Mycobacteriophage MP2445 exhibits siphovirus morphology and infects Mycobacterium smegmatis, Mycobacterium bovis BCG, and Mycobacterium fortuitum. Its genome is 50,711 bp in length, has a GC content of 63.34%, and contains 78 protein-coding sequences and one tRNA. MP2445 shares the highest nucleotide identity with phages in the A2 subcluster.

Vibrio parahaemolyticus is a significant zoonotic pathogen that severely threatens global public health and aquaculture. Manganese (Mn) acts as a cofactor for numerous enzymes and is crucial for bacterial oxidative stress resistance and virulence. However, excessive Mn is toxic, prompting bacteria to develop diverse Mn detoxification mechanisms. Herein, two genes (mneA and terC) located downstream of the Mn-sensing yybP-ykoY riboswitch were identified in V. parahaemolyticus. The expression of mneA and terC is specifically induced by Mn. The mneA deletion mutant (ΔmneA) exhibited impaired growth in standard (pH 7.3), alkaline (pH 8.4), or weakly acidic (pH 6.8) medium supplemented with Mn. By contrast, the terC deletion mutant (ΔterC) showed growth inhibition under high Mn concentrations in weakly acidic medium. Furthermore, the mneA and terC double mutant (ΔmneA ΔterC) was more sensitive to Mn stress than ΔmneA. Bacterial survival assays revealed that both MneA and TerC protect V. parahaemolyticus against Mn-induced bacteriostasis. Inductively coupled plasma-mass spectrometry showed that the deletion of mneA or terC significantly increases intracellular Mn content. A competitive infection assay using a zebrafish model revealed that mneA contributes to virulence in V. parahaemolyticus, while terC has no such effect. RNA sequencing analysis revealed that V. parahaemolyticus modulates the expression of multiple genes in response to Mn bacteriostatic activity. Collectively, these findings indicate that MneA and TerC exhibit distinct roles in Mn detoxification and virulence in V. parahaemolyticus. This work expands the understanding of Mn detoxification mechanisms in bacteria and highlights the role of Mn in the host-pathogen interface.IMPORTANCEManganese (Mn) is important for life, but is toxic in excess. Consequently, bacteria have evolved elaborate mechanisms to maintain intracellular Mn homeostasis. In this study, we identified and characterized two Mn exporters, that is, MneA and TerC, in Vibrio parahaemolyticus, a significant zoonotic pathogen. We showed that MneA and TerC protect V. parahaemolyticus against Mn-induced bacteriostasis via Mn efflux. Strikingly, unlike the findings in Escherichia coli that a TerC homolog Alx tunes intracellular Mn concentration at alkaline pH, V. parahaemolyticus TerC facilitates the bacterium's growth under high Mn concentrations in weakly acidic medium. Moreover, MneA is crucial for the virulence of V. parahaemolyticus, whereas TerC plays no apparent role. Collectively, while both MneA and TerC act as Mn exporters in V. parahaemolyticus, they exhibit distinct roles. This study also highlights the importance of exploring conserved proteins in different bacterial species.

