Resistance to cutaneous leishmaniasis: acquired ability of the host to kill parasites at the site of infection

J O Hill
1984 Infection and Immunity  
The purpose of these studies was to follow the development of acquired resistance in experimental cutaneous leishmaniasis by measuring changes, against time, in the ability of mice infected with Leishmania tropica to inhibit the growth of a challenge inoculum of parasites. In addition, the development of T lymphocytes that mediate the acquired response was followed by adoptive immunization. It was found that acquired resistance developed rapidly and reached a maximum level at the time when the
more » ... the time when the progressive multiplication of the parasites in the primary lesion stopped. Systemic immunity, however, as determined by the ability of splenic T cells to adoptively immunize normal recipients, did not develop fully until 4 weeks later. Acquired resistance is expressed systemically in the animal and probably nonspecifically, in that mice expressing resistance to a homologous challenge with L. tropica were also capable of destroying Listeria monocytogenes. These data, obtained from in vivo studies, are consistent with the hypothesis that the immunity which causes the destruction of L. tropica is mediated by T lymphocytes and expressed, nonspecifically, through macrophages. Cutaneous leishmaniasis in mice serves as a model of the human disease. By capitalizing on the differences in susceptibility to Leishmania tropica exhibited by various inbred strains, models of both the resolving and the progressive, lethal form of the disease have been developed (3, 4, 10, 19, 26) . Previous studies (3, 4, 26) have shown that the lesions that develop in resistant mouse strains at the site of inoculation of the parasites increase in size and then gradually heal over a period of months. On reinfection, however, the lesion that appears at the challenge site develops and resolves relatively rapidly (16, 25, 26) . In addition, the capacity to rapidly resolve the lesion that develops at the site of a challenge infection can be passively transferred with cells (1, 16, 27), but not serum (27) , taken from donors whose primary lesions have healed. These data, therefore, suggest that mice acquire a cell-mediated state of immunity as the result of a primary L. tropica infection. Since all of the previous studies of acquired resistance to L. tropica have relied almost exclusively on measurements of changes in lesion size, there are no data on how fast the acquired state of immunity develops and how it results in the destruction of the parasites in vivo. Based on the observation of Keppel and Janovy (13) that Leishmania donovani will grow as distinct colonies on rabbit blood agar, a technique was developed in this laboratory to quantitate the number of viable L. tropica parasites present in infected tissues (12; J. 0. Hill, J. Parasitol., in press). The previous use of this procedure to enumerate parasites revealed that data on lesion size can be misleading in that resolution of the disease, as determined by the onset of destruction of the parasites, begins before the lesions begin to heal (12). Furthermore, marked swelling can occur as the result of challenging a previously infected animal, despite the fact that the parasites in the challenge inoculum are rapidly eliminated. The purpose of the studies summarized here was to follow the development of acquired resistance in mice by measuring changes, against time of infection, in the ability of an animal to destroy a challenge inoculum of parasites. In addition, the development of T cells which mediate this acquired resistance in the animal was followed by adoptive immunization. This paper shows that acquired resistance develops rapidly, in proportion to the number of organisms present in the primary lesion. However, the systemic resistance that develops as the result of the local cutaneous infection is probably expressed nonspecifically, since Leishmania tropica-infected mice are also capable of rapidly inactivating a lethal, intravenous challenge with the facultative intracellular bacterium Listeria monocytogenes. MATERIALS AND METHODS Mice. Specific-pathogen-free, male C3H/He mice were used. The animals were 8 to 12 weeks old and were obtained from the Trudeau Institute Animal Breeding Facility, Saranac Lake, N.Y. The mouse colony is routinely monitored for the presence of pathogenic bacteria and mycoplasmas by standard bacteriological techniques. In addition, the colony is monitored for virus infections by serological tests (Mouse. Virus Profile 80-211; Microbiological Associates, Bethesda, Md.). Microorganisms and their quantitation. Leishmania tropica major 173 was used. The source and passage history of the organism has been described previously (7, 12) . Primary infections were initiated by injecting 5 x 105 amastigotes in 50 1jl of Graces Insect Tissue Culture Medium subcutaneously into the left hind footpad. All media were obtained from GIBCO Laboratories, Grand Island, N.Y. At various weeks of the primary infection, four or five animals were killed and the infected foot was rinsed with 70% ethanol and then aseptically removed. The lesions were disrupted as previously described (12) , except instead of manually cutting the tissue into small pieces, the samples were minced with a M122 Biohomogenizer (Biospec Products, Bartlesville, Okla.). The homogenates were then appropriately diluted in Graces medium, and 50-,ul samples were plated on rabbit blood agar plates (Hill, in press). After 5 to 7 days of incubation at 26°C, the L. tropica colonies were counted and the number of viable parasites present in the lesions was calculated. Listeria monocytogenes EGD, serotype 1/2a, was grown in Trypticase soy broth (BBL Microbiology Systems, Cockeysville, Md.), harvested while still in log phase (2.5 x 108/ ml), and dispensed in 1.0-ml lots. The aliquots were then frozen and stored at -70°C until needed. For each experi-127 on May 7, 2020 by guest http://iai.asm.org/ Downloaded from
doi:10.1128/iai.45.1.127-132.1984 fatcat:5is24e6ksndn3d7hw3uqirwxny