reduction programs was conducted to find advantages and disadvantages and implications of the policies to bring out implications of the programs. A model for foodborne pathogen reduction program was developed based on both the CODEX risk management scheme and the case studies as follows; 1) preliminary risk management activities, 2) planing a foodborne pathogen reduction program, 3) option identification and selection, 4) implementation (conducting the each stake-holders role and applying the intervention methods), 5) monitoring activities, 6) interim review, 7) continuation or amendment of implementation method by the interim review before achieving the goal, and 8) final review and additional cost-benefit analysis if necessary. This proposed model according to the role of the stake-holders can be used to conduct microbial risk management programs in Korea in the near future.

Vibrio choleraeandVibrio parahaemolyticusDetected in Seafood Products from Senegal

The detection of pathogenic Vibrio in seafood from Senegal has generated five food alerts in the European Union. To investigate the presence and abundance Vibrio cholerae and Vibrio parahaemolyticus in seafood and coastal and estuarine waters, 123 seafood samples and 52 water samples were collected during 2007-2009 from two large seafood markets in Dakar, and from different oceanic and estuarine areas of the country. V. parahaemolyticus was detected in 30.1% of seafood samples, whereas presence of V. cholerae was only found in 1.6%. In water samples, V. parahaemolyticus and V. cholerae were detected in 28.8% and 5.7% of the samples, respectively. Abundance of V. parahaemolyticus in seafood from the fishing areas ranged from <0.3 to 7.5 most probable number (MPN) per gram. In samples from markets, densities of V. parahaemolyticus showed higher values ranging from 0.61 to >110 MPN/g. Densities of V. cholerae in the two positive seafood samples reached values of 0.36 and 0.61 MPN/g, repectively. V. parahaemolyticus strains were found to possess tlh, but not tdh and trh by polymerase chain reaction, and all the strains of V. cholerae were non-O1 or non-O139. These results suggest that the prevalence of high salinities in coastal and estuarine environments of Senegal limits the occurrence of V. parahaemolyticus and V. cholerae, despite warmer temperatures prevailing in seawater environments throughout the year. Furthermore, temperature abuse driven by a deficient cold chain over the distribution and retail sales may represent a major risk due to the postharvest multiplication of these Vibrio pathogens.

Prevalence and characterisation of potentially virulent Vibrio parahaemolyticus in seafood in Malaysia using conventional methods, PCR and REP-PCR

A previously developed multiplex PCR targeting gyrB of Vibrios at genus level and pntA genes for specific detection of Vibrio cholerae, Vibrio parahaemolyticus and Vibrio vulnificus was evaluated. The sensitivity of the multiplex PCR on spiked seafood was 1.5 × 103 CFU g−1. One hundred and fifty seafood samples were collected from retail stores and hypermarkets in different locations in Kuala Lumpur, Petaling Jaya and Seri Kembangan. The prevalence of V. parahaemolyticus was 29% (43/150). The pntA primers for V. parahaemolyticus detection were 100% specific and comparable to the toxR gene-based PCR. Six (12%) and 2 (4%) isolates contained trh and tdh genes, respectively. Repetitive Extragenic Palindromic PCR (REP-PCR) was used to genetically characterize the V. parahaemolyticus isolates in which 41 REP profiles were observed and all the isolates were categorized into 11 distinct clusters at the similarity of 80%. tdh-positive isolates shared a low level of similarity with trh-positive isolates. The prevalence of V. parahaemolyticus and particularly the presence of virulent gene such as trh and tdh among the isolates reiterate a high risk of contamination for seafood consumers in Malaysia. DNA fingerprinting of V. parahaemolyticus in this study indicates a high genetic diversity among the isolates and REP-PCR was able to distinguish the isolates with different virulotypes.