Association of Vitamin D Receptor Gene Polymorphism (FokI) with Susceptibility to Musculoskeletal Tuberculosis: A Case–Control Study from Central India

Introduction

Materials and Methods

This study was approved by the Institutional Human Ethics Committee (IHEC) of the institution (IHEC Ref Number: 2020/PG/July/30) on February 24, 2021. All participants provided written informed consent before enrolment in the study. This research was conducted ethically in accordance with the World Medical Association Declaration of Helsinki. Participants were recruited from the outdoor and indoor facilities of the orthopaedics department of a tertiary institute in Central India from February 2021 to December 2023. All patients with MSKTB confirmed by histopathology or molecular diagnostic methods (e.g., Cartridge-Based Nucleic Acid Amplification Test) were included, excluding those who did not provide consent. Controls were recruited from admitted patients of the orthopedics ward with no current clinical features or history of TB.

Demographic information of cases and controls was collected in a standardized data collection form. Age, gender, and residence were recorded. Body weight and height were recorded for the calculation of body mass index (BMI), level of education, occupation, and monthly family income to determine socioeconomic status (Modified Kuppuswamy Socioeconomic Scale, 2020), and site of involvement.

Results

Demographic profile of MSKTB patients is shown in Table 1. Among the cases (n = 110), the majority were females (61.8%). Most patients belonged to the age group of 41–60 years (40.9%). A large proportion of cases were from lower socioeconomic strata, particularly the lower middle class (36.4%) and rural backgrounds (61.8%). In addition, 30% of patients were underweight (BMI <18.5 kg/m2).

The distribution of VDR FokI polymorphism genotypes among cases and controls is shown in Table 2. Among 110 cases of MSKTB, 45 (40.9%) had the FF genotype, 46 (41.8%) had the Ff genotype, and 19 (17.3%) had the ff genotype. In comparison, among 112 controls, 60 (53.6%) had the FF genotype, 41 (36.6%) had the Ff genotype, and 11 (9.8%) had the ff genotype.

The spine was the most frequently affected site, with other commonly involved locations including the knee, distal femur, hip joint, and sacroiliac joint.

The frequency of the mutant f allele was higher in cases (38.2%) compared to controls (28.1%). Using the FF genotype as the reference, the Ff genotype showed a non-significant increase in odds of MSKTB (OR = 1.49, P = 0.17). In contrast, the ff genotype was associated with significantly increased odds (OR = 2.30, 95% CI: 1.03–5.15, P = 0.04).

In the dominant model (Ff + ff vs. FF), the OR was 1.67 (P = 0.06), whereas in the recessive model (ff vs. Ff + FF), the OR was 1.92 (P = 0.10). Allelic analysis demonstrated that the f allele was significantly associated with MSKTB (OR = 1.58, 95% CI: 1.05–2.37, P = 0.023).

Discussion

This case–control study evaluated the demographic profile and the FokI polymorphism of the VDR gene in 110 patients with MSKTB and 112 controls. The demographic data revealed that most patients were females, belonged to rural backgrounds, and were from lower socioeconomic strata. A significant proportion were also underweight. These findings reinforce the well-established concept that TB is not only a biological disease but also strongly influenced by social determinants such as nutrition, living conditions, and access to healthcare.

With respect to the site of involvement, the spine was the most affected site, followed by the knee, distal femur, sacroiliac joint, hip, and other joints. This pattern of distribution is largely similar to previously published studies on MSKTB ^{[15, 16]}.

On molecular evaluation, our findings demonstrate that the mutant f allele was significantly more frequent among cases than controls, with the ff genotype showing a significant association with MSKTB. This suggests that the FokI polymorphism of the VDR gene may contribute to increased susceptibility to MSKTB.

Similar findings have been reported by Panda et al. ^[17] in patients with pulmonary TB, where the ff genotype was significantly associated with the disease. Further, various meta-analyses including genetic studies in pulmonary and extra-pulmonary TB patients also showed a significant correlation with the polymorphic allele (ff) and recessive model (ff vs. Ff+FF) of FokI polymorphism ^{[18, 19, 20]}

A similar study conducted at All India Institute of Medical Sciences Delhi involving 150 patients with active pulmonary TB, 150 TB contacts, and 150 healthy controls reported comparable findings, demonstrating a higher frequency of the “ff” genotype among patients, whereas the “FF” genotype was significantly more prevalent in the control group (χ2 = 25.22, df of 4, and P = 0.0001) ^[17].

