Molecular analysis of streptococcal infections diseases by the functional genomics.

A群链球菌（GAS）是一种革兰氏阳性细菌病原体，可引起各种化脓性感染和非化脓性后遗症。自20世纪80年代末以来，全球已经报道了链球菌中毒性休克样综合征（STSS）和严重侵袭性GAS感染。本文对从日本STSS患者分离的M3血清型SSI-1进行了基因组测序，并与其他GAS菌株进行了比较。SSI-1基因组全长1,884,275 bp，其中1.7 Mb序列相对于菌株SF370 （M1血清型）和MGAS8232 （M18血清型）高度保守，相对于菌株MGAS315 （M3血清型）几乎完全保守。然而，在同源rrn-comX1区域和复制轴上的两个噬菌体编码区域之间的复制轴上发生了大规模的基因组重排。这种基因组重排发生在1990年之后收集的65%的临床分离株（64/94）中，而在1985年之前收集的临床分离株中仅发现25%（7/ 94）。这些观察结果表明，链球菌噬菌体是GAS染色体中重要的可塑性区域，同源噬菌体基因之间可以发生重组，不仅可以产生新的噬菌体衍生物，还可以导致大规模的染色体重排。GAS表达多种与细菌细胞壁相关的结构蛋白和酶蛋白。已经证明，几种表面蛋白具有与人类宿主蛋白的结合能力，如纤维连接蛋白（Fn）、层粘连蛋白、纤溶蛋白、胶原蛋白、免疫球蛋白和C4b。GAS的fn结合蛋白也被报道为黏附蛋白和侵入蛋白，包括蛋白F1/SfbI、蛋白F2、SfbII/SOF、PFBP和Fba（更名为FbaA）。通过对来自TSLS患者的M3菌株的全基因组序列进行检索，在推测的表面蛋白中发现了一个开放的阅读框。它具有LPXTG基序和c端fn结合重复结构域，被命名为FbaB （A群链球菌B型fn结合蛋白）。fbaB基因在所有M3和M18菌株中均有发现，但在其他M血清型中未发现。此外，FbaB蛋白在TSLS菌株的细胞表面表达，而在非TSLS菌株的细胞表面不表达。酶联免疫吸附试验和配体印迹显示重组FbaB具有较强的fn结合能力。fbab缺陷突变株对HEp-2细胞的粘附和侵袭效率比野生型低6倍。此外，与野生型相比，感染突变株的小鼠死亡率降低。这些数据表明，FbaB在病因学上参与了侵袭性链球菌疾病的发展。在过去的二十年中，侵袭性化脓性链球菌感染在世界范围内的明显死灰复燃仍未得到解释。目前，化脓性脓球菌感染后引起中毒性休克样综合征或坏死性筋膜炎的动物模型尚不完善。我们在此证明，在鼻内感染非致死剂量的化脓性葡萄球菌菌株前2天感染非致死剂量的甲型流感病毒可导致小鼠死亡率超过90%，其中10%表现为坏死性筋膜炎。甲型流感病毒感染肺泡上皮细胞导致细胞表面表达病毒血凝素，促进化脓性脓毒杆菌内化。然而，用单克隆血凝素抗体治疗可显著降低这种内化。我们的研究结果表明，先前感染甲型流感病毒可诱导致死性协同作用，导致小鼠侵袭性化脓性s.c ogenes感染。我们发现自噬机制可以有效地清除非吞噬细胞内的致病性A群链球菌（GAS）。从核内体逃逸到细胞质后，GAS被类似自噬体的室室包裹，并在这些室室与溶酶体融合时被杀死。在自噬缺陷的Atg5-/-细胞中，GAS存活、繁殖并从细胞中释放出来。因此，自噬机制可以作为对抗入侵病原体的先天防御系统。少

Group A streptococcus(GAS) is a gram-positive bacterial pathogen that causes various suppurative infections and nonsuppurative sequelae. Since the late 1980s, streptococcal toxic-shock like syndrome(STSS) and severe invasive GAS infections have been reported globally. Here we sequenced the genome of serotype M3 strain SSI-1, isolated from an STSS patient in Japan, and compared it with those of other GAS strains. The SSI-1 genome is composed of 1,884,275 bp, and 1.7 Mb of the sequence is highly conserved relative to strain SF370 (serotype M1) and MGAS8232 (serotype M18), and almost completely conserved relative to strain MGAS315 (serotype M3). However, a large genomic rearrangement has been shown to occur across the replication axis between the homologous rrn-comX1 regions and between two prophage-coding regions across the replication axis. This genomic rearrangement occurs in 65% of clinical isolates (64/94) collected after 1990, whereas it is found in only 25% of clinical isolates (7/ … More 28) collected before 1985. These observations indicate that streptococcal phages represent important plasticity regions in the GAS chromosome where recombination between homologous phage genes can occur and result not only in new phage derivatives, but also in large chromosomal rearrangements.GAS expresses a wide variety of structural and enzyme proteins that are associated with the bacterial cell wall. It has been demonstrated that several surface proteins have binding abilities to human host proteins, such as fibronectin(Fn), laminin, plasmin, collagen, immunoglobulins, and C4b. Fn-binding proteins of GAS have also been reported to be adhesins and invasins, including protein F1/SfbI, protein F2, SfbII/SOF, PFBP, and Fba (renamed FbaA). By searching the whole genome sequence of an M3 strain from a TSLS patient, an open reading frame was found among the putative surface proteins. It possessed an LPXTG motif and Fn-binding repeat domains in the C-terminal region and was designated as FbaB (Fn-binding protein of group A streptococci type B). The fbaB gene was found in all M3 and M18 strains examined, although not in other M serotypes. Furthermore, FbaB protein was expressed on the cell surface of TSLS strains but not on non-TSLS ones. Enzyme-linked immunosorbent assay and ligand blotting revealed that recombinant FbaB exhibits a strong Fn-binding ability. An FbaB-deficient mutant strain showed 6-fold lower adhesion and invasion efficiencies to HEp-2 cells than the wild type. Moreover, mortality was decreased in mice infected with the mutant strain in comparison to the wild type. These data suggest that FbaB is etiologically involved in the development of invasive streptococcal diseases.The apparent worldwide resurgence of invasive Streptococcus pyogenes infection in the last two decades remains unexplained. At present, animal models in which toxic shock-like syndrome or necrotizing fasciitis is induced after S.pyogenes infection are not well developed. We demonstrate here that infection with a nonlethal dose of influenza A virus 2 days before intranasal infection with a nonlethal dose of S.pyogenes strains led to a death rate of more than 90% in mice, 10% of which showed necrotizing fasciitis. Infection of lung alveolar epithelial cells by the influenza A virus resulted in viral hemagglutinin expression on the cell surface and promoted internalization of S.pyogenes. However, treatment with monoclonal antibodies to hemagglutinin markedly decreased this internalization. Our results indicate that prior infection with influenza A virus induces a lethal synergism, resulting in the induction of invasive S.pyogenes infection in mice.We found that the autophagic machinery could effectively eliminate pathogenic group A Streptococcus(GAS) within nonphagocytic cells. After escaping from endosomes into the cytoplasm, GAS became enveloped by autophagosome-like compartments and were killed upon fusion of these compartments with lysosomes. In autophagy-deficient Atg5-/- cells, GAS survived, multiplied, and were released from the cells. Thus, the autophagic machinery can act as an innate defense system against invading pathogens. Less