1. Introduction
Leishmaniasis, a neglected vector-borne disease caused by the intracellular protozoan parasites Leishmania spp., remains a significant public health issue in the Americas, East Africa, North Africa and West and South-East Asia. According to the WHO, 99 countries were considered endemic for leishmaniasis in 2022, and, among these, 90 were endemic for cutaneous leishmaniasis (CL), of which 71 were also endemic for visceral leishmaniasis (VL) [1]. In 2022, 205,986 cases of CL and 12,842 cases of VL were reported [1]. In 2020, 16,813 new cases of tegumentary leishmaniasis (TL) were reported in Brazil [2], of which 1690 were from the state of Amazonas [2].
Leishmaniasis is transmitted by phlebotomine sandflies during blood meals, introducing Leishmania promastigotes into the host. The clinical manifestations of leishmaniasis range from asymptomatic cases and self-healing or non-healing CL to severe mucosal involvement (ML) or life-threatening VL. The host’s clinical response to Leishmania infection is closely linked to their immunological response. Disease outcomes depend on the infecting Leishmania species, intrinsic virulence factors and the genetic and immune profile of the host [3,4]. More than 20 Leishmania species can cause leishmaniasis. In Brazil, the most common species causing CL are L. braziliensis, L. guyanensis, L. lainsoni, L. naiffi and L. lindenbergi.
The nucleotide-binding domain and leucine-rich repeat-containing receptor protein (NLRP) family comprises a group of intracellular pathogen recognition receptors with diverse functions in regulating innate immunity and inflammation. The activation of NLRP1, NLRP3 and NLRC4 triggers the assembly of inflammasome complexes through the association of apoptosis-associated speck-like protein containing a caspase-1 recruiting domain (ASC) and pro-CASP-1 via Pyrin–Pyrin and CARD–CARD homotypic interactions, respectively. Upon activation, pro-CASP-1 is converted to CASP-1, which processes pro-IL-1β and pro-IL-18 into their active secreted forms, IL-1β and IL-18 [5,6]. Caspase recruitment domain-containing family member 8 (CARD8) acts as a negative regulator of the NLRP3 inflammasome [7]. The FIIND domain of CARD8 interacts with the nucleotide-binding domain (NOD) of NLRP3, while the C-terminal CARD domain interacts with CASP-1 via CARD–CARD homophilic interactions [8,9]. Silencing CARD8 expression through small interfering RNA (siRNA) increases the production of IL-1β [7]. CARD8 also inhibits nuclear factor kappa B (NFkB) activity, leading to the reduced expression of IL-1 β and TNFα [10,11].
The NLRP3 inflammasome is activated during Leishmania infection [12,13]. The activation of the NLRP3 inflammasome requires a priming signal to activate the transcription factor nuclear factor kappa B (NF-kB) and the extracellular signal-related kinase (ERK) pathways for the transcription of NLRP3, pro-IL-1β and pro-IL-18. Leishmania lipophosphoglycan, glycosphingophospholipids and DNA are recognized by Toll-like receptors (TLR2, TLR4 and TLR9, respectively). During Leishmania infection, these TLRs promote the transcription of inflammatory cytokines such as TNFα, IL-6, IL-1β and IL-12. This signaling may function as a primary trigger for NLRP3 inflammasome assembly by facilitating the transcription of NLRP3 via NFκB [12].
In studies with C57BL6 mice deficient in NLRP3, ASC or CASP1, an increase in the lesion size and parasite burden was observed upon infection with L. amazonensis [14]. In vitro analyses using macrophages infected with L. braziliensis, L. amazonensis or L. mexicana demonstrated that the NLRP3 inflammasome’s activation is required for IL-1β secretion and host resistance [14], while these mice exhibited protection against the L. major Seidman strain, which induces chronic lesions in C57BL6 mice [15]. In contrast, susceptible BALB/c mice showed resistance to L. major infection in the absence of the NLRP3 inflammasome components [16]. Similarly, mice lacking CASP1 or NLRP3 exhibited reduced pathology when infected with L. braziliensis [17].
