Candida albicans is a major cause of fungal diseases in humans, and its resistance to available drugs is of concern. In an attempt to identify novel antifungal agents, we initiated a small-scale screening of a library of 199 natural plant compounds (i.e., natural products [NPs]). In vitro susceptibility profiling experiments identified 33 NPs with activity against C. albicans (MIC50s ≤ 32 μg/ml). Among the selected NPs, the sterol alkaloid tomatidine was further investigated. Tomatidine originates from the tomato (Solanum lycopersicum) and exhibited high levels of fungistatic activity against Candida species (MIC50s ≤ 1 μg/ml) but no cytotoxicity against mammalian cells. Genome-wide transcriptional analysis of tomatidine-treated C. albicans cells revealed a major alteration (upregulation) in the expression of ergosterol genes, suggesting that the ergosterol pathway is targeted by this NP. Consistent with this transcriptional response, analysis of the sterol content of tomatidine-treated cells showed not only inhibition of Erg6 (C-24 sterol methyltransferase) activity but also of Erg4 (C-24 sterol reductase) activity. A forward genetic approach in Saccharomyces cerevisiae coupled with whole-genome sequencing identified 2 nonsynonymous mutations in ERG6 (amino acids D249G and G132D) responsible for tomatidine resistance. Our results therefore unambiguously identified Erg6, a C-24 sterol methyltransferase absent in mammals, to be the main direct target of tomatidine. We tested the in vivo efficacy of tomatidine in a mouse model of C. albicans systemic infection. Treatment with a nanocrystal pharmacological formulation successfully decreased the fungal burden in infected kidneys compared to the fungal burden achieved by the use of placebo and thus confirmed the potential of tomatidine as a therapeutic agent.

The oxidative burst generated by the host immune system can restrict intracellular parasite entry and growth. While this burst leads to the induction of antioxidative enzymes, the molecular mechanisms and the consequences of this counter-response on the life of intracellular human parasites are largely unknown. The transcription factor NF-E2-related factor (NRF2) could be a key mediator of antioxidant signaling during infection due to the entry of parasites. Here, we showed that NRF2 was strongly upregulated in infection with the human Leishmania protozoan parasites, its activation was dependent on a NADPH oxidase 2 (NOX2) and SRC family of protein tyrosine kinases (SFKs) signaling pathway and it reprogrammed host cell metabolism. In inflammatory leishmaniasis caused by a viral endosymbiont inducing TNF-α in chronic leishmaniasis, NRF2 activation promoted parasite persistence but limited TNF-α production and tissue destruction. These data provided evidence of the dual role of NRF2 in protecting both the invading pathogen from reactive oxygen species and the host from an excess of the TNF-α destructive pro-inflammatory cytokine.

More than 35 million people remain infected with HIV-1. Upon antiretroviral therapy cessation, HIV-1-positive individuals systematically fail to achieve sustained virological remission, revealing the presence of a reservoir. This reservoir takes into account anatomical sanctuaries where HIV-1 continues to replicate, and latently infected cells also known as the latent reservoir (LR). A better understanding of the nature and features of the LR and its quantification are crucial to evaluate the efficiency of therapeutic strategies aiming at purging HIV-1. Culture- and PCR-based assays have already been implemented to measure the LR, and new assays are continuously being developed. In this review, we will discuss these methods highlighting the difficulties to accurately measure the LR, one main obstacle in curing HIV-1.

Leishmania parasites preferentially invade macrophages, the professional phagocytic cells, at the site of infection. Macrophages play conflicting roles in Leishmania infection either by the destruction of internalized parasites or by providing a safe shelter for parasite replication. In response to invading pathogens, however, macrophages induce an oxidative burst as a mechanism of defense to promote pathogen removal and contribute to signaling pathways involving inflammation and the immune response. Thus, oxidative stress plays a dual role in infection whereby free radicals protect against invading pathogens but can also cause inflammation resulting in tissue damage. The induced oxidative stress in parasitic infections triggers the activation in the host of the antioxidant response to counteract the damaging oxidative burst. Consequently, macrophages are crucial for disease progression or control. The ultimate outcome depends on dangerous liaisons between the infecting Leishmania spp. and the type and strength of the host immune response.

