Variations in macrophage migration inhibitory factor gene are not associated with visceral leishmaniasis in India.

BACKGROUND: The host genetic factors play important role in determining the outcome of visceral leishmaniasis (VL). Macrophage migration inhibitory factor (MIF) is an important host cytokine, which is a key regulator of innate immune system. Genetic variants in MIF gene have been found to be associated with several inflammatory and infectious diseases. Role of MIF is well documented in leishmaniasis diseases, including Indian visceral leishmaniasis, where elevated level of serum MIF has been associated with VL phenotypes. However, there was no genetic study to correlate MIF variants in VL, therefore, we aimed to study the possible association of three reported MIF gene variants -794 CATT, -173G > C and non-coding RNA gene LOC284889 in Indian VL phenotype. METHODS: Study subjects comprised of 214 VL patients along with ethnically and demographically matched 220 controls from VL endemic regions of Bihar state in India. RESULTS: We found no significant difference between cases and controls in allelic, genotypic and haplotype frequency of the markers analysed [-794 CATT repeats (χ2=0.86; p=0.35; OR=0.85; 95% CI=0.61-1.19); -173 G>C polymorphism (χ2=1.11; p=0.29; OR=0.83; 95% CI=0.59-1.16); and LOC284889 (χ2=0.78; p=0.37; OR=0.86; 95% CI=0.61-1.20)]. CONCLUSION: Since we did not find any significant differences between case and control groups, we conclude that sequencing of complete MIF gene and extensive study on innate and adaptive immunity genes may help in identifying genetic variations that are associated with VL susceptibility/resistance among Indians.

Identification of the Rare, Four Repeat Allele of IL-4 Intron-3 VNTR Polymorphism in Indian Populations.

BACKGROUND: Cytokines are cell signaling molecules which upon release by cells facilitate the recruitment of immune-modulatory cells towards the sites of inflammation. Genetic variations in cytokine genes are shown to regulate their production and affect the risk of infectious as well as autoimmune diseases. Intron-3 of interleukin-4 gene (IL-4) harbors 70-bp variable number of tandem repeats (VNTR) that may alter the expression level of IL-4 gene. OBJECTIVE: To determine the distribution of IL-4 70-bp VNTR polymorphism in seven genetically heterogeneous populations of Chhattisgarh, India and their comparison with the finding of other Indian and world populations. METHODS: A total of 371 healthy unrelated individuals from 5 caste and 2 tribal populations were included in the present study. The IL-4 70-bp VNTR genotyping was carried out using PCR and electrophoresis. RESULTS: Overall, 3 alleles of IL-4 70-bp VNTR (a2, a3 and a4) were detected. The results demonstrated the variability of the IL-4 70-bp VNTR polymorphism in Chhattisgarh populations. Allele a3 was the most common allele at the 70-bp VNTR locus in all populations followed by a2 allele. This study reports the presence four repeat allele a4 at a low frequency in the majority of the Chhattisgarh populations studied. Further, the frequency of the minor allele (a2) in Chhattisgarh populations showed similarity with the frequencies of European populations but not with the East Asian populations where the a2 allele is a major allele. CONCLUSIONS: Our study provides a baseline for future research into the role of the IL-4 locus in diseases linked to inflammation in Indian populations.

Mannose-binding Lectin (MBL) as a susceptible host factor influencing Indian Visceral Leishmaniasis.

Visceral Leishmaniasis (VL), caused by Leishmania donovani is endemic in the Indian sub-continent. Mannose-binding Lectin (MBL) is a complement lectin protein that binds to the surface of Leishmania promastigotes and results in activation of the complement lectin cascade. We utilized samples of 218 VL patients and 215 healthy controls from an Indian population. MBL2 functional variants were genotyped and the circulating MBL serum levels were measured. MBL serum levels were elevated in patients compared to the healthy controls (adjusted P=0.007). The MBL2 promoter variants -78C/T and +4P/Q were significantly associated with relative protection to VL (-78C/T, OR=0.7, 95% CI=0.5-0.96, adjusted P=0.026 and +4P/Q, OR=0.66, 95% CI=0.48-0.9, adjusted P=0.012). MBL2*LYQA haplotypes occurred frequently among controls (OR=0.69, 95% CI=0.5-0.97, adjusted P=0.034). MBL recognizes Leishmania and plays a relative role in establishing L. donovani infection and subsequent disease progression. In conclusion, MBL2 functional variants were associated with VL.

