Manganese acquisition is essential for virulence of Enterococcus faecalis

Abstract

Manganese (Mn) is an essential micronutrient that is not readily available to pathogens during infection due to an active host defense mechanism known as nutritional immunity. To overcome this nutrient restriction, bacteria utilize high-affinity transporters that allow them to compete with host metal-binding proteins. Despite the established role of Mn in bacterial pathogenesis, little is known about the relevance of Mn in the pathophysiology of E. faecalis. Here, we identified and characterized the major Mn acquisition systems of E. faecalis. We discovered that the ABC-type permease EfaCBA and two Nramp-type transporters, named MntH1 and MntH2, work collectively to promote cell growth under Mn-restricted conditions. The simultaneous inactivation of EfaCBA, MntH1 and MntH2 (ΔefaΔmntH1ΔmntH2 strain) led to drastic reductions (>95%) in cellular Mn content, severe growth defects in body fluids (serum and urine) ex vivo, significant loss of virulence in Galleria mellonella, and virtually complete loss of virulence in rabbit endocarditis and murine catheter-associated urinary tract infection (CAUTI) models. Despite the functional redundancy of EfaCBA, MntH1 and MntH2 under in vitro or ex vivo conditions and in the invertebrate model, dual inactivation of efaCBA and mntH2 (ΔefaΔmntH2 strain) was sufficient to prompt maximal sensitivity to calprotectin, a Mn- and Zn-chelating host antimicrobial protein, and for the loss of virulence in mammalian models. Interestingly, EfaCBA appears to play a prominent role during systemic infection, whereas MntH2 was more important during CAUTI. The different roles of EfaCBA and MntH2 in these sites could be attributed, at least in part, to the differential expression of efaA and mntH2 in cells isolated from hearts or from bladders. Collectively, this study demonstrates that Mn acquisition is essential for the pathogenesis of E. faecalis and validates Mn uptake systems as promising targets for the development of new antimicrobials.

Adaptation of the Alphaproteobacterium Rhodobacter sphaeroides to stationary phase

“…Some bacteria even have multiple Mn uptake systems. Enterococcus faecalis , for example, has three Mn uptake systems, at least two of which are dedicated to Mn uptake (Colomer‐Winter et al ., ). The knockout of all three systems resulted in poor growth.…”

Manganese Uptake, Mediated by SloABC and MntH, Is Essential for the Fitness of Streptococcus mutans

“…In contrast, genes encoding hypothetical proteins accounted for more than 50% of the downregulated genes followed by genes involved in transport and binding. The genes that were most highly upregulated during growth under Mn-restricted conditions were those of the dual Fe and Mn transporter sloABC operon (Ն56-fold to 99-fold), a small open (32,39). Other genes upregulated in the absence of Mn were several belonging to the CRISPR2-cas operon (smu1753c to smu1764c; Ͼ4-fold) as well as 3 of 4 genes of the smu995 to smu998 operon (Ͼ2-fold), recently shown to code for an Fe transport system (40).…”

“…Beyond the oral environment, the ability to scavenge Mn in environments in which availability of this metal is known to be restricted, such as the bloodstream and internal organs, has proven to be an essential trait for bacterial pathogens. In fact, a growing number of Mn transport systems have been identified as major virulence factors, including examples where loss of Mn transporters rendered organisms closely related to S. mutans, such as Streptococcus pneumoniae and Enterococcus faecalis, virtually avirulent in animal infection models (32,33). In S. mutans, previous characterization of pathways associated with Mn homeostasis has been restricted to the metalloregulator SloR and the ABC-type transporter SloABC (34)(35)(36)(37)(38).…”

Genomic, Phenotypic, and Virulence Analysis of Streptococcus sanguinis Oral and Infective-Endocarditis Isolates

“…Our bioinformatic analysis indicated that all of the strains in our collection have the manganese transport system encoded by ssaACB (22), with some strains also having NRAMP-family transporters (55). We therefore expected that none of the strains would be deficient for manganese to the same extent as transport mutants examined in past studies (21,22,56). Nevertheless, we wondered whether manganese levels would vary among strains and, if so, whether levels would correlate with virulence.…”