Low Vitamin-D Levels Combined with PKP3-SIGIRR-TMEM16J Host Variants Is Associated with Tuberculosis and Death in HIV-Infected and -Exposed Infants

Abstract

Background: This study examined the associations of 25-hydroxyvitamin D and specific host genetic variants that affect vitamin D levels or its effects on immune function, with the risk of TB or mortality in children.

Methods: A case-cohort sample of 466 South African infants enrolled in P1041 trial (NCT00080119) underwent 25-hydroxyvitamin D testing by chemiluminescent immunoassay. Single nucleotide polymorphisms (SNPs) that alter the effect of vitamin D [e.g. vitamin D receptor (VDR)], vitamin D levels [e.g. vitamin D binding protein (VDBP)], or toll like receptor (TLR) expression (SIGIRR including adjacent genes PKP3 and TMEM16J) were identified by real-time PCR. Outcomes were time to TB, and to the composite of TB or death by 192 weeks of follow-up. Effect modification between vitamin D status and SNPs for outcomes was assessed.

Findings: Median age at 25-hydroxyvitamin D determination was 8 months; 11% were breastfed, 51% were HIV-infected and 26% had low 25-hydroxyvitamin D (<32ng/mL). By 192 weeks, 138 incident TB cases (43 definite/probable, and 95 possible) and 26 deaths occurred. Adjusting for HIV status and potential confounders, low 25-hydroxyvitamin D was associated with any TB (adjusted hazard ratio [aHR] 1.76, 95% CI 1.01-3.05; p = 0.046) and any TB or death (aHR 1.76, 95% CI 1.03-3.00; p = 0.038). Children with low 25-hydroxyvitamin D and TMEM 16J rs7111432-AA or PKP3 rs10902158-GG were at increased risk for probable/definite TB or death (aHR 8.12 and 4.83, p<0.05) and any TB or death (aHR 4.78 and 3.26, p<0.005) respectively; SNPs in VDBP, VDR, and vitamin D precursor or hydroxylation genes were not. There was significant interaction between low 25-hydroxyvitamin D and, TMEM 16J rs7111432-AA (p = 0.04) and PKP3 rs10902158-GG (p = 0.02) SNPs.

Conclusions: Two novel SNPs, thought to be associated with innate immunity, in combination with low vitamin D levels were identified as increasing a young child's risk of developing TB disease or death. Identifying high-risk children and providing targeted interventions such as vitamin D supplementation may be beneficial.

Trial registration: ClinicalTrials.gov NCT00080119.

Fig 1. Consort diagram for case-cohort study.

Fig 2. Cox Proportional Hazard Models for TB disease or death, looking at (A) low vitamin D (LVD) status as risk factor ignoring genetic variant, (B) TMEM16J rs7111432 variant as risk factor ignoring LVD status, and (C) PKP3 rs10902158 variant as risk factor ignoring LVD status.

Model 1 adjusts for HIV infection status. Model 2 adjusts for HIV status, season, site, sex, weight z-score, type of house, mother prev TB diagnosis, age (in months) at plasma draw date, and INH/Placebo group. Model 3 adjusts for HIV status, site, sex, weight z-score, type of house, mother prev TB diagnosis, and INH/Placebo group. Evts/LVD = no. of events/no. of subjects with low vitamin D (%). Evts/NotLVD = no. of events/no. of subjects without low vitamin D (%). Evts/AA = no. of events/no. of subjects with AA genotype (%). Evts/NotAA = no. of events/no. of subjects without AA genotype (%). Evts/GG = no. of events/no. of subjects with GG genotype (%). Evts/NotGG = no. of events/no. of subjects without GG genotype (%).

Fig 3. Kaplan-Meier Plots on Tuberculosis Free Survival (from all available data in case-cohort) by (A) Vitamin D status, (B) Vitamin D status / rs7111432 variant and (C) Vitamin D status / rs10902158 variant.

Risk differences were computed as the differences in Kaplan-Meier survival estimates at end of follow-up period for this analysis, and were not adjusted for covariates.

Fig 4. Cox Proportional Hazard Models for TB disease or death, looking at (A) combination of low Vitamin D (LVD) status and TMEM 16J rs7111432 variant [AA genotype or not] as risk factor, and (B) combination of LVD status and PKP3 rs10902158 variant [GG genotype or not] as risk factor.

A: Model adjusts for HIV status, season, site, sex, weight z-score, type of house, mother with previous TB diagnosis, age (in months) at plasma draw date, and INH/Placebo group. B: Model adjusts for HIV status, season, site, sex, weight z-score, type of house, mother with previous TB diagnosis, age (in months) at plasma draw date, and INH/Placebo group.