Antibiotic resistance trends among Vibrio vulnificus and Vibrio parahaemolyticus isolated from the Chesapeake Bay, Maryland: a longitudinal study.

Antibiotics are often used to treat severe Vibrio infections, with third-generation cephalosporins and tetracyclines combined or fluoroquinolones alone being recommended by the US Centers for Disease Control and Prevention. Increases in antibiotic resistance of both environmental and clinical vibrios are of concern; however, limited longitudinal data have been generated among environmental isolates to inform how resistance patterns may be changing over time. Hence, we evaluated long-term trends in antibiotic resistance of vibrios isolated from Chesapeake Bay waters (Maryland) across two 3-year sampling periods (2009-2012 and 2019-2022). Vibrio parahaemolyticus (n = 134) and Vibrio vulnificus (n = 94) toxR-confirmed isolates were randomly selected from both sampling periods and tested for antimicrobial susceptibility against eight antibiotics using the Kirby-Bauer disk diffusion method. A high percentage (94%-96%) of V. parahaemolyticus isolates from both sampling periods were resistant to ampicillin and only 2%-6% of these isolates expressed intermediate resistance or resistance to third-generation cephalosporins, amikacin, tetracycline, and trimethoprim-sulfamethoxazole. Even lower percentages of resistant V. vulnificus isolates were observed and those were mostly recovered from 2009 to 2012, however, the presence of multiple virulence factors was observed. The frequency of multi-drug resistance was relatively low (6%-8%) but included resistance against antibiotics used to treat severe vibriosis in adults and children. All isolates were susceptible to ciprofloxacin, a fluoroquinolone, indicating its sustained efficacy as a first-line agent in the treatment of severe vibriosis. Overall, our data indicate that antibiotic resistance patterns among V. parahaemolyticus and V. vulnificus recovered from the lower Chesapeake Bay have remained relatively stable since 2009.IMPORTANCEVibrio spp. have historically been susceptible to most clinically relevant antibiotics; however, resistance and intermediate-resistance have been increasingly recorded in both environmental and clinical isolates. Our data showed that while the percentage of multi-drug resistance and resistance to antibiotics was relatively low and stable across time, some Vibrio isolates displayed resistance and intermediate resistance to antibiotics typically used to treat severe vibriosis (e.g., third-generation cephalosporins, tetracyclines, sulfamethoxazole-trimethoprim, and aminoglycosides). Also, given the high case fatality rates observed with Vibrio vulnificus infections, the presence of multiple virulence factors in the tested isolates is concerning. Nevertheless, the continued susceptibility of all tested isolates against ciprofloxacin, a fluoroquinolone, is indicative of its use as an effective first-line treatment of severe Vibrio spp. infections stemming from exposure to Chesapeake Bay waters or contaminated seafood ingestion.

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

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

Preparation of valine-curcumin conjugate and its in vitro antibacterial and antitumor activity and in vivo biological effects on American eels (Anguilla rostrata).

Curcumin (Cur) exhibits diverse natural pharmacological activities, despite its limited water solubility (hydrophobicity) and low bioavailability. In this investigation, a valine-curcumin conjugate (Val-Cur) was synthesized through amino acid side chain modification, and its solubility increased to 1.78 mg/mL. In vitro experimental findings demonstrated that the antibacterial activity of Val-Cur against Escherichia coli, Staphylococcus aureus, Aeromonas hydrophila, and Vibrio parahaemolyticus was significantly superior to that of Cur. The inhibition rate of Val-Cur against HepG2 (human hepatocellular carcinoma) cells was higher than that of Cur at low concentrations (below 25 µmol/L), although the IC50 value of Val-Cur did not differ significantly from that of Cur. In vivo biological effects of Val-Cur were assessed by adding it into the feed (150 mg/kg) of American eels (Anguilla rostrata). Val-Cur significantly improved the growth performance (↑weight gain rate, ↑specific growth rate, and ↓feed conversion rate) and activities of intestinal digestive enzymes (amylase and lipase) and antioxidant enzymes (superoxide dismutase) in American eels. Additionally, Val-Cur significantly improved serum biochemical indices (↑high-density lipoprotein cholesterol, ↓low-density lipoprotein cholesterol, ↓aspartate and alanine aminotransferases). Furthermore, Val-Cur increased intestinal microbial diversity, reduced the abundance of potentially pathogenic bacteria (Spiroplasma, Clostridium, and Pseudomonas), and elevated the abundance of beneficial digestion-promoting bacteria (Romboutsia, Phyllobacterium, Romboutsia sedimentorum, and Clostridium butyricum) conducive to glucose metabolism (P < 0.05). To the best of our knowledge, this study is the first to explore water-soluble curcumin in aquaculture, and the findings will lay the groundwork for the potential application of water-soluble curcumin in the field of aquaculture.

