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PeerJ 2024is the leading cause of bacterial seafood-borne gastroenteritis in humans worldwide. To ensure seafood safety and to minimize the occurrence of seafood-borne diseases,...
BACKGROUND
is the leading cause of bacterial seafood-borne gastroenteritis in humans worldwide. To ensure seafood safety and to minimize the occurrence of seafood-borne diseases, early detection of total (pathogenic and non-pathogenic strains) and pathogenic ( and/or and/or ) is required. This study further improved a loop-mediated isothermal amplification (LAMP) assay using xylenol orange (XO), a pH sensitive dye, to transform conventional LAMP into a one-step colorimetric assay giving visible results to the naked eye. LAMP-XO targeted for species specificity and , and for pathogenic strains. Multiple hybrid inner primers (MHP) of LAMP primers for detection to complement the main primer set previously reported were designed by our group to maximize sensitivity and speed.
METHODS
Following the standard LAMP protocol, LAMP reaction temperature for , , , and detection was first determined using a turbidimeter. The acquired optimal temperature was subjected to optimize six parameters including dNTP mix, betaine, MgSO, 2.0 WarmStart DNA polymerase, reaction time and XO dye. The last parameter was done using a heat block. The color change of the LAMP-XO result from purple (negative) to yellow (positive) was monitored visually. The detection limits (DLs) of LAMP-XO using a 10-fold serial dilution of gDNA and spiked seafood samples were determined and compared with standard LAMP, PCR, and quantitative PCR (qPCR) assays. Subsequently, the LAMP-XO assay was validated with 102 raw seafood samples and the results were compared with PCR and qPCR assays.
RESULTS
Under optimal conditions (65 °C for 75 min), -LAMP-XO and -LAMP-XO showed detection sensitivity at 10 copies of gDNA/reaction, or 10 folds greater than -LAMP-XO and -LAMP-XO. This level of sensitivity was similar to that of standard LAMP, comparable to that of the gold standard qPCR, and 10-100 times higher than that of PCR. In spiked samples, -LAMP-XO, -LAMP-XO, and -LAMP-XO could detect at 1 CFU/2.5 g spiked shrimp. Of 102 seafood samples, LAMP-XO was significantly more sensitive than PCR ( < 0.05) for and detection and not significantly different from qPCR for all genes determined. The reliability of -LAMP-XO and -LAMP-XO to detect pathogenic was at 94.4% and 100%, respectively.
CONCLUSIONS
To detect total and pathogenic , at least -LAMP-XO and -LAMP-XO should be used, as both showed 100% sensitivity, specificity, and accuracy. With short turnaround time, ease, and reliability, LAMP-XO serves as a better alternative to PCR and qPCR for routine detection of in seafood. The concept of using a one-step LAMP-XO and MHP-LAMP to enhance efficiency of diagnostic performance of LAMP-based assays can be generally applied for detecting any gene of interest.
Topics: Humans; Colorimetry; Vibrio parahaemolyticus; Reproducibility of Results; Gastroenteritis
PubMed: 38188160
DOI: 10.7717/peerj.16422 -
Microbiology Spectrum Aug 2023Bacteria belonging to the genus include many known and emerging pathogens. Horizontal gene transfer of pathogenicity islands is a major contributor to the emergence of...
Bacteria belonging to the genus include many known and emerging pathogens. Horizontal gene transfer of pathogenicity islands is a major contributor to the emergence of new pathogenic strains. Here, we use the brine shrimp as a model and show that the marine bacterium Vibrio proteolyticus uses a horizontally shared type VI secretion system, T6SS3, to intoxicate a eukaryotic host. Two T6SS3 effectors, which were previously shown to induce inflammasome-mediated pyroptotic cell death in mammalian phagocytic cells, contribute to this toxicity. Furthermore, we find a novel T6SS3 effector that also contributes to the lethality mediated by this system against . Therefore, our results reveal a T6SS that is shared among diverse vibrios and mediates host lethality, indicating that it can lead to the emergence of new pathogenic strains. The rise in sea surface temperature has been linked to the spread of bacteria belonging to the genus and the human illnesses associated with them. Since vibrios often share virulence traits horizontally, a better understanding of their virulence potential and determinants can prepare us for new emerging pathogens. In this work, we showed that a toxin delivery system found in various vibrios mediates lethality in an aquatic animal. Taken together with previous reports showing that the same system induces inflammasome-mediated cell death in mammalian phagocytic cells, our findings suggest that this delivery system and its associated toxins may contribute to the emergence of pathogenic strains.
