Clinical exome sequencing unravels the diverse spectrum of genetic heterogeneity and genotype-phenotype correlations in hypertrophic cardiomyopathy.

BACKGROUND: Catalogues of pathogenic genetic mutations in hypertrophic cardiomyopathy (HCM) are disproportionately small when compared to that of the size of the population with South Asian ancestry and their collective increased risk of heart disease. METHODS: We conducted clinical exome sequencing of 200 HCM patients to identified cardiomyopathy-associated genetic mutations. The clinical and echocardiographic characteristics of genotype-positive and genotype-negative patients were compared, and the likelihood of detecting a positive genetic test result was evaluated. Allelic burden analysis was done to compare the minor allele frequencies (MAF) of the pathogenic or likely pathogenic (P/LP) variants and variants of uncertain significance (VUSs) identified in the cohort against various population genomics databases. RESULTS: The genetic yield was 40% for P/LP variants, with MYBPC3 and MYH7 as the predominant sarcomere genes. Younger age-at-diagnosis, family history of HCM, asymmetric hypertrophic (ASH) pattern, the ratio of the interventricular septum to posterior wall thickness (IVS/PW ratio), left atrial (LA) dimensions, severe mitral regurgitation grade (MR grade), late gadolinium enhancement (LGE) detected fibrosis and absence of hypertension were associated with an increased likelihood of HCM-associated variants. Patients who experienced ventricular tachycardia and premature cardiovascular death were significantly likely to carry MYBPC3 or loss-of-function variants. LA and interventricular septal (IVS) dimensions were associated with MYH7 variants. The rare variant burden for P/LP variants and VUSs was significantly enriched in HCM cases compared to population controls. CONCLUSION: Our study provides a comprehensive evaluation of HCM-associated genetic mutations from an Indian population. The identified genotype-phenotype associations could improve the yield of targeted genetic testing in HCM.

Identification of novel endogenous control miRNAs in heart failure for normalization of qPCR data.

MicroRNAs (miRNAs), a class of non-coding RNAs, are utilized as biomarkers for a wide range of disorders. Circulating miRNAs are proposed as potential markers in the clinical identification of heart failure (HF). However, identifying miRNA biomarkers in HF requires identification of robust endogenous control miRNAs for normalization in differential expression analysis. Hence, this study aimed to identify circulating miRNAs that can be utilized as endogenous controls in HF. We evaluated the expression of eight miRNAs, which were previously reported as endogenous controls in different pathological conditions. Total RNA, including miRNA, was extracted from the serum samples of 30 HF patients (15 HFrEF and 15 HFpEF) and their matched controls (n = 15). We used quantitative PCR to determine the miRNA expression. The stability of the selected endogenous miRNAs was assessed and compared using a standard set of criteria with the RefFinder software. Six of the eight miRNAs analyzed showed consistent expression among all sample groups. Stability analysis ranked hsa-let-7i-5p, hsa-miR-148b-3p, and hsa-miR-484 as the most stable miRNAs, indicating their potential as reliable endogenous controls.

Charting a roadmap for heart failure research in India: Insights from a qualitative survey.

Background & objectives: Heart failure (HF) is emerging as a major health problem in India. The profile of HF in India is divergent from elsewhere in the world. While cardiologists must equip themselves with the requisite clinical management tools, scientists and health policymakers would need epidemiological data on HF and information on the resources required to meet the challenges ahead. The aim of this study was to identify the lacunae and to suggest recommendations to improve HF research. Methods: We surveyed a multidisciplinary group of HF experts using a two stage process. An email-based survey was conducted using a structured questionnaire, followed by an online discussion. The experts prioritized the major challenges in convergence research in India and inter-rater agreement values were calculated. In addition, they enlisted potential research gaps and barriers in the domains of epidemiology, diagnostics, management and technology and suggested recommendations to overcome those barriers. Results: The experts identified a paucity of data on HF burden, lack of state-of-the-art diagnostic facilities and trained personnel, overt dependence on imported devices/equipment/reagents, lack of interaction/awareness/information among stakeholders and lack of biobanks, as major barriers in HF research. Three fourths of the experts agreed that lack of interaction among stakeholders was the major challenge with the highest inter-rater agreement in both stages (19 out of 25 and 11 out of 17, respectively). The experts recommended the creation of multidisciplinary taskforces dedicated to population sciences, data sciences, technology development and patient management with short-, intermediate- and long-term strategies. Interpretation & conclusions: The study generated a wish list for advances in HF research and management, and proposed recommendations for facilitating convergence research as a way forward to reduce the burden of HF in India.

