Parahaemolyticus In Shrimp Research Articles

Heat shock protein 70 (HSP70) regulates innate immunity and intestinal microbial homeostasis against Vibrio parahaemolyticus in shrimp

Heat shock proteins (HSPs) are essential for maintaining cellular homeostasis and regulating immune responses across diverse species. In aquaculture animals, the HSP70 family plays a crucial role in regulating innate immunity against pathogenic infections, although the mechanisms remain unclear. In this study, we investigate the role of HSP70 from Litopenaeus vannamei (LvHSP70) in regulating immune responses and intestinal microbiota composition during Vibrio parahaemolyticus infection. Using RNA interference (RNAi) to knock down LvHSP70 in shrimp, we observed significant impacts on homeostasis, including reduced activity, loss of feed intake, hepatopancreas damage, and individual death. During V. parahaemolyticus infection, shrimp with silenced LvHSP70 exhibited increased bacterial loads and elevated tissue damage. Transcriptomic analysis revealed that LvHSP70 knockdown significantly downregulated the expression of a broad range of immune-related genes, including antimicrobial peptides, cytokines, and components of the prophenoloxidase (ProPO) system. Additionally, 16S rDNA sequencing demonstrated a marked increase in the relative abundance of Vibrio in the intestines of LvHSP70-deficient shrimp. These findings underscore the essential role of LvHSP70 beyond its classical heat shock response in maintaining shrimp homeostasis, regulating immune responses, and controlling intestinal microbiota.

Passive immunoprophylaxis with Ccombodies against Vibrio parahaemolyticus in Pacific white shrimp (Penaeus vannamei)

The Vibrio parahaemolyticus strain causing acute hepatopancreatic necrosis disease (AHPND) in shrimp secretes toxins A and B (PirAVp/PirBVp). These toxins have been implicated in pathogenesis and are targets for developing anti-AHPND therapeutics or prophylactics that include passive immunization. We have previously reported that Ccombodies (recombinant hagfish variable lymphocyte receptor B antibodies; VLRB) targeting PirBVp conferred protection against V. parahaemolyticus in shrimp when administered as a feed supplement. In this study, we screened a phage-displayed library of engineered VLRBs for PirAVp-targeting Ccombodies that were mass-produced in a bacterial expression system. We then introduced these Ccombodies into the diet of Pacific white shrimp (Penaeus vannamei) over a seven-day period. Subsequently, the shrimp were exposed to a challenge with V. parahaemolyticus. Mortality rates were then observed and recorded over the following seven days. Administering shrimp feed supplemented with Ccombodies at a high dose (100 mg per 100 g feed) reduced mortality in recipient animals (2.96–5.19 %) statistically similar to mock-challenged control (1.48 %), but significantly different from the Ccombody-deficient control (74.81 %). This suggests that the Ccombodies provided strong protection against the bacterium. Feeding shrimp with a median dose (10 mg/100 g feed) gave statistically comparable low mortality (5.93–6.67 %) as the high dose. Reducing the Ccombody dose to 1 mg/100 g feed showed variable effects. Ccombody A2 showed mortality (11.85 %) significantly lower than that of the Ccombody-deficient group (74.81 %), suggesting that it can effectively protect against the bacterial challenge at a low dose. Our results demonstrate the ability of the phage-displayed VLRB library to generate antigen-specific Ccombodies rapidly and simply, with the expression of high protein levels in bacteria. The protective effect provided by these Ccombodies aligns with our earlier results, strongly supporting the use of VLRB antibodies as a substitute for IgY in passive immunoprophylaxis against AHPND in shrimp.

Meta-Analysis and Systematic Review of Phenotypic and Genotypic Antimicrobial Resistance and Virulence Factors in Vibrio parahaemolyticus Isolated from Shrimp.