Efficacy of Annona glabra extract against acute hepatopancreatic necrosis disease in white-leg shrimp (Penaeus vannamei).

This study aims to investigate the use of pond apple (Annona glabra) compounds as a novel strategy to prevent and treat acute hepatopancreatic necrosis disease (AHPND) as well as to better understand the mechanism of health improvement in shrimp. The A. glabra leaf extracts were extracted using various solvents and examined for in vitro and in vivo activity against Vibrio parahaemolyticus strains. In comparison with ethanol and water extracts, methanol extract showed the strongest bactericidal effect (MBC/MIC ratio of 2.50 ± 1.00), with minimal inhibitory concentration (MIC) of 0.023 ± 0.012 mg ml-1 and minimum bactericidal concentration (MBC) of 0.065 ± 0.062 mg ml-1. White leg shrimp (P. vannamei, body weight 10.37 ± 0.27 g) fed A. glabra methanol extracts-containing diets (AMEDs) at 1 %, 1.5 %, and 2.0 % demonstrated no deleterious effects on survival and were significantly increased in length and weight after 30 days of feeding. The level of total haemocyte, hyaline haemocyte on day 15 and granulocyte on day 30 remarkably increased (p < 0.05) in shrimps fed AMEDs groups compared to those in the control group. The finding demonstrates that granulocyte was induced time dependently. In particular, the survival rate of V. parahaemolyticus challenged shrimps under medication with AMEDs at 1.5 % and 2.0 % was significantly higher (p < 0.05) than that of the control group. The decrease in bacterial load of Vibrio spp. and V. parahaemolyticus was obviously recorded in hepatopancreas shrimp given AMEDs 1.5 % and 2.0 % and may be linked to herb characteristics such as antibacterial activity, enhancing innate immunity, and its potential to maintain the integrity of hepatopancreatic tissue. Our findings suggest that A. glabra extract might be used as a health enhancer in commercial farmed shrimp.

Phytochemical analysis and antibacterial activity of palm waste extract against Vibrio harveyi and Vibrio parahaemolyticus.

Vibrio harveyi and Vibrio parahaemolyticus are species of the Vibrio genus that often cause disease and mass mortality in crustaceans. If not handled quickly and appropriately, these diseases can cause considerable losses to farmers. Therefore, it is necessary to find a solution with safe and environmentally friendly disease prevention technology using natural ingredients, among others from plants, namely oil palm. Some parts of oil palm, namely leaves, fronds, fibres and oil palm pulp, which are palm waste, contain antibacterial compounds. This study aimed to assess the antibacterial activity of palm waste extracts, namely pulp, leaves, fronds and fibres using n-hexane, ethyl acetate, chloroform, ethanol and water maceration solvents against pathogenic bacteria V. harveyi and V. parahaemolyticus, and identify active compounds contained in palm waste. The results of the research are expected to produce innovative and sustainable solutions to control diseases in shrimp farming, contribute to the development of a sustainable fishing industry and open up the potential for utilizing palm waste as a value-added resource in the field of aquatic health. The results of observations on antibacterial activity tests and identifying the content of palm waste extract compounds were analysed descriptively displayed in the form of figures, tables and graphs. The results showed that palm waste extracts (pulp, leaves, fronds and fibres) with ethyl acetate and ethanol maceration solvents had very strong antibacterial potential, namely 20.14 ± 0.31 mm-25.52 ± 1.42 mm on V. harveyi bacteria and 20.41 ± 0.55 mm-25.00 ± 0.51 mm on V. parahaemolyticus bacteria. Palm extracts with n-hexane (>20 mm) and chloroform solvents generally have strong category antibacterial potential (10-20 mm), and palm extracts in water solvents have medium category potential (5-10 mm) against V. harveyi and V. parahemolyticus bacteria. The results of phytochemical tests on palm waste extracts with ethyl acetate and ethanol maceration solvents contain bioactive compounds of flavonoids, saponins, polyphenols and alkaloid tannins, steroids and triterpenoids. Palm extracts with n-hexane and chloroform solvents generally contain saponins, alkaloids, steroids and triterpenoids, while palm waste extracts with water solvents contain saponins.