Topics: Animals; Humans; Inflammasomes; Vibrio; Eukaryota; Virulence; Phagocytes; Mammals
PubMed: 37310285
DOI: 10.1128/spectrum.01093-23 -
Frontiers in Cellular and Infection... 2023To evaluate the diagnostic value of metagenomic next-generation sequencing (mNGS) in () infection.
OBJECTIVE
To evaluate the diagnostic value of metagenomic next-generation sequencing (mNGS) in () infection.
METHODS
A retrospective analysis of patients with infection at the Fifth Affiliated Hospital of Sun Yat-Sen University from January 1, 2020 to April 23, 2023 was conducted. 14 enrolled patients were diagnosed by culture or mNGS. The corresponding medical records were reviewed, and the clinical data analyzed included demographics, epidemiology laboratory findings, physical examination, symptoms at presentation, antibiotic and surgical treatment, and outcome.
RESULTS
In this study, 78.6% (11/14) patients had a history of marine trauma (including fish stab, shrimp stab, crab splints and fish hook wounds), 7.1% (1/14) had eaten seafood, and the remaining 14.3% (2/14) had no definite cause. Isolation of from clinical samples including blood, tissue, fester and secreta. 9 cases were positive for culture, 5 cases were detected synchronously by mNGS and got positive for . 85.7% (12/14) cases accepted surgical treatment, with 1 patient suffering finger amputated. 14 enrolled patients received appropriate antibiotic therapy, and all of them had recovered and discharged. 9 strains isolated in this study were sensitive to most beta-lactam antibiotics, aminoglycosides, quinolones, etc.
CONCLUSION
infection is a common water-exposed disease in Zhuhai, which requires identification of a number of pathogens. Of severe infections with unknown pathogen, mNGS can be used simultaneously, and the potential to detect multiple pathogens is of great help in guiding treatment.
Topics: Animals; Humans; Retrospective Studies; Vibrio Infections; Vibrio vulnificus; Anti-Bacterial Agents; High-Throughput Nucleotide Sequencing
PubMed: 38035326
DOI: 10.3389/fcimb.2023.1210919 -
Cell Reports Nov 2023The losses caused by Vibrio infections in the aquaculture industry are challenging to quantify. In the face of antibiotic resistance, a natural and environmentally...
The losses caused by Vibrio infections in the aquaculture industry are challenging to quantify. In the face of antibiotic resistance, a natural and environmentally friendly alternative is urgently needed. In this study, we identify E3 ubiquitin-protein ligase RNF103 (rnf103) as a crucial target involved in immune evasion by Vibrio anguillarum. Our research demonstrates that Rnf103 promotes immune escape by inhibiting Traf6. Interestingly, we discover a circular RNA (circRNA), circRnf103, formed by reverse splicing of the Rnf103 gene. Predictive analysis and experimentation reveal that circRnf103 encodes Rnf103-177aa, a protein that competes with Rnf103 and binds to Traf6, preventing its degradation. Notably, circRnf103 therapy induces Rnf103-177aa protein production in zebrafish. In zebrafish models, circRnf103 exhibits significant effectiveness in treating V. anguillarum infections, reducing organ burden. These findings highlight the potential of circRNA therapy as a natural and innovative approach to combat infectious diseases sustainably, particularly in aquaculture and environmental management.
Topics: Animals; RNA, Circular; Zebrafish; TNF Receptor-Associated Factor 6; Vibrio Infections; Vibrio; Fish Diseases
PubMed: 37874674
DOI: 10.1016/j.celrep.2023.113314 -
Microbiome Oct 2023Shrimp cultured in a biofloc system (BFS) have a lower disease incidence than those farmed in a water exchange system (WES). Although a number of studies have reported...
BACKGROUND
Shrimp cultured in a biofloc system (BFS) have a lower disease incidence than those farmed in a water exchange system (WES). Although a number of studies have reported that the gut bacterial community induced by BFS is highly associated with shrimp disease resistance, the causal relationship remains unknown. Here, the promotive roles of gut bacterial community induced by BFS in pathogenic Vibrio infection resistance and its potential micro-ecological and physiological mechanisms were investigated by gut bacterial consortium transplantation and synthetic community (SynCom) construction.
RESULTS
The BFS induced a more stable and resistant gut bacterial community, and significantly enriched some beneficial bacterial taxa, such as Paracoccus, Ruegeria, Microbacterium, Demequina, and Tenacibaculum. Transplantation of a gut bacterial consortium from BFS shrimp (Enrich) greatly enhanced the stability of the bacterial community and resistance against pathogenic V. parahaemolyticus infection in WES shrimp, while transplantation of a gut bacterial consortium from WES shrimp significantly disrupted the bacterial community and increased pathogen susceptibility in both WES and BFS shrimp. The addition of Enrich in shrimp postlarvae also improved the pathogen resistance through increasing the relative abundances of beneficial bacterial taxa and stability of bacterial community. The corresponding strains of five beneficial bacterial taxa enriched in BFS shrimp were isolated to construct a SynCom. The addition of SynCom could not only suppress disease development, but also improve shrimp growth, boost the digestive and immune activities, and restore health in diseased shrimp. Furthermore, the strains of SynCom well colonized shrimp gut to maintain a high stability of bacterial community.