Sarcomeric gene variants among Indians with hypertrophic cardiomyopathy: A scoping review.

Hypertrophic cardiomyopathy (HCM) is a genetic heart muscle disease that frequently causes sudden cardiac death (SCD) among young adults. Several pathogenic mutations in genes encoding the cardiac sarcomere have been identified as diagnostic factors for HCM and proposed as prognostic markers for SCD. The objective of this review was to determine the scope of available literature on the variants encoding sarcomere proteins associated with SCD reported among Indian patients with HCM. The eligibility criteria for the scoping review included full text articles that reported the results of genetic screening for sarcomeric gene mutations in HCM patients of Indian south Asian ancestry. We systematically reviewed studies from the databases of Medline, Scopus, Web of Science core collection and Google Scholar. The electronic search strategy included a combination of generic terms related to genetics, disease and population. The protocol of the study was registered with Open Science Framework (https://osf.io/53gde/). A total of 19 articles were identified that reported pathogenic or likely pathogenic (P/LP) variants within MYH7, MYBPC3, TNNT2, TNNI3 and TPM1 genes, that included 16 singletons, one de novo and one digenic mutation (MYH7/ TPM1) associated with SCD among Indian patients. Evidence from functional studies and familial segregation implied a plausible mechanistic role of these P/LP variants in HCM pathology. This scoping review has compiled all the P/LP variants reported to-date among Indian patients and summarized their association with SCD. Single homozygous, de novo and digenic mutations were observed to be associated with severe phenotypes compared to single heterozygous mutations. The abstracted genetic information was updated with reference sequence ID (rsIDs) and compiled into freely accessible HCMvar database, available at https://hcmvar.heartfailure.org.in/. This can be used as a population specific genetic database for reference by clinicians and researchers involved in the identification of diagnostic and prognostic markers for HCM.

Pharmacogenetic variants influence vitamin K anticoagulant dosing in patients with mechanical prosthetic heart valves.

Background: Vitamin K antagonists (VKAs) are class I oral anticoagulants that are widely prescribed following surgical heart valve implantation. The objective of this study was to quantify the relative effects of VKORC1, CYP2C9 and CYP4F2 genotypes in predicting VKA dosing. Materials & methods: A total of 506 South Indian patients with mechanical prosthetic heart valves who were prescribed oral VKAs, such as warfarin or acenocoumarol, were genotyped. The discriminatory ability of mutant genotypes to predict dose categories and bleeding events was assessed using regression analysis. Results: The VKORC1 rs9923231, CYP2C9*3 and CYP4F2*3 mutant genotypes significantly influenced VKA-dose requirements and explained 27.47% of the observed dose variation. Conclusion: These results support pharmacogenetic screening for initial VKA dosing among South Indian patients with mechanical prosthetic heart valves.

1, 25-(OH)2D3 protects against ER stress and miRNA dysregulation in Mus musculus neurons.

BACKGROUND: The pathophysiology of neurodegenerative diseases (NDDs) is closely associated with cellular oxidative stress which can result in the accumulation of toxic proteins in the endoplasmic reticulum (ER) leading to ER stress and subsequent unfolded protein response (UPR) signaling, a mechanism that aggravate these disorders. Vitamin D has been suggested to have important neuroprotective role and its administration has been shown to reduce neuronal injury, neurotoxicity and oxidative stress in various animal systems. OBJECTIVE: The current study was undertaken to examine the effect of vitamin D3 on UPR in ER stress induced Mus musculus neuronal cells. METHODS: Mus musculus cortical and hippocampal primary neuronal cultures were pretreated with 1,25-dihydroxyvitamin D3 (1, 25-(OH)2D3), the active form of vitamin D, followed by ER stress induction with a chemical ER stress inducer thapsigargin and with an advanced glycated protein, AGE-BSA. The UPR genes and related microRNAs (miRNA) expressions were analyzed mainly using real-time PCR. RESULTS: The experiment resulted in the suppression of ER stress marker BiP and UPR pathway genes such as Perk, Ire1α, Chop and Puma which mediate cellular apoptosis indicating the protective effect of 1, 25-(OH)2D3 against neuronal ER stress. Further studies into the molecular aspects showed that ER stress mediated down-regulated expression of microRNAs (miRNAs) such as mmu-miR-24, 27b, 124, 224, 290, 351 and 488 which are known to regulate the UPR pathway genes were also reduced with vitamin pretreatment, of which the miRNAs miR-24 and 27b which shares the same cluster are potentially involved in various human diseases. CONCLUSION: This study emphasizes the therapeutic role of vitamin D in reducing neuronal ER stress and the need for maintaining sufficient amount of this vitamin in our diet.