Indole-3-acetic acid increases the survival of brine shrimp challenged with vibrios belonging to the Harveyi clade.

Vibrios belonging to the Harveyi clade (including closely related species such as Vibrio campbellii, Vibrio harveyi and Vibrio parahaemolyticus) are important pathogens of aquatic organisms. In this study, we investigated the use of indole-3-acetic acid to control disease caused by Harveyi clade vibrios. Indole-3-acetic acid, which can be produced by various seaweeds and microalgae, was added to the rearing water of brine shrimp larvae challenged with 12 different Harveyi clade Vibrio strains. Indole-3-acetic acid significantly decreased the virulence of 10 of the strains without any effect on their growth. The latter is important as it will minimize the selective pressure for resistance development. The survival rate of brine shrimp larvae increased from 1.2-fold to 4.8-fold upon treatment with 400 µM indole-3-acetic acid. Additionally, indole-3-acetic acid significantly decreased the swimming motility in 10 of the strains and biofilm formation in eight of the strains. The mRNA levels of the pirA and pirB toxin genes were decreased to 46% and 42% by indole-3-acetic acid in the AHPND-causing strain V. parahaemolyticus M0904. Hence, our data demonstrate that indole-3-acetic acid has the potential to be an effective virulence inhibitor to control infections in aquaculture.

Global metabolic regulation in Vibrio parahaemolyticus under polymyxin B stimulation.

Vibrio parahaemolyticus is responsible for infection diseases of people who consume the contaminated seafood, but its metabolic regulation profile in response to colistin, the last treatment option for multidrug-resistant Gram-negative bacteria, remains unclear. In this study, the metabolic regulation profile of V. parahaemolyticus ATCC33846 under polymyxin B stimulation has been investigated. V. parahaemolyticus exposed to polymyxin B resulted in 4597 differentially transcribed genes, including 673 significantly up-regulated genes and 569 significantly down-regulated genes. In V. parahaemolyticus under polymyxin B stimulation, the cellular antioxidant systems to prevent bacteria from oxidant stress was activated, the synthesis of some nonessential macromolecules was reduced, and the assembly and modification of lipopolysaccharide and peptidoglycan to resist the attack from other antibiotics were promoted. These findings provide new insights into polymyxin B-related stress response in V. parahaemolyticus which should be useful for developing novel drugs for infection.

Antibiofilm activities of the cinnamon extract against Vibrio parahaemolyticus and Escherichia coli.

Vibrio parahaemolyticus and Escherichia coli are two major foodborne pathogens. In this paper, the antibiofilm activities of the ethanol extract of cinnamon against these two bacteria were studied in detail. The antibacterial activity and the MIC of the extract were determined, and the inhibition and removing effects of the extract on the biofilms of V. parahaemolyticus and E. coli were investigated. The biofilms stained with fluorescein isothiocyanate (FITC) and concanavalin A (Con A) were also observed by confocal laser scanning microscope (CLSM). The results indicated that the extract exhibited high antibacterial activity, with the MIC against V. parahaemolyticus and E. coli was 6.25 mg/mL. The effects on V. parahaemolyticus biofilm were significant with the inhibition rate of 75.46% at MIC, and the eradication rate of 93.26% at 32MIC, respectively. As to E. coli, the inhibition rate was 48.18% at MIC, and the eradication rate was 46.16% at 8MIC. Meanwhile, the extract could notably reduce the metabolic activities and the secretion of EPS in biofilm, it inhibited 78.57% EPS formation in V. parahaemolyticus biofilm at MIC, and eliminated 61.28% EPS in mature biofilm at 4MIC. CLSM images showed that the EPS of the treated biofilm became thinner and biofilm structure was looser, when compared with the untreated control. This study elucidated that the cinnamon extract was effective to prevent biofilm formation and eradicate mature biofilms of V. parahaemolyticus and E. coli.