The double-stranded Leishmania RNA virus (LRV) found in L. guyanensis inhibits CASP-1 activation and IL-1β production by inducing type-1 interferon signaling via TLR3, leading to the autophagy-mediated degradation of NLRP3 [18]. Bone marrow-derived macrophages (BMDMs) infected with LRV-positive L. guyanensis displayed reduced levels of CASP-1 and IL-1β compared to those infected with LRV-negative L. guyanensis in vitro. C57BL6 mice infected with LRV-positive L. guyanensis exhibited more pronounced ear swelling and higher parasite burdens than those infected with LRV-negative L. guyanensis. This difference was diminished in NLRP3-deficient C57BL6 mice infected with LRV-negative L. guyanensis, suggesting that NLRP3 and IL-1β play protective roles against L. guyanensis infection [18].
Apoptotic bodies released from dying cells can act as danger-associated molecular patterns (DAMPs), activating inflammasomes to trigger CASP-1 activation for the processing of pro-IL-1β [19]. CASP-1 has been shown to be critical in inducing CD8+T cell-mediated pathology but not in controlling parasite multiplication in C57BL/6 mice co-infected with L. major and lymphocytic choriomeningitis virus (LCMV) [20]. Similarly, wild-type C57BL/6 mice co-infected with L. major and LCMV developed severe lesions compared to NLRP3-deficient C57BL/6 mice co-infected under the same conditions [20]. CD8+ T cells have been shown to promote inflammation through granule-mediated cytotoxicity in L. braziliensis patients [21,22,23,24,25].
Given the dual role of NLRP3 inflammasome activation in leishmaniasis, where it may be protective or pathogenic depending on the infecting Leishmania species, and the levels of pro-IL-1β processed into active IL-1β by CASP-1, we investigated whether genetic variants of CASP1 influenced the plasma levels of IL-1 β in patients with CL caused by L. guyanensis and healthy individuals with no history of leishmaniasis from the same endemic region.
2. Results
2.1. Basic Characteristics of the Study Population
The basic characteristics of the study population have been previously described [26]. Briefly, the study comprised 850 patients with Lg-CL and 891 HCs. Among the Lg-CL patients, 639 were male, with a mean age of (mean ± standard deviation) 34.4 ± 13.7 years, while the remaining 211 female participants had a mean age of 37.5 ± 15.7 years. In the HC group, there were 608 male (42 ± 17.5 years) and 283 female (40 ± 17.4 years) participants. The male HCs were significantly older than the male Lg-CL patients (p < 0.0001).
2.2. Genotype Frequencies and Statistical Comparison of the Genotypes Between Patients with L. guyanensis and Healthy Controls
This case–control study compared unrelated Lg-CL patients to unrelated healthy individuals, following the Strengthening the Reporting of Genetic Association Studies (STREGA) guidelines. The CASP1 gene is located on chromosome 11q22.3 and comprises nine exons, as shown in Figure 1A. The variants rs530537A>G, rs531542C>T, rs531604A>T and rs560880G>T are located in intron 8. The genotype frequencies of the CASP1 variants (rs530537A>G; rs531542C>T; rs531604A>T; rs560880G>T) are presented in Table 1. All four variants studied conformed to the Hardy–Weinberg equilibrium expectation. Based on the success rates of direct nucleotide sequencing, 780, 745, 745 and 725 Lg-CL patients had high-quality genotypic data for the variants rs530537, rs531542, rs531604 and rs560880, respectively. For the HC group, the corresponding success rates were 760, 770, 765 and 752, respectively. The genotype distribution for the four variants did not significantly differ between Lg-CL patients and HCs.
Figure 1B displays the linkage disequilibrium (LD) between the four variants in Lg-CL patients, the HCs and all study participants. The degree of pairwise correlation between the nucleotides was very high, as indicated by the D’ values, suggesting that all four variants are in strong LD. Only 717 Lg-CL patients and 730 HCs had complete genotypic data for the four variants. Six haplotypes were generated using the Haploview software, as shown in Table 2. The GTTT haplotype is associated with resistance to the progression of Lg-CL development. Individuals with the GTTT haplotype have a 19% reduced likelihood of developing CL upon infection with L. guyanensis.