This systematic review and meta-analysis investigates the prevalence of Vibrio parahaemolyticus, its virulence factors, antimicrobial resistance (AMR), and its resistance determinants in shrimp. This study was conducted following the Preferred Reporting Items for Systematic Reviews and Meta-Analysis (PRISMA) guidelines, to identify and select relevant peer-reviewed articles published between January 2020 and December 2022. The search strategy involved multiple online databases, including Google Scholar, PubMed, ScienceDirect, and Scopus. The inclusion criteria focused on studies that examined V. parahaemolyticus prevalence, virulence factors, and AMR in shrimp from farms to retail outlets. A total of 32 studies were analyzed, revealing a pooled estimate prevalence of V. parahaemolyticus in shrimp at 46.0%, with significant heterogeneity observed. Subgroup analysis highlighted varying prevalence rates across continents, emphasizing the need for further investigation. Virulence factor analysis identified thermostable direct hemolysin (tdh) and tdh-related hemolysin (trh) as the most common. Phenotypic AMR analysis indicated notable resistance to glycopeptides, nitrofurans, and beta-lactams. However, the correlation between antimicrobial usage in shrimp farming and observed resistance patterns was inconclusive. Funnel plots suggested potential publication bias, indicating a need for cautious interpretation of findings. This study underscores the urgency of coordinated efforts to address AMR in V. parahaemolyticus to safeguard public health and to ensure sustainable aquaculture practices.

Responses of growth performance, immunity, disease resistance of shrimp and microbiota in Penaeus vannamei culture system to Bacillus subtilis BSXE-1601 administration: Dietary supplementation versus water addition

This study evaluated the effects of Bacillus subtilis BSXE-1601, applied either as dietary supplementation or water addition, on growth performance, immune responses, disease resistance of Penaeus vannamei, and microbiota in shrimp gut and rearing water. During the 42-day feeding experiment, shrimp were fed with basal diet (CO and BW group), basal diet supplemented with live strain BSXE-1601 at the dose of 1 × 109 CFU kg−1 feed (BD group) and 15 mg kg−1 florfenicol (FL group), and basal diet with strain BSXE-1601 added to water at the concentration of 1 × 107 CFU L−1 every five days (BW group). Results showed that dietary supplementation of strain BSXE-1601 significantly promoted growth performance of shrimp, both in the diet and water, enhanced disease resistance against Vibrio parahaemolyticus (P < 0.05). The BD and BW groups exhibited significant increases in acid phosphatase, alkaline phosphatase, lysozyme, peroxidase, superoxide dismutase activities, phenonoloxidase content in the serum of shrimp compared to the control (P < 0.05). Meanwhile, the expression of immune-related genes proPO, LZM, SOD, LGBP, HSP70, Imd, Toll, Relish, TOR, 4E-BP, eIF4E1α, eIF4E2 were significantly up-regulated compared to the control (P < 0.05). When added in rearing water, strain BSXE-1601 induced greater immune responses in shrimp than the dietary supplement (P < 0.05). Chao1 and Shannon indices of microbiota in rearing water were significantly lower in BD group than in the control. The microbiota in rearing water were significantly altered in BD, BW and FL groups compared to the control, while no significant impacts were observed on the microbiota of shrimp gut. When supplemented into the feed, strain BSXE-1601 obviously reduced the number of nodes, edges, modules in the ecological network of rearing water. The results suggested that dietary supplementation of BSXE-1601 could be more suitable than water addition in the practice of shrimp rearing when growth performance, non-specific immunity, disease resistance against V. parahaemolyticus in shrimp were collectively considered.

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

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

Decreased Severity of Acute Hepatopancreatic Necrosis Disease in White Shrimp (Litopenaeus vannamei) by Mixed Culture of Bacillus subtilis, Bacillus licheniformis and Bacillus megaterium.

The objective of this study was to investigate the mixed culture of Bacillus subtilis, B. licheniformis and B. megaterium to control acute hepatopancreatic necrosis disease (AHPND) or EMS (Early Mortality Syndrome) in white shrimp Litopenaeus vannamei as a model. The infected shrimps with Vibrio parahaemolyticus AHPND strain were divided into tanks and different feeding of either B. subtilis, B. licheniformis, B. megaterium or all Bacillus strains. The infected shrimps that were fed with a mixed culture of Bacillus showed significantly highest survival rate and revealed lower percent detection of V. parahaemolyticus AHPND strain by Polymerase Chain Reaction (PCR) (57.14%) with a small amount of viability count in their hepatopancreas. In contrast, the infected shrimps that were fed with each of B. subtilis, B. licheniformis or B. megaterium, revealed the spread of V. parahaemolyticus AHPND strain in all tissue by PCR detection (86.67%-100%) with a large amount of viability count (3.53 - 4.24 × 103 CFU/g). This study indicated that the mixed culture of Bacillus subtilis, B. licheniformis and B. megaterium could control the dissemination of V. parahaemolyticus in shrimps, especially in hepatopancreatic that is the target tissue of AHPND in white shrimp (L. vannamei). The result of this study revealed the efficiency and mechanism of the mixed culture of B. subtilis, B. licheniformis and B. megaterium to control the virulence of AHPND and support the application of this mixed culture in aquaculture of shrimp farms to avoid chemical and antibiotic treatment by using it as a biological control.

