Experimental transmission of human leprosy bacilli in foot pads of severe combined immunodeficient mice
To the Editor:
After the discovery of Mycobacterium leprae as the etiologic agent of human leprosy, it soon became clear that this mycobacterium cannot be grown in vitro. Hence, the search for a suitable animal model began. Animal models of leprosy used by investigators between 1879 and 1986 have been reviewed by Johnstone (1). Of the several animal models so far employed, only armadillos and nude mice are currently used for the production of M. leprae to be used in all fields of leprosy research. After infection, the maintenance of these animals for 12-18 months under controlled conditions is quite expensive. Recently, a mouse with severe combined immunodeficiency (SCID) reconstituted with human peripheral blood leukocytes has been developed (4). In an attempt to determine if SCID mice are susceptible to human leprosy and whether higher yields of M. leprae can be obtained in a relatively shorter period of time, studies on the transmission of human leprosy to SCID mice were carried out.
A bacillary suspension of M. leprae containing 1 x 108/ml acid-fast bacilli (AFB) was prepared from a foot-pad lesion of nude mice previously infected with human leprosy bacilli. Three groups of 10 SCID mice (females, 6 weeks of age) were inoculated in the hind foot pads with a 20 µ l bacillary suspension containing 1 x 105, 1 x 106 and 1 x 107 bacilli. In parallel, three groups of 10 nude mice (as controls) were also infected the same way. Both types of mice were kept at 22ºC in the same specific pathogen free vinyl plastic isolator. Food, water (ad libitum) and bedding after sterilization were provided under aseptic conditions. Following the inoculation of the foot pads with M. leprae both SCID and nude mice were sacrificed at various time intervals and AFB were counted according to the method of Shepard and McRae (5).
Regardless of the number of bacilli used in the inocula, about 5 months' postinfection a slight swelling in all foot pads of both types of mice started to appear; although more visible in SCID mice. The swelling gradually continued and became quite apparent after 7 to 8 months of infection. Our results have shown that in the foot pads of SCID mice infected with 1 x 105, 1 x 106 and 1 x 107 AFB maximum yields of 1.2 x 108, 4.3 x 108 and 9.0 x 108 bacilli were found after 11,9 and 8 months of infection, respectively. Thereafter, the number of bacilli gradually decreased upon further incubation, and only some degenerated bacilli were found at the inoculation site after 15 months of incubation. In the foot pads of nude mice infected with 1 x 105 and 1 x 106 bacilli, at 10 months' postinfection 7.8 x 107 and 2.5 x 108 bacilli/foot pad were obtained, respectively. These results show that up to 10 months postinfection the total number of bacilli in the foot pads of nude mice were lower than estimated in the foot pads of SCID mice. However, in the foot pads of nude mice multiplication of M. leprae continued progressively at all three inocula used and about 12 months postinfection remarkable swelling of the infected foot pads was noted. In the foot pads of nude mice infected with 1 x 105, 1 x 106 and 1 x 107 bacilli, maximum yields of 1.7 x 1010, 2.0 x 1010 and 2.1 x 1010 bacilli per foot pad were estimated, respectively, after 13, 12 and 11 months of infection. Dissemination of M. leprae in foot pads of nude mice is well established (2,3). These results of a comparative study show that, like nude mice, SCID mice were also susceptible to M. leprae infection and the onset of the lepromatoid lesions in the foot pads of SCID mice occurred earlier than that observed in the nude mice. However, an interesting aspect of this study is that the progress of M. leprae infection in SCID mice is different than the progress observed in the nude mice. Although a rapid multiplication of M. leprae occurred in the foot pads of SCID mice after reaching a maximum of about 4 to 9 x 108 bacilli/foot pad, the number of bacilli decreased upon further incubation and eventually, about 15 months' postinfection, only a few degenerated bacilli were found at the site of infection and there was no sign of dissemination. On the other hand, the number of bacilli in the foot pads of nude mice after reaching about 1.5 to 2.0 x 1010/ foot pad remained nearly the same up to 15 months of incubation. Since the total number of bacilli in the foot pads of nude mice is considerably higher, such mice should continue to be used for the production of M. leprae for leprosy research. SCID mice possibly could be used for screening the antileprosy drugs in a relatively shorter period of time.
The infection of M. leprae in SCID mice has not been investigated extensively. The phenomenon of decline and eventual clearing of millions of bacilli in the foot pads of SCID mice in a relatively shorter time period is not clear and is worthy of investigation. Additional studies such as histopathology and the status of the immune response of SCID mice at various stages of infection, lepromin reactivity as well as DNA homology, should be carried out.
- Muhammad Ishaque, Ph.D.
Professor
Applied Microbiology Research Center
Inst it ut Armand-Frappier
University of Quebec
CP. 100
Laval, Quebec, Canada H7N 4Z3
- Veronika Sticht-Groh, M.D., F.R.C.P. (C).
Professor of Microbiology
Armauer-Hansen-Institut/DAHW
Hermann-Schell Str. 7
97074 Wurzburg, Germany
Acknowledgment. This investigation was generously supported by the Military and Hospitaller Order of Saint Lazarus of Jerusalem, Canada and Le Secours aux Lepreux, Inc., Canada.
REFERENCES
1. JOHNSTONE, P. A. S. The search for animal models of leprosy. Int. J. Lepr. 55(1987)535-547.
2. KOHSAKA, K., MORI, T. and ITO, T. Lepromatoid lesion developed in nude mouse inoculated with Mycobacterium leprae. Lepro 45(1976)177-187.
3. LANCASTER, R. D., MCDOUGALL, A., HILSON, G. R. F. and COLSTON, M. J. Leprosy in nude mice. Exp. Cell Biol. 52(1984)154-157.
4. MILMAN, G.and SOUZA, D. HIV infections in SCID mice: safety consideration. ASM News 56(1990)639-642.