A comparative genomics approach identifies contact-dependent growth inhibition as a virulence determinant

Abstract

Emerging evidence suggests thePseudomonas aeruginosaaccessory genome is enriched with uncharacterized virulence genes. Identification and characterization of such genes may reveal novel pathogenic mechanisms used by particularly virulent isolates. Here, we utilized a mouse bacteremia model to quantify the virulence of 100 individualP. aeruginosabloodstream isolates and performed whole-genome sequencing to identify accessory genomic elements correlated with increased bacterial virulence. From this work, we identified a specific contact-dependent growth inhibition (CDI) system enriched among highly virulentP. aeruginosaisolates. CDI systems contain a large exoprotein (CdiA) with a C-terminal toxin (CT) domain that can vary between different isolates within a species. Prior work has revealed that delivery of a CdiA-CT domain upon direct cell-to-cell contact can inhibit replication of a susceptible target bacterium. Aside from mediating interbacterial competition, we observed our virulence-associated CdiA-CT domain to promote toxicity against mammalian cells in culture and lethality during mouse bacteremia. Structural and functional studies revealed this CdiA-CT domain to have in vitro tRNase activity, and mutations that abrogated this tRNAse activity in vitro also attenuated virulence. Furthermore, CdiA contributed to virulence in mice even in the absence of contact-dependent signaling. Overall, our findings indicate that thisP. aeruginosaCDI system functions as both an interbacterial inhibition system and a bacterial virulence factor against a mammalian host. These findings provide an impetus for continued studies into the complex role of CDI systems inP. aeruginosapathogenesis.

A Genome-based Model to Predict the Virulence ofPseudomonas aeruginosaIsolates

“…From these data, a modified 50% lethal dose (mLD 50 ) was estimated for each of the 115 P. aeruginosa isolates (Supplementary Table 3). The isolates showed a median mLD 50 of 6.9 log 10 CFU but with a wide range of pathogenicity in mice, varying by over 100-fold in the dose required to cause severe disease, as was previously reported for the NMH isolates 15 . For the purpose of this study, we classified those isolates with an estimated mLD 50 below the median value for the group as “high virulence” and the remainder as “low virulence” (Figure 1A).…”

“…We performed whole genome sequencing for each of the isolates that had not been previously sequenced. Likewise, previously reported virulence data 15,32 were supplemented with new experiments (Supplementary Table 2) to approximate the colony forming units (CFU) of each bacterial isolate necessary to cause pre-lethal illness in 50% of mice using a bacteremia model. From these data, a modified 50% lethal dose (mLD 50 ) was estimated for each of the 115 P. aeruginosa isolates (Supplementary Table 3).…”

“…Several genomic islands in P. aeruginosa , including those containing the type III secretion system (T3SS) effector gene exoU , have been shown to enhance pathogenicity in multiple infection models 12-14 . We recently mined the accessory genome and identified multiple novel factors important in virulence in a mouse model of bacteremia 15 . Conversely, a study using a Caenorhabditis elegans model identified several P. aeruginosa accessory genes whose presence reduced virulence 16 .…”

“…This is not necessarily as simple as detecting individual virulence or anti-virulence factors. For example, exoU is a recognized virulence factor whose disruption dramatically attenuates a strain’s ability to cause disease 18,19 , but some strains naturally lacking exoU are more virulent than those possessing the gene 15 . As virulence is a complex and combinatorial phenotype, the strategy taken to study it must be appropriately robust to that complexity.…”

Emergent Properties of Antiagglomerant Films Control Methane Transport: Implications for Hydrate Management

“…Furthermore, one must avoid inefficient sampling in the CV orthogonal degrees of freedom (either X− or Y − direction), despite ensuring good overlap in the Z− direction. As others indicated [112,118,119], this numerical issue could cause a discontinuity in configurational space that would introduce PMF errors. To ensure the results are reliable, we controlled the efficiency of the sampling along the CV orthogonal degrees of freedom with the implementation of the flat-bottomed potential [112,118,119]…”