2.3. Genetic Combination of Variant rs530537 of CASP1 and rs2043211 of CARD8
CARD8, a negative regulator of the NLRP3 inflammasome, inhibits the transcription factor NF-ĸB, leading to the reduced expression of IL-1β and TNFα [9,10]. The variant rs2043211 A>T is a transversion from Adenine to Thymine, resulting in a premature stop codon at position ten (Cys10stop codon) [27]. Recent findings from our study population demonstrate that this variant is not associated with the development of Lg-CL [26]. The rs2043211 A>T genotypes (AA, AT and TT) had frequencies of 49%, 42% and 9% among the Lg-CL patients, respectively, and 52%, 41% and 7% among the HCs [26]. The T allele of rs2043211 has been suggested to correlate with increased NFĸB activity [27]. We conducted a combined analysis of the CADR8 variant rs2043211 with CASP1 rs530537, as shown in Table 3. Individuals homozygous for both variants (GG/TT) demonstrated a 38% reduced risk of developing Lg-CL (OR = 0.62 [95%CI: 0.46–0.83]) in comparison to individuals with other combined genotypes.
2.4. CASP1 Variants and Plasma IL-1β Levels
CASP-1 is responsible for processing pro-IL-1β into the active form, IL-1β. We investigated whether the genotypes of the CASP1 variants were associated with differences in the plasma IL-1β levels. Figure 2 illustrates the plasma IL-1β levels across different genotypes. None of the studied variants showed a significant correlation with the plasma IL-1β levels.
We also investigated whether individuals homozygous for genetic combination GG/TT of variants CASP1 rs530537 and CARD8 rs2043211 correlated with the plasma IL-1β levels, given that the rs2043211 TT genotype of CARD8 has been associated with high NF-ĸB activity [27]. As shown in Figure 3A, no significant differences were observed among the combined genotypes, despite the mean plasma IL-1β levels for AG/AT being 1.39 ± SD2.31 compared to AA/TT (0.87 ± 0.87); GG/TT (0.88 ± 1.54); AA/AA (0.94 ± 2.03); and GG/AA (0.74 ± 0.65). Figure 3B compares the plasma IL-1β levels of the GG/TT genotype to those of the other the combined genotypes (REST), showing no significant differences.
3. Discussion
The immune response in leishmaniasis has been extensively studied in animal models, particularly in resistant C57BL/6 and susceptible BALB/c mouse strains infected with L. major, to understand the cellular and molecular mechanisms underlying the disease. BALB/c mice display a Th2-type immune response to L. major, leading to disease development through the production of Th2 cytokines such as IL-4, IL-13 and IL-10, while C57BL/6 mice mount a robust Th1-type response, conferring resistance that depends primarily on the IL-12/IFN-γ/TNF axis [28]. The multiplication of Leishmania parasites is controlled by IFN-γ, which activates macrophages to mediate parasite killing.
In humans, progress has been made in the understanding of the cellular and molecular mechanisms driving leishmaniasis progression, but further research is required to understand why only a subset of individuals in endemic areas develop the disease. Understanding the host predisposition to the development of the disease, particularly in immune response genes, can lead to the development of alternative therapies such as immunotherapy, since chemotherapeutic drugs for leishmaniasis are often toxic and may be ineffective, with treatment success ranging from only 53 to 70% [29]. Indeed, efforts are being made to test plant metabolites that increase reactive oxygen species, which are potent inhibitors of Leishmania multiplication, in vitro in human cells [30]. Recently, two metabolites, deoxyalpinoid B and sulforaphane, have shown promising results in controlling L. donovani and L. infantum multiplication in vitro [31].
This study aimed to contribute to and advance our understanding of the genetic mechanisms underlying disease progression, as leishmaniasis remains a major neglected disease, affecting millions of people worldwide, with approximately 2.4 million disability-adjusted life years (DALYs) lost globally due to CL and VL [32]. Leishmaniasis is the second-leading cause of mortality and fourth-leading cause of morbidity among tropical diseases [33]. CL alone accounts for approximately 41,700 DALYs [34]. Currently, there is no effective vaccine or highly efficacious treatment.
