*Department of Pediatrics & Children's Hospital of Philadelphia & University of Pennsylvania School of Medicine, Philadelphia, Pennsylvania
†Botswana-UPenn Partnership, Gaborone, Botswana
‡Duke University School of Medicine, Durham North Carolina
§Department of Pediatrics, Faculty of Health Sciences, University of the Witwatersrand, Johannesburg, South Africa
‖Department of Medicine, University of Pennsylvania School of Medicine, Philadelphia, Pennsylvania
Funding was provided by the Doris Duke Charitable Foundation (ID: 200787) and in part by the University of Pennsylvania Center for AIDS Research (P30-AI45008). The author A.R. is a recipient of the Clinical Research Fellowship at the University of Pennsylvania from the Doris Duke Charitable Foundation.
Authors' contributions: A.S., A.R., J.P., and R.G. were involved in the study conception, design, implementation, data acquisition and analysis, drafting, and final approval of version to be published. G. B. and C.K. were involved in aiding the study design, article revision and final approval of version to be published. P.P., R.T., M.M., and J.H. were involved in data acquisition, article revision, and final approval of the version to be published.
The authors have no conflicts of interest to disclose.
The funding bodies played no role in the study design; in the collection, analysis and interpretation of data; in the writing of the article; and in the decision to submit the article for publication.
To the Editors:
Tuberculosis (TB) is a leading cause of death in HIV-infected individuals. Efforts to reduce the burden of TB include isoniazid prophylactic therapy for latent TB infection (LTBI).1 However, isoniazid prophylactic therapy confers a risk of hepatotoxicity and requires at least 6 months of therapy.2 Therefore, additional strategies to reduce the burden of active TB are needed.
Vitamin D supplementation may decrease the progression of LTBI to active TB. Primarily, in vitro studies demonstrate that 1,25-dihydroxyvitamin D [1,25(OH)2D] enhances macrophage function, thereby augmenting immunologic control of mycobacteria.3 The benefit of vitamin D supplementation is suggested by the observation of increased rates of clearance of TB from sputum in HIV-uninfected individuals.4
Vitamin D, however, may have detrimental effects in HIV-infected patients. In vitro, vitamin D depresses cell-mediated immune function5 which could hasten progression of LTBI to active disease. This may limit vitamin D supplementation as an adjunct for TB control. Given vitamin D's conflicting mechanisms of action on the immune system, we conducted a study in Botswana, an area of high TB and HIV prevalence,6 to determine if there was evidence for differences in 25-hydroxyvitamin D (25-OHD) levels in HIV-infected individuals with and without active TB. We hypothesized that 25-OHD levels would be significantly lower in HIV-infected individuals with active TB as compared with those without active TB.
STUDY POPULATION AND METHODS
Between December 2008 and January 2010, we conducted a case–control pilot study in Gaborone, Botswana with HIV-infected Batswana subjects recruited from HIV and TB clinics. Participants were at least 21 years old with Bacillus Calmette-Guerin vaccination (documented by medical record or scar) and HIV infection (diagnosed by enzyme-linked immunosorbent assay).
Cases of TB had either active pulmonary or extrapulmonary disease defined as having at least 1 positive sputum smear for acid-fast bacilli, positive culture, or a biopsy consistent with Mycobacterium tuberculosis as documented in the medical records. Cases were also antiretroviral therapy naive and had been receiving antituberculosis therapy (ATT) for less than 21 consecutive days before enrollment, a time period chosen to limit the effect of TB treatment on 25-OHD levels.7 ATT consisted of rifampicin, isoniazid, pyrazinamide, and ethambutol.
Controls were antiretroviral therapy naive without active TB and were assumed to be latently infected. Controls were matched to cases by sex and CD4 counts in a 1:1 ratio to achieve the smallest difference possible in CD4 count between cases and controls, given the strong relationship between CD4 count and active TB in HIV-infected individuals.8
This included CD4 count and treatment history abstracted from clinical records. Serum 25-OHD was measured in 2 separate batches using the Diasorin Liaison assay at the Medical Research Council Mineral Metabolism Research Unit of the University of the Witwatersrand, South Africa, a member of the International Vitamin D External Quality Assessment Program (http://www.deqas.org/).
Comparison of the 25-OHD levels was performed by calculating the difference in 25-OHD levels for each case–control pair, then the median difference and interquartile range among case–control pairs were determined. The overall median (and interquartile range) 25-OHD level for subjects with and without active TB was also calculated.
The study was approved by the Ethics Boards of the Botswana Ministry of Health, Princess Marina Hospital and the University of Pennsylvania.