Analysis of genetic variants in the IL4 promoter and VNTR loci in Indian patients with Visceral Leishmaniasis.

Visceral Leishmaniasis (VL) is the most severest form of Leishmaniasis and resistance to infection is mediated by cellular immune responses. Interleukin 4 (IL-4) orchestrates of Th2 and Th1 immune responses during infections. In this study, we aimed to investigate possible association between three functional IL-4 polymorphisms -590C/T (rs2243250), -34C/T (rs2070874) and 70bp VNTR (rs79071878 in intron3) with VL in an Indian cohort comprising of 197 VL patients and 193 healthy controls. The three investigated IL-4 polymorphisms were in strong linkage disequilibrium. The investigated IL-4 alleles, genotypes and the reconstructed haplotypes were not significantly distributed between the VL patients and healthy controls. Our study signifies no possible association of functional IL-4 polymorphisms with Indian VL and postulate other vital genes involved in the IL-4 pathway may provide genetic clues to elucidate of IL-4 regulation and immune-pathogenesis during VL.

MBL2 variations and malaria susceptibility in Indian populations.

Human mannose-binding lectin (MBL) encoded by the MBL2 gene is a pattern recognition protein and has been associated with many infectious diseases, including malaria. We sought to investigate the contribution of functional MBL2 gene variations to Plasmodium falciparum malaria in well-defined cases and in matched controls. We resequenced the 8.7 kb of the entire MBL2 gene in 434 individuals clinically classified with malaria from regions of India where malaria is endemic. The study cohort included 176 patients with severe malaria, 101 patients with mild malaria, and 157 ethnically matched asymptomatic individuals. In addition, 830 individuals from 32 socially, linguistically, and geographically diverse endogamous populations of India were investigated for the distribution of functional MBL2 variants. The MBL2 -221C (X) allelic variant is associated with increased risk of malaria (mild malaria odds ratio [OR] = 1.9, corrected P value [P(Corr)] = 0.0036; severe malaria OR = 1.6, P(Corr) = 0.02). The exon1 variants MBL2*B (severe malaria OR = 2.1, P(Corr) = 0.036; mild versus severe malaria OR = 2.5, P(Corr) = 0.039) and MBL2*C (mild versus severe malaria OR = 5.4, P(Corr) = 0.045) increased the odds of having malaria. The exon1 MBL2*D/*B/*C variant increased the risk for severe malaria (OR = 3.4, P(Corr) = 0.000045). The frequencies of low MBL haplotypes were significantly higher in severe malaria (14.2%) compared to mild malaria (7.9%) and asymptomatic (3.8%). The MBL2*LYPA haplotypes confer protection, whereas MBL2*LXPA increases the malaria risk. Our findings in Indian populations demonstrate that MBL2 functional variants are strongly associated with malaria and infection severity.

Relic excavated in western India is probably of Georgian Queen Ketevan.

History has well documented the execution of Queen Ketevan of Georgia by the Persian Emperor of modern day Iran. Based on historical records, in 1624 two Augustinian friars unearthed the queen's remains and one of them brought the relic to the St. Augustine convent in Goa, India. We carried out ancient DNA analysis on the human bone remains excavated from the St. Augustine convent by sequencing and genotyping of the mitochondrial DNA. The investigations of the remains revealed a unique mtDNA haplogroup U1b, which is absent in India, but present in Georgia and surrounding regions. Since our genetic analysis corroborates archaeological and literary evidence, it is likely that the excavated bone belongs to Queen Ketevan of Georgia.

LRRK2 and RIPK2 variants in the NOD 2-mediated signaling pathway are associated with susceptibility to Mycobacterium leprae in Indian populations.

In recent years, genome wide association studies have discovered a large number of gene loci that play a functional role in innate and adaptive immune pathways associated with leprosy susceptibility. The immunological control of intracellular bacteria M. leprae is modulated by NOD2-mediated signaling of Th1 responses. In this study, we investigated 211 clinically classified leprosy patients and 230 ethnically matched controls in Indian population by genotyping four variants in NOD2 (rs9302752A/G), LRRK2 (rs1873613A/G), RIPK2 (rs40457A/G and rs42490G/A). The LRRK2 locus is associated with leprosy outcome. The LRRK2 rs1873613A minor allele and respective rs1873613AA genotypes were significantly associated with an increased risk whereas the LRRK2 rs1873613G major allele and rs1873613GG genotypes confer protection in paucibacillary and leprosy patients. The reconstructed GA haplotypes from RIPK2 rs40457A/G and rs42490G/A variants was observed to contribute towards increased risk whereas haplotypes AA was observed to confer protective role. Our results indicate that a possible shared mechanisms underlying the development of these two clinical forms of the disease as hypothesized. Our findings confirm and validates the role of gene variants involved in NOD2-mediated signalling pathways that play a role in immunological control of intracellular bacteria M. leprae.

