Correlation between infection mechanisms and virulence plasmid of Salmonella choleraesuis

1.猪霍乱沙门氏菌毒力质粒pKDSC50与其小鼠毒力的相关性研究：将含有霍乱沙门氏菌毒力质粒pKDSC50的菌株(亲本株和再导入株)和治愈株接种ICR小鼠，连续14天进行病理学检查。亲本菌株和重新引入的菌株引起小鼠菌血症，并从脾和肝脏中回收接种的细菌。结果：1.小鼠腹腔液、肠系膜和内脏器官组织均有明显的病理改变，并持续到第14天。相比之下，感染质粒治愈株的小鼠未出现菌血症，仅在腹腔液、肠系膜和内脏有轻微和一过性的病变。携带pKDSC50的霍乱沙门氏菌和治愈株细胞表面成分的差异：脂多糖、外膜蛋白和亲本磷脂，再导入和治愈株是ChE…在化学和生物化学方面更具可比性。两者差异无统计学意义，表明pKDSC50不介导细菌细胞表面可检测到的变化。转座子插入失活导致小鼠菌血症能力的区域定位：在病理研究结果的基础上建立小鼠菌血症试验，用于筛选100株转座子TnL插入失活构建的突变株。将小鼠菌血症试验阴性菌株的质粒DNA通过转化导入治愈菌株，并再次检测转化子的小鼠菌血症能力。有9株菌株二次检测为阴性，其质粒上的TnL定位于pKDSC50的6.2kb区域。这一地区被指定为MBA地区。PKDSC50基因MBA区的遗传学研究：在大肠杆菌中克隆MBA区的DNA片段，并用大肠杆菌微型细胞系统或异丙基硫代半乳糖苷诱导的Lac-启动子系统对该区域编码的蛋白质进行鉴定。检测到4个表观分子量分别为29K、70K、32K和32K的蛋白质，通过核苷酸序列分析，在MBA区找到了与这4个蛋白质相对应的开放阅读框。其他研究小组最近也报道了类似的编码在鼠伤寒沙门氏菌毒力质粒中的蛋白质，这表明我们发现的蛋白质在携带沙门氏菌的强毒质粒中是共同的，在沙门氏菌的致病机制中起着重要作用。较少

1. Correlation between virulence plasmid of Salmonella choleraesuis, pKDSC50, and its mouse virulence: S. choleraesuis strains harboring virulence plasmid, pKDSC50 (parent and re-introduced strains), and cured strains were inoculated intraperitoneally into ICR mice, and the mice were examined pathologically for 14 days. Parent and re-introduced strains caused bacteremia in mice, and inoculated bacteria were recovered from spleen and liver. Obvious pathological changes were observed at individual tissues of peritoneal fluid, mesenterium, and visceral organs, and those changes persisted to day 14. In contrast, mice infected with plasmid-cured strain showed no bacteremia, and had only mild and transient pathological changes at peritoneal fluid, mesenterium, and viscera.2. Difference of cell surface components between pKDSC50-harbouring and -cured strains of S. choleraesuis : Lipopolysaccharide, outer membrane proteins, and phospholipids of parent, re-introduced, and cured strains were che … More mically and biochemically compared. There was no significant difference between them, indicating that pKDSC50 did not mediate the detectable change of bacterial cell surface.3. Mapping of the region responsible for the ability to cause mouse bacteremia by transposoninsertional inactivation: Mouse bacteremia test was established in the base of the results of pathological study, and used for screening 100 mutant strains constructed by transposon Tnl-insertional inactivation. Plasmid DNA of mouse bacteremia test-negative strains were introduced to cured strain by transformation, and the transformants were again tested for mouse bacteremia ability. Nine strains were negative for the second test, and Tnl on their plasmids were localized in 6.2 kb region of pKDSC50. This region was designated mba region.4. Genetical study on mba region of pKDSC50: DNA fragments of mba region were cloned in E. coli, and proteins encoded in this region were identified by E. coli minicell system, or by lac-promoter system with induction of isopropylthiogalactoside. Four proteins with apparent molecular weights of 29k, 70k, 32k, and 32k were detected, and open reading frames corresponding to these four proteins were found in mba region by nucleotide sequential analysis. Other research groups has recently reported similar proteins encoded in virulence plasmid of S. typhimurium, suggesting that the proteins we found were common among virulence plasmid-harboring Salmonella, and playing important role in pathogenesis of Salmonella. Less