Parahaemolyticus Detection Research Articles

Sensitive detection of Vibrio parahaemolyticus via a dual-recognition colorimetric biosensor comprising Cefe-PEG-MNPs and apt-Fe@PDA

Highly accurate and sensitive detection of Vibrio parahaemolyticus is crucial for ensuring seafood safety and preventing the transmission of foodborne diseases. Herein, a dual-recognition strategy-driven colorimetric biosensor for the detection of V. parahaemolyticus was developed. This biosensor was based on the use of PEG-mediated cefepime-functionalized magnetic nanoparticles (Cefe-PEG-MNPs) and aptamer-modified Fe-doped polydopamine (Fe@PDA) nanozymes. The magnetic platform Cefe-PEG-MNPs was introduced to enhance the enrichment efficiency of V. parahaemolyticus, achieving a capture efficiency exceeding 90%. The Fe@PDA nanozyme exhibited notable peroxidase mimetic enzyme activity. The colorimetric biosensor developed for V. parahaemolyticus detection demonstrated good sensitivity, with a detection range spanning from 2.1 × 101 to 2.1 × 106 CFU/mL, and a limit of detection of 21 CFU/mL. Moreover, the biosensor demonstrated remarkable specificity, achieving good recovery rates from 95.80% to 103.80% in Penaeus vannamei with a relative standard deviation of less than 1.38%. The newly developed colorimetric biosensor offers a remarkable opportunity for the accurate and sensitive detection of V. parahaemolyticus in seafood.

Contamination of Vibrio parahaemolyticus in crayfish for sale.

Crayfish (Procambarus clarkii) are economically important freshwater crustaceans. With the growth of the crayfish industry, the associated food-safety risks should be seriously considered. Although Vibrio parahaemolyticus is commonly recognized as a halophilic foodborne pathogen associated with seafood, it has been found to be a major pathogen in crayfish-associated food poisoning cases. In this study, the V. parahaemolyticus contamination level in crayfish production-sale chain was investigated using crayfish and environmental samples collected from crayfish farms and markets. Serious V. parahaemolyticus contamination (detection rate of 66%) was found in the entire crayfish production-sale chain, while the V. parahaemolyticus contamination level of the market samples was extremely high (detection rate of 92%). The V. parahaemolyticus detection rate of crayfish surface was similar to that of whole crayfish, indicating that crayfish surface was important for V. parahaemolyticus contamination. The simulation experiments of crayfish for sale being contaminated by different V. parahaemolyticus sources were performed. All the contamination sources, containing V. parahaemolyticus-positive tank, water, and crayfish, were found to be efficient to contaminate crayfish. The crayfish tank displayed the most significant contaminating role, while the water seemed to inhibit the V. parahaemolyticus contamination. The contamination extent of the crayfish increased with the number of V. parahaemolyticus cells the tank carried and the contact time of the crayfish and the tank, but decreased with the time that the crayfish were maintained in the water. It was also confirmed that the crayfish surface was more susceptible to V. parahaemolyticus contamination than the crayfish intestine. Furthermore, the adsorption of V. parahaemolyticus onto the crayfish shell was analyzed. Over 90% of the V. parahaemolyticus cells were adsorbed onto the crayfish shell in 6 h, indicating a significant adsorption effect between V. parahaemolyticus and the crayfish shell. In conclusion, within a water-free sale style, the fresh crayfish for sale in aquatic products markets uses its shell to capture V. parahaemolyticus cells from the V. parahaemolyticus-abundant environments. The V. parahaemolyticus contamination in crayfish for sale exacerbates the crayfish-associated food-safety risk. This study sheds light on V. parahaemolyticus control and prevention in crayfish industry.