Regulatory Nod-like receptors (NLRs) are a subgroup of the cytosolic NLR family of pathogen recognition receptors (PRRs). These receptors can tune the innate immune responses triggered by the activation of other PRRs by either augmenting or attenuating the activated pro-inflammatory signaling cascades. Nod-like receptor X1 (NLRX1) is the only known mitochondria-associated negative regulatory NLR. NLRX1 attenuates several inflammatory pathways and modulates cellular processes such as autophagy and mitochondrial function following infection or injury. Using both in vitro expression and in vivo experimental models, NLRX1 is extensively described in the context of anti-viral signaling and host-defense against invading pathogens. More recently, NLRX1 has also gained interest in the field of cancer and metabolism where NLRX1 functions to attenuate overzealous inflammation in various inflammatory and autoimmune diseases. However, the exact function of this novel receptor is still under debate and many, often contradictory, mechanisms of action together with cellular localizations have been proposed. Thus, a better understanding of the underlying mechanism is crucial for future research and development of novel therapeutical approaches. Here, we summarize the current findings on NLRX1 and discuss its role in both infectious and inflammatory context.

Mitochondria regulate steroid hormone synthesis, and in turn sex hormones regulate mitochondrial function for maintaining cellular homeostasis and controlling inflammation. This crosstalk can explain sex differences observed in several pathologies such as in metabolic or inflammatory disorders. Nod-like receptor X1 (NLRX1) is a mitochondria-associated innate receptor that could modulate metabolic functions and attenuates inflammatory responses. Here, we showed that in an infectious model with the human protozoan parasite, Leishmania guyanensis, NLRX1 attenuated inflammation in females but not in male mice. Analysis of infected female and male bone marrow derived macrophages showed both sex- and genotype-specific differences in both inflammatory and metabolic profiles with increased type I interferon production, mitochondrial respiration, and glycolytic rate in Nlrx1-deficient female BMDMs in comparison to wild-type cells, while no differences were observed between males. Transcriptomics of female and male BMDMs revealed an altered steroid hormone signaling in Nlrx1-deficient cells, and a “masculinization” of Nlrx1-deficient female BMDMs. Thus, our findings suggest that NLRX1 prevents uncontrolled inflammation and metabolism in females and therefore may contribute to the sex differences observed in infectious and inflammatory diseases.

Inducible nitric oxide synthase (iNOS) is essential to the production of nitric oxide (NO), an efficient effector molecule against intracellular human pathogens such as Leishmania protozoan parasites. Some strains of Leishmania are known to bear a viral endosymbiont termed Leishmania RNA virus 1 (LRV1). Recognition of LRV1 by the innate immune sensor Toll-like receptor-3 (TLR3) leads to conditions worsening the disease severity in mice. This process is governed by type I interferon (type I IFNs) arising downstream of TLR3 stimulation and favoring the formation of secondary metastatic lesions. The formation of these lesions is mediated by the inflammatory cytokine IL-17A and occurs in the absence, or low level of, protective cytokine IFN-γ. Here, we described that the presence of LRV1 led to the initial expression of iNOS and low production of NO that failed to control infection. We subsequently showed that LRV1-triggered type I IFN was essential but insufficient to induce robust iNOS induction, which requires strong activation of nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB). Leishmania guyanensis carrying LRV1 (LgyLRV1+) parasites mitigated strong iNOS production by limiting NF-kB activation via the induction of tumor necrosis factor-alpha-induced protein 3 (TNFAIP3), also known as A20. Moreover, our data suggested that production of LRV1-induced iNOS could be correlated with parasite dissemination and metastasis via elevated secretion of IL-17A in the draining lymph nodes. Our findings support an additional strategy by which LRV1-bearing Leishmania guyanensis evaded killing by nitric oxide and suggest that low levels of LRV1-induced NO might contribute to parasite metastasis.

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