Pharmacological properties of some mangrove sediment-associated bacillus isolates.

Mangrove sediment-associated bacteria are of significantly important in the field of medicine and pharmaceuticals as new promising sources of biologically active pharmacophores due to the extreme conditions, such as high salt concentration and soil anoxia. The sediment bacteria associated with Acanthus ilicifolius and Avicennia officinalis collected from the Mangalavanam mangrove ecosystem of the Kerala State of India were evaluated using various in vitro models for the assessment of their pharmacological properties. The bacteria exhibiting significant antioxidant and antimicrobial activities were isolated, identified, and characterized by the integrated microbiological, biochemical, and 16S rRNA sequencing. Among the varied bacteria isolated from mangrove sediments, Bacillus amyloliquefaciens MBMS5 (GenBank accession number MK765025) exhibited significant antimicrobial activities against various pathogenic bacteria, such as Aeromonas caviae, Vibrio parahemolyticus, and methicillin-resistant Staphylococcus aureus. The extracellular extracts of B. amyloliquefaciens MBMS5 exhibited potential antioxidant activity against free radical species coupled with anti-inflammatory property as displayed by the attenuation activity against pro-inflammatory 5-lipoxygenase.

The inhibitory effect of Ulva fasciata on culturability, motility, and biofilm formation of Vibrio parahaemolyticus ATCC17802.

The outbreak of vibriosis from Vibrio parahaemolyticus (V. parahaemolyticus) is one of common pathogenic diseases found in the mariculture environment. In this study, the inhibitory effect of Ulva fasciata (U. fasciata) on the culturability, motility, and biofilm formation of V. parahaemolyticus ATCC17802 was examined by co-culturing system. Results showed that both of secretion and live tissue of U. fasciata could convert culturable V. parahaemolyticus ATCC17802 to non-culturable, both reaching more than 99% of inhibition rate after 3-day co-culture, and higher density (12 g L-1) of U. fasciata exhibited stronger inhibition. The twitching behavior of V. parahaemolyticus ATCC17802 was more easily affected by U. fasciata than the swimming behavior after 3-day co-culture, with the inhibitory rates varying at the ranges of 1.70-30.29% (twitching behavior) and 10.06-44.86% (swimming behavior) under the different environmental factors (salinity, NO3--N and PO43--P concentrations), but no significant correlation was found. The greatest inhibition effect on V. parahaemolyticus ATCC17802 biofilm formation occurred at 12 h, with inhibition rates at the range of 11.03-67.10 %, while there was still no significant correlation between inhibition rate and the three environmental factors. The different environmental factors might induce U. fasciata to excrete different levels of secondary metabolites, which caused the various inhibitory effect on the cultivability, motility, and biofilm formation of V. parahaemolyticus ATCC17802.

Talarodrides A-F, Nonadrides from the Antarctic Sponge-Derived Fungus Talaromyces sp. HDN1820200.

Six new nonadride derivatives, named talarodrides A-F (1-6), were isolated from the Antarctic sponge-derived fungus Talaromyces sp. HDN1820200. All structures including the absolute configurations were deduced by extensive spectroscopic analysis and computational ECD calculations. Compounds 1-4 share a rare caged bicyclo[4.3.1]-deca-1,6-diene with a bridgehead olefin and maleic anhydride core skeleton, while compounds 5 and 6 possess the first case of a naturally occurring 5/7/6 methanocyclonona[c]furan skeleton. Talarodride A (1) and talarodride B (2) showed selective inhibitory effects against Proteus mirabilis and Vibrio parahemolyticus with MICs of 3.13-12.5 µM.