Construction of an Electrochemical Molecularly Imprinted Sensor Based on L-carnitine Doped Polypyrrole for Vibrio Parahaemolyticus Detection

This study addresses the pressing need for rapid detection of pathogenic microorganisms by designing and fabricating an L-carnitine-doped polypyrrole (L-CN/PPy) electrochemical molecularly imprinted sensor, with Vibrio parahaemolyticus as the model target. Employing L-carnitine as a dual-functional dopant and auxiliary recognition element in the electropolymerization process, the sensor not only enhances the conductive properties of polypyrrole but also significantly improves its specific recognition capability towards the target bacteria. The sensor preparation encompasses the electrochemical polymerization of L-CN/PPy composite material, immobilization and elution of template bacteria, followed by bacterial recognition and detection steps. Through systematic optimization of L-CN concentration, polymerization conditions, eluent composition, elution time, and operational parameters, the sensor's performance is maximized. Experimental results confirm the sensor's efficacy in rapidly completing template bacteria elution within just 30 minutes and achieving efficient recognition of Vibrio parahaemolyticus within 20 minutes, demonstrating high sensitivity and selectivity. Moreover, the straightforward and rapid construction method of the sensor introduces a novel technological strategy for practical microbial detection applications, highlighting extensive application prospects.

A 3D-Printed Integrated Handheld Biosensor for the Detection of Vibrio parahaemolyticus.

Vibrio parahaemolyticus (V. parahaemolyticus) is one of the important seafood-borne pathogens that cause a serious gastrointestinal disorder in humans. Recently, biosensors have attracted serious attention for precisely detecting and tracking risk factors in foods. However, a major consideration when fabricating biosensors is to match the low cost of portable devices to broaden its application. In this study, a 3D-printed integrated handheld biosensor (IHB) that combines RPA-CRISPR/Cas12a, a lateral flow strip (LFS), and a handheld device was developed for the ultrasensitive detection of V. parahaemolyticus. Using the preamplification of RPA on tlh gene of V. parahaemolyticus, a specific duplex DNA product was obtained to activate the trans-cleavage activity of CRISPR/Cas12a, which was then utilized to cleave the ssDNA probe. The ssDNA probe was then detected by the LFS, which was negatively correlated with the content of amplified RPA products of the tlh gene. The IHB showed high selectivity and excellent sensitivity for V. parahaemolyticus detection, and the limit of detection was 4.9 CFU/mL. The IHB also demonstrated great promise for the screening of V. parahaemolyticus in samples and had the potential to be applied to the rapid screening of other pathogen risks for seafood and marine environmental safety.

Boron nitride nanoplate-based improvement of the specificity and sensitivity in loop-mediated isothermal amplification for Vibrio parahaemolyticus detection

BackgroundNucleic acid testing based on DNA amplification is gradually entering people's modern life for clinical diagnosis, food safety monitoring and infectious disease prevention. Polymerase chain reaction (PCR) and loop-mediated isothermal amplification (LAMP) are the most powerful techniques that have been the gold standard for quantitative nucleic acid analysis. However, the high nonspecific amplification rate caused by the formation of primer dimers, hairpin structures and mismatched hybridization severely restricts their real-world applications. It is highly desirable to explore a way for improving the specificity and sensitivity of PCR and LAMP assays. ResultsIn this work, we demonstrated that a nanomaterial boron nitride nanoplate (BNNP), due to its unique surface properties, can interact with the main components of the amplification reaction, such as single stranded primers and Bst DNA polymerase, and increase the thermal conductivity of the solution. As a result, the presence of BNNPs dramatically improved the specificity of PCR and LAMP. And BNNPs maintained the specificity even after five rounds of PCR. Moreover, the sensitivity of LAMP was also enhanced by BNNPs, and the detection limit of BNNP-based LAMP was two orders of magnitude lower than that of classical LAMP. Then the BNNP-based LAMP was applied to detect Vibrio parahaemolyticus in contaminated seafood samples with high specificity and a 10-fold increase in sensitivity. SignificanceThis is the first systematic demonstration of BNNPs as a promising additive to enhance the efficiency and fidelity of PCR and LAMP amplification reactions, thereby greatly expanding the application of nucleic acid detection in a wide range of laboratory and clinical settings.

