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Vibrio alginolyticus is a Gram-negative pathogen of both marine animals and humans, resulting in significant losses for the aquaculture industry. Emerging evidence indicates that V. alginolyticus manipulates cell death for its pathogenicity, but the underlying molecular mechanisms remain unclear. Here, a gene designated vopS in V. alginolyticus HY9901 was identified, which was predicted to encode the T3SS effector protein. To determine whether VopS contributes to the pathogenesis of V. alginolyticus, the ΔvopS mutant strain was constructed and phenotypically characterized. The deletion of VopS not only reduced the ability to secrete extracellular proteases and virulence but also affected the expression of the T3SS genes. Furthermore, VopS was cytotoxic and induced apoptosis, as confirmed by elevated LDH and the activation of caspase-3. Metabolomic analysis revealed considerable metabolomic disruptions upon V. alginolyticus infection. The VopS effector induced host cell ferroptosis by promoting the synthesis of adrenic acid, depleting cellular glutathione, and subsequently increasing the accumulation of ferrous (Fe2+). Taken together, our findings provide that the VopS effector is an essential virulence factor of V. alginolyticus, which can lead to ferroptosis.

Acetylation modification has become one of the most popular topics in protein post-translational modification (PTM) research and plays an important role in bacterial virulence. A previous study indicated that the virulence-associated caseinolytic protease proteolytic subunit (ClpP) is acetylated at the K165 site in Vibrio alginolyticus strain HY9901, but its regulation regarding the virulence of V. alginolyticus is still unknown. We further confirmed that ClpP undergoes lysine acetylation (Kace) modification by immunoprecipitation and Western blot analysis and constructed the complementation strain (C-clpP) and site-directed mutagenesis strains including K165Q and K165R. The K165R strain significantly increased biofilm formation at 36 h of incubation, and K165Q significantly decreased biofilm formation at 24 h of incubation. However, the acetylation modification of ClpP did not affect the extracellular protease (ECPase) activity. In addition, we found that the virulence of K165Q was significantly reduced in zebrafish by in vivo injection. To further study the effect of lysine acetylation on the pathogenicity of V. alginolyticus, GS cells were infected with four strains, namely HY9901, C-clpP, K165Q and K165R. This indicated that the effect of the K165Q strain on cytotoxicity was significantly reduced compared with the wild-type strain, while K165R showed similar levels to the wild-type strain. In summary, the results of this study indicate that the Kace of ClpP is involved in the regulation of the virulence of V. alginolyticus.

Lysine acetylation modification is a dynamic and reversible post-translational modification, which plays an important role in the metabolism and pathogenicity of pathogenic bacteria. Vibrio alginolyticus is a common pathogenic bacterium in aquaculture, and bile salt can trigger the expression of bacterial virulence. However, little is known about the function of lysine acetylation in V. alginolyticus under bile salt stress. In this study, 1,315 acetylated peptides on 689 proteins were identified in V. alginolyticus under bile salt stress by acetyl-lysine antibody enrichment and high-resolution mass spectrometry. Bioinformatics analysis found that the peptides motif ****A*Kac**** and *******Kac****A* were highly conserved, and protein lysine acetylation was involved in regulating various cellular biological processes and maintaining the normal life activities of bacteria, such as ribosome, aminoacyl-tRNA biosynthesis, fatty acid metabolism, two-component system, and bacterial secretion system. Further, 22 acetylated proteins were also found to be related to the virulence of V. alginolyticus under bile salt stress through secretion system, chemotaxis and motility, and adherence. Finally, comparing un-treated and treated with bile salt stress lysine acetylated proteins, it was found that there were 240 overlapping proteins, and found amino sugar and nucleotide sugar metabolism, beta-Lactam resistance, fatty acid degradation, carbon metabolism, and microbial metabolism in diverse environments pathways were significantly enriched in bile salt stress alone. In conclusion, this study is a holistic analysis of lysine acetylation in V. alginolyticus under bile salt stress, especially many virulence factors have also acetylated.