Previous studies have shown that leishmaniasis may be influenced by the genetic factors of the host [35]. Here, we did not observe any association of the four CASP1 variants studied with either susceptibility to or protection against Lg-CL. In Chagas disease, caused by the protozoan Trypanosoma cruzi, another CASP1 variant, rs12417050, located at nucleotide 105065283, was not associated with the disease [36]. However, we showed that a haplotype (GTTT) derived from the four variants located in intron eight of CASP1 is associated with protection against Lg-CL. Furthermore, a combined analysis of the CARD8 variant rs2043211 with CASP1 rs530537 revealed that individuals homozygous for both variants (GG/TT) had a 38% reduced risk of developing Lg-CL (OR = 0.62 [95%CI: 0.46–0.83]) in comparison to individuals with other combined genotypes. CARD8 has been shown to inhibit the major transcription factor controlling innate immunity and inflammation, NF-ĸB [10,11]. The CARD8 variant rs2043211 generates a premature stop codon in the presence of the T allele; interestingly, one study has showed that the TT genotype correlates with the high activity of NF-ĸB in vitro [27]. A plausible explanation is that the loss of inhibition of NF-ĸB activity by CARD8 may provide an advantage in a subset of individuals carrying the combined genotypes GG/TT, generating sufficient proinflammatory cytokines to control the infection.
Several studies have cited a deleterious effect of NLRP3 [15,16,17], while others have indicated a protective role during Leishmania infection [14]. The activation of the NLRP3 inflammasome promotes the activation of CASP-1 to process pro-IL-1β into active IL-1β. The signaling pathway of IL-1β, a strong pro-inflammatory cytokine, subsequently induces more inflammation [37]. Recently, we have shown that IL1B variant genotypes –511T/C (rs16944) and +3954C/T (rs1143634) did not correlate with the plasma IL-1β levels [38]. In animal models, CASP1 knock-out mice have significantly lower levels of IL-1β compared to wild-type mice [39,40]. The reduced expression of the inflammasome adaptor molecule ASC has been shown to limit the proteolytic processing of CASP-1 in murine hepatocytes, leading to low levels of IL-1β and promoting Plasmodium infection [41]. IL-1β upregulates the potent neutrophil chemoattractants CXCL1, CXCL2 and CCL4 [42]. Neutrophils have been shown to promote pathogenesis in murine Leishmania infection [43], while neutropenic mice are resistant to the L. major Seidman strain [15].
In light of these studies, IL-1β regulation must be finely tuned to eliminate the pathogen without contributing to immunopathogenesis. However, in LRV-positive Lg-CL patients, sufficiently high plasma IL-1β levels may provide an advantage, as the virus inhibits CASP-1 to contain parasite replication. In the current study, the plasma IL-1β levels did not correlate with any genotype of the four CASP1 variants studied. Further studies should explore additional variants in the CASP1 gene to better understand the impact of the genetic variation on the plasma IL-1β levels. Notably, other genes in the inflammasome complex are also critical to study, as IL-1β regulation does not depend solely on CASP1. Additionally, a comprehensive genetic analysis of all inflammasome components is essential to elucidate the biological significance of these variants. Indeed, only individuals heterozygous for both variants of CASP1 rs530537 and CARD8 rs2043211 tended to show elevated plasma IL-1β levels.
Our study was limited by the absence of screening for the LRV virus to stratify the patients into LRV-positive and LRV-negative groups, as well as the lack of delayed-type hypersensitivity skin testing with Leishmania antigens in the control group to confirm parasite exposure. However, the LD structure was similar between the patients and HCs, indicating that both groups were drawn from the same population. Another limitation was the lack of a plasma IL-18 assay.
Resistance or susceptibility to pathogens often depends on the genetic make-up of the host in terms of natural immunity. Here, we show that a CASP1 haplotype, as well as combined homozygous CASP1 and CARD8 genotype variants, are associated with protection against the development of Lg-CL.