The Prebiotic Effect of an Organic Acid Mixture on Faecalibacterium prausnitzii Metabolism and Its Anti-Pathogenic Role against Vibrio parahaemolyticus in Shrimp.

Increasing the abundance of probiotic bacteria in the gut requires either direct dietary supplementation or the inclusion of feed additives able to support the growth of beneficial commensal bacteria. In crustaceans, the increased presence of probiotic-like bacteria in the gut, including of Faecalibacterium prausnitzii (F. prausnitzii), will guarantee a positive health status and a gut environment that will ensure enhanced performance. The aim of this study was to investigate if a mixture of organic acids, AuraAqua (Aq) can stimulate the growth and the anti-pathogenic efficacy of F. prausnitzii through a combination of in vitro and ex vivo models. The results showed that 0.5% Aq was able to improve the growth rate of F. prausnitzii in vitro and in an ex vivo shrimp gut model. Moreover, we were able to demonstrate that Aq increases butyrate production and cellulose degradation in culture or in the shrimp gut model. The growth-stimulating effect of Aq also led to an improved and anti-pathogenic effect against Vibrio parahaemolyticus in a co-culture experiment with shrimp gut primary epithelial cells (SGP). In conclusion, our work demonstrates that Aq can stimulate the growth of F. prausnitzii, increase the production of short-chain fatty acid (SCFA) butyrate, improve substrate digestion, and prevent V. parahaemolyticus invasion of SGP cells.

Phytochemical and Antibacterial Effects of Leaf Extract from Mangrove Plant (Avicennia Marina) on Vibrio Parahaemolyticus in Shrimps

Recently, there has been a tremendous increase in the studies addressing the application of bioactive compounds from the natural ecosystem, particularly for medical purposes. Hence, the present study investigated the antibacterial properties of the secondary metabolites possibly contained in the leaves of Avicennia marina (A. marina) for possible prevention of Vibrio parahaemolyticus (V. parahaemolyticus), a devastating bacterial pathogen in shrimp aquaculture. In the current study, secondary metabolites were extracted from the leaves of mangrove plant using ethanol extraction method. The ethanolic extracts were then subjected to phytochemical and antibacterial activity tests. The results from the phytochemical analysis demonstrated that the ethanolic extract from the mangrove plant contained varying amounts of flavonoids, tannins, saponins, polyphenols, alkaloids, steroids, and triterpenoids. However, the number of flavonoids and alkaloids seemed to be higher than the other metabolites. The antibacterial activity analysis through the agar diffusion method has shown that different concentrations (50 ppm, 100 ppm, 200 ppm, and 300 ppm) of the ethanolic extract of A. marina inhibited the V. parahaemolyticus. At 300 ppm, the plant extract exhibited 17.3% antibacterial effectiveness, compared to the antibacterial activity of chloramphenicol. The findings indicated that the secondary metabolites of A. marina have the potential that can be developed as an alternative treatment for aquatic animal diseases in the future.

Quantitative Risk Assessment of Vibrio parahaemolyticus Toxi Infection Associated with the Consumption of Roasted Shrimp (Penaeus monodon)