CONCLUSIONS
Our study reveals an important role for native microbiota in protecting shrimp from bacterial pathogens and provides a micro-ecological regulation strategy towards the development of probiotics to ameliorate aquatic animal diseases. Video Abstract.
Topics: Animals; Vibrio parahaemolyticus; Gastrointestinal Microbiome; Penaeidae; Bacteria; Vibrio Infections; Aquaculture
PubMed: 37858205
DOI: 10.1186/s40168-023-01663-2 -
Scientific Reports Apr 2024Food-related illnesses have become a growing public concern due to their considerable socioeconomic and medical impacts. Vibrio parahaemolyticus and Staphylococcus...
Food-related illnesses have become a growing public concern due to their considerable socioeconomic and medical impacts. Vibrio parahaemolyticus and Staphylococcus aureus have been implicated as causative organisms of food-related infections and poisoning, and both can form biofilms which confer antibiotic resistance. Hence, the need for continuous search for compounds with antibiofilm and antivirulence properties. In this study, 22 iodinated hydrocarbons were screened for their antibiofilm activity, and of these, iodopropynyl butylcarbamate (IPBC) was found to effectively control biofilm formation of both pathogens with a MIC of 50 µg/mL which was bactericidal to V. parahaemolyticus and S. aureus. Microscopic studies confirmed IPBC inhibits biofilm formation of both bacteria and also disrupted their mixed biofilm formation. Furthermore, IPBC suppressed virulence activities such as motility and hemolytic activity of V. parahaemolyticus and the cell surface hydrophobicity of S. aureus. It exhibited a preservative potential against both pathogens in a shrimp model. IPBC disrupted the cell membrane of S. aureus and V. parahaemolyticus and differentially affected gene expressions related to biofilm formation and virulence. Additionally, it displayed broad-spectrum antibiofilm activities against other clinically relevant pathogens. These findings indicate IPBC offers a potential means of controlling infections mediated by Vibrio and Staphylococcus biofilms.
Topics: Biofilms; Vibrio parahaemolyticus; Staphylococcus aureus; Anti-Bacterial Agents; Microbial Sensitivity Tests; Animals; Virulence
PubMed: 38644387
DOI: 10.1038/s41598-024-55479-7 -
BMC Veterinary Research Dec 2023Parasitic and bacterial co-infections have been associated with increasing fish mortalities and severe economic losses in aquaculture through the past three decades. The...
Oxidative stress, gene expression and histopathology of cultured gilthead sea bream (Sparus aurata) naturally co-infected with Ergasilus sieboldi and Vibrio alginolyticus.
BACKGROUND
Parasitic and bacterial co-infections have been associated with increasing fish mortalities and severe economic losses in aquaculture through the past three decades. The aim of this study was to evaluate the oxidative stress, histopathology, and immune gene expression profile of gilthead sea bream (Sparus aurata) co-infected with Ergasilus sieboldi and Vibrio alginolyticus.
RESULTS
Vibrio alginolyticus and Ergasilus sieboldi were identified using 16 S rRNA and 28 S rRNA sequencing, respectively. The collagenase virulence gene was found in all Vibrio alginolyticus isolates, and the multiple antimicrobial resistance index ranged from 0.286 to 0.857. Oxidant-antioxidant parameters in the gills, skin, and muscles of naturally infected fish revealed increased lipid peroxidation levels and a decrease in catalase and glutathione antioxidant activities. Moreover, naturally co-infected gilthead sea bream exhibited substantial up-regulation of il-1β, tnf-α, and cyp1a1. Ergasilus sieboldi encircled gill lamellae with its second antennae, exhibited severe gill architectural deformation with extensive eosinophilic granular cell infiltration. Vibrio alginolyticus infection caused skin and muscle necrosis in gilthead sea bream.
CONCLUSION
This study described some details about the gill, skin and muscle tissue defense mechanisms of gilthead sea bream against Ergasilus sieboldi and Vibrio alginolyticus co-infections. The prevalence of co-infections was 100%, and no resistant fish were detected. These co-infections imbalance the health status of the fish by hampering the oxidant-antioxidant mechanisms and proinflammatory/inflammatory immune genes to a more detrimental side. Our results suggest that simultaneous screening for bacterial and parasitic pathogens should be considered.