ABSTRACT Bovine tuberculosis (bTB), caused by Mycobacterium bovis, causes major economic losses in the global livestock industry and threatens public health. Macrophages are essential for pathogen clearance, yet serve as the primary intracellular niche for mycobacterial survival. These characteristics emphasize the importance of identifying host factors that regulate macrophage functions during infection. To systematically uncover such factors, we constructed a genome-wide CRISPR-KO library in Bomac cells (the bovine macrophage cell line) and performed iterative infection screening under high selective pressure. Here, the host protein SH3PXD2B was identified as a key host factor to promote mycobacterial invasion: SH3PXD2B was significantly enriched in the screening. M. bovis infection significantly increases the expression of SH3PXD2B. The phagocytic activity of macrophages is enhanced in an SH3PXD2B-dependent manner, and the high expression of SH3PXD2B inhibits the migration of macrophages, causing macrophages to remain at the site of infection and promoting the entry of more mycobacteria into macrophages to establish infection. Further results indicate that macrophage migration inhibitory factor is positively regulated by SH3PXD2B and plays an important role in SH3PXD2B-mediated macrophage migration and phagocytic activity. These results indicate that SH3PXD2B is a critical host factor manipulated by M. bovis to promote infection. It provides a new potential target for genetic breeding against bTB.IMPORTANCEThere is a complex interaction between Mycobacterium bovis and the host, which has an important impact on the occurrence and development of bTB. However, the host gene regulatory network for M. bovis infection is still incomplete, and the specific genes that play a key role in the infection process are still unclear. Bovine tuberculosis remains a major economic and public health burden worldwide. This study identifies SH3PXD2B as a novel host factor that is exploited by M. bovis to promote macrophage phagocytosis and inhibit migration, thereby facilitating bacterial invasion. These findings reveal a new pathogenic strategy and suggest that SH3PXD2B may represent a potential target for host-directed therapy and genetic breeding against bTB.

IntroductionMycobacterium tuberculosis (M. tuberculosis) is the causative agent of tuberculosis (TB), which continues to be a leading cause of death from infectious diseases globally. Lung epithelial cells play a crucial role in the infection process of M. tuberculosis. However, the specific M. tuberculosis proteins that regulate lung epithelial cells remain to be identified, and the mechanisms underlying the interaction between M. tuberculosis and lung epithelial cells are still not fully understood.MethodsIn this study, CCK8 assay and xenograft tumor models were employed to investigate the effect of M. tuberculosis protein Rv0927c on the proliferation of lung epithelial cells (line A549). Flow cytometry was used to detect cell apoptosis, Western blot analysis was performed to examine the cleavage levels of apoptosis-related proteins, and JC-1 staining assay was conducted to assess mitochondrial membrane potential. Additionally, the interaction between Rv0927c and host TUFM molecules was verified by immunoprecipitation, and the role of this interaction in Rv0927c-induced apoptosis was also explored.ResultsThe results showed that Rv0927c inhibits the proliferation of lung epithelial cells both in vitro and in vivo. Flow cytometry analysis demonstrated that Rv0927c significantly increased apoptosis in A549 cells. Additionally, Rv0927c facilitated the cleavage of caspase-3, caspase-9, and PARP, while having no effect on the cleavage level of caspase-8, and it led to a decrease in mitochondrial membrane potential. Furthermore, Rv0927c interacts with host TUFM molecules, which is necessary for Rv0927c to promote apoptosis in host cells.DiscussionOur findings provide evidence that Rv0927c inhibits proliferation and regulates apoptosis by targeting TUFM in A549 cells, which contributes to the understanding of the mechanisms underlying the interaction between M. tuberculosis and lung epithelial cells.

Mycobacterium tuberculosis (M. tuberculosis) deploys effector proteins to reprogram host immunity, but how it perturbs mitochondrial homeostasis during infection remains incompletely understood. Here, Rv0927c is identified as an M. tuberculosis effector that targets the host mitochondrial translation elongation factor TUFM. Rv0927c associates with TUFM and induces mitochondrial dysfunction, including membrane depolarization, oxidative stress, and PINK1 accumulation, followed by leakage of mitochondrial DNA (mtDNA) into the cytosol. Cytosolic mtDNA is accompanied by activation of STING-IRF3 signaling and increased production of IFN-β, which promotes intracellular mycobacterial survival. In infection models, genetic deletion of Rv0927c attenuates interferon induction, whereas Rv0927c expression enhances type I interferon responses. Consistently, in a mouse tail vein infection model using H37Ra, Rv0927c is associated with elevated IFN-β transcripts in spleen and liver and exacerbated tissue pathology. Together, these findings define an Rv0927c–TUFM axis that triggers mitochondrial stress and PINK1 accumulation. This response activates STING dependent type I interferon signaling and promotes disease associated inflammation.