A rare non-synonymous c.102C>G SNP in the IFNB1 gene might be a risk factor for cerebral malaria in Indian populations.

Interferon beta1 (IFNB1) is a type I interferon that is mainly known for its antiviral activity, but it also regulates a number of anti-inflammatory and immunomodulatory functions. Studies on mouse models of cerebral malaria have established that IFNB1 regulates severe malaria pathogenesis and increases overall survival against malaria. It down-regulates pro-inflammatory cytokines: TNF, IFNG and ICAM-1, resulting in decreased adherence of Plasmodium falciparum parasitized RBC to capillary wall, entry into the brain and delayed onset of death. Therefore, we hypothesized that variations in IFNB1 gene could regulate malarial pathogenesis. We re-sequenced the complete IFNB1 gene along with 900bp of 5' up-stream and 500bp of 3'-UTR in 437 individuals from malaria endemic regions of the Orissa and Chhattisgarh states of India. The subjects comprised of 173 cases of severe malaria, 101 of mild malaria, and 156 ethnically matched asymptomatic controls. Data were statistically compared between cases and controls for their possible association with P. falciparum malarial outcome. Two single nucleotide polymorphisms (SNPs): a synonymous c.153C>T (rs1051922) and a non-synonymous substitution c.102C>G (rs139262191, p.Ser34Arg) were identified. The genotype and allele distribution of c.153C>T did not differ significantly between the study groups [mild, χ(2)2=4.10, p-value<0.13 and severe χ(2)2=0.06, p-value<0.97]. Interestingly, the rare non-synonymous SNP (rs139262191) was observed only in malaria patients. The differences between all cases and controls did not reach statistical significance, however, a statistically significant difference was observed between the asymptomatic control group and the cerebral malaria group [OR=20.32, 95% CI=1.08-382.63, p-value=0.044]. Moreover, the genotypes between cerebral malaria positive and negative groups were not significantly different [OR=5.58, 95% CI=0.61-50.97, p-value=0.123]. Our findings suggest that the IFNB1 variant, p.Ser34Arg, might be a risk factor for cerebral malaria in Indian populations.

IL-4 haplotype -590T, -34T and intron-3 VNTR R2 is associated with reduced malaria risk among ancestral indian tribal populations.

BACKGROUND: Interleukin 4 (IL-4) is an anti-inflammatory cytokine, which regulates balance between T(H)1 and T(H)2 immune response, immunoglobulin class switching and humoral immunity. Polymorphisms in this gene have been reported to affect the risk of infectious and autoimmune diseases. METHODS: We have analyzed three regulatory IL-4 polymorphisms; -590C>T, -34C>T and 70 bp intron-3 VNTR, in 4216 individuals; including: (1) 430 ethnically matched case-control groups (173 severe malaria, 101 mild malaria and 156 asymptomatic); (2) 3452 individuals from 76 linguistically and geographically distinct endogamous populations of India, and (3) 334 individuals with different ancestry from outside India (84 Brazilian, 104 Syrian, and 146 Vietnamese). RESULTS: The -590T, -34T and intron-3 VNTR R2 alleles were found to be associated with reduced malaria risk (P<0.001 for -590C>T and -34C>T, and P = 0.003 for VNTR). These three alleles were in strong LD (r²>0.75) and the TTR2 (-590T, -34T and intron-3 VNTR R2) haplotype appeared to be a susceptibility factor for malaria (P = 0.009, OR = 0.552, 95% CI = 0.356 -0.854). Allele and genotype frequencies differ significantly between caste, nomadic, tribe and ancestral tribal populations (ATP). The distribution of protective haplotype TTR2 was found to be significant (χ²3 =  182.95, p-value <0.001), which is highest in ATP (40.5%); intermediate in tribes (33%); and lowest in caste (17.8%) and nomadic (21.6%). CONCLUSIONS: Our study suggests that the IL-4 polymorphisms regulate host susceptibility to malaria and disease progression. TTR2 haplotype, which gives protection against malaria, is high among ATPs. Since they inhabited in isolation and mainly practice hunter-gatherer lifestyles and exposed to various parasites, IL-4 TTR2 haplotype might be under positive selection.