4. Materials and Methods
4.1. Area of the Study Population
This study was carried out in the surrounding regions of the city of Manaus, the capital of the state of Amazonas, Brazil, near the BR-174 and AM-010. Over the years, there has been significant deforestation due to settlement expansion, agriculture and livestock farming. These activities have transformed these areas into endemic zones for L. guyanensis infection, particularly in the communities of Pau-Rosa, Cooperativa, Água-Branca, Leao, and Brasileirinho. The state of Amazonas is situated in the extreme north of Brazil at latitude −19,9657 and longitude −44,0413. Patients with active CL (presenting with ≤six lesions), affected by L. guyanensis, were recruited between November 2012 and April 2017 at the Fundação de Medicina Tropical Dr. Heitor Vieira Dourado (FMT-HVD), a regional reference center for tropical diseases. The control group consisted of healthy subjects (HCs) from the same endemic area as the patients. These healthy subjects had no history of leishmaniasis and most worked in agriculture, similarly to the patients. A considerable proportion of the study participants reported a history of malaria, as these regions also are endemic for Plasmodium vivax malaria. This population represents an admixture group of Native American ancestry, commonly referred to as caboclo, comprising 50–60% Native American, 40–50% European and approximately 10% African ancestry [44]. This study involved 1714 unrelated individuals (855 patients with Lg-CL and 859 HCs), as previously described [38,45,46,47]. Patients were diagnosed with CL for the first time and presented with six or fewer lesions, with most having a single lesion. All patients had active CL. HCs were not stratified as asymptomatic, as we did not perform the delayed hypersensitivity test for Leishmania antigens. All participants tested negative for HIV and had no history of cardiac, renal or diabetes diseases. These HCs underwent a thorough physical examination by physicians to rule out any doubts regarding their previous history of leishmaniasis. They shared similar socio-epidemiological conditions.
4.2. Sample Size Calculations
The effective sample size was determined using the Genetic Power calculator of Harvard University (
4.3. Ethical Statement
This study adhered to the principles outlined in the Declaration of Helsinki and received approval from the Research Ethics Committee of the Fundação de Medicina Tropical Dr. Heitor Vieira Dourado under the file number CAAE:09995212.0.0000.0005. Written informed consent was obtained from all participants for biological sample collection and subsequent analysis. For participants under 18 years of age, written informed consent was obtained from a parent or guardian.
4.4. Identification of Leishmania Species
All CL patients who had microscopic confirmation of Leishmania parasites in Giemsa-stained lesion scarifications provided biopsy specimens from their skin ulcer lesions for DNA extraction to identify the Leishmania species. Species identification was performed through the direct nucleotide sequencing of a fragment of the HSP70 gene (233bp) and mini exon gene (227 bp) amplified by polymerase chain reaction (PCR). PCR products were purified using 20% PEG, following previously described protocols [38]. Sequencing was performed using an ABI 3130XL automated DNA sequencer (Applied Biosystem, Life technologies, Waltham, MA, USA) with the POP-7 polymer. Nucleotide readings were carried out using the Sequencing Analysis software (Applied Biosystems, v5.3.1). The sequencing was conducted using the BigDye Terminator v3.1 Cycle Sequencing Kit (Thermo Fisher, Waltham, MA, USA), following the manufacturer’s protocols.
4.5. Collection of Biological Samples and DNA Isolation
For genomic DNA isolation using the protein K salting-out method [48] and for the measurement of circulating plasma cytokines, each participant provided five milliliters of peripheral blood, collected into EDTA-containing Vacutainers (Becton Dickinson, Sao Paulo, Brazil) through venipuncture.
4.6. Genotyping of CASP-1 Variants
A pair of primers (CASP1F 5′-CTCTGTATGTAATGACAGACAC-3′ and CASP1R 5′-CCTTACTCCTACCTTCTAGATG-3′) was designed to flank four single-nucleotide variants (SNVs) (rs530537; rs531542; rs531604; rs560880) located in intron 8 of the CASP-1 gene. The forward and reverse primers spanned nucleotide positions 105,027,602 to 105,027,631 and 105,027,930 to 105,027,951, respectively, on chromosome 11q22.3 build Grch38p14, respectively, generating a 349bp fragment. PCR reactions were performed in the Applied Biosystem Veriti Thermal Cycler (Life technologies, Waltham, MA, USA), containing 50 ng of genomic DNA in a final volume of 25 μL with 1.5 mmol/L of MgCl2, 0.25 pmol/L of the forward and reverse primers, 40 μmol/L of each dNTP and 1U of Taq polymerase in buffer containing 100 mmol/L of Tris–HCL (pH 8.3) and 500 mmol/L KCL. The PCR cycling conditions consisted of initial denaturation at 95 °C for 5 min, followed by 35 cycles at 95 °C for 15 s, 15 s at 58 °C, 30 s at 72 °C and a final extension step at 72 °C for 7 min.