Vibrio alginolyticus, a Gram-negative bacterium, is an opportunistic pathogen of both marine animals and humans, resulting in significant losses in the aquaculture industry. Type III secretion system (T3SS) is a crucial virulence mechanism of V. alginolyticus. In this study, the T3SS regulatory gene exsA, which was cloned from V. alginolyticus wild-type strain HY9901, is 861 bp encoding a protein of 286 amino acids. The ΔexsA was constructed by homologous recombination and Overlap-PCR. Although there was no difference in growth between HY9901 and ΔexsA, the ΔexsA exhibited significantly decreased extracellular protease activity and biofilm formation. Besides, the ΔexsA showed a weakened swarming phenotype and an ~100-fold decrease in virulence to zebrafish. Antibiotic susceptibility testing showed the HY9901ΔexsA was more sensitive to kanamycin, minocycline, tetracycline, gentamicin, doxycycline and neomycin. Compared to HY9901 there were 541 up-regulated genes and 663 down-regulated genes in ΔexsA, screened by transcriptome sequencing. qRT-PCR and β-galactosidase reporter assays were used to analyze the transcription levels of hop gene revealing that exsA gene could facilitate the expression of hop gene. Finally, Danio rerio, vaccinated with ΔexsA through intramuscular injection, induced a relative percent survival (RPS) value of 66.7% after challenging with HY9901 wild type strain. qRT-PCR assays showed that vaccination with ΔexsA increased the expression of immune-related genes, including GATA-1, IL6, IgM, and TNF-α in zebrafish. In summary, these results demonstrate the importance of exsA in V. alginolyticus and provide a basis for further investigations into the virulence and infection mechanism.

Crassostrea hongkongensis (C. hongkongensis) is an important economic shellfish in marine aquaculture, but suffers from severe infection of Vibrio parahaemolyticus (V. Parahemolyticus). The study on immune strategy against Vibrio infection will help to enhance the disease resistance of C. hongkongensis. In this study, after V. parahaemolyticus challenge, the immune enzyme activities and transcriptome were analyzed to explain the immune strategy of C. hongkongensis. The immune enzyme activities analysis showed that all of superoxide dismutase (SOD), catalase (CAT), lysozyme (LZM) activities and malondialdehyde (MDA) content were significantly increased among 3 and 12 h post-infection, indicating that V. parahaemolyticus had stimulated the immune response of C. hongkongensis. Transcriptome analysis showed that a total of 636,295,318 clean readings were generated from the libraries of control and infection groups. Comparing with the control groups (C0h), 147 differentially expressed genes (DEGs) were identified in T3h group, 392 DEGs in T6h group, 47 DEGs in T12h group, and 423 DEGs in T24h group. The time-series expression profiles of DEGs showed that the rising and failing trends were significantly enriched in 2 clusters, including 158 DEGs. Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis showed that immune-related DEGs were annotated into 20 categories, including Toll and Imd signaling pathway, IL-17 signaling pathway, Toll like receptor (TLR) pathway and c-type lectin receptor signaling pathway. Since multiple immune-related DEGs were enriched in TLR signal pathway, qRT-PCR was employed to validate the expression of genes in TLR pathway. The qRT-PCR results showed that all of six detected genes were significantly upregulated, including TLR4, MyD88, IRAK4, TRAF6, IL17 and COX2. The data indicated that TLR4-MyD88 pathway took part in the immune response of C. hongkongensis against Vibrio infection.

Vibrio alginolyticus, a zoonotic bacterial pathogen, expresses a type III secretion system (T3SS) that is critical for pathogen virulence and disease development. In this study, the mutant HY9901 ΔaraC was obtained from the laboratory and its biological characteristics were analyzed. The swimming ability of ΔaraC decreased and exhibited a 2,600 times reduction in virulence to zebrafish. However, ΔaraC showed no difference in growth and extracellular protease activity compared to wild type. Biofilm-forming ability was improved at 24 h, but no difference was observed at other time points. The results of drug sensitivity testing showed that compared with the wild-type HY9901 strain, ΔaraC was sensitive to amikacin, tetracycline, neomycin, minocycline, and gentamicin. The transcription levels of T3SS effector proteins HopPmaJ, VopS, VcrV, and VopN were analyzed by qRT-PCR. The results showed that ΔaraC had significantly upregulated the mRNA expression of VopS, VcrV, and VopN, but significantly downregulated the mRNA expression of HopPmaJ at each stage compared with HY9901. Western blotting and the β-galactosidase reporter gene experiment also showed that the deletion of araC gene significantly downregulated the expression of HopPmaJ. Finally, the relative percent survival (RPS) rate of grouper inoculated by intramuscular (IM) injection of HY9901 ΔaraC was 61.3% after being challenged with HY9901. Real-time qPCR analysis showed that vaccination of HY9901 ΔaraC could enhance the expression of immune-related genes, including gata-1, il6, IgM, il-1β, and lyz in liver and spleen, indicating that ΔaraC applied as a live-attenuated vaccine effectively induced an immune response in the zebrafish. This study provides evidence for the subsequent development of an effective live-attenuated V. alginolyticus vaccine.