Prioritizing rs7294 as a mirSNP contributing to warfarin dosing variability.

Aim: The role of mirSNPs in the 3'UTR of VKORC1, CYP2C9 and CYP4F2 genes that could influence warfarin dose variability via a discrete miRNA-mediated mechanism remains unexplained. Methods: Genotypic data in the 1000 Genomes dataset were analyzed for pair-wise linkage disequilibrium and allelic enrichment. Results: MirSNP rs7294 in the 3'UTR of VKORC1 gene displayed varying strengths of linkage disequilibrium with rs9923231 and rs9934438 across populations, albeit consistently associated with higher warfarin dose requirements based on genome-wide association studies, meta-analysis and population-based association studies. In silico analysis predicted altered hybrid stability for the hsa-miR-133a-3p conserved binding site, providing evidence for miRNA-mediated gene regulation. Conclusion: The results support the inclusion of rs7294 as a functional variable for population-specific dosing algorithms to improve dosing accuracy.

Lack of association of mirSNP rs11174811 in AVPR1A gene with arterial blood pressure and hypertension in South Indian population.

Epigenetic regulation of arterial blood pressure mediated through mirSNPs in renin-angiotensin aldosterone system (RAAS) genes is a less explored hypothesis. Recently, the mirSNP rs11174811 in the 3'UTR of the AVPR1A gene was associated with higher arterial blood pressure in a large study population from the Study of Myocardial Infarctions Leiden (SMILE). The aim of the present study was to replicate the association of mirSNP rs11174811 with blood pressure outcomes and hypertension in a south Indian population. Four hundred and fifteen hypertensive cases and 416 normotensive controls were genotyped using a 5' nuclease allelic discrimination assay. Logistic regression was used to test the association of mirSNP rs11174811 with the hypertension phenotype. Censored normal regression was used to test the association of the polymorphism with continuous blood pressure outcomes such as systolic and diastolic blood pressure. The mirSNP rs11174811 did not show any significant association with hypertension. The adjusted odds ratio was 1.02, with 95% CI of 0.72 to 1.45 (p = 0.909). The mean systolic and diastolic blood pressure values were not significantly different across the three genotypic groups, between hypertensives and normotensives, or when stratified by gender. Despite having a similar minor allele frequency (MAF) of 14.5% compared with the SMILE cohort, our results did not support an association of the mirSNP rs11174811 with the hypertension phenotype or with continuous blood pressure outcomes in the south Indian population.

Value of VKORC1 (-1639G>A) rs9923231 genotyping in predicting warfarin dose: A replication study in South Indian population.

OBJECTIVE: Warfarin is the most commonly prescribed oral anticoagulant, although having a narrow therapeutic index and wide interindividual variability. The aim of this study was to replicate the utility of VKORC1 (-1639G>A) rs9923231 genotyping in predicting the mean daily dose and to evaluate its ability to categorize warfarin-treated patients to high-, intermediate-, or low-dose categories in the South Indian population. MATERIALS AND METHODS: A cohort of 222 warfarin-treated patients was genotyped using restriction fragment length polymorphism method. The influence of the rs9923231 polymorphism on the variations in the mean daily dose was compared using one-way analysis of variance and linear regression analysis. Discriminatory ability of the rs9923231 polymorphism to group the patients into ordered dose categories was assessed by estimating the proportional odds ratios using the ordered logit regression analysis. RESULTS: The frequency of AA genotype and A allele in the study sample was found to be 1.8% and 9.23%, respectively, which was similar to reports from other South Indian populations. The mean daily dose required to achieve the optimum international normalized ratio was significantly lower in AA homozygous genotype carriers (3.99 ± 1.67 mg/day) and GA heterozygous (4.26 ± 1.57 mg/day) compared to the GG genotype carriers (5.51 ± 2.13 mg/day), p = 0.003. The A allele carriers (GA+AA genotypes) had a 3.23 higher odds of being grouped as a low-dose requiring category compared to non-carriers (95% CI 1.49-6.98, p = 0.003). CONCLUSIONS: These preliminary results strongly support the use of VKORC1 (-1639G>A) rs9923231 polymorphism for genetically guided initial warfarin dosing in South Indian patients with heart valve replacements.