In this study, a risk assessment on Vibrio parahaemolyticus infections was carried out in order to estimate the likelihood of gastroenteritis for Cameroonians after consumption of roasted shrimp (Penaeus monodon). The Codex Alimentarius Commission framework was used in this study. Based on the distribution of total V. parahaemolyticus in shrimp and literature information indicating that nonhaemolysing carrier strains could be pathogenic to humans, the cooking, and consumption patterns, the daily exposure level generated in this study, and the dose-response model from other studies, the infectious risk was evaluated and quantified by the Monte Carlo simulation. This simulation was realized based on 10,000 iterations using the Model Risk software, version 4.0, in combination with Microsoft Excel. To better quantify the exposure of consumers and the resulting risk of infection, several scenarios reflecting the minimal, average, and maximal exposures were undertaken. According to the results, the 90% confidence intervals for minimum and maximum exposures ranged from 15 to 24 colony-forming units per day (cells/day) and from 160 to 228 cells/day, respectively. Based on the modal scenario, 90% of the population consuming this shrimp is exposed to V. parahaemolyticus loads ranging from 74 to 110 cells/day, indicating a risk of infection ranging from 1.2 to 1.8 cases per million of consumption. The estimated number of annual disease cases based on annual production is between 1 and 10 cases. This reflects a relatively low risk of infection for roasted shrimp. Good hygiene practices during handling, cooking, and storage may help reduce the actual risk.

A kelch motif-containing protein KLHDC2 regulates immune responses against Vibrio parahaemolyticus and white spot syndrome virus in Penaeus vannamei

The kelch motif-containing proteins are widely present in organisms and known to be involved in various biological processes, but their roles in immunity remain unclear. In this study, a kelch motif-containing protein KLHDC2 was identified from Pacific white shrimp Penaeus vannamei and its immune function was investigated. The klhdc2 gene was widely expressed in shrimp tissues and its protein product was mainly present in the nucleus. Expression of klhdc2 was regulated by shrimp NF-κB family members Dorsal and Relish, and changed after immune stimulation. KLHDC2 could enhance the immune defense against Vibrio parahaemolyticus in shrimp but inhibit that against white spot syndrome virus (WSSV). Further analyses showed that KLHDC2 did not affect the phagocytosis of hemocytes but regulated the expression of a series of immune effector genes. KLHDC2 has a complex regulatory relationship with Dorsal and Relish, which may partly contribute to its positive role in antibacterial response by regulating humoral immunity. Moreover, the regulatory effect of KLHDC2 on WSSV ie1 gene contributed to its negative effect on antiviral response. Therefore, the current study enrichs the knowledge on the Kelch family and helps to learn more about the regulatory mechanism of shrimp immunity.

Protective effects of the biofloc system in Pacific white shrimp (Penaeus vannamei) culture against pathogenic Vibrio parahaemolyticus infection

The mitigation of vibriosis in shrimp culture could be accomplished by reducing the virulence of the pathogen or by increasing the immune response of the shrimp. This study aims to evaluate the application of a biofloc system in protecting Pacific white shrimp (Penaeus vannamei) from pathogenic Vibrio parahaemolyticus infection. Shrimp post-larvae (PL 20) with an average body weight of 0.041 ± 0.019 g were reared in an aquarium with a working volume of 33 L at a density of 3 L−1 for 21 days using two rearing systems, i.e., the biofloc system and the regular clear water system as control. In each system, the shrimp post-larvae were challenged by adding V. parahaemolyticus at different densities, 103, 105, and 107 CFU mL−1, while the negative control was performed by maintaining shrimp post-larvae in the clear water system without the addition of V. parahaemolyticus. The results of the in vitro experiment showed that the density and biofilm activity of V. parahaemolyticus reared in biofloc suspension was lower than that of the positive control (p < 0.05). The density of V. parahaemolyticus in shrimp rearing water was lower than that in the control at 105 CFU mL−1, especially on the 3rd day post-challenge, but there was no significant difference in the total presumptive Vibrio count between the biofloc treatment and the control. The survival, growth, and immune response parameters, such as total hemocyte count, phagocytic activity, respiratory burst, and phenoloxidase activity, of the shrimp, reared in the biofloc system were also higher than those of the positive control (p < 0.05), regardless of the density of V. parahaemolyticus. The present study demonstrated that the application of biofloc could significantly protect and increase the resistance of Pacific white shrimp against pathogenic V. parahaemolyticus infection.