Protein lysine acetylation is an evolutionarily conserved post-translational modification (PTM), which is dynamic and reversible, playing a crucial regulatory role in almost every aspect of metabolism, of both eukaryotes and prokaryotes. Several global lysine acetylome studies have been carried out in various bacteria, but thus far, there have been no reports of lysine acetylation for the commercially important aquatic animal pathogen Vibrio mimicus. In the present study, we used anti-Ac-K antibody beads to highly sensitive immune-affinity purification and combined high-resolution LC-MS/MS to perform the first global lysine acetylome analysis in V. mimicus, leading to the identification of 1,097 lysine-acetylated sites on 582 proteins, and more than half (58.4%) of the acetylated proteins had only one site. The analysis of acetylated modified peptide motifs revealed six significantly enriched motifs, namely, KacL, KacR, L(-2) KacL, LKacK, L(-7) EKac, and IEKac. In addition, bioinformatic assessments state clearly that acetylated proteins have a hand in many important biological processes in V. mimicus, such as purine metabolism, ribosome, pyruvate metabolism, glycolysis/gluconeogenesis, the TCA cycle, and so on. Moreover, 13 acetylated proteins were related to the virulence of V. mimicus. To sum up, this is a comprehensive analysis whole situation protein lysine acetylome in V. mimicus and provides an important foundation for in-depth study of the biological function of lysine acetylation in V. mimicus.

Biofloc technology (BFT) is one of the most promising technologies in global aquaculture for the purpose of improving water quality, waste treatment, and disease prevention in intensive aquaculture systems. However, characterization of the microbial species and antibiotic resistance potentially present in biofloc-based aquaculture environments is needed. In this study, we used high-throughput sequencing technology to comprehensively compare the bacterial communities in mariculture ponds of Penaeus monodon (P. monodon), by testing of water, biofloc, and intestine of P. monodon. Operational taxonomic units (OTUs) cluster analysis showed that the nine samples tested divided into 45 phyla and 457 genera. Proteobacteria was the dominant bacteria in water, biofloc and prawn intestine. In biofloc and intestine, the Ruegeria (2.23–6.31%) genus represented the largest proportion of bacteria, with Marivita (14.01–20.94%) the largest group in water. Microbial functional annotation revealed that in all the samples, genes encoding metabolism were predominant. The antibiotic resistance gene annotation showed the highest absolute abundance of patB, adeF, OXA-243, and Brucella_suis_mprF from Proteobacteria. PatB (11.33–15.01%), adeF (15.79–18.16%), OXA-243 (35.65%), and Brucella_suis_mprF (10.03%) showed the highest absolute abundance of antibiotic resistance genes in water, biofloc, and intestines, respectively. These findings may greatly increase our understanding of the characteristics of the microbiota of shrimp biofloc-based aquaculture systems and the complex interactions among shrimp, ambient microflora, and environmental variables. It provides a reference basis for policy on breeding, environmental safety, and maintaining food safety in the production of P. monodon.

Antibiotic resistance among aquatic bacterial pathogens has become a serious concern in aquaculture environments, which has increased research interest to develop solutions to overcome this problem. Moreover, the activities of several Aeromonas hydrophila gene pathways have remained elusive concerning antibiotic resistance evolution. Therefore, in this study, we have performed a transcriptomic analysis to compare differentially expressed genes between oxytetracycline (OXY) susceptible and resistant strains of A. hydrophila. Compared to the A. hydrophila susceptible strain, a total of 22 and 185 genes were differentially expressed in the 4-fold minimal inhibitory concentration (MIC) and 8-fold MIC resistant strains, respectively. Furthermore, the bioinformatics analysis revealed that the sulfur metabolism-related genes were down-regulated. The genes responsible for mannitol metabolism and the efflux pump system were up-regulated in resistant strains, compared to the susceptible strain. Therefore, it suggests that these three pathways may be involved in the OXY resistance evolution in A. hydrophila. The outcome of the transcriptomic data was further validated through quantitative reverse transcription-PCR (qRT-PCR) and Western blot analysis. Overall, the obtained data provides a deeper insight into the intrinsic molecular mechanism of OXY resistance evolution in A. hydrophila.

Recent studies have shown that a key strategy of many pathogens is to use post-translational modification (PTMs) to modulate host factors critical for infection. Lysine succinylation (Ksuc) is a major PTM widespread in prokaryotic and eukaryotic cells, and is associated with the regulation of numerous important cellular processes. Vibrio alginolyticus is a common pathogen that causes serious disease problems in aquaculture. Here we used the affinity enrichment method with LC-MS/MS to report the first identification of 2082 lysine succinylation sites on 671 proteins in V. alginolyticus, and compared this with the lysine acetylation of V. alginolyticus in our previous work. The Ksuc modification of SodB and PEPCK proteins were further validated by Co-immunoprecipitation combined with Western blotting. Bioinformatics analysis showed that the identified lysine succinylated proteins are involved in various biological processes and central metabolism pathways. Moreover, a total of 1,005 (25.4%) succinyl sites on 502 (37.3%) proteins were also found to be acetylated, which indicated that an extensive crosstalk between acetylation and succinylation in V. alginolyticus occurs, especially in three central metabolic pathways: glycolysis/gluconeogenesis, TCA cycle, and pyruvate metabolism. Furthermore, we found at least 50 (7.45%) succinylated virulence factors, including LuxS, Tdh, SodB, PEPCK, ClpP, and the Sec system to play an important role in bacterial virulence. Taken together, this systematic analysis provides a basis for further study on the pathophysiological role of lysine succinylation in V. alginolyticus and provides targets for the development of attenuated vaccines.