Mycobacterium bovis (M. bovis) is a major pathogen that causes zoonotic tuberculosis. It is transmitted via aerosols, colonizes the lungs of the host and causes oxidative stress in macrophages, resulting in tissue damage. The mechanisms underlying the oxidative damage to cells induced by this infection have not yet been fully elucidated. This study investigated the effects of SH3PXD2B on the production of reactive oxygen species (ROS) and antioxidant mechanisms during M. bovis infection. The results showed that SH3PXD2B expression was significantly upregulated after M. bovis infection, and promoted the production of cellular NOX-dependent ROS and the lipid peroxidation marker MDA. Additionally, overexpression of SH3PXD2B inhibited the Nrf2 antioxidant pathway and reduced the activities of antioxidant enzymes such as CAT, SOD, and GPx. Conversely, knockdown of SH3PXD2B exerted the opposite effect and improved cell viability. In summary, SH3PXD2B orchestrates M. bovis-induced oxidative stress in macrophages through a dual mechanism: amplifying ROS production while simultaneously crippling the host’s antioxidant capacity. These findings provide novel mechanistic insights into M. bovis pathogenesis and suggest that SH3PXD2B may be a potential molecular target for future research into disease resistance in cattle and the development of new therapies.

To evaluate the potential of Mycobacterium tuberculosis Rv0394c protein for use in the diagnosis of bovine tuberculosis, the Rv0394c protein was expressed and purified via a prokaryotic expression system. Its immunogenicity was assessed in mouse and guinea pig models, and its stimulatory effect in peripheral blood IFN-γ release assays was analyzed. The gene fragment of Rv0394c was introduced into the prokaryotic expression vector pColdI via homologous recombination to construct the recombinant plasmid pColdI-Rv0394c, which was transformed into Escherichia coli BL21(DE3) for induced expression. The purified recombinant protein was used to immunize mice to evaluate its immunogenicity. Its diagnostic efficacy as a skin hypersensitivity antigen was assessed in guinea pigs sensitized with inactivated Mycobacterium bovis. Additionally, its potential as a specific stimulating antigen was evaluated by the peripheral blood IFN-γ release assay for bovine tuberculosis. The Rv0394c fusion protein with good immunogenicity was successfully constructed and purified. Mouse experiments showed that Rv0394c stimulated splenic lymphocytes from immunized mice to produce high levels of specific IFN-γ. Skin test results demonstrated that Rv0394c elicited strong positive reactions in sensitized guinea pigs. In the bovine peripheral blood IFN-γ release assay, Rv0394c stimulated high levels of IFN-γ production in tuberculosis-positive cattle. When being used in combination with the fusion protein CFP10-ESAT6 (CE), Rv0394c significantly improved the diagnostic sensitivity for bovine tuberculosis. Rv0394c is an immunodominant antigen with the potential to serve as an allergen for skin test diagnosis and as a stimulating antigen for the peripheral blood IFN-γ release assay in bovine tuberculosis, which provides important biological materials for the development of novel diagnostic technologies for bovine tuberculosis.

Vibrio parahaemolyticus is a zoonotic pathogen that poses a serious threat to shrimp aquaculture and public health worldwide. Copper (Cu) serves as a catalytic cofactor and structural element in bacteria, but is toxic in excess. Herein, we report that the CueR-regulated transporters CopA and CusFABC coordinate Cu detoxification in V. parahaemolyticus. Through RNA sequencing, we show that most genes in the CueR and Fur regulons are significantly upregulated during V. parahaemolyticus response to external Cu. Quantitative real-time PCR (qRT-PCR) showed that copA and cusA are strongly induced by Cu. Growth curve analyses and spot dilution assay revealed that CopA and CusFABC protect V. parahaemolyticus against Cu-induced bactericidal effects. Inductively coupled plasma-mass spectrometry revealed that CopA and CusFABC are involved in maintaining Cu and Fe homoeostasis. Additionally, CopA promotes V. parahaemolyticus resistance to pyrithione. In a competitive-infection assay, the copA and cusFABC double mutant, but not the single mutants, was significantly attenuated in colonization of the zebrafish intestine. Lastly, qRT-PCR, RNA sequencing, β-galactosidase activity, and AIDmut-Seq analyses revealed that CueR can bind to the promoters of copA and cusFABC, and positively regulate their expression. Collectively, V. parahaemolyticus utilizes the CueR-regulated transporters CopA and CusFABC to coordinate Cu detoxification and colonization of the zebrafish intestine.