Enhancement of loop mediated isothermal amplification's sensitivity and speed by multiple inner primers for more efficient identification of Vibrio parahaemolyticus

The modified loop-mediated isothermal amplification (LAMP), called multiple hybrid, inner primers (MHP)-LAMP, was developed to enhance the efficiency of the existing LAMP-based assay for Vibrio parahaemolyticus detection. The method was built on a conventional LAMP assay by employing 2 newly designed extra sets of primers to increase the initial binding sites of core primers on the V. parahaemolyticus’s rpoD gene from 8 to 12. With this strategy, the assay detection sensitivity was increased by 10 folds, with the detection limit (DL) approaching 100 copies of purified target genomic DNA (gDNA) as analyzed by real-time turbidity measurement and gel electrophoresis. The MHP also accelerated the rate of DNA amplification by 30%, rendering the assay faster. The MHP-LAMP assay did not cross- react with other pathogens, indicating that it was highly specific for V. parahaemolyticus detection. Whilst V. parahaemolyticus was used as a study model herein, our idea of using MHP to maximize assay sensitivity and speed is considered as a universal strategy that can be applied to enhance efficiency of LAMP-based assays for detecting any DNA and RNA of interest.•The strategy of using multiple hybrid, inner primers (MHP) to enhance LAMP assay's efficiency was demonstrated with success.•The MHP enhanced the sensitivity and speed of the existing LAMP assay, designed to detect V. parahaemolyticus, by 10 times and 30%, respectively.•The proposed strategy can be applied to boost up any other LAMP-based assay's diagnostic performance.

D-tartaric acid doping improves the performance of whole-cell bacteria imprinted polymer for sensing Vibrio parahaemolyticus

Whole-cell bacteria imprinted polymer-based sensors still face challenges in the form of the difficulty of removing the template entirely, low affinity, and poor sensitivity. To further improve their performance, it is pivotal to modulate the morphology and chemical properties of imprintied polymer by taking advantage of doping engineering. Here we introduced D-tartaric acid (D-TA) as a dopant and employed pyrrole as a functional monomer to construct D-TA/polypyrrole (PPy)-based bacteria imprinted polymer (DPBIP) sensor for Vibrio parahaemolyticus (VP) detection. It is demonstrated that D-TA doping can synergistically accelerate the removal of template bacteria from imprinted polymers (1.5 h), improve bacteria affinity of imprinted sites (the recognition time of 30 min), and enhance the sensitivity of DPBIP sensor (a detection limit of 19 CFU mL−1). The DPBIP sensor had a linear range of 102∼106 CFU mL−1 and exhibited high selectivity and good repeatability. Moreover, a recovery of 94.8%–105.3% was achieved in drinking water and oyster samples. Therefore, small functional molecules doping opens a new avenue to engineering BIP-based sensors with high performance, holding potential applications in securing food safety.

Emergence and predominance of a new serotype of Vibrio parahaemolyticus in Huzhou, China

BackgroundDuring our surveillance period, we found a new type of Vibrio parahaemolyticus (V. parahaemolyticus) with serotype O10:K4, which had increased over the past 2 years in Huzhou and became the second most common serotype after O3:K6. MethodsStrains were isolated from stool samples of diarrhea patients in the First People's Hospital in Huzhou. The serotypes, virulence-associated genes, antimicrobial susceptibility, and pulsed field gel electrophoresis (PFGE) of these strains were analyzed. ResultsBetween January 2017 and December 2021, there were 598 (5.36%) V. parahaemolyticus-positive samples of 11,166 stool specimens. The V. parahaemolyticus detection rate was high in summer months. The O3:K6 was the dominant serotype in 2017–2020 and a new serotype, O10:K4, was the predominant serotype in 2021. The majority of isolates tested were resistant to ampicillin (86.8%). We randomly chose the strains with serotype O3:K6 and O10:K4 for PFGE to compare the genetic relationship between these two serotypes. The results showed that the PFGE profiles of V. parahaemolyticus O3:K6 and O10:K4 were genetically similar. The strains showed a tendency to cluster on the basis of their serotype profiles. However, some O3:K6 strains showed 100% similarity with O10:K4 strains. ConclusionA new serotype with pandemic potential of V. parahaemolyticus, O10:K4, was detected in 2020 and became dominant in 2021 in Huzhou.