Comparison of Radial Access, Guided Femoral Access, and Non-Guided Femoral Access Among Women Undergoing Percutaneous Coronary Intervention.

OBJECTIVES: This study was conducted to determine the association between radial access, guided femoral access, and non-guided femoral access on postprocedural bleeding and vascular complications after percutaneous coronary intervention (PCI). BACKGROUND: Bleeding events and major vascular complications after PCI are associated with increased morbidity, mortality, and cost. While the radial approach has been shown to be superior to the femoral approach in reducing bleeding and vascular complications, whether the use of micropuncture, fluoroscopy, or ultrasound mitigates these differences is unknown. METHODS: We conducted a post hoc analysis of women in the SAFE-PCI for Women trial who underwent PCI and had the access method identified (n = 643). The primary endpoint of postprocedure bleeding or vascular complications occurring within 72 hours or at discharge was adjudicated by an independent clinical events committee and was compared based on three categories of access technique: radial, guided femoral (fluoroscopy, micropuncture, ultrasound), or non-guided femoral (none of the aforementioned). Differences between the groups were determined using multivariate logistic regression using radial access as the reference. RESULTS: Of the PCI population, 330 underwent radial access, 228 underwent guided femoral access, and 85 underwent non-guided femoral access. There was a statistically significant lower incidence of the primary endpoint with radial access vs non-guided femoral access; however, there was no significant difference between radial approach and femoral access guided by fluoroscopy, micropuncture, or ultrasound. CONCLUSIONS: This post hoc analysis demonstrates that while radial access is safer than non-guided femoral access, guided femoral access appears to be associated with similar bleeding events or vascular complications as radial access.

Evaluating genomic DNA extraction methods from human whole blood using endpoint and real-time PCR assays.

The extraction of genomic DNA is the crucial first step in large-scale epidemiological studies. Though there are many popular DNA isolation methods from human whole blood, only a few reports have compared their efficiencies using both end-point and real-time PCR assays. Genomic DNA was extracted from coronary artery disease patients using solution-based conventional protocols such as the phenol-chloroform/proteinase-K method and a non-phenolic non-enzymatic Rapid-Method, which were evaluated and compared vis-a-vis a commercially available silica column-based Blood DNA isolation kit. The appropriate method for efficiently extracting relatively pure DNA was assessed based on the total DNA yield, concentration, purity ratios (A260/A280 and A260/A230), spectral profile and agarose gel electrophoresis analysis. The quality of the isolated DNA was further analysed for PCR inhibition using a murine specific ATP1A3 qPCR assay and mtDNA/Y-chromosome ratio determination assay. The suitability of the extracted DNA for downstream applications such as end-point SNP genotyping, was tested using PCR-RFLP analysis of the AGTR1-1166A>C variant, a mirSNP having pharmacogenetic relevance in cardiovascular diseases. Compared to the traditional phenol-chloroform/proteinase-K method, our results indicated the Rapid-Method to be a more suitable protocol for genomic DNA extraction from human whole blood in terms of DNA quantity, quality, safety, processing time and cost. The Rapid-Method, which is based on a simple salting-out procedure, is not only safe and cost-effective, but also has the added advantage of being scaled up to process variable sample volumes, thus enabling it to be applied in large-scale epidemiological studies.

Association of Transforming Growth Factor-ß Superfamily Genes with Non-Regression of Pulmonary Artery Hypertension Following Balloon Mitral Valvotomy: A Pilot Study.

BACKGROUND AND AIM OF THE STUDY: Pulmonary arterial hypertension (PAH) is a common accompaniment of rheumatic mitral stenosis (MS), with 70% of patients showing evidence of different grades of PAH. The latter condition is found to be a prognostic factor influencing disease outcome even after interventional or surgical therapy. The cause of the non-regression of PAH following successful balloon mitral valvotomy (BMV) is not clear. Hence, the study aim was to determine if there is an association of mutations in the genes of the TGF-ß superfamily and non-regression of PAH in patients who undergo a successful BMV. METHODS: Forty-six patients who underwent BMV and fulfilled the recruitment criteria were enrolled prospectively in this case-control study. Among the patients, 27 had non-regression of PAH while 19 had regression of PAH and served as controls. The mean age of the population was 32.63 ± 10.65 years. RESULTS: No statistically significant differences were identified in any of the baseline parameters between the two groups. None of the samples had BMPR2 or ACVRL1 mutations. Ten of the patients and four of the controls were positive for Endoglin mutation, but the inter-group difference was not statistically significant (p = 0.25) CONCLUSIONS: The present study - the first of its kind - showed that deletion-duplication mutations in the BMPR2 or ACVRL1 genes may not be associated with non-regression of PAH, even after successful BMV, or in a wider sense serve as a contributor to PAH in rheumatic MS. The association of Endoglin mutation and non-regression of PAH warrants further investigation in a larger population.