Objective To prepare monoclonal antibodies against bovine CD4 and identify their basic biological characteristics.Methods Recombinant bovine CD4(rHis-BoCD4 and rGST-BoCD4)was successfully expressed and purified by constructing a prokaryotic plasmid of bovine CD4 gene.The bovine CD4 monoclonal antibody was produced using hybridoma technology.The subtype and potency of the monoclonal antibody were identified and analyzed by ELISA,while specificity was analyzed through indirect immunofluorescence assay(IFA)and Western-blot.Results Four hybridoma cell lines,namely,1H4,6A10,3F9 and 4G10,stably secreting monoclonal antibodies against BoCD4 were successfully obtained.The subclasses of the monoclonal antibodies subclass 6A10 was IgG2b and the rest of the monoclonal antibodies were of IgM type.Western-blot results showed that the four anti-bovine CD4 mAb strains were able to specifically bind to the bovine CD4 protein expressed in vitro.Indirect immunofluorescence assay showed that four monoclonal antibodies were able to specifically recognize the natural bovine CD4 protein.Flow cytometry assay showed that 3F9 was best to recognize bovine natural CD4 molecules.Conclusion Four monoclonal antibody strains with high specificity to natural bovine CD4 protein were successfully prepared,which lays the foundation for the subsequent studies on the function of bovine CD4 and diagnosis and treatment of bovine T-lymphocyte diseases.

Brucellosis is classified as a class II animal disease in China, with recent years seeing an increasing prevalence of Brucella infections in livestock, posing a significant threat to public health. In this study, a novel IFN-γ enzyme-linked immunospot (ELISpot; Brucella purified protein derivative, Br-PPD) assay specifically tailored for detecting Brucella-infected cattle and goats was developed. This assay employed bovine and goat IFN-γ monoclonal antibodies, 3E3 and biotinylated 8D3, respectively, for capturing and detecting IFN-γ. This method demonstrated high sensitivity and specificity. When 10 spot-forming units was selected as the cut-off value, the sensitivity and specificity of the method were 96.9% and 90.6%, respectively. Compared with the ELISA method, the IFN-γ ELISpot assay showed ∼30 times greater sensitivity in detecting IFN-γ release from peripheral blood mononuclear cells. When applied to clinical samples from both cattle and goats, the ELISpot results strongly correlated with traditional antibody-based diagnostic methods, including the serum agglutination test (SAT), rose bengal plate test (RBPT), and competitive ELISA. The positive agreement rate exceeded 80%, and the negative agreement rate surpassed 90%. Notably, employing Brucella-vaccinated goat models, our study has confirmed that the ELISpot assay can detect Brucella infections earlier than the SAT and offers a more extended diagnostic window. As a cellular immunology-based diagnostic tool, the IFN-γ ELISpot (Br-PPD) assay holds considerable potential to improve early detection of Brucella-infected cattle and goats, addressing existing diagnostic shortcomings.