A total of 43 HIV-infected patients were recruited, and the 25-OHD levels of 38 patients were compared. Nineteen patients with active TB (cases) were matched by CD4 count to 19 patients without TB (controls). A total of 5 patients were not included in the analysis as follows: 2 withdrew from the study, CD4 information could not be located on 1, and 2 patients' data were excluded because the matched control had a CD4 count that was too high (174 cells/mm3) compared with the potential case (10 cells/mm3). The Table 1 displays the characteristics of study subjects and shows the 25-OHD levels and CD4 counts and their respective differences. Of the patients enrolled, 78% (15 of 19) of both cases and controls were vitamin D deficient (<50 nmol/L).
The absolute difference in 25-OHD levels of 1.3 nmol/L between HIV patients with and without TB was not consistent with previous studies of HIV-uninfected individuals. These studies had found significantly lower 25-OHD levels in those with active TB versus those without. For example, 375 HIV-uninfected sub Saharan African immigrants to Australia with TB had a median 25-OHD level of 21.2 nmol/L lower than those without TB.9 A Tanzanian study with 655 subjects found that 25-OHD levels were 7.4 nmol/L lower in patients with sputum culture positive pulmonary TB than in those who were sputum culture negative.10
The absence of such differences in our study may be the lack of a vitamin D immune-enhancing effect in immunocompromised individuals. Conversely, the immunosuppressive effect of vitamin D on cell-mediated immunity may dampen the enhancing effect on innate immunity in the setting of pre-existing T-cell suppression, as is the case with HIV.
Interestingly, there was a global vitamin D deficiency in both cases and controls and perhaps the controls had not yet developed TB because of other differences from the cases. However, current definitions of vitamin D deficiency are based on bone health, not immunological health. Future steps should include ascertaining 25-OHD levels that enhance TB immunity in HIV-infected patients and then evaluating how vitamin D supplementation may optimize immunity against TB.
This study had several limitations. First, the sample size was small, and thus the power to detect small differences was likely low. Yet the very small observed differences suggest that a larger study may not have detected either clinically or statistically significant differences either. Second, 25-OHD levels in cases may have been affected by ATT since rifampicin and isoniazid induce hepatic enzymes to metabolize 25-hydroxyvitamin D, although we mitigated this to some extent by only including individuals with <3 weeks of therapy.7 Third, sun exposure, diet, and nutritional status were not assessed.
The insignificant absolute difference in 25-OHD levels between HIV-infected patients with and without TB was not consistent with similar previous studies of HIV-uninfected individuals. This suggests that hypovitaminosis D is not a major risk factor for TB in HIV-infected as compared with HIV-uninfected individuals. Further studies are needed to explore the role of vitamin D as a factor in the development of TB in HIV-infected individuals.
The authors thank the doctors, nurses, and staff at the Princess Marina Hospital Infectious Disease Care Clinic, Bontleng Clinic, and Old Naledi Clinic who assisted with this study. They also greatly appreciate the extensive assistance of the staff of the Botswana-UPenn Partnership with carrying out this project.
2. Fountain FF, Tolley E, Chrisman CR, et al.. Isoniazid hepatotoxicity associated with reatment of latent tuberculosis infection: a 7-year evaluation from a public health tuberculosis clinic. Chest. 2005;128:116–123.
3. Liu PT, Stenger S, Tang DH, et al.. Vitamin D-mediated human antimicrobial activity against Mycobacterium tuberculosis is dependent on the induction of cathelicidin. J Immunol. 2007;179:2060–2063.
4. Nursyam EW, Amin Z, Rumende CM. The effect of vitamin D as supplementary treatment in patients with moderately advanced pulmonary tuberculous lesion. Acta Med Indones. 2006;38:3–5.
5. Baeke F, Takiishi T, Korf H, et al.. Vitamin D: modulator of the immune system. Curr Opin Pharmacol. 2010;10:482–496.
7. Williams SE, Wardman AG, Taylor GA, et al.. Long term study of the effect of rifampicin and isoniazid on vitamin D metabolism. Tubercle. 1985;66:49–54.
8. Akksilp S, Karnkawinpong O, Wattanaamornkiat W, et al.. Antiretroviral therapy during tuberculosis treatment and marked reduction in death rate of HIV-infected patients in Thailand. Emerg Infect Dis. 2007;13:1001–1007.
9. Gibney KB, MacGregor L, Leder K, et al.. Vitamin D deficiency is associated with tuberculosis and latent tuberculosis infection in immigrants from Sub-Saharan Africa. Clin Infect Dis. 2008;46:443–446.
10. Friis H, Range N, Pedersen ML, et al.. Hypovitaminosis D is common among pulmonary tuberculosis patients in Tanzania but is not explained by the acute phase response. J Nutr. 2008;138:2474–2480.