The 349bp amplicons were sequenced using either the forward (CASP1F) or reverse (CASP1R) primers with the BigDye Terminator v3.1 Cycle Sequencing Kit (Thermo Fisher, Waltham, MA, USA), according to the manufacturer’s protocol, in the Applied Biosystem Veriti Thermal Cycler, as described above [38]. Only high-quality sequences were used for variant analysis.
4.7. Measurement of Plasma IL-1β Levels by Luminex
Plasma samples were stored at −80 °C until cytokine level measurement. The collection of plasma from patients and controls for cytokine analysis took place between January 2013 and March 2016 and included 354 patients with Lg-CL (264 males and 90 females) and 376 (269 males and 107 females) healthy individuals. Plasma IL-1β levels were measured in 5 µL of plasma with the Human Cytokine Grp I Panel 27-Plex kit (Bio-Rad, Hercules, California. USA) through a multiplex cytokine assay on the Bio-plex 200 Protein Array System (Luminex Corporation, Austin, Texas, USA).
4.8. Statistical Analyses
The statistical comparison of the genotypes between patients with Lg-CL and HCs was performed using logistic regression analysis, calculating the odd ratio (OR) and 95% confidence interval (CI), with adjustments for sex and age, using the R software version 4.3.1 of the SNPassoc package 2.1-0, which also supports different inheritance models (codominant, dominant, recessive and overdominant). Allele frequencies were calculated using the gene counting method, and the Hardy–Weinberg equilibrium was assessed by comparing the observed and expected genotype frequencies. Linkage disequilibrium analysis, haplotype construction and comparisons were conducted using the Haploview software 4.2. The plasma IL-1β levels by genotype were analyzed using a Generalized Linear Model (GLM) for quantitative traits in the R software with the SNPassoc package and visualized using the ggplot2 package. Post hoc analysis following ANOVA was conducted using the postHoc package version 0.1.3 in the R software (
Conceptualization, R.R. and M.V.d.F.G.; methodology, L.S.d.S., L.A.d.S., F.M.M., J.d.E.S.J., H.D.G.S., L.S.F.M., J.d.C.T., S.K.P. and H.K.A.K.; validation, J.L.d.S. and T.G.R.d.M.; formal analysis, J.L.d.S., T.G.R.d.M. and R.R.; investigation, R.R.; resources, R.R.; data curation, R.R. and M.V.d.F.G.; writing—original draft preparation, J.L.d.S., A.S., M.M.O. and J.P.d.M.N.; writing—review and editing, J.L.d.S. and R.R.; supervision, R.R.; project administration, M.L.G.d.S. and T.G.R.d.M.; funding acquisition, R.R. All authors have read and agreed to the published version of the manuscript.
This study adhered to the principles outlined in the Declaration of Helsinki and received approval from the Research Ethics Committee of the Fundação de Medicina Tropical Dr. Heitor Vieira Dourado under the file number CAAE:09995212.0.0000.0005.
Written informed consent was obtained from all participants for biological sample collection and subsequent analysis. For participants under 18 years of age, written informed consent was obtained from a parent or guardian.
All data are included in the manuscript.
The authors thank all of the participants of the study.
The authors declare no conflicts of interest.
Footnotes
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Enlarge this image.Figure 1. Structure of the CASP1 gene as adapted from the NCBI (Build GRCh38.p14) showing the localization of the four variants in (A) and linkage disequilibrium between the variants among patients with cutaneous leishmaniasis (cases), healthy controls and all study participants (total) in (B).
Enlarge this image.Figure 2. Plasma IL-1β levels by genotype (circle—common homozygote, triangle—heterozygote and square—rare homozygote) for the four variants (rs530537, rs531542, rs531604 and rs560880) in patients with Leishmania guyanensis cutaneous leishmaniasis, healthy controls and all participants in the cytokine assay.