Vibrio alginolyticus is a major cause of Vibriosis in farmed marine aquatic animals and has caused large economic losses to the Asian aquaculture industry in recent years. Therefore, it is necessary to control V. alginolyticus effectively. The virulence mechanism of V. alginolyticus, the Type III secretion system (T3SS), is closely related to its pathogenicity. In this study, the T3SS gene tyeA was cloned from V. alginolyticus wild-type strain HY9901 and the results showed that the deduced amino acid sequence of V. alginolyticus tyeA shared 75–83% homology with other Vibrio spp. The mutant strain HY9901ΔtyeA was constructed by Overlap-PCR and homologous recombination techniques. The HY9901ΔtyeA mutant exhibited an attenuated swarming phenotype and an ~40-fold reduction in virulence to zebrafish. However, the HY9901ΔtyeA mutant showed no difference in growth, biofilm formation and ECPase activity. Antibiotic susceptibility test was observed that wild and mutant strains were extremely susceptible to Amikacin, Minocycline, Gentamicin, Cefperazone; and resistant to oxacillin, clindamycin, ceftazidime. In contrast wild strains are sensitive to tetracycline, chloramphenicol, kanamycin, doxycycline, while mutant strains are resistant to them. qRT-PCR was employed to analyze the transcription levels of T3SS-related genes, the results showed that compared with HY9901 wild type, ΔtyeA had increased expression of vscL, vscK, vscO, vopS, vopN, vscN, and hop. Following vaccination with the mutant strain, zebrafish had significantly higher survival than controls following infection with the wild-type HY9901 (71.2% relative percent survival; RPS). Analysis of immune gene expression by qPCR showed that vaccination with HY9901ΔtyeA increased the expression of IgM, IL-1β, IL-6, and TNF-α in zebrafish. This study provides evidence of protective efficacy of a live attenuated vaccine targeting the T3SS of V. alginolyticus which may be facilitated by up-regulated pro-inflammatory and immunoglobulin-related genes.

Aquaculture is a discipline system that focuses on exploring the growth, development, reproduction, aquaculture, and resources of aquatic animals and plants, and their complex relationships. Under the tide of the "high-quality development" strategy, the aquaculture discipline is also facing new opportunities and challenges for transformation, upgrading, and deepening development. Therefore, exploring and practicing an effective path for the high-quality development of graduate education in aquaculture is not only the key to promoting the transformation of graduate education in aquaculture from scale expansion to quality improvement, but also has immeasurable value for implementing the strategy on developing a quality workforce and the strategy of scientific and technological powerhouse. The high-quality development of graduate education in aquaculture can be promoted from the following aspects; optimizing and improving the construction of the graduate education system, focusing on enhancing the quality of high-level talent cultivation, strengthening the overall strength of the graduate supervisor team, actively promoting the adjustment and upgrading of the disciplinary and professional structure, strengthening the construction of resource platforms and deepening the implementation of collaborative education mechanisms, and continuously expanding and deepening the new pattern of international exchange and cooperation. Through the comprehensive promotion of the above paths, the aim is to fully build a model for the improvement and governance of graduate education quality. [ABSTRACT FROM AUTHOR]

[Objectives] This study was conducted to explore the biological functions of cyaA gene of Vibrio alginolyticus. [Methods] With DNA of V. alginolyticus HY 9901 as a template, primers were designed according to the sequence of cyaA gene, and the cyaA gene was amplified by PCR. Bioinformatics analysis was performed. [Results] The cyaA gene of V. alginolyticus HY9901 was 2 529 bp in size, and encoded 842 amino acids. The molecular structure of CyaA protein was C4 358H6 745N1 171O1 286S35. Its theoretical molecular weight was 97. 241 67 kDa and the theoretical pi value was 5. 56. It had no signal peptide and transmembrane domain. CyaA protein had three N-terminal glycosylation sites, one cAMP and cGMP-dependent protein kinase phosphorylation site, nine protein kinase C phospho-rylation sites, nine casein kinase II phosphorylation sites, one tyrosine kinase phosphorylation site, seven N-terminal myristoylation sites, one pentenyl binding site and ten microbody C-terminal localization signal sites. Subcellular localization prediction showed that CyaA protein was mainly located in the nucleus and cytoplasm. Through multi-sequence alignment and phylogenetic tree construction, it was concluded that V. alginolyticus had high CyaA homology with other Vibrio species. cyaA of V. alginolyticus was clustered with Vibrio fluminensis and Vibrio marinisedimini, and they were closely related. The secondary structure of CyaA protein consisted of a-helixes (43.11%), random coils (38.00%) and extended strands (14.49%). In protein network interaction, it was found that the proteins adjacent to CyaA protein were Crp-2, CpdA, Crr, PtsG-2, ANP67209.1, Crp-1, PykF, Pyk, RelA and Ndk. [Conclusions] This study provides a new idea for formulating strategies for the prevention and control of vibriosis. [ABSTRACT FROM AUTHOR]

