Animal models of tuberculosis
Introduction
Tuberculosis (TB) caused by Mycobacterium tuberculosis is the leading bacterial cause of death. TB remains a global threat to public health, with approximately 2 million people dying from M. tuberculosis infections every year and one-third of the world's population latently infected with M. tuberculosis. Every year, 8 million individuals develop active disease.1 In India, TB remains one of the most pressing health problems, accounting for 30% of the global cases.2 It has been estimated in India that every year 2 million people develop TB and nearly 25% of them succumb to the disease, accounting for more than 1000 deaths every day. A vaccine against TB was developed about 80 years ago; various drugs for the control of the disease were developed about 40–50 years ago and combination chemotherapy has been in place for about 2 decades. Yet today we do not seem to be any closer to eliminating TB than we were a century ago.
Complete eradication of TB requires a vaccine that is cost effective and can be used for mass immunization. BCG (live attenuated Mycobacterium bovis BCG), the only TB vaccine presently being used, has succeeded only in some countries but has shown very limited usefulness in countries like India. It is the most widely used of all the vaccines in the WHO Expanded Programme for immunization,3 but has been estimated to prevent only 5% of all potentially vaccine-preventable deaths due to TB.4 It has been found to be protective against disseminated and meningeal TB in young children.5 The protective efficacy of BCG vaccine against adult pulmonary TB has varied widely in different geographical areas and different populations.6 Thus, a TB vaccine that protects consistently against adult TB in all populations is needed. Emergence of drug resistance has also led to renewed interest in development of alternate drug formulations. To study alternate modes of treatment and new vaccines and to better understand the host–parasite relationship, animal models are necessary. In the present article, we discuss the different animal models currently available for TB, and propose improvements of the existing models.
Section snippets
Mouse model
Robert Koch, who discovered the tubercle bacillus, first used the mouse as an experimental model. He showed that inoculation with M. tuberculosis induced lesions like those seen in the natural disease in humans.7 Subsequently, investigators established infection in a variety of animals (rabbits, guinea pigs, rats and mice) with pure cultures of M. tuberculosis. Due to interest in progressive pulmonary infection, these studies tended to involve guinea pigs and rabbits because of their higher
TB in immunodeficient mouse models
Because immunity to TB is T-cell mediated, the nude mouse and severe combined immunodeficiency (SCID) mouse that cannot produce T-cells are of interest as models in TB. The nude mouse is furless and suffers from thymic aplasia. These mice cannot produce functional T-cells, therefore they have substantially less resistance to TB and other mycobacterial infections. On the other hand, SCID mice lack both T- and B-cells owing to a defect in the variable region gene recombination61 and lack
Gene-disrupted and transgenic mice
A new model of TB using targeted gene disruption was developed by Dalton et al.64 In this model, the IFN-γ gene was specifically disrupted by the insertion of a neomycin resistance gene after the second exon. Mice in which the IFN-γ gene has been disrupted (IFN-γ−/−) are unable to control a normally sublethal dose of M. tuberculosis, delivered either by intravenous or aerosol route.
These IFN-γ KO mice have been shown to be extremely susceptible to M. tuberculosis, and a progressive and
Immunosenescent mice
Another form of immunodeficiency is the decline in immunological response with age. It was observed that young mice infected intravenously with a sub-lethal dose (105) of M. tuberculosis bacilli could contain and control infection but that the same dose was fatal in mice of 24–28 months age.79 Initially, it was thought that this mortality was due to lack of protective T-cells in old animals, however Orme80 showed that T-cells that are capable of recognizing mycobacterial antigens in old animals
Immunology of TB in the mouse model
It has become clear from the research of the last few years that all three subsets of T-cells, namely CD4, CD8 and γδ T-cells, play a crucial role in the acquired cellular immune response to experimental M. tuberculosis infection in the mouse.20 The use of gene-disrupted mice70, 81 confirmed the role of CD4+ T-cells in the murine model. The main role that the CD4+ T-cell population plays in the combat against the pathogen is secretion of the cytokine IFN-γ. Other cytokines that activate
Modelling persistence and reactivation in the mouse
Little is known about the basic mechanism involved in maintaining a latent M. tuberculosis infection or the causes of reactivation. In large part, this is due to the difficulty in developing and manipulating an animal model of latent TB. The design of an adequate animal model of latent M. tuberculosis infection is hampered by lack of knowledge about the biological characteristics of both the tubercle bacilli and host immunity during human latent TB. Two murine models of latent M. tuberculosis
Guinea pig model of TB
In the 1800s and 1900s, the guinea pig was the most widely used experimental animal for infectious disease studies. The classic experiments that established a microorganism as the aetiological agent of TB were carried out with guinea pigs infected with M. tuberculosis.92 Koch's principle reason for choosing this species for studies of M. tuberculosis in those days is still valid today—the susceptibility of this species to infection with human tubercle bacilli.93
Sisk94 outlined a number of
Modelling other clinical forms of TB
Besides primary pulmonary TB, the guinea pig model may be useful for other forms of the disease such as tuberculous pleuritis, exogenous re-infection and endogenous reactivation. Pleuritis can be induced in previously immunized guinea pigs by the intrapleural injection of either living BCG148 or heat-killed M. tuberculosis.149 Phalen and McMurray150 reported enhanced TNF-α levels in effusion fluid as well as in supernatant fluids from pleural fusion lymphocytes stimulated with PPD. Exogenous
Studies of cytokines and chemokines in guinea pigs
Several guinea pig cytokine and chemokine genes have been cloned in recent years.156, 157, 158, 159 Using Northern blot and real-time RT-PCR methodologies, it was demonstrated that splenocytes from BCG-vaccinated guinea pigs stimulated with whole mycobacteria or purified antigens responded with higher levels of IFN-γ mRNA.160 BCG vaccination augmented TNF-α protein production in splenocytes, resident peritoneal cells, and bronchoalveolar lavage cells following exposure to virulent and
Rabbit model of TB
In rabbits, TB is a disease in which lung tissue is destroyed, largely as a result of the host's own reaction to bacillary antigens. The most extensive studies on TB using the rabbit as an experimental model were made by Lurie170, 171, 172 and Lurie and Dannenberg.173 Inbred lines of rabbit were developed for resistance and susceptibility to TB. Resistant rabbits developed cavitary TB like adult immunocompetent humans when infected with the virulent Ravenel strain of M. bovis. The susceptible
TB in monkeys
There are a number of reports that suggest that rhesus monkeys (Macaca mulatta) are highly susceptible to virulent M. tuberculosis and virulent M. bovis.194, 195, 196, 197 The disease in monkeys is usually a rapidly progressive pulmonary disease with both haematogenous and bronchial spread of the bacilli. There are reports that suggest extensive caseous necrosis along with liquefaction of the caseous material with cavity formation.194, 195, 198, 199 The walls of the cavities harbour numerous
Conclusions
Non-human primates (monkeys) appear to have significant advantages over conventional laboratory animals in terms of modelling pulmonary TB for the purpose of vaccine evaluation201 as they are closely related to humans, are quite susceptible to infection by the aerosol route, develop human-like disease, exhibit antigen-induced T-lymphocyte activity both in vivo and in vitro, and can be protected quite effectively by BCG. In addition, major advances in availability of immunologic and other
Acknowledgements
I thank Dr. David N. McMurray, Department of Medical Microbiology and Immunology, Texas A & M University System Health Science Center, 407 Reynolds Medical Building, College Station, TX 77843-1114, USA for critical reading of the manuscript.
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