Effect of bovine lactoferricin on the growth performance, digestive capacity, immune responses and disease resistance in Pacific white shrimp, Penaeus vannamei

The present study evaluated the growth performance, digestive enzyme activity, non-specific immunity, immunity and growth genes in Penaeus vannamei fed diets supplemented with Bovine lactoferricin (the basal diet without Bovine lactoferricin, the control; 1.0‰ Bovine lactoferricin，LCB1; 1.5‰ Bovine lactoferricin，LCB1.5; 2.0‰ Bovine lactoferricin, LCB2; 2.5‰ Bovine lactoferricin, LCB2.5) for 56 days. The feeding trial showed that the final weight, weight gain rate, and specific growth rate of the shrimp were improved significantly, while the feed conversion ratio was reduced significantly in the LCB1.5 group compared to the control (P < 0.05). The challenge test of Vibrio parahaemolyticus showed that the cumulative mortalities of shrimp in the LCB1.5, LCB2 and LCB2.5 groups were significantly lower than that in the control (P < 0.05). Compared with the control, Lipase and Trypsin activities in the hepatopancreas of LCB1.5 and LCB2 groups were significantly enhanced (P < 0.05). Compared with the control, alkaline phosphatase, acid phosphatase activities in the hepatopancreas and the relative expression levels of Relish, Toll, JAK, STAT, TOR, Raptor, 4E-BP, eIF4E1α, eIF4E2 genes in the hepatopancreas of LCB1.5, LCB2 and LCB2.5 groups were all significantly enhanced (P < 0.05). These results suggested that dietary Bovine lactoferricin could improve the growth performance, digestive capacity and immune responses of shrimp. When resistance against Vibrio parahaemolyticus in shrimp is considered, high dosage of Bovine lactoferricin showed a better effect than low dosage of Bovine lactoferricin. However, high dosage of Bovine lactoferricin can have a negative impact on the growth performance of shrimp. Considering collectively the above, Bovine lactoferricin could improve the growth performance, digestive enzymes activities, immune responses and disease resistance of P. vannamei.

Nano-SiO2 inhibits the marine aquatic pathogen Vibrio parahaemolyticus

In this study, we investigated the effects of Nano-SiO2, ultraviolet (UV) irradiation, and hydrogen peroxide on the growth of four marine aquatic pathogens as well as on their polysaccharide, biofilm formation, protein, and DNA. The results indicated that among all concentrations, 100 μg/L Nano-SiO2 demonstrated the most inhibition of Edwardsiella tarda, Vibrio anguillarum, and Vibrio parahaemolyticus. Ultraviolet irradiation for 3 min had the most obvious inhibitory effect on E. tarda, but neither inhibitory nor promoting effects on the other three strains. High hydrogen peroxide concentrations had obvious inhibitory effects on all bacterial growth. Moreover, Nano-SiO2 had some influence on bacterial polysaccharide properties and some amount of DNA splicing effect, but no effect was noted on proteins. At present, the recognized pathogenic genes of V. parahaemolyticus include tdh, trh, and tlh. We thus studied the effect of Nano-SiO2 on these genes and then conducted infection experiments. The results showed that Nano-SiO2 further reduced infection with V. parahaemolyticus in shrimp.

Evaluation of paraprobiotic applicability of Clostridium butyricum CBG01 in improving the growth performance, immune responses and disease resistance in Pacific white shrimp, Penaeus vannamei

The present study evaluated the growth performance, non-specific immunity and disease resistance in Penaeus vannamei fed diets supplemented with live or dead cells of Clostridium butyricum CBG01 (live cells, CB; sonication-killed cell-free extracts, UI; heat-killed whole-cell, HI; fermentation supernatant, FS; the control, the basal diet without C. butyricum, DZ) for 42 days. Results indicated that the final weight, specific growth rate, survival rate and feed efficiency rate of shrimp in the treatment groups were significantly improved versus the control (P < 0.05). The challenge test of Vibrio parahaemolyticus showed that the cumulative mortalities of shrimp in the CB and UI groups were significantly lower than that in the control (P < 0.05). Compared with the control, alkaline phosphatase, acid phosphatase, total nitric oxide synthase, lysozyme, peroxidase, superoxide dismutase activities, total antioxidant capacity, and phenonoloxidase content in the serum and the relative expression levels of SOD, LZM, proPO, LGBP, HSP70, Imd, Toll, Relish, TOR, 4E-BP, eIF4E1α, eIF4E2 genes in the hepatopancreas of CB and HI shrimp groups were all significantly enhanced, and those were significantly improved in the UI group as well, except for phenonoloxidase content, relative expression levels of SOD, Imd and eIF4E2 genes (P < 0.05). However, immune responses were induced partially in the FS shrimp group. These results suggested that dietary both live and dead cells of C. butyricum CBG01 could improve the growth performance and immune responses of shrimp. When resistance against Vibrio parahaemolyticus in shrimp is considered, sonication-killed cell-free extracts of C. butyricum showed a better effect than heat-killed whole-cells of probiotic. Considering collectively the above, sonication-killed cell-free extracts of C. butyricum could be applied as a potential paraprobiotic to enhance the growth performance, immunity capacity and disease resistance of P. vannamei.