[Objectives] This study was conducted to understand the structure and function of MsrA protein. [Methods] With Vibrio alginolyticus HY9901 as the object of study, primers were designed to amplify the full-length gene of msrA, and its bioinformatics analysis was carried out. [Results] The full length of msrA gene was 639 bp, encoding 212 amino acids, and its theoretical molecular weight was about 23 729.60 Da. The protein had a stable structure, and it was hydropho-bic overall. The structure of signal peptides at the N terminal of the amino acid sequence was predicted, and it was found that there was no signal peptide cleavage site and no transmembrane region. The amino acid sequence of MsrA contained multiple signal binding sites. Protein subcellular localization showed that MsrA protein was most likely located in the cytoplasm. Homology analysis showed that MsrA of V. alginolyticus had high homology with other Vibrio species, and the highest homology with V. alginolyticus. In the prediction of functional domains, MsrA had the function of methionine sulfoxide reduction. In secondary structure prediction, MsrA contained random coils at a proportion of 46.70%, which was the highest. The similarity between the tertiary structure model of MsrA and template Q87SW6. 1. A was 89. 15%. PTM analysis showed that MsrA protein had many PTM modification sites such as phosphorylation and glycosylation sites. [Conclusions] This study provides some reference value for further study on the role of MsrA in bacterial antioxidant stress. [ABSTRACT FROM AUTHOR]

PhoR is a histidine kinase in a two-component regulatory system that regulates phosphorus metabolic pathways and undertakes the key mission of information transmission in pathogenic bacteria. The full-length phoR gene was successfully cloned from the Vibrio alginolyticus HY9901 strain. A comprehensive analysis of the cloned gene was conducted using bioinformatics. Sequence analysis revealed that the total length of the phoR gene (GenBank accession No.: KJ958404.1) is 1299 bp, with the coding region containing a total of 432 amino acid residues. The phylogenetic tree of PhoR revealed that it belongs to the same subclade as V. diabolicus. The SMART program was employed for the purpose of functional domain prediction, which revealed that PhoR possesses three major functional domains: PAS (amino acids 98 -166), His-KA (amino acids 205-272), and HATPase_c (amino acids 317429). [ABSTRACT FROM AUTHOR]

Molecular Biology and Experiment is considered fundamental for graduate students specializing in aquaculture at Guangdong Ocean University. This discipline focuses on the examination of the structure and function of macromolecules, including proteins and nucleic acids. Moreover, it elucidates biological phenomena and principles at the molecular level, making it an essential foundational course for students pursuing various biology majors. As a foundational course for the basic application of aquaculture, Molecular Biology and Experiment requires guidance through numerous examples and cases. However, there are several challenges to address in developing the case library. Consequently, a case library has been established to meet the course requirements of Molecular Biology and Experiment for modern graduate students, with the central goal of reforming the educational model of higher education institutions and enhancing the effectiveness and quality of talent development. This strategy is designed to nurture highly skilled professionals who can address the current needs of the industry. [ABSTRACT FROM AUTHOR]

[Objectives] To explore the function of hep gene in Aeromonas hydrophila. [Methods] A pair of specific primers was designed referring to the hep gene sequence of A hydrophila. The hep gene was amplified by PCR, and performed bioinformatics analysis. [Results] The hep gene had a total length of 1 650 bp and encoded 549 amino acids. The theoretical molecular weight of the protein predicted was about 59 476.44 kDa. After predicting the N-terminal signal peptide structure of the amino acid sequence, neither obvious signal peptide cleavage site nor signal peptide was found, and the protein had no transmembrane region. The amino acid sequence had a N-glycosylation site, 4 protein kinase C phosphorylation sites, 7 casein kinase II phosphorylation sites, 9 N-myristoylation sites, 4 isoprene binding sites, 10 microbody C-terminal target signal sites, and an ATP/GTP binding site motif A (P-ring). The amino acid sequence of hep gene of A hydrophila was performed homology analysis with other Aeromonas strains, and it showed higher homology with A. veronii. In the secondary structure, the ct-he-lix, [3-sheet, random coil and extended strand accounted for 45.36%,6.01 %, 37.52% and 11. 11%, respectively. The tertiary structure model consisted of 18 a-helix and 22 [3-sheet. Analysis of protein-protein network interaction demonstrated that the proteins interacting with Hep protein were AHA_3407, nrfA, nirB-1, nirB-2 and AHA_1112. [Conclusions] Through the bioinformatics prediction results, the basic information of hep gene of A. hydrophila is preliminarily understood, and the possible function of this protein is predicted, in order to provide guidance for subsequent vaccine research. [ABSTRACT FROM AUTHOR]