BACKGROUNDS:Streptococcus suis is an emerging zoonotic bacterial disease with increasing prevalence in the human population and is one of the most important bacterial infections in pig husbandry. There is still a lack of a thorough understanding of S. suis metabolism and the connection between metabolism and virulence. RESULTS:A genome-scale metabolic model iNX525, which included 525 genes, 708 metabolites, and 818 reactions, was manually constructed with a 74% overall MEMOTE score. The flux balance analysis results of the model exhibited good agreement with growth phenotypes under different nutrient conditions and genetic disturbances. The model predictions aligned with 71.6%, 76.3%, and 79.6% of the gene essentiality predictions from three mutant screens. The model was then used to analyze virulence factors and related synthetic pathways. One hundred and thirty-one virulence-linked genes were found by comparing to virulence factor databases, and among them, seventy-nine virulence-linked genes were in 167 metabolic reactions in model iNX525. One hundred and one of the metabolic genes were predicted to affect the formation of nine virulence-linked small molecules. Complex interrelationships between growth- and virulence-associated pathways were evaluated, and 26 genes were found to be essential for both cell growth and virulence factor production. Among these, eight enzymes and metabolites were identified as antibacterial drug targets, focusing on the biosynthesis of capsular polysaccharides and peptidoglycans. CONCLUSION:Overall, the metabolic model iNX525 provides a high-quality platform for systematic elucidation of the metabolism of S. suis.

Although metals are essential for life, they are toxic to bacteria in excessive amounts. Therefore, the maintenance of metal homeostasis is critical for bacterial physiology and pathogenesis. Vibrio parahaemolyticus is a significant food-borne pathogen that mainly causes acute gastroenteritis in humans and acute hepatopancreatic necrosis disease in shrimp. Herein, we report that ZntA functions as a zinc (Zn) and cadmium (Cd) homeostasis mechanism and contributes to oxidative stress resistance and virulence in V. parahaemolyticus. zntA is remarkably induced by Zn, copper, cobalt, nickel (Ni), and Cd, while ZntA promotes V. parahaemolyticus growth under excess Zn/Ni and Cd conditions via maintaining Zn and Cd homeostasis, respectively. The growth of ΔzntA was inhibited under iron (Fe)-restricted conditions, and the inhibition was associated with Zn homeostasis disturbance. Ferrous iron supplementation improved the growth of ΔzntA under excess Zn, Ni or Cd conditions. The resistance of ΔzntA to H2O2-induced oxidative stress also decreased, and its virulence was attenuated in zebrafish models. Quantitative real-time PCR, mutagenesis, and β-galactosidase activity assays revealed that ZntR positively regulates zntA expression by binding to its promoter. Collectively, the ZntR-regulated ZntA is crucial for Zn and Cd homeostasis and contributes to oxidative stress resistance and virulence in V. parahaemolyticus.

ABSTRACT Metals are nutrients essential for almost all lifeforms. Bacteria have evolved several mechanisms to overcome the metal restrictions imposed by the host. Vibrio parahaemolyticus causes severe threats to public health and significant economic losses in shrimp aquaculture. Herein, we report that ZrgA contributes to zinc acquisition in this pathogen. The operon VP_RS01455 to VP_RS01475 of V. parahaemolyticus encodes the putative Zn transporter ZrgABCDE, whose homologs are widely distributed in Vibrionaceae. RNA sequencing analysis revealed that V. parahaemolyticus modulates the transcriptome in response to Zn limitation. Genes in the Zinc uptake regulator (Zur) regulon are upregulated during Zn limitation, including three genes annotated to encode Zn-binding proteins. Significant upregulation of these three genes during Zn limitation was also confirmed by quantitative real-time PCR (qRT-PCR) analysis. However, only the mutants containing a VP_RS01470 (zrgA) deletion exhibited impaired growth under Zn-deficient conditions, indicating that VP_RS01470 plays the predominant role in V. parahaemolyticus Zn acquisition. The VP_RS01470 deletion mutant displayed a false appearance of decreased swimming motility under Zn-deficient conditions, as revealed by the fact that the polar flagellar-related genes were not downregulated in the mutant. Moreover, VP_RS01470 deletion produced no noticeable impact on the swarming motility and virulence in mice. qRT-PCR analysis and β-galactosidase activity assays indicated that Zur negatively regulates VP_RS01470 expression in V. parahaemolyticus. Collectively, our findings suggest that ZrgA is required for Zn acquisition in V. parahaemolyticus and highlight the importance of detecting the expression of flagellar genes during analysis of motility of a mutant deficient in growth.