Identification of Vibrio parahaemolyticus by PCR Method in Fish and Water Samples from Tigris River, Turkey

Climate change has caused increasing Vibrio parahaemolyticus infections in areas unaffected by this pathogen. Water temperature is an important ecological factor. It is consequently of high importance to follow V. parahaemolyticus in aquatic environments and foods. The aim of this work was to detect the presence of V. parahaemolyticus in Tigris River from eight different sites namely Malabadi Bridge, Fiskaya, Ongözlü Bridge, Bağıvar, Pamukçay, Ambarçay, Seyrantepe and Silvan. Total 480 samples of water and fish were used for V. parahaemolyticus detection. Thiosulphatecitrate‐bile salt sucrose (TCBS) Agar was employed for bacteria isolation for 24 h at 37◦C. The dominant uniform bacterial colonies were purified by streaking onto the TCBS agar plates three times. Identification of this pathogen was made by PCR. A total of 480 water and fish samples were analyzed. According to the results, V. parahaemolyticus was detected in 40 (6.6%) fish samples taken from the Seyrantepe region.

Development of real‐time fluorescence saltatory rolling circle amplification for rapid detection of Vibrio parahaemolyticus in seafood

AbstractAs a marine food‐borne pathogen, Vibrio parahaemolyticus (V. parahaemolyticus) can cause human gastrointestinal disorders and pose a serious threat to food safety and public health worldwide. Using a portable scanner, herein, a real‐time fluorescence saltatory rolling circle amplification (RF‐SRCA) has been established for the rapid detection of V. parahaemolyticus in seafood targeting toxR gene. The RF‐SRCA results could be effectively determined within 20–60 min via real‐time fluorescence curve. Significant specificity of RF‐SRCA was exhibited against 12 V. parahaemolyticus strains and 29 non‐V. parahaemolyticus strains. The sensitivity and detection limit of V. parahaemolyticus in artificially contaminated raw oyster by RF‐SRCA were 3.2 × 100 fg μL−1 and 2.3 × 100 CFU g−1, respectively. Compared with SRCA by electrophoresis, RF‐SRCA has higher sensitivity, lower detection limit, and avoids complicated electrophoresis. Compared with visual SRCA, the RF‐SRCA takes shorter time, provides more accurate results and can eliminate the risk of cross‐contamination. Moreover, 236 seafood samples were investigated for V. parahaemolyticus contamination, and the results showed 100% sensitivity, 95.88% specificity and 98.31% accuracy compared with the ISO method. Therefore, RF‐SRCA possesses great potential as an accurate, specific, sensitive, rapid and convenient molecular diagnostic method for V. parahaemolyticus detection from various seafoods.

A reversible valve-assisted chip coupling with integrated sample treatment and CRISPR/Cas12a for visual detection of Vibrio parahaemolyticus

Vibrio parahaemolyticus (V. parahaemolyticus) is regarded as a major cause of seafood-associated illnesses, which has aroused widespread public concern. Here, a rapid and convenient detection method for V. parahaemolyticus detection was established by a reversible valve-assisted chip coupling with CRISPR/Cas12a. With optimized lysis buffer, loop mediated isothermal amplification (LAMP) reagents and CRISPR reagents, the whole detection process from sampling to results could be finished within 50 min. The structure of chip was simple and the cost was low. By relying on three reversible rotary valves and the rotation direction-dependent Coriolis pseudo force, the flow order of liquid and the direction of liquid flow could be precisely controlled. The LAMP amplicons were specifically and sensitively identified by CRISPR/Cas12a. Positive amplification would produce green fluorescent signal while negative amplification generated no fluorescent signal, which could be clearly distinguished by the naked eye. With 600 μL of samples processed, the limit of detection (LOD) for both pure cultured V. parahaemolyticus or spiked shrimp samples could achieve 30 copies/reaction. These illustrated the established method displayed great feasibility for real samples detection. In the future, the chip could also combine with other amplification reactions, like PCR or recombinase polymerase amplification reaction (RPA), to conduct detection by changing the corresponding lyophilized amplification reagents. Overall, the proposed detection platform displays great potential for food safety analysis and clinical diagnostics, especially in resource-limited areas.

Simple Colorimetric Assay for Vibrio parahaemolyticus Detection Using Aptamer-Functionalized Nanoparticles.