Multifunctional carbonized nanogels to treat lethal acute hepatopancreatic necrosis disease.

BACKGROUND: Shrimp aquaculture has suffered huge economic losses over the past decade due to the outbreak of acute hepatopancreatic necrosis disease (AHPND), which is mainly caused by the bacteria Vibrio parahaemolyticus (V. parahaemolyticus) with the virulence pVA1 plasmid, which encodes a secretory photorhabdus insect-related (Pir) toxin composed of PirA and PirB proteins. The Pir toxin mainly attacks the hepatopancreas, a major metabolic organ in shrimp, thereby causing necrosis and loss of function. The pandemic of antibiotic-resistant strains makes the impact worse. METHODS: Mild pyrolysis of a mixture of polysaccharide dextran 70 and the crosslinker 1,8-diaminooctane at 180 â„ƒ for 3 h to form carbonized nanogels (DAO/DEX-CNGs) through controlled cross-linking and carbonization. The multifunctional therapeutic CNGs inherit nanogel-like structures and functional groups from their precursor molecules. RESULTS: DAO/DEX-CNGs manifest broad-spectrum antibacterial activity against Vibrio parahaemolyticus responsible for AHPND and even multiple drug-resistant strains. The polymer-like structures and functional groups on graphitic-carbon within the CNGs exhibit multiple treatment effects, including disruption of bacterial membranes, elevating bacterial oxidative stress, and neutralization of PirAB toxins. The inhibition of Vibrio in the midgut of infected shrimp, protection of hepatopancreas tissue from Pir toxin, and suppressing overstimulation of the immune system in severe V. parahaemolyticus infection, revealing that CNGs can effectively guard shrimp from Vibrio invasion. Moreover, shrimps fed with DAO/DEX-CNGs were carefully examined, such as the expression of the immune-related genes, hepatopancreas biopsy, and intestinal microbiota. Few adverse effects on shrimps were observed. CONCLUSION: Our work proposes brand-new applications of multifunctional carbon-based nanomaterials as efficient anti-Vibrio agents in the aquatic industry that hold great potential as feed additives to reduce antibiotic overuse in aquaculture.

Surveillance of disease incidence in shrimp farms located in the east coastal region of India and in vitro antibacterial efficacy of probiotics against Vibrio parahaemolyticus.

We surveyed 130 shrimp farms located on the eastern coast of India to determine the prevalence of emerging diseases in Litopenaeus vannamei and Penaeus monodon. Live shrimps were collected from the farms based on external symptoms. The biochemical, molecular, and histopathology results confirmed infection with Enterocytozoon hepatopenaei (32.4%), Vibrio parahaemolyticus (27.7%), White Spot Syndrome Virus (25.4%), Vibrio alginolyticus (16.1%), Vibrio harveyi (13.1%), Monodon-type baculovirus (4.61%), and infectious Hematopoietic Necrosis Virus (2.3%) in the collected shrimps. Enterocytozoon hepatopenaei (EHP) occurred more frequently in L. vannamei than P. monodon, with the microsporidian spores in the hepatopancreas. In P. monodon, Monodon-type Baculovirus infection (33.3%) was dominant and small percentages of WSSV, IHHNV, V. alginolyticus, and V. harveyi were observed. A few ponds were observed with co-infection of EHP and WSSV (7.6%), V. parahaemolyticus and WSSV (4.6%) and also V. parahaemolyticus and EHP (6.1%). Among the Vibrio spp, V. parahaemolyticus showed the highest percentage of infection in L. vannamei. Overall, we found that shrimp were chiefly infected with EHP and V. parahaemolyticus. The impact of water quality parameters on shrimp diseases was not addressed in this study. In an antibiotic susceptibility study, V. parahaemolyticus isolated from L. vannamei ponds was susceptible to nitrofurantoin, chloramphenicol, oxytetracycline and tetracycline, but resistant to erythromycin and nalidixic acid. In a preliminary in vitro antibacterial activity assay, probiotics against V. parahaemolyticus showed high inhibitory activity and the results encourage further in-depth studies on the efficacy of probiotics for disease control and prevention in shrimp farms.