Topics: Animals; Sea Bream; Vibrio alginolyticus; Antioxidants; Coinfection; Vibrio Infections; Gene Expression; Oxidative Stress; Oxidants; Fish Diseases
PubMed: 38104092
DOI: 10.1186/s12917-023-03840-9 -
Microbial Genomics Dec 2023This study presents the assembly and comparative genomic analysis of luminous strains isolated from the light organs of 12 fish species using Oxford Nanopore...
This study presents the assembly and comparative genomic analysis of luminous strains isolated from the light organs of 12 fish species using Oxford Nanopore Technologies (ONT) sequencing. The majority of assemblies achieved chromosome-level continuity, consisting of one large (>3 Mbp) and one small (~1.5 Mbp) contig, with near complete BUSCO scores along with varying plasmid sequences. Leveraging this dataset, this study significantly expanded the available genomes for and its subspecies , enabling a comparative genomic analysis between the two lineages. An analysis of the large and small chromosomes unveiled distinct patterns of core and accessory genes, with a larger fraction of the core genes residing on the large chromosome, supporting the hypothesis of secondary chromosome evolution from megaplasmids in Vibrionaceae. In addition, we discovered a proposed new species, sp. nov., isolated from an acropomatid host, with an average nucleotide identify (ANI) of 93 % compared to the and strains. A comparison of the and lineages revealed minimal differences in gene content, yet highlighted the former's larger genome size and potential for horizontal gene transfer. An investigation of the operon, responsible for light production, indicated congruence between the presence of and host family, challenging its role in differentiating from . Further insights were derived from the identification of metabolic differences, such as the presence of the NADH:quinone oxidoreductase respiratory complex I in as well as variations in the type II secretion system (T2S) genes between the lineages, potentially impacting protein secretion and symbiosis. In summary, this study advances our understanding of genome evolution, highlighting subtle differences between closely related lineages, specifically and . These findings highlight the benefit of long read sequencing for bacterial genome assembly and pangenome analysis and provide a foundation for exploring early bacterial speciation processes of these facultative light organ symbionts.
Topics: Animals; Photobacterium; Symbiosis; DNA, Bacterial; Genomics; Genome, Bacterial
PubMed: 38112751
DOI: 10.1099/mgen.0.001161 -
Proceedings of the National Academy of... May 2024Several microbial genomes lack textbook-defined essential genes. If an essential gene is absent from a genome, then an evolutionarily independent gene of unknown...
Several microbial genomes lack textbook-defined essential genes. If an essential gene is absent from a genome, then an evolutionarily independent gene of unknown function complements its function. Here, we identified frequent nonhomologous replacement of an essential component of DNA replication initiation, a replicative helicase loader gene, in . Our analysis of genomes revealed two genes with unknown function, named and , that were substantially enriched in genomes without the known helicase-loader genes. These genes showed no sequence similarities to genes with known function but encoded proteins structurally similar with a viral helicase loader. Analyses of genomic syntenies and coevolution with helicase genes suggested that encodes a helicase loader. The in vitro assay showed that VdhL1 and VdhL2 promote the helicase activity of DnaB. Furthermore, molecular phylogenetics suggested that / were derived from phages and replaced an intrinsic helicase loader gene of over 20 times. This high replacement frequency implies the host's advantage in acquiring a viral helicase loader gene.
Topics: Vibrionaceae; DNA Helicases; DNA Replication; Phylogeny; Bacterial Proteins; Viral Proteins; Bacteriophages; Evolution, Molecular; Genome, Bacterial; DnaB Helicases; Vibrio
PubMed: 38683976
DOI: 10.1073/pnas.2317954121 -
Microorganisms Nov 2023Bacterial communities associated with fish larvae are highly influenced by the microbiota of live prey used as feed (rotifers or ), generally dominated by bacterial...
Bacterial communities associated with fish larvae are highly influenced by the microbiota of live prey used as feed (rotifers or ), generally dominated by bacterial strains with a low degree of specialization and high growth rates, (e.g., ), which can be detrimental to larvae. Co-cultivation of microalgae used in the enrichment of (e.g., , or ) with -antagonistic probiotics belonging to the clade bacteria (e.g., spp. or spp.) was studied. The introduction of the probiotics did not affect microalgae growth or significantly modify the composition of bacterial communities associated with both microalgae, as revealed by DGGE analysis. The inoculation of with ALR6 allowed the maintenance of the probiotic in the scale-up of the microalgae cultures, both in axenic and non-axenic conditions. Using -inoculated cultures in the enrichment of reduced the total count in by 2 Log units, therefore preventing the introduction of opportunistic or pathogenic bacteria to fish larvae fed with them.
PubMed: 38004727
DOI: 10.3390/microorganisms11112715