Simple, rapid, and sensitive screening methods are the key to prevent and control the spread of foodborne diseases. In this study, a simple visual colorimetric assay using magnetic nanoparticles (MNPs) and gold nanoparticles (AuNPs) was developed for the detection of Vibrio parahaemolyticus. First, the aptamer responding to V. parahaemolyticus was conjugated onto the surface of MNPs and used as a specific magnetic separator. In addition, the aptamer was also immobilized on the surface of AuNPs and used as a colorimetric detector. In the presence of V. parahaemolyticus, a sandwich structure of MNP–aptamer–bacteria–aptamer–AuNPs is formed through specific recognition of the aptamer and V. parahaemolyticus. The magnetic separation technique was then applied to generate a detection signal. Owing to the optical properties of AuNPs, a visual signal could be observed, resulting in an instrument-free colorimetric detection. Under optimal conditions, this assay shows a linear response toward V. parahaemolyticus concentration through the range of 10–106 cfu/mL, with a limit of detection of 2.4 cfu/mL. This method was also successfully applied for V. parahaemolyticus detection in spiked raw shrimp samples.

Production of Phage Display-Derived Peptide and the Application for Detecting Vibrio parahaemolyticus by Combined PCR Technology

Vibrio parahaemolyticus (V. parahaemolyticus) is one of the most common and widely distributed food-borne bacteria. Thus, it is necessary to establish a rapid, specific, and sensitive detection approach for V. parahaemolyticus. The strategy of combining immunomagnetic separation method (IMS) and polymerase chain reaction (PCR) measure was established successfully for V. parahaemolyticus detection, because ordinary IMS always has restriction on employing conventional antibodies for their labor-intensive procedure, batch-to-batch variation in functionality, and limited shelf-life. Herein, we have generated a novel phage display-derived peptide with an amino acid sequence of MPRLPPA and use it as bio-recognition probe in IMS. The peptide was evaluated to have the advantages of high specificity, sensitivity, easy reproduction, and well-defined ability. When compared with the rabbit polyclone IgG antibodies (IgG)-IMS and chicken egg yolk antibodies (IgY)-IMS, peptide-IMS has absolute superiorities in terms of the dosages, detection time, and capture efficiency. It has the ability to take the place of traditional antibody and unquestionable value of reusing. Furthermore, the detection system of peptide-IMS-PCR possessed high specificity with a limit of detection (LOD) of 102 CFU/mL in pure culture and 103 CFU/mL in oyster. Thus, we successfully synthesize peptide against V. parahaemolyticus and applied it to establish a sensitive and specific detecting method for V. parahaemolyticus.

Pre-enrichment step, incubation temperature and type of selective media affect the pathogenic Vibrio parahaemolyticus detection efficiency in frozen prawns

This study evaluates the performance of different detection protocols of Vibrio parahaemolyticus in frozen prawns. The effect of enrichment and incubation temperature on the selectivity of thiosulfate citrate bile sucrose (TCBS) and CHROMagar™ Vibrio (CAV) media were determined. Enumeration of V. parahaemolyticus in frozen prawns by direct-plating on CAV at 37 °C yielded the highest count compared to the other tested direct-plating protocols (p < 0.05). The enrichment step was found to increase the selectivity of TCBS for V. parahaemolyticus but did not affect the selectivity of CAV. Furthermore, trh + V. parahaemolyticus isolates were only successfully recovered from CAV, suggesting that CAV is the better media for the detection of virulent V. parahaemolyticus compared to TCBS. Repetitive Extragenic Palindromic PCR typing (REP-PCR) indicated that CAV and TCBS, as well as direct-plating or enrichment-plating are selective for different subsets of V. parahaemolyticus present in a sample. In conclusion, the current work suggests that CAV is a better selective media for detection and isolation of V. parahaemolyticus in frozen prawns. This study also suggests that direct-plating on CAV with an incubation temperature of 35–37 °C could be an alternative to Most Probable Number (MPN)-PCR for enumeration of V. parahaemolyticus in frozen prawn samples.