Using Bacillus subtilis as a Host Cell to Express an Antimicrobial Peptide from the Marine Chordate Ciona intestinalis.

Ciona molecule against microbes-A24 (CiMAM) isolated from the marine chordate Ciona intestinalis is an antimicrobial peptide. To generate CiMAM-expressing transgenic Bacillus subtilis, we constructed a plasmid expressing recombinant CiMAM (rCiMAM) and introduced it into B. subtilis. Transgenic strains C117 and C166 were selected since they were able to highly and stably express rCiMAM. We studied the bactericidal activity of pepsin-digested extracts from rCiMAM-expressing strains against freshwater and euryhaline pathogens that commonly occur in aquaculture ponds and found no difference from that of lactoferricin-expressing strains. The bactericidal activity of 1-µL aliquot from a total 5.5 mL extracted from 5 mL of cultured C117 (1.45 × 108 CFU·mL-1) and C166 (2.17 × 108 CFU·mL-1) against halophilic bacteria was equivalent to the efficacy of 57.06 and 32.35 ng of Tetracycline against Vibrio natriegens, 47.07 and 25.2 ng against V. parahaemolyticus, and 58.17 and 36.55 ng against V. alginolyticus, respectively, indicating higher bactericidal activity of pepsin-extracts from rCiMAM-containing strains against halophilic bacteria compared to that from lactoferricin-containing strains. Since the antibacterial activity of rCiMAM-expressing B. subtilis strains shows higher competence against halophilic pathogens compared to that against freshwater and euryhaline pathogens, these strains are promising candidates to protect marine fish and shellfish from halophilic bacterial infection.

A Lymphoid Organ Specific Anti-Lipopolysaccharide Factor from Litopenaeus vannamei Exhibits Strong Antimicrobial Activities.

Different shrimp species are known to possess apparent distinct resistance to different pathogens in aquaculture. However, the molecular mechanism underlying this finding still remains unknown. One kind of important antimicrobial peptides, anti-lipopolysaccharide factors (ALF), exhibit broad-spectrum antimicrobial activities. Here, we reported a newly identified ALF from the shrimp Litopenaeus vannamei and compared the immune function with its counterpart in the shrimp Fenneropenaeus chinensis. The ALF, designated as LvALF8, was specifically expressed in the lymphoid organ of L. vannamei. The expression level of LvALF8 was apparently changed after white spot syndrome virus (WSSV) or Vibrio parahaemolyticus challenges. The synthetic LBD peptide of LvALF8 (LvALF8-LBD) showed strong antibacterial activities against most tested Gram-negative and Gram-positive bacteria. LvALF8-LBD could also inhibit the in vivo propagation of WSSV similar as FcALF8-LBD, the LBD of LvALF8 counterpart in F. chinensis. However, LvALF8-LBD and FcALF8-LBD exhibited apparently different antibacterial activity against V. parahaemolyticus, the main pathogen causing acute hepatopancreatic necrosis disease (AHPND) of affected shrimp. A structural analysis showed that the positive net charge and amphipathicity characteristics of LvALF8-LBD peptide were speculated as two important components for its enhanced antimicrobial activity compared to those of FcALF8-LBD. These new findings may not only provide some evidence to explain the distinct disease resistance among different shrimp species, but also lay out new research ground for the testing and development of LBD-originated antimicrobial peptides to control of shrimp diseases.

Characteristics of Antimicrobial-Resistant Vibrio parahaemolyticus Strains and Identification of Related Antimicrobial Resistance Gene Mutations.