[Objectives] The paper was to clone and analyze bioinformatics of ndk gene from Vibrio alginolyticus. [Methods] A pair of specific primers was designed based on the ndk gene sequence of V. alginolyticus HY9901. The full length of ndk gene was amplified by PCR and bio informatics analysis was performed. MEGA 5.0 software was used to construct NDK phylogenetic tree by neighbor-joining method. SWISS-MODEL program was used to obtain the three-dimensional structural model of single subunit from NDK protein. [Results] The ndk gene, molecular structural formula C7CB Hlow N]92 02J4 S7, was 426 bp in total, encoding 141 amino acids, with the theoretical molecular weight of 15.871 99 kD and the theoretical pl value of 5.13. The prediction results of protein subcellular localization, Signal? 5. 0, TMHMM Server 2.0 and SoftBerry-Psite showed that NDK mainly existed in the cytoplasm, and the protein was unstable and hydrophobic. There was neither signal peptide cleavage site, nor transmembrane region and KEGG metabolic pathway. The amino acid sequence had two protein kinase C phosphorylation sites, a casein kinase II phosphorylation site, a N-myristoylation site, three microbody C-terminal target signal sites, and a nucleoside diphosphate kinase active site. Homology analysis showed that the NDK of V. alginolyticus had high homology with that of V. diabolicus, with a similarity of 98.58%. Analysis of the structural functional domain revealed that the protein had one NDK structural functional domain. The prediction results of secondary structure showed that the a-helix, random coil, [3-sheet and extended strand accounted for 53.19%, 28.37%, 7.09% and 11.35%, respectively. Analysis of NDK protein via STRING database demonstrated that the proteins interacting with NDK protein were NrdA, NrdB, GmK, CmK, TmK, PyrG, PyrH, RelA, FolE and SpoT. [Conclusions] The study plays a positive role in the prevention and control of vibriosis and the improvement of the current aquaculture environment. [ABSTRACT FROM AUTHOR]

[Objectives] To clone and analyze the vscB gene of Vibrio alginolyticus HY9901 by bioinformatics. [Methods] A pair of specific primers were designed according to the vscB gene sequence of Vibrio alginolyticus HY9901. The full length of the primers was cloned by PCR and analyzed by bioinformatics. [Results] The vscB gene was 429 bp long, encoding 142 amino acids, with a theoretical molecular weight of 16.4 kDa and a pl value of 5.48. Amino acid sequence analysis of VscB showed that VscB was not a secretory protein, without signal peptide and transmembrane region, and there were protein kinase C phosphorylation site and casein kinase II phosphorylation site in the sequence. Homologous comparison of amino acid sequences showed that VscB of V. alginolyticus had the highest protein similarity with Vibrio Parahaemolyticus, reaching 91%. Phylogenetic tree analysis showed that the corresponding proteins of V. alginolyticus VscB, Vibrio Parahaemolyticus and Vibrio diabolicus were clustered in the same subfamily. Functional domain analysis showed that it had CesT family domain. Tertiary structure prediction showed that there were 3 a-helices and 5 ß-turns in VscB protein. [Conclusions] This study provided a theoretical basis for further study on the function of chaperone of V. alginolyticus. [ABSTRACT FROM AUTHOR]

[Objectives] To amplify the h-ns gene of Vibrio alginolyticus and analyze it by bioinformatics. [Methods] According to the h-ns gene sequence of V. alginolyticus HY9901, a pair of specific primers were designed and amplified by PCR. [Results] The h-ns gene was 408 bp in length and 135 amino acids were encoded. The predicted theoretical protein molecular weight was about 14. 98 kD, and the isoelectric point was 4.99. Protein subcellular localization, SignalP 5. 0, TMHMM Seiver 2.0 and SoftBerry-Psite predictions showed that H-NS was located outside the cell membrane, and the protein was unstable and hydrophobic. There was no signal peptide cleavage site, no transmembrane region and no KEGG metabolic pathway. The amino acid sequence contained three phosphorylation sites, one N-terminal myristoylation site and three microsomal C-terminal target signal sites. Using MEGA 5.0, H-NS phylogenetic tree was constructed by ortho-connection method. The results showed that H-NS of V. alginolyticus was closer to H-NS of Vibrio diabolicus. Using SWISS-MODEL, the three-dimensional structure model of H-NS subunit was simulated, which was similar to the crystal structure of Salmonella typhimurium H-NS1-83. [Conclusions] This study lays a foundation for exploring the regulation mechanism of V. alginolyticus H-NS protein on bacterial virulence in the future. [ABSTRACT FROM AUTHOR]