Although cobalt (Co) is indispensable for life, it is toxic to cells when accumulated in excess. The DmeRF system is a well-characterized metal-response system that contributes to Co and nickel resistance in certain bacterial species. The Vibrio parahaemolyticus RIMD 2210633 genome also harbors a dmeRF operon that encodes a multiple antibiotic resistance regulator family transcriptional regulator and a cation diffusion facilitator family protein. Quantitative real-time PCR, growth curves analysis, inductively coupled plasma-mass spectrometry, β-galactosidase activity assays, electrophoretic mobility shift assays, and a mouse infection experiment were performed to characterize the function of the DmeRF system in V. parahaemolyticus. Zinc, copper, and Co significantly increase dmeF expression, with Co inducing the greatest increase. DmeF promotes V. parahaemolyticus growth under high-Co conditions. Additionally, increased accumulation of cellular Co in the ΔdmeF mutant indicates that DmeF is potentially involved in Co efflux. Moreover, DmeR represses the dmeRF operon by binding directly to its promoter in the absence of Co. Finally, the DmeRF system was not required for V. parahaemolyticus virulence in mice. Collectively, our data indicate that the DmeRF system is involved in maintaining Co homeostasis in V. parahaemolyticus and DmeR functioning as a repressor of the operon.

Metals are necessary elements for bacteria. Typically, vertebrate hosts restrict invading bacterial pathogens from accessing metals. Therefore, bacteria have evolved high-affinity metal importers to acquire metals. Streptococcus suis is a major swine pathogen and an emerging zoonotic agent that endangers the swine industry and human health worldwide. Herein, we aimed to identify the zinc acquisition systems in S. suis and evaluate their roles in bacterial virulence. Bioinformatic analyses revealed that S. suis encodes homologues of AdcA and AdcAII, two well-characterised Zn-binding lipoproteins in certain streptococci. Quantitative reverse transcription PCR (qRT-PCR) analysis revealed that the expressions of adcA and adcAII were significantly upregulated in response to Zn limitation, with a higher expression level of adcAII than adcA. Gene deletion mutants and complementation strains were constructed; their growth characteristics under Zn-deficient and Zn-replete conditions indicated that AdcA and AdcAII have overlapping functionality in Zn acquisition. A mouse infection model was used to evaluate the roles of AdcA and AdcAII in S. suis virulence. Mice infected with the double mutant ΔadcAΔadcAII exhibited a significantly higher survival rate, decreased bacterial burden, and lower production of inflammatory cytokines compared to those infected with the wild type (WT) strain. Furthermore, ΔadcAΔadcAII showed reduced competitiveness in infection establishment compared with the WT strain. RNA sequencing, qRT-PCR, and electrophoretic mobility shift assays revealed that AdcR negatively regulates the expressions of adcA and adcAII. Collectively, our results demonstrated that AdcA and AdcAII, which are negatively regulated by AdcR, contribute additively to zinc acquisition and virulence in S. suis.

Toxin-antitoxin (TA) systems, composed of a stable toxin and a cognate unstable antitoxin, are ubiquitous in the genomes of bacteria and archaea. Under suitable growth conditions, an antitoxin prevents its cognate toxin from inducing toxicity; nonetheless, under stress or plasmid loss, it is either rapidly degraded or downregulated, thereby freeing the toxin to exert its activity toward various targets. Currently, TA systems are classified into eight types based on the nature and mode of action of antitoxins. TA expression is tightly regulated at multiple levels. These systems have various biological roles, including genetic element maintenance, virulence, stress resistance, and phage inhibition. Because of the toxic property of toxins, TA systems have been exploited for biotechnological (e.g., DNA cloning, plasmid maintenance, and counterselection) and medical (e.g., antibacterial drugs, antivirals, and anticancer therapies) applications. Herein, we provided an updated overview of TA systems by focusing on their classification, biological roles, and applications. We also described recent advances in research on TA systems and discussed research perspectives in this field.

