The oxidative stress response of pathogenic Leptospira is controlled by two peroxide stress regulators which putatively cooperate in controlling virulence

Abstract

Pathogenic Leptospira are the causative agents of leptospirosis, the most widespread zoonotic infectious disease. Leptospirosis is a potentially severe and life-threatening emerging disease with highest burden in sub-tropical areas and impoverish populations. Mechanisms allowing pathogenic Leptospira to survive inside a host and induce acute leptospirosis are not fully understood. The ability to resist deadly oxidants produced by the host during infection is pivotal for Leptospira virulence. We have previously shown that genes encoding defenses against oxidants in L. interrogans are repressed by PerRA (encoded by LIMLP_10155), a peroxide stress regulator of the Fur family. In this study, we describe the identification and characterization of another putative PerR-like regulator (LIMLP_05620) in L. interrogans. Protein sequence and phylogenetic analyses indicated that LIMLP_05620 displayed all the canonical PerR amino acid residues and is restricted to pathogenic Leptospira clades. We therefore named this PerR-like regulator PerRB. In L. interrogans, the PerRB regulon is distinct from that of PerRA. While a perRA mutant had a greater tolerance to peroxide, inactivating perRB led to a higher tolerance to superoxide, suggesting that these two regulators have a distinct function in the adaptation of L. interrogans to oxidative stress. The concomitant inactivation of perRA and perRB resulted in a higher tolerance to both peroxide and superoxide and, unlike the single mutants, to the loss of Leptospira virulence. Interestingly, this correlated with major changes in gene and non-coding RNA expression, only observed in the double perRAperRB mutant. Notably, several virulence-associated genes (clpB, ligA/B, and lvrAB) were repressed. By obtaining the first double mutant in a pathogenic Leptospira strain, our study has uncovered for the first time the interplay of two PerRs, not only in the adaptation of Leptospira to oxidative stress, but also in their virulence and pathogenicity, most likely through the transcriptional control of a complex regulatory network.

The FUR-like regulators PerRA and PerRB integrate a complex regulatory network that promotes mammalian host-adaptation and virulence ofLeptospira interrogans

“…To investigate the molecular basis for the phenotypic differences between PerRA and PerRB single and double mutants and identify putative virulence-related genes dysregulated by the loss of both regulators in vivo , we performed comparative RNA-Seq on all three mutant strains cultivated within DMCs. Similar to RNA-seq data for in vitro -cultivated organisms [91, 92], we saw very little overlap between the PerRA and PerRB regulons within mammals. Interestingly, the PerRB DMC regulon was substantially larger than its in vitro counterpart [91].…”

“…Similar to RNA-seq data for in vitro -cultivated organisms [91, 92], we saw very little overlap between the PerRA and PerRB regulons within mammals. Interestingly, the PerRB DMC regulon was substantially larger than its in vitro counterpart [91]. Importantly, by RNA-Seq, we identified 90 genes that are dysregulated only in the double mutant cultivated in DMCs.…”

“…[90], suggest that PerRA functions as a metal-dependent peroxide stress regulator. Consistent with its repressor function in other bacteria, a L. interrogans perRA transposon mutant expresses increased levels of catalase, AhpC and cytochrome c-peroxidase and enhanced survival following exposure to peroxide in vitro [89–91]. Recently, Zavala-Alvarado et al .…”

“…[91] demonstrated that expression of perRB also was increased by H 2 O 2 in vitro . Inactivation of perRB increased survival to superoxide but not H 2 O 2 [91]. These data suggest that PerRA and PerRB likely are functionally distinct.…”

“…[91, 92] saw little overlap between the PerRA and PerRB regulons by RNA-Seq analysis of in vitro -cultivated organisms. Interestingly, while perRA and perRB single mutants are virulent in hamsters [89, 92], a perRA/B double mutant was avirulent [91]. Collectively, these data argue that L. interrogans requires at least one functional PerR-like regulator for infection in mammals.…”