一株猪葡萄球菌的分离鉴定、生物学特性研究及全基因组测序分析

Isolation,identification,biological characterization,and whole genome sequencing analysis of a Staphylococcus hyicus strain

【背景】2021年6月,广东省茂名市某散养户送检了一头发病仔猪,猪身上长有脓疱,四肢关节肿大,关节内可见脓液。【目的】确定引起仔猪发病的病原菌,分析其药物敏感性,为临床用药提供指导;对分离菌株进行全基因组序列分析,挖掘其毒力因子和耐药基因,揭示该菌致病和耐药的分子机制。【方法】取关节脓液分离细菌;通过革兰氏染色、16S rRNA基因和全基因组测序分析,鉴定细菌种类;通过溶血试验、血浆凝固酶试验和生长曲线测定,确定分离菌株的溶血活性、血浆凝固酶活性和生长特性;用小鼠感染模型评估分离菌株的致病性;用纸片扩散法测定分离菌株的药物敏感性;通过全基因组序列分析挖掘分离菌株的毒力因子和耐药基因。【结果】分离菌株被鉴定为猪葡萄球菌(Staphylococcus hyicus);该菌不溶血,无血浆凝固酶活性,在胰蛋白胨大豆肉汤培养基中于37°C、120 r/min条件下生长良好;小鼠感染试验结果显示,该菌具有高致病性;药敏试验结果显示,该菌对苯唑西林、大观霉素等7种药物敏感,对青霉素G、红霉素等9种药物耐药;全基因组序列分析结果显示,该菌携带多个毒力因子和耐药基因。【结论】从发病仔猪的关节脓液中分离到一株猪葡萄球菌,可用苯唑西林、大观霉素等药物防控该菌感染;解析了该菌的基因组信息,为后续深入研究该菌致病和耐药的分子机制奠定了基础。

[Background]In June 2021,a sick piglet in Maoming,Guangdong,which had pustules,swollen joints of limbs,and pus in the joints,was detected.[Objective]To identify the pathogen for the piglet,analyze its antimicrobial susceptibility,and thus to guide the clinical medication.Moreover,to analyze the whole genome sequence of the isolated strain,explore its virulence factors and antimicrobial resistance genes,thereby revealing its molecular mechanism of pathogenicity and antimicrobial resistance.[Methods]Bacteria were isolated from the joint pus.The bacterial species was identified by Gram staining,16S rRNA gene and whole genome sequencing.The hemolysis activity,coagulase activity,and growth characteristics of the isolated strain were determined based on hemolysis test,coagulase test,and growth curve,respectively.The pathogenicity of the strain was evaluated with a mouse model.The antimicrobial susceptibility of the strain was determined by disk diffusion test.Whole genome sequence analysis was performed to explore virulence factors and antimicrobial resistance genes of the strain.[Results]The isolated strain was identified as Staphylococcus hyicus.The strain was non-hemolytic,displayed no coagulase activity,and grew well in Tryptic Soy Broth at 37°C under shaking conditions(120 r/min).The strain exhibited high pathogenicity,and was sensitive to seven antimicrobials including oxacillin and spectinomycin and resistant to nine antimicrobials including penicillin G and erythromycin.It carried a number of virulence factors and antimicrobial resistance genes.[Conclusion]A S.hyicus strain was isolated from the joint pus of a diseased piglet.Antimicrobials such as oxacillin and spectinomycin can be used for the prevention and control of the infection caused by this strain.Furthermore,whole genome sequencing analysis was performed for this strain,which laid a foundation for further research on its molecular mechanism of pathogenicity and antimicrobial resistance.