A novel FRET biosensor based on four-way branch migration HCR for Vibrio parahaemolyticus detection

A versatile and powerful fluorescence resonance energy transfer (FRET) biosensor based on four-way branch migration hybridization chain reaction (HCR) was proposed for the detection of Vibrio parahaemolyticus (V. parahaemolyticus). In the system, two partly complementary hairpin structures of H1 and H2, and an assistant DNA strand (R) were designed. H1 was labeled with carboxyfluorescein (FAM) as fluorophore donor, and H2 was labeled with tetramethylrhodamine (TAMRA) as fluorophore acceptor. When target DNA was presented in the system, it would firstly bind with the assistant DNA to form a short double-strand DNA (dsDNA), and subsequently trigger the four-way branch migration HCR, which would produce long nicked dsDNA concatamer and bring a good deal of FAM and TAMRA in close proximity, so that dramatic FRET signals could be achieved. With the assistance of well-designed four-way branch migration HCR circuits, this FRET biosensor exhibited a superior detection capability, which could detect 0.067 nM target DNA and as low as 10 CFU mL−1 of V. parahaemolyticus. To our knowledge, it is the first time that a four-way branch migration HCR strategy for signal amplification has been adapted to the FRET biosensor, which might show great potential in food safety and clinical diagnosis.

Sensitive colorimetric immunoassay of Vibrio parahaemolyticus based on specific nonapeptide probe screening from a phage display library conjugated with MnO2 nanosheets with peroxidase-like activity.

Pathogen detection continues to receive significant attention due to the harmful effects of pathogens on public health. Herein, specific nonapeptide-fusion proteins pVIII (pVIII fusion) were isolated from phage VQTVQIGSD (designated by the sequence of a fused foreign peptide), which was specifically screened from the f8/9 landscape phage library against Vibrio parahaemolyticus (V. parahaemolyticus) in a high-throughput way. The as-prepared V. parahaemolyticus-specific recognition element is cheaper and more available than antibodies. Further, a highly sensitive colorimetric immunoassay for V. parahaemolyticus was established using pVIII fusion as capture probes coupled with protein-templated MnO2 nanosheets (NSs) as signal probes. In the presence of a target bacterium, V. parahaemolyticus, a sandwich-type complex of pVIII fusion-V. parahaemolyticus-MnO2 NS@pVIII fusion was formed through specific recognition of pVIII fusion and V. parahaemolyticus. The signal probes (MnO2 NSs) could catalyze the reaction of 3,3',5,5'-tetramethylbenzidine and H2O2 to generate a colorimetric change. The proposed V. parahaemolyticus detection method demonstrated a wide detection range (20-104 colony-forming units (CFU) mL-1), low limit of detection (15 CFU mL-1), excellent selectivity, and high reliability for real marine samples, showing potential application in marine microbiological detection and control.

Selective turn-on fluorescence detection of Vibrio parahaemolyticus in food based on charge-transfer between CdSe/ZnS quantum dots and gold nanoparticles

Vibrio parahaemolyticus (V. parahaemolyticus) is one of the most common food-borne pathogens found along the east coast of China, Japan, and several southeast Asian countries, where marine foods are frequently consumed. Thus it is of great interest to establish a fast, sensitive and robust analytical assay for V. parahaemolyticus detection. Herein, we report a V. parahaemolyticus-specific egg yolk antibody (IgY), extracted from the egg yolks of immunized hens with inactivated V. parahaemolyticus bacteria. By immobilizing IgY onto the surface of gold nanoparticles (AuNPs) through electrostatic self-assembly, a detection system was developed based on charge-transfer quenching fluorescence, which contains core/shell CdSe/ZnS quantum dots (QDs), IgY-AuNPs and free AuNPs. In this strategy, the IgY-AuNPs were induced to aggregate quickly in the presence of target V. parahaemolyticus, resulting in a rapid fluorescence response of the QDs. More importantly, the convenient turn-on fluorescent detection system exhibited excellent selectivity over the other bacteria. Furthermore, this facile one-step method can detect V. parahaemolyticus at low concentrations of 10 cfu/mL without pre-enrichment and can be applicable for the detection of V. parahaemolyticus in real food samples. With the advantage of simplicity, rapidness, sensitivity, and specificity, this proposed approach shows promise as a successful application of V. parahaemolyticus in bioanalysis.

Ecological fitness and virulence features of Vibrio parahaemolyticus in estuarine environments.