[Objectives] To study the effects of microplastics on antibiotic resistance genes and virulence genes of Vibrio alginolyticus, so as to provide a certain reference for controlling marine pollution, curbing the spread of environmental antibiotic resistance genes and virulence genes, formulating environmental policies, and maintaining food safety. [Methods] After adding V. alginolyticus into the artificial seawater, they were divided into three groups, namely blank control group (BLK), polyvinyl chloride microplastic group (PVC group) and polyvinyl alcohol microplastic group (PVA group). Aerated culture experiments were carried out, and the effects of microplastics on the expression of resistance genes and virulence genes of V. alginolyticus were studied by PCR and qPCR methods. [Results] The presence of microplastics significantly changed the resistance gene structure of V. alginolyticus. Compared with the control group, the cfxA and cfr resistance genes were detected in the microplastic group. However, only PVC group detected blaZ resistance gene, and only PVA group did not detect aaC resistance gene. In addition, compared with the control group, the expressions of virulence genes in the microplastic group were all down-regulated (P <0.01). [Condusions] This study provides some reference for curbing the spread of environmental antibiotic resistance genes and virulence genes, formulating environmental policies, and maintaining food safety, but the specific mechanisms of drug resistance and virulence need further research. [ABSTRACT FROM AUTHOR]

This paper intends to combine the development trend of the subject, restructure and optimize the course content, and construct the sendee learning model of Pathogenic Biology of Aquatic Animals for postgraduates. At the same time, through service practice, it is needed to further consolidate curriculum knowledge and skills, stimulate the learning initiative and enthusiasm of postgraduates, expand professional knowledge, improve professional quality, and lay a solid foundation for serving the national rural revitalization strategy in the future. [ABSTRACT FROM AUTHOR]

[Objectives] To clone and analyze the TpiA gene of Vibrio alginolyticus HY9901. [Methods] According to the TpiA gene sequence of V. alginolyticus, a pair of specific primers was designed, and its full length was amplified by PCR. [Results] The full length of TpiA gene is 771 bp, encoding 256 amino acid residues in total, and the NCBI accession number is OM906798. According to the deduced amino acid sequence, its molecular weight was predicted to be about 26. 975 48 kDa, and its isoelectric point was 4.78. The amino acid sequence of the N-terminal signal peptide structure was predicted, and it was found that there was no obvious signal peptide cleavage site, no signal pep-tide, and no transmembrane region; the amino acid sequence contained 3 N-glycosylation sites, 4 protein kinase C phosphorylation sites, 2 casein kinase II phosphorylation sites, 6 N-myristoylation sites, 7 microbody C-terminal target signal site, and 1 triose phosphate isomerase active site. The prediction results of protein subcellular localization showed that TpiA may be located in mitochondria or cytoplasm, with probability of 39.1 % and 34.8%, respectively. The amino acid sequence of the TpiA gene of V. alginolyticus shared 98.83% - 99.61 % homology with other Vibrio species, and it was clustered into the same subfamily with Vibrio parahaemolyticus and had a close relationship. In the secondary structure prediction, the proportions of α-helix, random coil and extended chain were 44.53%, 41.41% and 14. 06%, respectively, and the similarity of its tertiary structure model to template lawl. 1. A was 85.16%. [Conclusions] This study is intended to provide a basis for further research on the role of TpiA gene in the type III secretion system and related research on antibiotic resistance. [ABSTRACT FROM AUTHOR]

[Objectives] The crp gene in Vibrio alginolyticus was studied. [Methods]The crp gene of V. alginolyticus HY9901 was cloned and analyzed by bioinformatics. [Results] The ORF of the crp gene is 633 bp long and the predicted amino acid sequence encompasses 210 amino acid residues. The physicochemical property analysis indicated that the chemical formula of CRP is C1051 H1704N290 O308S10 with a molecular weight of 23. 651 4 kDa, and its theoretical pl is 7. 74. Besides, the protein is stable and hydrophilic. The protein had three protein kinase C phosphorylation site, three casein kinase II phosphorylation site, one N-terminal myristoylation site. The BLAST analysis on the sequence revealed high homology with CRPs in other Vibrio species, and particularly the sequence shares about a homology of 99. 52 % with the CRP in V. parahaemolyticus. The SWlSS-MODEL software simulated the subunit tertiary structural model of the CRP, and the similarity with template 3hif. 1. A was 95. 71 % . [Conclusions ] This study provides a reference for the search for efficient protective antigens against vibrosis. [ABSTRACT FROM AUTHOR]

