Shen Kai, Sayeed Sameera, Antalis Patricia, Gladitz John, Ahmed Azad, Dice Bethany, Janto Benjamin, Dopico Richard, Keefe Randy, Hayes Jay, Johnson Sandra, Yu Sujun, Ehrlich Nathan, Jocz Jennifer, Kropp Laura, Wong Ray, Wadowsky Robert M, Slifkin Malcolm, Preston Robert A, Erdos Geza, Post J Christopher, Ehrlich Garth D, Hu Fen Z
Center for Genomic Sciences, Allegheny-Singer Research Institute, Allegheny General Hospital, 320 East North Ave., 11th Floor South Tower, Pittsburgh, PA 15212, USA.
Infect Immun. 2006 Sep;74(9):5272-83. doi: 10.1128/IAI.00546-06.
The distributed genome hypothesis (DGH) states that each strain within a bacterial species receives a unique distribution of genes from a population-based supragenome that is many times larger than the genome of any given strain. The observations that natural infecting populations are often polyclonal and that most chronic bacterial pathogens have highly developed mechanisms for horizontal gene transfer suggested the DGH and provided the means and the mechanisms to explain how chronic infections persist in the face of a mammalian host's adaptive defense mechanisms. Having previously established the validity of the DGH for obligate pathogens, we wished to evaluate its applicability to an opportunistic bacterial pathogen. This was accomplished by construction and analysis of a highly redundant pooled genomic library containing approximately 216,000 functional clones that was constructed from 12 low-passage clinical isolates of Pseudomonas aeruginosa, 6 otorrheic isolates and 6 from other body sites. Sequence analysis of 3,214 randomly picked clones (mean insert size, approximately 1.4 kb) from this library demonstrated that 348 (10.8%) of the clones were unique with respect to all genomic sequences of the P. aeruginosa prototype strain, PAO1. Hypothetical translations of the open reading frames within these unique sequences demonstrated protein homologies to a number of bacterial virulence factors and other proteins not previously identified in P. aeruginosa. PCR and reverse transcription-PCR-based assays were performed to analyze the distribution and expression patterns of a 70-open reading frame subset of these sequences among 11 of the clinical strains. These sequences were unevenly distributed among the clinical isolates, with nearly half (34/70) of the novel sequences being present in only one or two of the individual strains. Expression profiling revealed that a vast majority of these sequences are expressed, strongly suggesting they encode functional proteins.
分布式基因组假说（DGH）指出，细菌物种内的每个菌株从一个基于群体的超基因组中获得独特的基因分布，该超基因组比任何给定菌株的基因组大许多倍。自然感染群体通常是多克隆的，并且大多数慢性细菌病原体具有高度发达的水平基因转移机制，这些观察结果提示了DGH，并提供了解释慢性感染如何在哺乳动物宿主的适应性防御机制面前持续存在的方法和机制。我们之前已经确定了DGH对专性病原体的有效性，现在希望评估其对机会性细菌病原体的适用性。这是通过构建和分析一个高度冗余的混合基因组文库来实现的，该文库包含约216,000个功能克隆，由12株低传代铜绿假单胞菌临床分离株、6株耳漏分离株和6株来自其他身体部位的分离株构建而成。对该文库中随机挑选的3214个克隆（平均插入片段大小约为1.4 kb）进行序列分析表明，其中348个（10.8%）克隆相对于铜绿假单胞菌原型菌株PAO1的所有基因组序列而言是独特的。对这些独特序列内开放阅读框的假设翻译显示，其蛋白质与许多细菌毒力因子和其他先前未在铜绿假单胞菌中鉴定出的蛋白质具有同源性。进行了基于PCR和逆转录PCR的分析，以分析这些序列的一个包含70个开放阅读框的子集在11株临床菌株中的分布和表达模式。这些序列在临床分离株中分布不均，近一半（34/70）的新序列仅存在于一两个菌株中。表达谱分析表明，这些序列中的绝大多数都有表达，强烈提示它们编码功能蛋白。
