Groupe de Recherche en Ecologie Buccale, Faculté de Médecine Dentaire, Université Laval, Quebec City, Quebec, Canada.
BMC Microbiol. 2010 Feb 10;10:42. doi: 10.1186/1471-2180-10-42.
Streptococcus suis is a major swine pathogen and zoonotic agent that mainly causes septicemia, meningitis, and endocarditis. It has recently been suggested that proteinases produced by S. suis (serotype 2) are potential virulence determinants. In the present study, we screened a S. suis mutant library created by the insertion of Tn917 transposon in order to isolate a mutant deficient in a cell surface proteinase. We characterized the gene and assessed the proteinase for its potential as a virulence factor.
Two mutants (G6G and M3G) possessing a single Tn917 insertion were isolated. The affected gene coded for a protein (SSU0757) that shared a high degree of identity with Streptococccus thermophilus PrtS (95.9%) and, to a lesser extent, with Streptococcus agalactiae CspA (49.5%), which are cell surface serine proteinases. The SSU0757 protein had a calculated molecular mass of 169.6 kDa and contained the catalytic triad characteristic of subtilisin family proteinases: motif I (Asp200), motif II (His239), and motif III (Ser568). SSU0757 also had the Gram-positive cell wall anchoring motif (Leu-Pro-X-Thr-Gly) at the carboxy-terminus, which was followed by a hydrophobic domain. All the S. suis isolates tested, which belonged to different serotypes, possessed the gene encoding the SSU0757 protein. The two mutants devoid of subtilisin-like proteinase activity had longer generation times and were more susceptible to killing by whole blood than the wild-type parent strain P1/7. The virulence of the G6G and M3G mutants was compared to the wild-type strain in the CD1 mouse model. Significant differences in mortality rates were noted between the P1/7 group and the M3G and G6G groups (p < 0.001).
In summary, we identified a gene coding for a cell surface subtilisin-like serine proteinase that is widely distributed in S. suis. Evidences were brought for the involvement of this proteinase in S. suis virulence.
猪链球菌（Streptococcus suis）是一种主要的猪病原体和人畜共患病原，主要引起败血病、脑膜炎和心内膜炎。最近有研究表明，猪链球菌（血清型 2）产生的蛋白酶可能是潜在的毒力决定因素。在本研究中，我们筛选了由 Tn917 转座子插入创建的猪链球菌突变体文库，以分离出一种缺乏细胞表面蛋白酶的突变体。我们对该基因进行了特征描述，并评估了该蛋白酶作为毒力因子的潜力。
分离到两个带有单个 Tn917 插入的突变体（G6G 和 M3G）。受影响的基因编码一种蛋白（SSU0757），与嗜热链球菌 PrtS（95.9%）和程度稍低的无乳链球菌 CspA（49.5%）具有高度同源性，这两种蛋白均为细胞表面丝氨酸蛋白酶。SSU0757 蛋白的计算分子质量为 169.6 kDa，含有枯草杆菌蛋白酶家族蛋白酶的特征性催化三联体： motif I（Asp200）、motif II（His239）和 motif III（Ser568）。SSU0757 还在羧基末端具有革兰氏阳性细胞壁锚定基序（Leu-Pro-X-Thr-Gly），其后是一个疏水区。所有测试的猪链球菌分离株均属于不同的血清型，但都具有编码 SSU0757 蛋白的基因。这两种缺乏枯草杆菌蛋白酶样蛋白酶活性的突变体的代时更长，并且比野生型亲本 P1/7 菌株更容易被全血杀死。在 CD1 小鼠模型中比较了 G6G 和 M3G 突变体与野生型菌株的毒力。P1/7 组与 M3G 和 G6G 组之间的死亡率存在显著差异（p < 0.001）。
总之，我们鉴定了一个编码细胞表面枯草杆菌蛋白酶样丝氨酸蛋白酶的基因，该基因在猪链球菌中广泛分布。该蛋白酶参与猪链球菌毒力的证据被提出。