[Objectives] To amplify the DNA-binding response regulator PhoP in Vibrio alginolyticus and analyze its sequence characteristics and subunit structure. [Methods] According to the sequence of the DNA-binding response regulator PhoP in V. alginolyticus, a pair of specific primers was designed for PCR amplification, and the bioinformatics of the sequence amplified was analyzed. Using MEGA 5.0 software, the phoP phylogenetic tree was constructed by the neighbor-joining method. Using SWISS-MODEL software, the three-dimensional structural model of the PhoP subunit was simulated. [Results] Thefull-IengthphoP gene was732 bp, encoding a total of 243 amino acids. The predicted theoretical molecular weight of the protein is about 27.67 kD, and the isoelectric point is 5.09. The prediction results of protein subcellular localization, SignalP4.0, TMHMM Server 2.0 and SoftBeny-Psite show that PhoP is located in the cytoplasm, and is stable and hydrophobic; there is a signal peptide cleavage site between amino acids 29 and 30, and there is no transmembrane region. The amino acid sequence contains one Asn-glycosylation site, one protein kinase C phosphorylation site, seven casein kinase II phosphorylation sites, one tyrosine kinase phosphorylation site, three myristoylation sites, and seven C-terminal microbody targeting signal sites. The PhoP of V. alginolyticus has high homology with that of Vibrio campbellii. The PhoP subunit of V. alginolyticus has similar configuration to the single-subunit RegX3 protein of Mycobacterium tuberculosis. [Conclusions] This study has a positive effect on the prevention and control of vibriosis and the improvement of the current aquatic economic animal breeding environment. [ABSTRACT FROM AUTHOR]

Aquatic Animal Bacteriology is a basic course of Aquatic Animal Medicine discipline. Bacteriology is not only a frontier subject in the field of life sciences, but also one of the most rapidly developing scientific fields today. The Belt and Road Initiative has brought opportunities and challenges to the upgrading and development of this discipline, so this discipline faces many problems that need to be solved. In view of this, this study combined the development trend of the discipline, reorganized and optimized the course content, and prepared sophisticated multimedia courseware, to stimulate students' learning initiative and enthusiasm, expand the field of professional knowledge, improve their English translation and writing skills, so as to lay a solid foundation for their future sendee to the Belt and Road Initiative. [ABSTRACT FROM AUTHOR]

By making use of the technical advantages of Guangzhou Huibiao Testing Technology Center in the field of testing, the project analyzes the characteristics of undergraduate professional education in aquatic animal medicine, and intends to build a case library of Inspection and Quarantine of Aquatic Animal suitable for modern undergraduate teaching needs, with training applied talents to meet the requirements of the times as the goal, and strengthening the reform of college education mode and improving the quantity and quality of talent training as the guiding ideology. [ABSTRACT FROM AUTHOR]

[Objective] To clone araC gene of Vibrio alginolyticus HY9901 strain, and analyze bioinformatics. [Methods] the whole genome sequence of Vibrio alginolyticus on GenBank was used to design specific primers. According to the principle of PCR amplification sequence, the target gene araC was amplified, and then the sequence was further analyzed by bioinformatics method to establish the phylogenetic tree of araC gene and its corresponding subunit three-dimensional structure model. Results] Sequence analysis revealed araC gene is 711 bp and encodes. a putative protein of 236 amino acids. The predicted molecular mass of AraC was 26. 92 ku. Using Signal P 4.0 and TMHMM Server 2.0 soft ware for analysis, it was predicted that the AraC protein did not contain a signal peptide or a transmembranous region. The AraC protein had two cAMP and CGMP dependent protein kinase phosphorylation site, five protein kinase C phosphorylation sites, three casein kinase Il phos phorylation sites, one prenyl group binding site (CAAX box) and five microbodies C-terminal targeting signal. The predicted results of protein subcellular localization showed that AraC was located in the mitochondria, nucleus and cytoplasm. Its protein is unstable and hydrophilic. The AraC protein is a transcriptional regulatory protein which belongs to HTH 18 superfamily. According to the prediction, secondary structure: a-helix (Alpha helix) accounted for 52. 12%, random coil (31. 78%), extended strand (11.02%), b-fold (Beta turn) accounted for 5.08%. V. alginolyticus, Vibrio parahaemolyticus and Vibrio palustris were clustered together, which implies that the genetic relationship be tween these three species was the closest. [ABSTRACT FROM AUTHOR]