Toxicity of recombinant PirA and PirB derived from Vibrio parahaemolyticus in shrimp

Acute hepatopancreatic necrosis disease (AHPND), caused by emerging strains of Vibrio Parahaemolyticus, is of concern in shrimp aquaculture. Secreted proteins PirA and PirB, encoded by a plasmid harbored in V. parahaemolyticus, were determined to be the major virulence factors that induce AHPND. To better understand pathogenesis associated with PirA and PirB, recombinant proteins rPirA and rPirB were produced to evaluate their relative toxicities in shrimp. By challenging shrimp at concentration of 3 μM with reverse gavage method, rPirA and rPirB (approximately 0.4 and 1.5 μg per g of body weight, respectively) caused 27.8 ± 7.8% and 33.3 ± 13.6% mortality, respectively; combination of 3 μM rPirA and rPirB resulted in 88.9 ± 7.9% mortality. Analysis of protein mobility in native gel revealed that rPirB was apparently in the form of monomer while rPirA was oligomerized as an octamer-like macromolecule, suggesting that inter- and intra-molecular interactions between rPirA and rPirB enhanced the toxic effect. An attempt to block or reduce rPirA activity with a putative receptor, N-acetyl-galactosamine, was unsuccessful, implying that remodeling analysis of PirA molecule, such as the octamer observed in this study, is necessary. Results of this study provided new insight into toxic mechanism of PirA and PirB and shall help design strategic antitoxin methods against AHPND in shrimp.

Isolation and Characterization of Bacteriophages from Inland Saline Aquaculture Environments to Control Vibrio parahaemolyticus Contamination in Shrimp.

The online version contains supplementary material available at 10.1007/s12088-021-00934-6.

Complete genome sequence of highly pathogenic Vibrio parahaemolyticus isolated from mariculture Penaeus vannamei reveals virulence factor genes

The complete genomic sequence of Vibrio parahaemolyticus strain HN1 was determined. The HN1 genome consisted of two circular chromosomes, with a total length of 5,024,488 bp and a G + C content of 45.16%. In total, 4,580 coding genes with an average length of 944 bp were identified. Among them, 3,818 genes (83.36%) were annotated to the Clusters of Orthologous Groups (COG) database; 2,811 (61.38%) to the Gene Ontology (GO) database; and 2,493 (54.43%) to the Kyoto Encyclopedia of Genes and Genomes (KEGG) database. A genetic virulence analysis identified 577 virulence genes, including cya, flagellum-related genes, Bsa T3SS, p60 and the type IV secretion system in V. parahaemolyticus strain HN1. Meanwhile, hemolysin was identified as a virulence protein. In a further analysis, no thermostable direct hemolysin (TDH) or TDH-related hemolysin (TRH) was identified, but thermolabile hemolysin (TLH) was identified. In a secretory system analysis, genes associated with the type I, type II, type III, and type VI secretory systems were identified. The type III (located on chromosome 1) and type VI secretory systems (located on chromosome 1 and chromosome 2) were annotated by multiple genes and may play an important role in the strong pathogenicity of V. parahaemolyticus in shrimp. In addition, multiple drug resistance genes were identified and antibiotic-susceptibility assays were performed.

Comparative genomic analysis unravels the transmission pattern and intra-species divergence of acute hepatopancreatic necrosis disease (AHPND)-causing Vibrio parahaemolyticus strains.

