Experimental model of tuberculosis in the domestic goat after endobronchial infection with Mycobacterium caprae

Abstract

Caprine tuberculosis (TB) has increased in recent years, highlighting the need to address the problem the infection poses in goats. Moreover, goats may represent a cheaper alternative for testing of prototype vaccines in large ruminants and humans. With this aim, a Mycobacterium caprae infection model has been developed in goats. Eleven 6-month-old goats were infected by the endobronchial route with 1.5 × 10(3) CFU, and two other goats were kept as noninfected controls. The animals were monitored for clinical and immunological parameters throughout the experiment. After 14 weeks, the goats were euthanized, and detailed postmortem analysis of lung lesions was performed by multidetector computed tomography (MDCT) and direct observation. The respiratory lymph nodes were also evaluated and cultured for bacteriological analysis. All infected animals were positive in a single intradermal comparative cervical tuberculin (SICCT) test at 12 weeks postinfection (p.i.). Gamma interferon (IFN-γ) antigen-specific responses were detected from 4 weeks p.i. until the end of the experiment. The humoral response to MPB83 was especially strong at 14 weeks p.i. (13 days after SICCT boost). All infected animals presented severe TB lesions in the lungs and associated lymph nodes. M. caprae was recovered from pulmonary lymph nodes in all inoculated goats. MDCT allowed a precise quantitative measure of TB lesions. Lesions in goats induced by M. caprae appeared to be more severe than those induced in cattle by M. bovis over a similar period of time. The present work proposes a reliable new experimental animal model for a better understanding of caprine tuberculosis and future development of vaccine trials in this and other species.

Fig. 1.

Kinetics of IFN-γ responses in infected goats. The release of IFN-γ was measured by ELISA after in vitro stimulation of whole blood with different antigens. The results are expressed as mean ΔOD₄₅₀ responses with 95% CI. The dashed horizontal line is the cutoff point for positivity. (A) PPD-B and PPD-A. **, P < 0.005; significant differences were determined by a nonparametric Mann-Witney-Wilcoxon test. (B) PPD-B, ESAT-6/CFP-10, and Rv3615c. *, P < 0.05; significant differences were determined by a nonparametric Friedman test with a post hoc Mann-Whitney-Wilcoxon test.

Fig. 2.

Humoral responses to MBP83 and PPD-B at 14 weeks postinfection. Shown are the OD₄₅₀ absorbances of total IgG to MPB83 and PPD-B from individual goats infected or not with M. caprae. The results are expressed as ΔOD₄₅₀ (OD₄₅₀ of antigen-stimulated wells minus OD₄₅₀ of nonstimulated wells). Filled symbols, infected animals; open symbols, noninfected control animals; dashed horizontal line, cutoff point for positivity.

Fig. 3.

Gross pathology analysis of a goat case. (A) MDTC-3D representation of the whole lung after excluding air (H, head; F, foot, L, left, R, right). The total volume of the lung was calculated in cm³ and is shown in the red dashed box at the bottom. (B) Volume-rendering image of the lung showing different tissue densities discriminated by color: water, gray; air, black; calcium, white. The volume of affected lung is also shown. (C) Pathological areas identified by segmentation in axial (top; A, anterior; P, posterior), coronal, and sagittal (bottom) planes. (D) Formalin-fixed, 5-mm sections of the left diaphragmatic lobe showing a large cavitary lesion. Cranial to caudal sections are represented as bottom up and left-right. Bar = 3 cm.

Fig. 4.

Correlation between the volumes of lesions in lungs measured by two quantitative methods. Visible lesions in lungs were calculated by MDCT and by image analysis of photographs of lung sections (direct observation) in infected goats (n = 11). Statistical analysis was conducted with a nonparametric Spearman rank test.