Pathogenesis of intracranial aneurysm is mediated by proinflammatory cytokine TNFA and IFNG and through stochastic regulation of IL10 and TGFB1 by comorbid factors.

BACKGROUND: Intracranial aneurysm (IA) is often asymptomatic until the time of rupture resulting in subarachnoid hemorrhage (SAH).There is no precise biochemical or phenotype marker for diagnosis of aneurysm. Environmental risk factors that associate with IA can result in modifying the effect of inherited genetic factors and thereby increase the susceptibility to SAH. In addition subsequent to aneurismal rupture, the nature and quantum of inflammatory response might be critical for repair. Therefore, genetic liability to inflammatory response caused by polymorphisms in cytokine genes might be the common denominator for gene and environment in the development of aneurysm and complications associated with rupture. METHODS: Functionally relevant polymorphisms in the pro- and anti-inflammatory cytokine genes IL-1 complex (IL1A, IL1B, and IL1RN), TNFA, IFNG, IL3, IL6, IL12B, IL1RN, TGFB1, IL4, and IL10] were screened in radiologically confirmed 220 IA patients and 250 controls from genetically stratified Malayalam-speaking Dravidian ethnic population of south India. Subgroup analyses with genetic and environmental variables were also carried out. RESULTS: Pro-inflammatory cytokines TNFA rs361525, IFNG rs2069718, and anti-inflammatory cytokine IL10 rs1800871 and rs1800872 were found to be significantly associated with IA, independent of epidemiological factors. TGFB1 rs1800469 polymorphism was observed to be associated with IA through co-modifying factors such as hypertension and gender. Functional prediction of all the associated SNPs of TNFA, IL10, and TGFB1 indicates their potential role in transcriptional regulation. Meta-analysis further reiterates that IL1 gene cluster and IL6 were not associated with IA. CONCLUSIONS: The study suggests that chronic exposure to inflammatory response mediated by genetic variants in pro-inflammatory cytokines TNFA and IFNG could be a primary event, while stochastic regulation of IL10 and TGFB1 response mediated by comorbid factors such as hypertension may augment the pathogenesis of IA through vascular matrix degradation. The implication and interaction of these genetic variants under a specific environmental background will help us identify the resultant phenotypic variation in the pathogenesis of intracranial aneurysm. Identifying genetic risk factors for inflammation might also help in understanding and addressing the posttraumatic complications following the aneurismal rupture.

Association of Versican (VCAN) gene polymorphisms rs251124 and rs2287926 (G428D), with intracranial aneurysm.

Intracranial aneurysm (IA) accounts for 85% of Subarachnoid Hemorrhage (SAH) and is mainly caused due to the weakening of arterial wall. The structural integrity of the intracranial arteries is mainly influenced by the extracellular matrix (ECM) remodeling. The Proteoglycan Versican plays an important role in extracellular matrix assembly and plays a major role in the pathogenesis of IA. The linkage studies also indicated VCAN as a putative candidate gene for IA in the 5q22-31 region. Using a case-control study design, we tested the hypothesis whether the variants in VCAN gene, nonsynonymous variants in the coding region of Glycosaminoglycan α (GAG-α) and GAG-ß and two reported SNPs involved in splicing rs251124 and rs173686 can increase the risk of aSAH among South Indian patients, either independently, or by interacting with other risk factors of the disease. We selected 200 radiologically confirmed aneurysmal cases and 250 ethnically, age and sex matched controls from the Dravidian Malayalam speaking population of South India. The present study reiterated the earlier association of rs251124 with intracranial aneurysm (P = 0.0002) and also found a novel association with rs2287926 (G428D) in exon 7 coding for GAG-α with intracranial aneurysm (P = 0.0015). Interestingly, both these SNPs contributed to higher risk for aneurysm in males. In-silico analysis predicted this SNP to have the highest functional relevance in the gene which might have a potentially altered regulatory role in transcription and splicing. Using meta-analysis with available literature rs251124 was found to be the strongest intracranial aneurysm marker for global ethnicities. This study with a novel functional SNP rs2287926 (G428D) further substantiates the potential role of VCAN in the pathogenesis of IA.