Enlarge this image.Figure 3. Plasma IL-1β levels for combined genotypes of variants rs530537 of CASP1 and rs2043211of CARD8 in all participants with cytokine assay, including patients with Leishmania guyanensis cutaneous leishmaniasis and healthy controls (total) (A) and the GG/TT to other combined genotypes (REST) (B).
Distribution of genotypes and statistical comparison of the variants of CASP1 between patients with Leishmania guyanensis cutaneous leishmaniasis and healthy controls.
| Genotype Frequencies | Statistical Comparison of the Genotypes | |||||
|---|---|---|---|---|---|---|
| Cases (%) | HC (%) | Comparison | OR [95% CI] | p-Value | AIC | |
| rs530537 | n = 780 | n = 760 | AA vs. AG | 1.14 [0.89–1.47] | 0.12 | 2072 |
| AA | 215 (27.6) | 179 (23.6) | AA vs. GG | 1.35 [1.01–1.80] | ||
| AG | 384 (49.2) | 380 (50) | AA vs. AG + GG | 1.21 [0.96–1.53] | 0.11 | 2071 |
| GG | 181 (23.2) | 201 (26.4) | AA + AG vs. GG | 1.23 [0.97–1.56] | 0.08 | 2071 |
| AA + GG vs. AG | 1.00 [0.81–1.21] | 0.91 | 2074 | |||
| rs531542 | n = 745 | n = 770 | ||||
| CC | 426 (57.2) | 412 (53.5) | CC vs. CT | 1.10 [0.88–1.37] | 0.20 | 2034 |
| CT | 267 (35.8) | 289 (37.5) | CC vs. TT | 1.41 [0.95–2.10] | ||
| TT | 52 (7) | 69 (9) | CC vs. CT + TT | 1.15 [0.93–1.41] | 0.19 | 2033 |
| CC + CT vs. TT | 1.36 [0.93–2.00] | 0.11 | 2032 | |||
| CC + TT vs. CT | 1.05 [0.85–1.35 | 0.65 | 2035 | |||
| rs531604 | n = 745 | n = 765 | ||||
| AA | 462 (62) | 446 (58.3) | AA vs. AT | 1.17 [0.94–1.46] | 0.25 | 2027 |
| AT | 238 (32) | 264 (34.5) | AA vs. TT | 1.28 [0.84–1.96] | ||
| TT | 45 (6) | 55 (7.2) | AA vs. AT + TT | 1.19 [0.96–1.47] | 0.11 | 2025 |
| AA + AT vs. TT | 1.21 [0.80–1.84] | 0.36 | 2027 | |||
| AA + TT vs. AT | 1.14 [0.92–1.42] | 0.23 | 2026 | |||
| rs560880 | n = 725 | n = 752 | ||||
| GG | 359 (49.5) | 346 (46.1) | GG vs. GT | 1.06 [0.85–2.01] | 0.18 | 1982 |
| GT | 301 (41.5) | 318 (42.3) | GG vs. TT | 1.40 [0.98–2.01] | ||
| TT | 65 (9) | 87 (11.6) | GG vs. GT + TT | 1.12 [0.91–1.39] | 0.27 | 1982 |
| GG + GT vs. TT | 1.36 [0.96–1.93] | 0.07 | 1980 | |||
| GG + TT vs. GT | 1.00 [0.81–1.24] | 0.98 | 1983 | |||
Abbreviations: cases, patients with Leishmania guyanensis cutaneous leishmaniasis; HC, healthy controls; OR, odds ratio; CI, confidence interval; AIC, Akaike Information Criterion.