Multidrug-resistant (MDR) Vibrio parahaemolyticus strains have become a great threat to public health. The purpose of this study was to investigate differences in biological characteristics and antimicrobial resistance gene (ARG) mutations of V. parahaemolyticus that displayed different levels of antimicrobial resistance. The susceptibility of 74 V. parahaemolyticus strains to 9 common antimicrobials was investigated, of which 88% were resistant to 3-4 antimicrobials and 3% to 5-7 antimicrobials. Interestingly, only 9% were resistant to 1-2 antimicrobials. The MDR strains possessed longer growth lag time than the non-MDR strains and displayed weaker swimming abilities. Whole genome sequencing was performed on strains VP41, VP44, 460, and 469 that were resistant to two to three classes of antimicrobials. ARGs were identified and compared with that of reference strain ATCC17802, and some important mutations were deduced. The Val189Ile mutation emerged in qnr gene of a single strain. Besides, the nonsynonymous mutations existed in four ARGs in different strains, including CatB (Pro165Ser, Gly208Asp), VmeA (Ile313Thr), VmeC (Glu329Ala), and VmeD (Asn205Ser). These results linked resistance gene mutations to enhance resistance in V. parahaemolyticus strains and provide a reference for more effective monitoring and prevention of V. parahaemolyticus infections.

Rosmarinic Acid and Sodium Citrate Have a Synergistic Bacteriostatic Effect against Vibrio Species by Inhibiting Iron Uptake.

BACKGROUND: New strategies are needed to combat multidrug-resistant bacteria. The restriction of iron uptake by bacteria is a promising way to inhibit their growth. We aimed to suppress the growth of Vibrio bacterial species by inhibiting their ferric ion-binding protein (FbpA) using food components. METHODS: Twenty spices were selected for the screening of FbpA inhibitors. The candidate was applied to antibacterial tests, and the mechanism was further studied. RESULTS: An active compound, rosmarinic acid (RA), was screened out. RA binds competitively and more tightly than Fe3+ to VmFbpA, the FbpA from V. metschnikovii, with apparent KD values of 8 µM vs. 17 µM. Moreover, RA can inhibit the growth of V. metschnikovii to one-third of the control at 1000 µM. Interestingly, sodium citrate (SC) enhances the growth inhibition effect of RA, although SC only does not inhibit the growth. The combination of RA/SC completely inhibits the growth of not only V. metschnikovii at 100/100 µM but also the vibriosis-causative pathogens V. vulnificus and V. parahaemolyticus, at 100/100 and 1000/100 µM, respectively. However, RA/SC does not affect the growth of Escherichia coli. CONCLUSIONS: RA/SC is a potential bacteriostatic agent against Vibrio species while causing little damage to indigenous gastrointestinal bacteria.

Molecular characterization and antibiotic resistance of Vibrio parahaemolyticus from Indian oyster and their probable implication in food chain.

Vibrio parahaemolyticus is one of the leading causes of diarrhoea and gastroenteritis in human on consumption of raw or insufficiently cooked seafood. This study was aimed at isolating and characterizing the pathogenic and pandemic V. parahaemolyticus from oysters (n = 90) in coastal parts of West Bengal, India; their antibiotic resistance and potential for involvement in the food chain. During bacteriological culture, typical V. parahaemolyticus colony was recovered in 88.9% samples followed by presumptive identification in 71 (78.9%) samples by characteristic biochemical (K/A) test. All the presumptive isolates (n = 71) were confirmed by species specific Vp-toxR PCR assay. Of these, 10 (14.08%) were tdh+ and none for the trh. Further, 5 (50%) of these tdh+ isolates were found to carry the pandemic potential gene in PGS-PCR assay; however, none in GS-PCR. Majority (80%) of these pathogenic (tdh+) isolates belonged to pandemic serovars (OUT: KUT; OUT: K24; O1: KUT; O1:K25; O10: KUT) and only 20% to non-pandemic serovars (OUT: K15; O9:K17). All the isolates (100%) exhibited resistance to cefpodoxime followed by ampicillin and cefotaxime (90%), ceftizoxime (60%), tetracycline (50%), ceftriaxone (40%), ciprofloxacin and nalidixic acid (10% each). Overall, the study findings suggested that 11.1% (10/90) of commonly marketed oysters in this area were harbouring pathogenic V. parahaemolyticus. Moreover, 5.5% (5/90) of the oyster population were harbouring pandemic strains of this pathogen. Besides, the pathogenic isolates from oysters were exhibiting a considerable genetic relatedness (53 to 70%) to human clinical isolates in PFGE analysis that relates to a substantial public health risk. Further, their multidrug resistance added gravity to the antimicrobial resistance (AMR), a globally growing public health threat and this is a critical area of concern especially during the treatment of foodborne gastroenteritis.

