Preexisting Simian Immunodeficiency Virus Infection Increases Susceptibility to Tuberculosis in Mauritian Cynomolgus Macaques

Abstract

Tuberculosis (TB), caused by Mycobacterium tuberculosis, is the leading cause of death among human immunodeficiency virus (HIV)-positive patients. The precise mechanisms by which HIV impairs host resistance to a subsequent M. tuberculosis infection are unknown. We modeled this coinfection in Mauritian cynomolgus macaques (MCM) using simian immunodeficiency virus (SIV) as an HIV surrogate. We infected seven MCM with SIVmac239 intrarectally and 6 months later coinfected them via bronchoscope with ∼10 CFU of M. tuberculosis Another eight MCM were infected with M. tuberculosis alone. TB progression was monitored by clinical parameters, by culturing bacilli in gastric and bronchoalveolar lavages, and by serial [¹⁸F]fluorodeoxyglucose (FDG) positron emission tomography/computed tomography (PET/CT) imaging. The eight MCM infected with M. tuberculosis alone displayed dichotomous susceptibility to TB, with four animals reaching humane endpoint within 13 weeks and four animals surviving >19 weeks after M. tuberculosis infection. In stark contrast, all seven SIV⁺ animals exhibited rapidly progressive TB following coinfection and all reached humane endpoint by 13 weeks. Serial PET/CT imaging confirmed dichotomous outcomes in MCM infected with M. tuberculosis alone and marked susceptibility to TB in all SIV⁺ MCM. Notably, imaging revealed a significant increase in TB granulomas between 4 and 8 weeks after M. tuberculosis infection in SIV⁺ but not in SIV-naive MCM and implies that SIV impairs the ability of animals to contain M. tuberculosis dissemination. At necropsy, animals with preexisting SIV infection had more overall pathology, increased bacterial loads, and a trend towards more extrapulmonary disease than animals infected with M. tuberculosis alone. We thus developed a tractable MCM model in which to study SIV-M. tuberculosis coinfection and demonstrate that preexisting SIV dramatically diminishes the ability to control M. tuberculosis coinfection.

Keywords: Mauritian cynomolgus macaque; Mycobacterium tuberculosis; PET/CT imaging; bacterial burden; disease pathology; necropsy; simian immunodeficiency virus.

FIG 1

SIV plasma viremia and total CD4⁺ T cell counts are independent of TB disease progression. (A) Plasma SIV viral loads were quantified using quantitative RT-PCR as described in Materials and Methods. (B) CD4⁺ T cell counts were calculated from the CD3⁺ CD4⁺ frequency and the CBC data. Blue, SIV-naive animals; red, SIV⁺ animals. M. tuberculosis infection is designated by the gray bar. For this and subsequent figures, see Table 1 for symbols.

FIG 2

SIV⁺ coinfected MCM reach humane endpoint before SIV-naive animals. MCM were monitored after M. tuberculosis infection for clinical signs of advancing TB. If humane endpoint criteria (see the text) were met, the animal was humanely euthanized and necropsied. All other animals not meeting endpoint criteria were humanely euthanized at the planned endpoint, approximately 5 months after M. tuberculosis infection. No SIV⁺ coinfected MCM (red, open circles) survived to the planned endpoint, while half of the SIV-naive animals (blue, closed circles) did (P = 0.038, log rank test; hazard ratio = 4.0 [95% CI, 1.1 to 15.1]).

FIG 3

FDG PET/CT reveals extent of TB disease at final imaging time point before necropsy. Three-dimensionally rendered images of the final imaging time point for each animal are shown, with SIV-naive MCM on the top row and SIV⁺ coinfected MCM on the bottom row. Animals are ordered left to right by time after M. tuberculosis infection to necropsy, noted in weeks for each animal. As indicated, the final PET/CT scan for two SIV⁺ animals (6016 and 6716) preceded necropsy by 3 weeks due to scanner maintenance. The three SIV-naive MCM that exhibited the least disease survived to the study endpoint (12815, 6216, and 12915). The calibration bar in the lower right correlates color to actual SUV values.

FIG 4

Granuloma numbers increase more dramatically in SIV⁺ MCM between 4 and 8 weeks after M. tuberculosis infection than in SIV-naive MCM. Granulomas observed by PET/CT were enumerated over time after M. tuberculosis infection in SIV-naive animals (A) and SIV⁺ animals (B). (C) Change in the number of granulomas between weeks 4 and 8 (Mann-Whitney test, P = 0.0497).

FIG 5

Lung inflammation following M. tuberculosis infection tends to be higher in SIV⁺ MCM than in SIV-naive MCM. Total FDG avidity reflects FDG uptake in the lungs during PET/CT imaging and correlates with inflammation; avidity was quantified as described in the text. Total FDG avidities following M. tuberculosis infection of SIV-naive (A) and SIV⁺ (B) MCM are shown. (C) Total FDG avidity from the final scan before necropsy (unpaired t test on log₁₀-transformed data, P = 0.173).

FIG 6

SIV⁺ MCM exhibited more gross pathology scores than SIV-naive MCM following M. tuberculosis infection. (A) Overall necropsy scores (P = 0.056). (B) Subscore for the gross pathology score for lungs (P = 0.092). (C) Subscore for the gross pathology of the mediastinal lymph nodes (P = 0.494). (D) Subscore for extrapulmonary pathology (P = 0.119). The lines denote median scores for each group; unpaired t tests were used in all cases.

FIG 7

SIV⁺ MCM exhibited a higher thoracic bacterial burden than SIV-naive MCM. (A) SIV⁺ MCM had a slightly higher total M. tuberculosis burden than SIV-naive MCM (P = 0.054, unpaired t test on log₁₀-transformed data). (B) There was a 3-fold difference in median M. tuberculosis burden in just lungs between SIV⁺ and SIV-naive MCM (P = 0.072, Mann-Whitney test). (C) Lymph nodes from SIV⁺ animals had slightly more M. tuberculosis bacilli (P = 0.063, unpaired t test on log₁₀-transformed data) at necropsy than did those from SIV-naive animals. (D) The percentage of granulomas that yielded culturable M. tuberculosis reflects mycobacterial control within each animal. The medians of the SIV⁺ and SIV-naive groups were similar (P = 0.374, unpaired t test), even though >60% of the granulomas recovered from two SIV-naive animals were sterile.