Variants of CARD8 in Leishmania guyanensis-cutaneous leishmaniasis and influence of the variants genotypes on circulating plasma cytokines IL-1 beta, TNF alpha and IL-8

Nucleotide-binding oligomerization domain, leucine-rich repeat-containing protein family (NLR) are intracellular pathogen recognition receptors mediating innate immunity, releasing proinflammatory cytokines IL-1 beta and IL-18, and promoting pyroptotic cell death, upon sensing pathogenic or endogenous danger signals. In animal models, NLRP3 inflammasome has a dual role, pathogenic or protective in Leishmania-infection, depending on the Leishmania species and mice strain. Caspase recruitment containing domain 8 (CARD8) is a negative regulator of NLRP3 inflammasome and also an inhibitor of transcription factor NF kappa B, a major transcription factor of proinflammatory cytokines. We investigated whether single nucleotide variants in CARD8 may partially explain why only a proportion of individuals coming from the same area of endemicity of leishmaniasis develop cutaneous leishmaniasis caused by Leishmania guyanensis. We genotyped four single nucleotide variants of the CARD8 gene by direct nucleotide sequencing in 1741 individuals from an endemic area of leishmaniasis, constituting 850 patients with CL and 891 healthy controls. The frequencies of the genotypes of the variants rs2288877 T>C, rs73944113 C>T, and rs2043211 A>T are similar among the patients with CL and HC, while the variant rs2288876 A>G) reveals an excess of the genotype AA among the patients with CL (44%) compared to 37% in the HC group. Allele A of the variant rs2288876 A>G) is associated with susceptibility to CL (OR = 1.2 [95%CI 1.03-1.4]; P = 0.01). Haplotype analysis showed that individuals harboring the haplotype CCAA have 280% odds of developing CL caused by L. guyanensis (OR = 3.8 [95% CI 2.0-7.7]; p = 0.00004). The variants rs2288877 T>C and rs2288876 A>G correlate with the plasma level of IL-8. Spearman correlation showed a significant positive correlation between the rs2288876 A>G allele A and the level of IL-8 (rho = 0.22; p = 0.0002). CARD8 may partially contribute to the development of CL caused by L. guyanensis. Author summaryLeishmaniasis is a neglected tropical disease caused by species of the parasite Leishmania, transmitted by the sand fly bite. The disease manifests in different forms: asymptomatic, cutaneous, and visceral. The immunological response of the infected individual is critical in the clinical manifestation of the infection by Leishmania. Till today, we do not understand why some infected people with the parasite Leishmania, living in the same endemic area, develop the disease and others do not. Our work is to study the DNA of infected people developing the disease Cutaneous Leishmaniasis and compare it to those remaining and living in the same area. The gene CARD8 has a relevant role in the regulation of immunological response. In this work, we look for differences in a small piece of DNA fragment of the gene CARD8 to establish if the single nucleotide DNA sequence variation occurs more frequently in people developing the disease than people who do not. We showed here that some individuals who develop the disease possess a small fragment of DNA called haplotype of the CARD8 gene that may contribute to some people to progress to disease development. However, this does not mean that all individuals bearing this haplotype will develop the disease when infected with Leishmania.