Lack of association of lysyl oxidase (LOX) gene polymorphisms with intracranial aneurysm in a south Indian population.

Intracranial aneurysm (IA) accounts for 85 % of haemorrhagic stroke and is mainly caused due to weakening of arterial wall. Lysyl oxidase (LOX) is a cuproenzyme involved in cross linking structural proteins collagen and elastin, thus providing structural stability to artery. Using a case-control study design, we tested the hypothesis whether the variants in LOX gene flanking the two LD block, can increase risk of aSAH among South Indian patients, either independently, or by interacting with other risk factors of the disease. SNPs were genotyped by fluorescence-based competitive allele-specific PCR (KASPar) chemistry. We selected 200 radiologically confirmed aneurysmal cases and 235 ethnically and age and gender matched controls from the Dravidian Malayalam speaking population of South India. We observed marked interethnic differences in the genotype distribution of LOX variants when compared to Japanese and African populations. However, there was no significant association with any of the LOX variants with IA. This study also could not observe any significant role of LOX polymorphisms in influencing IA either directly or indirectly through its confounding factors such as hypertension and gender in South Indian population.

Risk factors for aneurysmal subarachnoid hemorrhage in an Indian population.

BACKGROUND: Aneurysmal subarachnoid hemorrhage (aSAH) has a mortality rate as high as 50%. The prevalence of intracranial aneurysms from various parts of India varies from 0.75 to 10.3%, with higher numbers of cases being diagnosed due to the increasing age of the population and improvements in imaging techniques. However, little is known about the attributable risk factors of aSAH in the Indian population. METHODS: Using a case-control study we estimated the risk of factors such as hypertension, cigarette smoking, alcohol consumption, diabetes mellitus and family history of aSAH in a South Indian population. The population-attributable risk (PAR) of smoking, hypertension and alcohol use was estimated for the South Indian as well as for the general Indian population. RESULTS: Our results showed that cigarette smoking (OR, 3.59; p < 0.001) and a history of hypertension (OR, 2.98; p < 0.001) were significant risk factors associated with aSAH. When patients were classified by gender, it was observed that being a smoker and having hypertension increased the risk for aSAH by nearly fourfold in men. Among women, hypertension and older age were significant risk factors. The PAR estimates indicated that smoking (OR, 3.59; 95% CI, 2.13-6.06) and hypertension (OR, 2.98; 95% CI, 1.73-5.12) are significant risk factors. CONCLUSIONS: Hypertension and smoking may be causal risk factors which might also modify the effect of genetic factors that could increase susceptibility to aSAH in the Indian population. Since these risk factors are amenable to effective modification, our findings will be useful for a gender-specific management of aSAH.

Evidence of association of serotonin transporter gene polymorphisms with schizophrenia in a South Indian population.

Serotonin (5-hydroxytryptamine (5-HT)) transporter (SLC6A4) is known to influence mood, emotion, cognition and efficacy of antidepressants, particularly that of selective serotonin reuptake inhibitors. Atypical antipsychotics exert their effects partially through serotinergic systems, and hence, variation in 5-HT uptake may affect antipsychotic action mediated through the serotinergic system. Therefore, investigating the role of SLC6A4 as a risk factor for developing schizophrenia and treatment response had been a point of concern for many investigators, but with variable outcome. In this study, we examined the genetic roles of five polymorphisms of SLC6A4, including those of the widely studied 44 base pair variable number of tandem repeat (VNTR) in the promoter region of SLC6A4 (the serotonin transporter gene-linked polymorphic region: 5HTTLPR) and a VNTR polymorphism (STin2) in the second intron, in schizophrenia and its influence on the severity of symptoms in a South Indian population from Kerala, comprising 586 individuals. We detected significant allelic and genotypic associations with rs2066713 (both allelic and genotypic P-value <0.001), 5HTTLPR (allelic P-value=0.008 and genotypic P-value=0.03) and STin2 polymorphisms (allelic P-value=0.001 and genotypic P-value=0.002). A haplotype linking these three risk alleles, 5HTTLPR/S-rs2066713/C-STin2/12-repeat (P-value=0.0059), was also significantly associated with disease in our population. Patients with STin2 12-repeat homozygotes showed a greater severity of blunted effect symptom. These results suggest a strong role of SLC6A4 in schizophrenia, possibly with a specific behavioral endophenotype in a South Indian population.