NKL-24: A novel antimicrobial peptide derived from zebrafish NK-lysin that inhibits bacterial growth and enhances resistance against Vibrio parahaemolyticus infection in Yesso scallop, Patinopecten yessoensis.

The extensive use of antibiotics in aquaculture has resulted in the prevalence of antibiotic-resistant bacteria and, consequently, new antibacterial strategies or drugs with clear modes of action are urgently needed. Antimicrobial peptides (AMPs) are currently widely considered as alternatives to antibiotics in the treatment of infections in aquatic animals. In this study, we aimed to evaluate the effects of NKL-24, a truncated peptide derived from zebrafish NK-lysin, against Yesso scallop (Patinopecten yessoensis) pathogen, Vibrio parahaemolyticus. The results showed that NKL-24 had a potent antibacterial effect against V. parahaemolyticus via a membrane active cell-killing mechanism. The in vitro study showed that sub-lethal levels of NKL-24 obviously reduced bacterial swimming ability and downregulated the transcription of the selected genes associated with V. parahaemolyticus virulence. Studies on NKL-24 biosafety in hemocytes and in Yesso scallop have shown no adverse effects from this peptide. Bacteria challenge test results demonstrated that NKL-24 significantly decreased the mortality and inhibited bacterial growth in the scallop infected with V. parahaemolyticus, while further in vivo examination revealed that NKL-24 could enhance non-specific immune parameters. Moreover, NKL-24 was capable of modulating a series of V. parahaemolyticus-responsive genes in the scallop. These results suggest the protective action of NKL-24 against V. parahaemolyticus and the potential of this peptide as a promising candidate for aquaculture applications.

First screening report of immune and protective effect of non-toxic Jatropha vernicosa stem bark against Vibrio parahaemolyticus in Longfin yellowtail Seriola rivoliana leukocytes.

In México, the infusion of Jatropha vernicosa stem bark has been used in folk medicine for many clinical situations, but no reports were available about this particular species of Jatropha in fish of mammals. In this first screening report, the phytochemical, antioxidant profile and antimicrobial properties of aqueous J. vernicosa stem bark extract were explored against Vibrio parahaemolyticus, an opportunist fish pathogen. To evaluate the cytotoxicity and immunological effect for the possible application of aqueous J. vernicosa stem bark in aquaculture, this study assessed it by using Longfin yellowtail Seriola rivoliana leukocytes. The results showed that phytochemical composition of the J. vernicosa extract was rich in phenol, flavonoid, saponin, and coumarin compounds. The antioxidant capacity of hydroxyl radical and superoxide anion scavenging activities, iron-chelation activity and ß-carotene bleaching coupled to linoleic acid showed that J. vernicosa extracts had a moderate antioxidant effect compared with synthetic antioxidants (BHT, BHA and EDTA). No adverse effects were observed on spleen leukocytes (viability > 98%). Interestingly, J. vernicosa stem bark extract has immunostimulant and antioxidant effects, increasing phagocytosis, respiratory burns activity, and nitric oxide production, as well as superoxide dismutase and catalase activities. Additionally, J. vernicosa extract increased pro-inflammatory cytokine IL-1ß and suppressed anti-inflammatory IL-10 gene expression upon stimuli and V. parahaemolyticus challenge. Finally, the data confirms that J. vernicosa stem bark extract is non-cytotoxic, rich in bioactive compounds with antioxidant effects, capable of enhancing the immune system in leukocytes and with great potential to fight against opportunistic diseases, such as vibriosis in fish.