Transposon-insertion sequencing in a clinical isolate of Legionella pneumophila to identify essential genes and determinants of natural transformation

Transposon-insertion sequencing (TIS) is an increasingly common component of the toolkit of microbiologists to explore the genotype-phenotype relationship in bacteria. In addition to its use in identification of the genetic determinants of selectable phenotypes, TIS can also identify fitness determinants and essential genes, possible antibiotic targets. The power of TIS to identify genetic determinants of growth and/or of specific functions is largely dependent on the complexity of the transposon insertion library that can be obtained. We here sought to obtain a high-saturation transposon insertion library for Legionella pneumophila, a human pathogen for which TIS has not yet been used to its full power. We obtained high-saturation libraries in the non-ST1 clinical isolate HL77S. We therefore identified essential genes in L. pneumophila, and also revealed that TIS could be used to identify genes controlling vertical transmission of mobile genetic elements. We then applied TIS to identified all 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 and the identification of the genetic basis other life traits of this species.