Distribution of haplotypes in patients with Leishmania guyanensis cutaneous leishmaniasis and healthy controls as derived by Haploview software 4.2.
| Haps | SNV 1 | SNV 2 | SNV 3 | SNV 4 | Cases n 1434 (%) | HC n 1460 (%) | X2 | OR | CI 95% | p-Value |
|---|---|---|---|---|---|---|---|---|---|---|
| 1 | A | C | A | G | 47,5 | 44,5 | 2.4 | 1.12 | 0.97–1.30 | 0.11 |
| 2 | G | C | A | G | 22,6 | 23 | 0.002 | 0.99 | 0.83–1.18 | 0.96 |
| 3 | G | T | T | T | 20,8 | 24,5 | 5.5 | 0.81 | 0.67–0.96 | 0.01 |
| 4 | G | T | A | T | 3,77 | 3,2 | 0.003 | 1.01 | 0.69–1.48 | 0.95 |
| 5 | A | C | A | T | 2,38 | 3,1 | 1.37 | 0.76 | 0.48–1.19 | 0.24 |
| 6 | G | C | A | T | 1,18 | 1,7 | 1.08 | 0.71 | 0.38–1.30 | 0.29 |
Abbreviations: SNV, single-nucleotide variant; 1, rs530537 A>G; 2, rs531542 C>T; 3, rs531504 A>T and n4 rs560880 G>T according to the position in the IL13 gene; χ2, chi-squared test; OR, odds ratio; CI, confidence interval.
Genetic combination of variants rs530537 of CASP1 and rs2043211 of CARD8 in patients with Leishmania guyanensis cutaneous leishmaniasis and healthy controls.
| Combined Genotypes | Patients with Lg-CL | Healthy Controls | Total |
|---|---|---|---|
| Rs530537/rs2043211 | |||
| AA/TT | 106 (14) | 81 (11.5) | 187 (13) |
| AG/TT | 185 (24) | 160 (23) | 345 (23.5) |
| GG/TT | 86 (11.2) | 120 (17) | 206 (14) |
| AA/AT | 83 (10.8) | 73 (10.3) | 156 (10.5) |
| AG/AT | 163 (21.5) | 152 (21.6) | 315 (21.5) |
| GG/AT | 73 (9.5) | 60 (8.5) | 133 (9) |
| AA/AA | 19 (2.5) | 16 (2.3) | 35 (2.5) |
| AG/AA | 29 (4) | 31 (4.5) | 60 (4) |
| GG/AA | 18 (2.5) | 9 (1.3) | 27 (2) |
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Details
; Marcus Vinitius de Farias Guerra 2 ; Tirza Gabrielle Ramos de Mesquita 3
; José do Espírito Santo Junior 4
; Hector David Graterol Sequera 3 ; Lener Santos da Silva 1 ; Larissa Almeida da Silva 5 ; Filipe Menezes Moura 3 ; Lizandra Stephanny Fernandes Menescal 6 ; Júlia da Costa Torres 5 ; Suzana Kanawati Pinheiro 3
; Herllon Karllos Athaydes Kerr 3 ; Ogusku, Mauricio Morishi 7 ; Mara Lúcia Gomes de Souza 5
; Jose Pereira de Moura Neto 6
; Sadahiro, Aya 8 ; Ramasawmy, Rajendranath 9 1 Programa de Pós-Graduação em Biodiversidade e Biotecnologia da Amazonia Legal (Rede Bionorte), Universidade do Estado do Amazonas, Manaus 69055038, Amazonas, Brazil;
2 Fundação de Medicina Tropical Doutor Heitor Vieira Dourado, Manaus 69040000, Amazonas, Brazil
3 Programa de Pós-Graduação em Medicina Tropical, Universidade do Estado do Amazonas, Manaus 69055038, Amazonas, Brazil;
4 Faculdade de Medicina, Universidade Nilton Lins, Manaus 69058030, Amazonas, Brazil;
5 Fundação de Medicina Tropical Doutor Heitor Vieira Dourado, Manaus 69040000, Amazonas, Brazil
6 Programa de Pós-Graduação em Imunologia Básica e Aplicada, Universidade Federal do Amazonas, Manaus 69060001, Amazonas, Brazil;
7 Laboratório de Micobacteriologia, Instituto Nacional de Pesquisas da Amazônia, Manaus 69060001, Amazonas, Brazil;
8 Programa de Pós-Graduação em Imunologia Básica e Aplicada, Universidade Federal do Amazonas, Manaus 69060001, Amazonas, Brazil;
9 Programa de Pós-Graduação em Biodiversidade e Biotecnologia da Amazonia Legal (Rede Bionorte), Universidade do Estado do Amazonas, Manaus 69055038, Amazonas, Brazil;