Genome-Wide Transposon Mutagenesis Indicates that Mycobacterium marinum Customizes Its Virulence Mechanisms for Survival and Replication in Different Hosts

Abstract

eThe interaction of environmental bacteria with unicellular eukaryotes is generally considered a major driving force for the evolution of intracellular pathogens, allowing them to survive and replicate in phagocytic cells of vertebrate hosts. To test this hypothesis on a genome-wide level, we determined for the intracellular pathogen Mycobacterium marinum whether it uses conserved strategies to exploit host cells from both protozoan and vertebrate origin. Using transposon-directed insertion site sequencing (TraDIS), we determined differences in genetic requirements for survival and replication in phagocytic cells of organisms from different kingdoms. In line with the general hypothesis, we identified a number of general virulence mechanisms, including the type VII protein secretion system ESX-1, biosynthesis of polyketide lipids, and utilization of sterols. However, we were also able to show that M. marinum contains an even larger set of host-specific virulence determinants, including proteins involved in the modification of surface glycolipids and, surprisingly, the auxiliary proteins of the ESX-1 system. Several of these factors were in fact counterproductive in other hosts. Therefore, M. marinum contains different sets of virulence factors that are tailored for specific hosts. Our data imply that although amoebae could function as a training ground for intracellular pathogens, they do not fully prepare pathogens for crossing species barriers.

iniBAC induction Is Vitamin B12- and MutAB-dependent in Mycobacterium marinum

“…In this work we established that M. marinum produces vitamin B12 in vitro and that MMAR_4563 is indeed a functional gene in this biosynthesis route, as previously suggested by Weerdenburg et al (33). Therefore, we propose to call this gene cobF.…”

“…Because PDIMs are important for virulence in mycobacteria (34), this may suggest a direct link between vitamin B12 biosynthesis, cell homeostasis, and virulence. Evidence for the effect of vitamin B12 homeostasis and virulence was presented by Weerdenburg et al (33), where it was shown that mutations in the vitamin B12 biosynthesis genes caused an increased fitness for M. marinum in Acanthamoeba infection experiments. Although there are two enzymes in M. marinum that need vitamin B12 as a cofactor, methionine synthase MetH and methylmalonyl-CoA mutase MutAB (24,25), based on our results the absence of MutAB could explain the observed virulence effects because the mycobacterial lipid profile is altered.…”

Global assessment ofMycobacterium aviumsubspecieshominissuisgenetic requirement for growth and virulence

“…None of the genes within the major MAH 11 insertions (relative to MAH 104, listed in Figure 3B) were required for infection, and only four were essential in vitro . It has been shown that M. marinum customizes its virulence mechanisms to infect different animal cells [19]. It is thus possible that, if subjected to infection of other animal models, a greater proportion of genes specific to MAH would be required.…”

A unique PE_PGRS protein inhibiting host cell cytosolic defenses and sustaining full virulence ofMycobacterium marinumin multiple hosts

“…When necessary, media were supplemented with the appropriate antibiotics, e.g. the MMAR_0242::Tn mutant (Weerdenburg et al , ) was grown in the presence of 25 µg/ml kanamycin. M. smegmatis overexpressing MMAR_0242 was grown in the presence of 50 µg/ml apramycin.…”

“…When necessary, media were supplemented with the appropriate antibiotics, e.g. the MMAR_0242::Tn mutant (Weerdenburg et al, 2015) was grown in the presence of 25 μg/ml kanamycin. M.…”