Acute hepatopancreatic necrosis disease (AHPND) is a recently discovered shrimp disease that has become a severe threat to global shrimp-farming industry. The causing agents of AHPND were identified as Vibrio parahaemolyticus and other vibrios harboring a plasmid encoding binary toxins PirAvp/PirBvp. However, the epidemiological involvement of environmental vibrios in AHPND is poorly understood. In this study, with an aim to reveal the possible transmission route of AHPND-causing V. parahaemolyticus, we sequenced and analyzed the genomes of four pairs of V. parahaemolyticus strains from four representative regions of shrimp farming in China, each including one strain isolated from diseased shrimp during an AHPND outbreak and one strain isolated from sediment before AHPND outbreaks. Our results showed that all the four shrimp-isolated and three of the sediment-isolated strains encode and secret PirAvp/PirBvp toxins and, therefore, are AHPND-causing strains. In silico multilocus sequence typing and high-resolution phylogenomic analysis based on single-nucleotide polymorphisms, as well as comparison of genomic loci in association with prophages and capsular polysaccharides (CPSs) consistently pointed to a close genetic relationship between the shrimp- and sediment-isolated strains obtained from the same region. In addition, our analyses revealed that the sequences associated with prophages, CPSs, and type VI secretion system-1 are highly divergent among strains from different regions, implying that these genes may play vital roles in environmental adaptation for AHPND-causing V. parahaemolyticus and thereby be potential targets for AHPND control. Summing up, this study provides the first direct evidence regarding the transmission route of AHPND-causing V. parahaemolyticus and underscores that V. parahaemolyticus in shrimp are most likely originated from local environment. The importance of environmental disinfection measures in shrimp farming was highlighted.

Microalgae species as potential inhibitors of the bioactivity of Vibrio parahaemolyticus in shrimps

Vibrio parahaemolyticus is a Gram-negative motile bacteria belonging to Vibrionaceae family causing cytotoxicity in cultured cells and enterotoxicity in animal models. Similarly, this bacterium is a major foodborne pathogen that causes life-threatening diseases in human, including gastroenteritis and septicemia, via consumption of contaminated seafood. Based on the increasing interest in developing novel environmental friendly solutions for the inhibition of the bioactivity of V. parahaemolyticus, microalgae species have emerged as potential candidates due to the microbiota associated with microalgae cultures, oxygen radicals produced during the photosynthetic process and the production of secondary metabolites, useful for antibacterial properties. In the present review, conventional treatments concerning V. parahaemolyticus bioactivity have been presented. However, in terms of effectiveness, numerous studies explaining the use of microalgae species for antibacterial activity against V. parahaemolyticus have been detailed suggesting that microalgae-based treatment are considerably better. Finally, we presented a proposed microalgae-based solution that can act as an inhibitor of V. parahaemolyticus bioactivity and therefore can be applied in the aquaculture industrial field to improve productivity and reduce costs.

Identification of the thioredoxin-related protein of 14 kDa (TRP14) from Litopenaeus vannamei and its role in immunity

The thioredoxin system plays essential roles in maintenance and regulation of the redox state of cysteine residues in cellular proteins. The thioredoxin-related protein of 14 kDa (TRP14) is an important member of the TRX superfamily which acts on various substrate proteins, some of which are not overlapped with those of thioredoxin. The knowledge on the function of TRP14 in invertebrates is limited to date. In this study, a TRP14 gene was identified from Pacific white shrimp Litopenaeus vannamei (LvTRP14) and its role in immune responses was investigated. We demonstrated that the expression level of LvTRP14 was high in hepatopancreas and intestine, low in eyestalk, and medium in other tissues of healthy shrimp. The transcription of LvTRP14 in vivo was significantly down-regulated in Relish-silencing shrimp but up-regulated in STAT-silencing shrimp, indicating a complex regulation of LvTRP14 expression. Although the LvTRP14 expression showed little change after immune stimulation with different type of pathogens, knockdown of LvTRP14 expression using RNAi strategy could significantly facilitate the infection of white spot syndrome virus (WSSV) and Vibrio parahaemolyticus in shrimp. Dual luciferase reporter assays demonstrated that LvTRP14 enhanced the transcription factor activity of Relish but attenuated that of Dorsal. Furthermore, silencing of LvTRP14 in vivo had opposite effects on expression of different type of antimicrobial peptides. These suggested that LvTRP14 could play a complex role in shrimp immunity.