Legionella pneumophila is a Gram-negative bacterium ubiquitous in freshwater environments which, if inhaled, can cause a severe pneumonia in humans. The emergence of L. pneumophila is linked to several traits selected in the environment, the acquisition of some of which involved intra- and interkingdom horizontal gene transfer events. Transposon insertion sequencing (TIS) is a powerful method to identify the genetic basis of selectable traits as well as to identify fitness determinants and essential genes, which are possible antibiotic targets. TIS has not yet been used to its full power in L. pneumophila, possibly because of the difficulty of obtaining a high-saturation transposon insertion library. Indeed, we found that isolates of sequence type 1 (ST1), which includes the commonly used laboratory strains, are poorly permissive to saturating mutagenesis by conjugation-mediated transposon delivery. In contrast, we obtained high-saturation libraries in non-ST1 clinical isolates, offering the prospect of using TIS on unaltered L. pneumophila strains. Focusing on one of them, we then used TIS to identify essential genes in L. pneumophila. We also revealed that TIS could be used to identify genes controlling vertical transmission of mobile genetic elements. We then applied TIS to identify all the genes required for L. pneumophila to develop competence and undergo natural transformation, defining the set of major and minor type IV pilins that are engaged in DNA uptake. This work paves the way for the functional exploration of the L. pneumophila genome by TIS and the identification of the genetic basis of other life traits of this species.

中文翻译：

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嗜肺军团菌临床分离株中的转座子插入测序确定必需基因和自然转化的决定因素。

嗜肺军团菌是在淡水环境中普遍存在的革兰氏阴性细菌，如果吸入，会导致人类严重的肺炎。嗜肺乳杆菌的出现与在环境中选择的若干性状有关，其中一些性状的获得涉及内部和内部的水平基因转移事件。转座子插入测序（TIS）是一种强大的方法，可以识别可选性状的遗传基础，并可以确定适合的决定簇和必需基因，这些可能是抗生素的靶标。TIS尚未在嗜肺乳杆菌中完全发挥作用，可能是因为难以获得高饱和度的转座子插入文库。实际上，我们发现序列类型1（ST1）的分离株（包括常用的实验室菌株）很难通过缀合介导的转座子传递来饱和诱变。相比之下，我们在非ST1临床分离株中获得了高饱和度文库，这为在未改变的嗜肺乳杆菌菌株上使用TIS提供了前景。专注于其中之一，然后我们使用TIS来鉴定嗜肺乳杆菌的必需基因。我们还揭示了TIS可以用于识别控制移动遗传元件垂直传播的基因。然后，我们应用TIS来鉴定嗜肺乳杆菌的所有基因发展能力并进行自然转化，从而确定了参与DNA吸收的主要和次要IV型菌毛素的集合。这项工作为通过TIS进行嗜肺乳杆菌基因组的功能探索以及确定该物种其他生命特征的遗传基础奠定了基础。