Interestingly, while the ff genotype showed a statistically significant association with MSKTB in our study, the dominant (P = 0.06) and recessive (P = 0.10) genetic models did not reach statistical significance, although a trend toward increased risk was observed. Given the relatively small cell sizes underlying the genotypic comparison (19 ff cases vs. 11 ff controls), this inconsistency across models means the precise mode of inheritance cannot be confirmed from the present data, and the genotypic and allelic associations should be interpreted with appropriate caution pending replication in larger cohorts. No correction for multiple comparisons was applied across the genotype, allele, and genetic-model analyses; under a conservative Bonferroni-adjusted threshold, neither association would retain statistical significance, and these findings should therefore be regarded as preliminary rather than definitive. Our study has several important limitations. As the research was carried out in a single tertiary care institution and included a modest number of participants, the study was not powered a priori for genotype-model or site-stratified comparisons, and the findings may not be representative of the wider population; given known regional and ethnic variation in VDR allele frequencies across India, formal assessment of population stratification (e.g. ethnicity) was also not performed, though cases and controls were drawn from the same hospital and control genotypes conformed to Hardy–Weinberg equilibrium. Larger multicentric studies involving diverse populations are required to further validate these observations. Furthermore, only a single polymorphism of the VDR gene was studied, whereas susceptibility to TB is likely influenced by multiple genetic and environmental factors, including other VDR polymorphisms (BsmI, ApaI, TaqI) and non-VDR susceptibility loci. Genotype–disease associations were assessed using unadjusted ORs; potential confounders such as age, sex, BMI, and socioeconomic status were not incorporated into a multivariable model, and several established TB risk factors – household TB contact, Bacille Calmette-Guérin vaccination status, HIV status, diabetes mellitus, smoking, and immunosuppressive drug use – were not systematically recorded, so residual confounding from these factors cannot be excluded. Controls were recruited from hospitalized orthopedic patients rather than a community-based healthy sample; while this minimized differences in healthcare access and documentation quality, it introduces the possibility of selection bias if any control-group orthopedic diagnoses are themselves associated with VDR genotype. Serum 25-hydroxyvitamin D levels were not measured, and no functional data (VDR expression, cathelicidin induction, or monocyte/macrophage antimycobacterial activity) were generated in our cohort; consequently, this study demonstrates a statistical association but does not establish the underlying biological mechanism, nor does it demonstrate causality, which cannot be inferred from a cross-sectional case–control design. Genotyping relied on PCR-Restriction Fragment Length Polymorphism without independent confirmation by an orthogonal method such as Sanger sequencing in a subset of samples. MSKTB was analyzed as a composite entity across multiple anatomical sites; due to small numbers at individual non-spinal sites, site-specific genetic associations could not be evaluated. Finally, this study did not include a pulmonary TB comparator arm, and therefore cannot determine whether the FokI association reflects a general susceptibility to M. TB infection or a susceptibility more specific to musculoskeletal or bone involvement, given Vitamin D’s additional independent role in bone metabolism.

In our literature search, we did not find any studies evaluating VDR gene polymorphisms exclusively in patients with MSKTB. This study contributes to the limited body of literature on genetic susceptibility to MSKTB, particularly in the Indian context.

Conclusion

This case–control study identifies a statistical association between the FokI polymorphism of the VDR gene and susceptibility to MSKTB, though the modest sample size, lack of correction for multiple comparisons, and other limitations discussed above mean these findings should be regarded as preliminary and hypothesis-generating rather than definitive. The mutant f allele and ff genotype were more frequent among patients with MSKTB compared with healthy controls; however, as a cross-sectional case–control design, this study can establish association only, not causation, and does not on its own demonstrate a functional or mechanistic link between the FokI variant and impaired host defense against Mycobacterium TB. The findings also highlight the influence of demographic and socioeconomic factors, including rural residence, lower socioeconomic status, and undernutrition, in the occurrence of MSKTB, underscoring that MSKTB susceptibility is likely multifactorial rather than attributable to a single genetic locus. To the best of our knowledge, this is among the very few studies to evaluate VDR FokI polymorphism exclusively in patients with MSKTB. Confirmation in larger, multicentric, multi-ethnic cohorts – incorporating multivariable adjustment for confounders, measurement of serum 25-hydroxyvitamin D and VDR expression, assessment of additional VDR and non-VDR genetic markers, and, where feasible, a pulmonary TB comparator arm – is warranted before these findings can inform clinical risk stratification or screening strategies for MSKTB.

Clinical Message

VDR FokI genotyping is not yet ready for routine clinical use, but our findings add to the evidence that impaired Vitamin D receptor signaling may influence susceptibility to MSKTB. Until larger, functionally validated, multicentric studies are available, VDR genotype should be regarded as a research tool rather than a diagnostic or prognostic marker for individual patient management in everyday practice.

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About the Authors

• 

  Dr. Hem Raj Gujar

• 

  Dr. Rajesh Ratan Goyal

• 

  Dr. Virendra Kr. Verma

• 

  Dr. Prashant Chaware

• 

  Dr. John Ashutosh Santoshi

• 

  Dr. Tharun Teja Aduri