Does ampicillin resistance or clade type determine GI colonization by E. faecium?

描述（由申请人提供）：屎肠球菌（Efm）过去引起的感染很少，但近年来显著增加，包括心内膜炎和其他严重感染，其中一些是医学上无法治疗的（例如，>80%的医院Efm是万古霉素耐药的（VRE），几乎所有的都是氨苄青霉素耐药的，氨苄青霉素是传统的药物选择）。这导致Efm被列入“坏bug”组，我们似乎“没有ESKAPE”。Efm从“传染性”转变为“致病性”的因素尚不清楚，但抗生素耐药性、移动的遗传元件和/或赋予致病性或适应性增加的基因已被归咎于此，因为这些基因更常见于几乎所有爆发/医院（医院相关（HA））分离株的Efm克隆复合体中。我们最近的基因组比较表明，物种Efm由2个不同的分支组成，其核心基因序列差异> 4%。几乎所有的HA Efm都属于一个（HA）分支，而几乎所有来自健康志愿者群体的社区相关（CA）分离株都属于另一个（CA）分支; faecalis，尽管它也具有HA克隆复合物。我们还发现pbp 5是HA进化枝菌株中编码PBP 5-R（氨苄青霉素抗性）的核心基因，并且与CA进化枝菌株的PBP 5-S（氨苄青霉素MIC <4）相差约5%。我们的分子钟估计表明，这两个分支在现代抗生素时代之前很久（300，000 ~ e100万年）就分化了。因此，最近增加的医院EFM是由于CA的HA菌株的替代，而不是，如已经提出的，最近的社区占主导地位的（CA）菌株内的演变。然而，目前尚不清楚是什么解释了这种替代，为什么HA菌株很少在健康志愿者中看到，也不清楚HA分离株是否更具致病性。也许CA菌株实际上更好地定殖，解释了它们在社区粪便中的巨大优势，也许仅仅是HA进化枝的抗生素耐药性使其分离株能够克服CA分离株，生长到高密度，然后在医疗环境中引起感染。这两个进化枝之间的固有差异是对氨苄青霉素的耐药性;值得注意的是，HA进化枝氨苄青霉素耐药菌株（也是高度头孢菌素耐药的）先于全球医院内VRE的出现。这个建议的假设是，核心基因组，也许完全是HA分支的pbp 5-R，已经允许这个分支在接受？内酰胺。在目标1中，我们将研究哪种更好地定殖：HA-进化枝Efm菌株或CA-进化枝Efm菌株，然后确定？在小鼠GI定殖模型中，使用内酰胺促进HA进化枝分离株（PBP 5-R）定殖超过CA分离株（PBP 5-S）。在目标2中，我们将研究pbp 5等位基因在Efm定植过程中的特定作用。通过在CA和HA菌株之间交换pbp 5-S和-R来使用内酰胺。这些结果可能为预防感染开辟新的途径，或者，如果HA菌株比CA菌株更好地定殖，即使没有？内酰胺类抗生素，新的途径，以追求在未来解开其他核心和/或后天基因的贡献。

DESCRIPTION (provided by applicant): Enterococcus faecium (Efm) caused very few infections in the past but these have increased markedly in recent years, including endocarditis and other serious infections, some of which are medically untreatable (e.g., >80% of nosocomial Efm are vancomycin resistant (VRE), almost all of which are ampicillin resistant, the traditional drugs of choice). This led to Efm's inclusion in the group of "bad bugs" from which we seem to have "no ESKAPE". The factors behind the shift of Efm from "commensal" to "pathogenic" are not well understood but antibiotic resistance, mobile genetic elements and/or genes conferring increased pathogenicity or fitness have been blamed, since such genes are more commonly found in the Efm clonal complexes that account for almost all outbreak/nosocomial (hospital-associated (HA)) isolates. Our recent genomic comparisons showed that the species Efm consists of 2 distinct clades whose core gene sequences differ by >4%. Almost all HA Efm belong to one (HA) clade whereas almost all community-associated (CA) isolates from healthy volunteers group in the other (CA) clade; such core gene differences are not observed in E. faecalis although it too has HA clonal complexes. We also found that pbp5 is a core gene that encodes, in HA-clade strains, PBP5-R (ampicillin resistance) and varies from PBP5-S (ampicillin MICs <4) of CA-clade isolates by ~5%. Our molecular clock estimates indicate that the 2 clades diverged long before (300,000 ~ e1 million years) the modern antibiotic era. Thus, the recent increase of nosocomial Efm is due to replacement of CA by HA strains, rather than, as had been suggested, recent evolution within community-predominant (CA) strains. However, it is not clear what explains this replacement, why HA strains are seldom seen in healthy volunteers nor if HA isolates are more pathogenic. Perhaps CA strains actually colonize better, explaining their vast predominance in community feces, and perhaps it is simply the antibiotic resistance of the HA clade that allows its isolates to overcome CA isolates, grow to high density and then cause infections in the healthcare setting. An inherent difference between these 2 clades is resistance to ampicillin; of note, HA-clade ampicillin-resistant strains (which are also highly cephalosporin resistant) preceded nosocomial VRE emergence worldwide. The hypothesis of this proposal is that the core genome, and perhaps entirely pbp5-R of the HA clade, has allowed this clade to replace CA strains in patients receiving ?-lactams. In Aim 1, we will investigate which colonize better: HA-clade Efm strains or CA-clade Efm strains, and then determine the impact of ?-lactam use on promoting colonization by HA-clade isolates (PBP5-R) over CA isolates (PBP5-S) in the mouse GI colonization model. In Aim 2, we will examine the specific role of pbp5 alleles in Efm colonization during ?-lactam use by swapping pbp5-S and -R between CA and HA strains. The results could open new avenues for preventing infection or, if HA strains colonize better than CA ones even without ?-lactam antibiotics, new avenues to pursue in the future to unravel the contributions of other core and/or acquired genes.