Streptococcus suis is an important zoonotic pathogen causing severe infections in swine and humans. Induction of the Vibrio parahaemolyticus YoeB toxin in Escherichia coli resulted in cell death, leading to the speculation that YoeBVp can be a counterselectable marker. Herein, the counterselection potential of YoeBVp was assessed in S. suis. The yoeBVp gene was placed under the copper-induced promoter PcopA. The PcopA-yoeBVp construct was cloned into the S. suis-E. coli shuttle vector pSET2 and introduced into S. suis to assess the effect of YoeBVp expression on S. suis growth. Reverse transcription quantitative PCR showed that copper induced yoeBVp expression. Growth curve analyses and spot dilution assays showed that YoeBVp expression inhibited S. suis growth both in liquid media and on agar plates, revealing that YoeBVp has the potential to be a counterselectable marker for S. suis. A SCIY cassette comprising the spectinomycin-resistance gene and copper-induced yoeBVp was constructed. Using the SCIY cassette and peptide-induced competence, a novel two-step markerless gene deletion method was established for S. suis. Moreover, using the ΔperR mutant generated by this method, we demonstrated that PmtA, a ferrous iron and cobalt efflux pump in S. suis, was negatively regulated by the PerR regulator.

Metals are essential nutrients for life; however, the accumulation of excess metals in cells can be toxic to bacteria. In the present study, we identified a metalloregulator, TroR, in Streptococcus suis , which is an emerging zoonotic pathogen.

[背景]副溶血弧菌是一种重要的食源性病原菌,给公众健康带来严重危害.毒素-抗毒素系统广泛存在于细菌和古生菌基因组中,具有重要的生物学功能.[目的]在副溶血弧菌中鉴定新的毒素-抗毒素系统,为从毒素-抗毒素系统角度探讨该菌致病性和耐药性的分子机制奠定基础.[方法]通过在线工具预测副溶血弧菌染色体上的假定Ⅱ型毒素-抗毒素系统;通过生长曲线分析和稀释点板实验检测假定毒素对大肠杆菌的毒性作用及相应抗毒素的抗毒性作用;通过反转录PCR确定毒素和抗毒素基因是否共转录;通过生物信息学分析确定新鉴定毒素-抗毒素系统的同源蛋白;通过LacZ报告实验确定抗毒素及毒素-抗毒素复合物对自身启动子的调控作用.[结果]副溶血弧菌染色体中编码6个假定Ⅱ型毒素-抗毒素系统;基因vp1820的表达产物(VP1820)对大肠杆菌具有杀菌活性,vp1821的表达产物(VP1821)能中和VP1820的毒性;基因vp1821和vp1820共转录;vp1821-vp1820编码YefM-YoeB毒素-抗毒素系统;抗毒素YefM正调控启动子,YefM-YoeB复合物负调控启动子.[结论]在副溶血弧菌中鉴定了一个新的Ⅱ型毒素-抗毒素系统,即YefM-YoeB,为进一步研究该系统对副溶血弧菌致病性和耐药性的影响奠定了基础.

为了确定扬州市某猪场发病猪的病原,对该猪场送检的病猪心脏进行了细菌分离与培养.通过16S rDNA测序和gdh基因PCR扩增,确定细菌种类;用血清型特异性引物进行PCR扩增,确定分离菌株的血清型;通过生长曲线测定、小鼠感染试验和药敏试验对分离菌株的生物学特性进行研究.结果表明,分离菌株为猪链球菌2型;分离菌株的生长情况与猪链球菌2型SC19菌株相似,但最高OD595值略低于SC19菌株;分离菌株对小鼠具有高致病性;分离菌株主要对青霉素类和头孢类药物敏感.这说明猪链球菌2型很可能是该猪场发病猪的病原,该猪场对猪链球菌感染的防控应优先选用青霉素类和头孢类药物.