Vibrio parahaemolyticus is a commonly encountered and highly successful organism in marine ecosystems. It is a fast-growing, extremely versatile copiotroph that is active over a very broad range of conditions. It frequently occurs suspended in the water column (often attached to particles or zooplankton), and is a proficient colonist of submerged surfaces. This organism is an important pathogen of animals ranging from microcrustaceans to humans and is a causative agent of seafood-associated food poisoning. This review examines specific ecological adaptations of V. parahaemolyticus, including its broad tolerances to temperature and salinity, its utilization of a wide variety of organic carbon and energy sources, and its pervasive colonization of suspended and stationary materials that contribute to its success and ubiquity in temperate and tropical estuarine ecosystems. Several virulence-related features are examined, in particular the thermostable direct hemolysin (TDH), the TDH-related hemolysin (TRH), and the type 3 secretion system, and the possible importance of these features in V. parahaemolyticus pathogenicity is explored. The impact of new and much more effective PCR primers on V. parahaemolyticus detection and our views of virulent strain abundance are also described. It is clear that strains carrying the canonical virulence genes are far more common than previously thought, which opens questions regarding the role of these genes in pathogenesis. It is also clear that virulence is an evolving feature of V. parahaemolyticus and that novel combinations of virulence factors can lead to emergent virulence in which a strain that is markedly more pathogenic evolves and propagates to produce an outbreak. The effects of global climate change on the frequency of epidemic disease, the geographic distribution of outbreaks, and the human impacts of V. parahaemolyticus are increasing and this review provides information on why this ubiquitous human pathogen has increased its footprint and its significance so dramatically.

Electrochemical Immunosensor for Detection of Vibrio parahaemolyticus Based on Faradaycage-Type Anodic Stripping Voltammetry

This paper aims to develop a Faradaycage-type immunoassay mode combined with anodic stripping voltammetry for Vibrio parahaemolyticus (VP) detection. Detection antibody (dAb) immobilized on GO-supported AgNPs (AgNPs@GO) was used as the detector unit, while capture antibody (cAb) immobilized on Fe3O4 particles cAb@Fe3O4 as the capture unit. The immunoassay was performed on a magnetic glassy carbon electrode via immunoreactions among the capture unit, VP and the detector unit. The detection sensitivity is expected to be considerably enhanced owing to two reasons: (1) large amounts of adsorbed AgNPs on the GO surface, made the electrochemical responses more sensitive; (2) thanks to the combined immune structure of Faradaycage-type, all AgNPs labeled on the detector unit could take part in the electrode reaction to emit signals. Under optimal conditions, the immunosensor exhibits a relatively wide linear range from 102 to 108 CFU mL−1 with a detection limit of 33 CFU mL−1. In addition, high selectivity and excellent recovery in seawater and seafood indicate that this immunosensor shows great prospect for practical application. Featured with high sensitivity, the proposed sensing scheme can thus offer new opportunities to achieve sensitive, selective, and simple detection of other bacteria.

The hot oyster: levels of virulent Vibrio parahaemolyticus strains in individual oysters.

Vibrio parahaemolyticus is the leading cause of seafood-associated gastroenteritis and is most commonly transmitted by raw oysters. Consequently, detection of virulent strains of this organism in oysters is a primary concern for seafood safety. Vibrio parahaemolyticus levels were determined in 110 individual oysters harvested from two sampling sites in SC, USA. The majority of oysters (98%) contained low levels of presumptive V. parahaemolyticus However, two healthy oysters contained presumptive V. parahaemolyticus numbers that were unusually high. These two 'hot' oysters contained levels of presumptive V. parahaemolyticus within the gills that were ∼100-fold higher than the average for other oysters collected at the same date and location. Current V. parahaemolyticus detection practices require homogenizing a dozen oysters pooled together to determine V. parahaemolyticus numbers, a procedure that would dilute out V. parahaemolyticus in these 'hot' oysters. This study demonstrates the variability of V. parahaemolyticus densities taken from healthy, neighboring individual oysters in the environment. Additionally, environmental V parahaemolyticus isolates were screened for the virulence-related genes, tdh and trh, using improved polymerase chain reaction primers and protocols. We detected these genes, previously thought to be rare in environmental isolates, in approximately half of the oyster isolates.