Safety and Immunogenicity of Recombinant Bacille Calmette-Guérin Strain VPM1002 and Its Derivatives in a Goat Model.

A more effective vaccine against tuberculosis than Bacille Calmette-Guérin (BCG) is urgently needed. BCG derived recombinant VPM1002 has been found to be more efficacious and safer than the parental strain in mice models. Newer candidates, such as VPM1002 Δpdx1 (PDX) and VPM1002 ΔnuoG (NUOG), were generated to further improve the safety profile or efficacy of the vaccine. Herein, we assessed the safety and immunogenicity of VPM1002 and its derivatives, PDX and NUOG, in juvenile goats. Vaccination did not affect the goats' health in regards to clinical/hematological features. However, all three tested vaccine candidates and BCG induced granulomas at the site of injection, with some of the nodules developing ulcerations approximately one month post-vaccination. Viable vaccine strains were cultured from the injection site wounds in a few NUOG- and PDX- vaccinated animals. At necropsy (127 days post-vaccination), BCG, VPM1002, and NUOG, but not PDX, still persisted at the injection granulomas. All strains, apart from NUOG, induced granuloma formation only in the lymph nodes draining the injection site. In one animal, the administered BCG strain was recovered from the mediastinal lymph nodes. Interferon gamma (IFN-γ) release assay showed that VPM1002 and NUOG induced a strong antigen-specific response comparable to that elicited by BCG, while the response to PDX was delayed. Flow cytometry analysis of IFN-γ production by CD4+, CD8+, and γδ T cells showed that CD4+ T cells of VPM1002- and NUOG-vaccinated goats produced more IFN-γ compared to BCG-vaccinated and mock-treated animals. In summary, the subcutaneous application of VPM1002 and NUOG induced anti-tuberculous immunity, while exhibiting a comparable safety profile to BCG in goats.

Vaccine-Induced Subcutaneous Granulomas in Goats Reflect Differences in Host-Mycobacterium Interactions between BCG- and Recombinant BCG-Derivative Vaccines.

Tuberculous granulomas are highly dynamic structures reflecting the complex host-mycobacterium interactions. The objective of this study was to compare granuloma development at the site of vaccination with BCG and its recombinant derivatives in goats. To characterize the host response, epithelioid cells, multinucleated giant cells (MNGC), T cell subsets, B cells, plasma cells, dendritic cells and mycobacterial antigen were labelled by immunohistochemistry, and lipids and acid-fast bacteria (AFB) were labelled by specific staining. Granulomas with central caseous necrosis developed at the injection site of most goats though lesion size and extent of necrosis differed between vaccine strains. CD4+ T and B cells were more scarce and CD8+ cells were more numerous in granulomas induced by recombinant derivatives compared to their parental BCG strain. Further, the numbers of MNGCs and cells with lipid bodies were markedly lower in groups administered with recombinant BCG strains. Microscopic detection of AFB and mycobacterial antigen was rather frequent in the area of central necrosis, however, the isolation of bacteria in culture was rarely successful. In summary, BCG and its recombinant derivatives induced reproducibly subcutaneous caseous granulomas in goats that can be easily monitored and surgically removed for further studies. The granulomas reflected the genetic modifications of the recombinant BCG-derivatives and are therefore suitable models to compare reactions to different mycobacteria or TB vaccines.

Video Endoscopy-Guided Intrabronchial Spray Inoculation of Mycobacterium bovis in Goats and Comparative Assessment of Lung Lesions With Various Imaging Methods.

Bovine tuberculosis (bTB) not only poses a zoonotic threat to humans but also has a significant economic impact on livestock production in many areas of the world. Effective vaccines for humans, livestock, and wildlife are highly desirable to control tuberculosis. Suitable large animal models are indispensable for meaningful assessment of vaccine candidates. Here, we describe the refinement of an animal model for bTB in goats. Intrabronchial inoculation procedure via video-guided endoscopy in anesthetized animals, collection of lungs after intratracheal fixation in situ, and imaging of lungs by computed tomography (CT) were established in three goats using barium sulfate as surrogate inoculum. For subsequent infection experiments, four goats were infected with 4.7 × 102 colony-forming units of M. bovis by intrabronchial inoculation using video-guided endoscopy with spray catheters. Defined amounts of inoculum were deposited at five sites per lung. Four age-matched goats were mock-inoculated. None of the goats developed clinical signs until they were euthanized 5 months post infection, but simultaneous skin testing confirmed bTB infection in all goats inoculated with M. bovis. In tissues collected at necropsy, M. bovis was consistently re-isolated from granulomas in lymph nodes, draining the lungs of all the goats infected with M. bovis. Further dissemination was observed in one goat only. Pulmonary lesions were quantified by CT and digital 2D radiography (DR). CT revealed mineralized lesions in all the infected goats ranging from <5 mm to >10 mm in diameter. Small lesions <5 mm predominated. The DR failed to detect small lesions and to determine the exact location of lesions because of overlapping of pulmonary lobes. Relative volume of pulmonary lesions was low in three but high in one goat that also had extensive cavitation. CT lesions could be correlated to gross pathologic findings and histologic granuloma types in representative pulmonary lobes. In conclusion, video-guided intrabronchial inoculation with spray catheters, mimicking the natural way of infection, resulted in pulmonary infection of goats with M. bovis. CT, but not DR, presented as a highly sensitive method to quantify the extent of pulmonary lesions. This goat model of TB may serve as a model for testing TB vaccine efficacy.