Genome-Wide Identification by Transposon Insertion Sequencing of Escherichia coli K1 Genes Essential for In Vitro Growth, Gastrointestinal Colonizing Capacity, and Survival in Serum.
Genome-Wide Identification by Transposon Insertion Sequencing of Escherichia coli K1 Genes Essential for In Vitro Growth, Gastrointestinal Colonizing Capacity, and Survival in Serum.
复制标题
DOI：
10.1128/jb.00698-17
复制
发表时间：
2018-04-01
影响因子：
3.2
通讯作者：
Taylor PW
中科院分区：
文献类型：
--
作者：
McCarthy AJ;Stabler RA;Taylor PW
大肠埃希菌K1株是新生儿侵袭性疾病的主要病原体。感染的年龄依赖性可以在新生大鼠身上复制。口服K1细菌后小肠的定植会迅速入侵血液循环；避免被肠系膜淋巴系统捕获并逃避血液中抗菌机制的细菌可能会传播，导致特定器官的感染，如脑膜炎。一些大肠杆菌K1的表面成分，特别是聚唾液酸胶囊，已知有助于侵袭潜力，但决定完全毒力表型的因素的全面图景迄今尚未出现。我们构建了大肠杆菌K1株A192PP的∼775,000个Tn5转座子突变体的文库和组成亚库，并使用转座子定向插入位点测序(TRADIS)来鉴定2日龄大鼠感染适应性所需的基因。在选择性琼脂上恢复后，357个基因中缺少转座子插入；这些基因被认为是在营养充足的培养基中生长所必需的。167个E.ColiK1基因产物促进了小肠中段的定植。限制细菌跨上皮屏障的移位排除了对肠道到血液和血液到脑转移的TRADIS分析；97个基因是在人血清中生存所必需的。这项研究表明，需要大量的细菌基因，其中许多以前没有与系统性大肠杆菌K1感染相关，以实现完全的入侵潜力。重要性大肠埃希氏菌K1株可导致新生儿发生危及生命的感染。它们在出生时从母亲那里获得，并在小肠定居，从那里它们入侵血液和中枢神经系统。很难从急病患者那里获得关于决定侵袭性疾病的生理和细菌因素的信息。自然发生的与年龄相关的人类感染的关键方面可以在新生大鼠身上复制。在这里，我们使用转座子定向的插入位点测序来识别对E.ColiK1的体外生长至关重要的基因，以及有助于易感大鼠定植的基因。由于血液和脑脊髓室存在侵袭的瓶颈，因此无法进行插入位点测序分析，但我们在血清中确定了存活基因。
Escherichia coli K1 strains are major causative agents of invasive disease of newborn infants. The age dependency of infection can be reproduced in neonatal rats. Colonization of the small intestine following oral administration of K1 bacteria leads rapidly to invasion of the blood circulation; bacteria that avoid capture by the mesenteric lymphatic system and evade antibacterial mechanisms in the blood may disseminate to cause organ-specific infections such as meningitis. Some E. coli K1 surface constituents, in particular the polysialic acid capsule, are known to contribute to invasive potential, but a comprehensive picture of the factors that determine the fully virulent phenotype has not emerged so far. We constructed a library and constituent sublibraries of ∼775,000 Tn5 transposon mutants of E. coli K1 strain A192PP and employed transposon-directed insertion site sequencing (TraDIS) to identify genes required for fitness for infection of 2-day-old rats. Transposon insertions were lacking in 357 genes following recovery on selective agar; these genes were considered essential for growth in nutrient-replete medium. Colonization of the midsection of the small intestine was facilitated by 167 E. coli K1 gene products. Restricted bacterial translocation across epithelial barriers precluded TraDIS analysis of gut-to-blood and blood-to-brain transits; 97 genes were required for survival in human serum. This study revealed that a large number of bacterial genes, many of which were not previously associated with systemic E. coli K1 infection, are required to realize full invasive potential. IMPORTANCE Escherichia coli K1 strains cause life-threatening infections in newborn infants. They are acquired from the mother at birth and colonize the small intestine, from where they invade the blood and central nervous system. It is difficult to obtain information from acutely ill patients that sheds light on physiological and bacterial factors determining invasive disease. Key aspects of naturally occurring age-dependent human infection can be reproduced in neonatal rats. Here, we employ transposon-directed insertion site sequencing to identify genes essential for the in vitro growth of E. coli K1 and genes that contribute to the colonization of susceptible rats. The presence of bottlenecks to invasion of the blood and cerebrospinal compartments precluded insertion site sequencing analysis, but we identified genes for survival in serum.