Isolation of cultivable mycobacteria from feces and lungs of armadillos infected with Mycobacterium leprae.

In the past, no cultivable mycobacteria were isolated from armadillos captured in the state of Florida, U.S.A. But recent findings of acid-fast bacilli (AFB) in the lungs of armadillos infected with Mycobacterium leprae prompted us to undertake this study to determine the correlation between systemic leprosy infection and the occurrence of cultivable mycobacteria in the lungs and stools of these animals. No AFB could be isolated from noninfected animals. Seventy percent of the infected animals developed disseminated infection, but no cultivable mycobacteria were isolated from their livers and spleens. However, cultivable mycobacteria were isolated from the lungs and stools of a large number of armadillos showing disseminated infection. The most common among these were M. gordonae, M. fortuitum, and M. avium. There was a close correlation between the development of disseminated leprosy infection and the occurrence of cultivable mycobacteria in their lungs and stools, perhaps due to the decline in the immune system in these animals in the later stages of infection.

Presence of "difficult to isolate" mycobacteria in armadillos.

Contrary to the findings with armadillos from Louisiana and Texas, armadillos from Florida are free of natural leprosy-like infection. Examination of ear clip, nasal, blood buffy coat, liver and spleen of inoculated armadillos from Florida did not reveal the presence of any acid fast bacteria. However, using massive inocula, 6 out of 77 armadillo tissues were found to contain very negligible proportions of cultivable mycobacteria. The significance of these isolates in relation to M. leprae and also to leprosy research is discussed.

Serum angiotensin-converting enzyme in leprosy.

Serum angiotensin-converting enzyme activity was measured in 91 adult healthy and lepromatous armadillos before inoculation with M. leprae and at necropsises. Mean ACE values were significantly elevated in armadillos with leprosy and the degree of elevation was roughly proportional to the extent of infection. There was also significant difference in the serum ACE levels between Florida and Louisiana armadillos. The dapsone treatment resulted in bringing these levels to normal. Serial assays of serum, ACE provided information on the response of armadillos to dapsone therapy.

Adenosine triphosphate content of Mycobacterium leprae: effect of purification procedures.

Various procedures to decontaminate and purify M leprae free of host tissue material resulted in total retention of their intracellular ATP and also infectiousness. The ATP content of one million M. leprae cells, isolated from either livers, spleens, or lymph nodes of infected armadillos, or a nude mouse foot pad or a human biopsy specimen, was in the range of 1.17 to 1.40 picograms. Suspensions could be decontaminated with 4% NaOH and all non-bacterial ATP could be eliminated by the combined action of trypsin, chymotrypsin, and collagenase initially, followed by Triton X-100 plus ATPase. These findings further assure that M. leprae are different from M. lepraemurium in that they can withstand even the severest purification procedures that are necessary in order for them to be used for sophisticated biochemical and metabolic studies.

Electron microscopy of peroxidase and acid phosphatase in leprous and uninfected armadillo macrophages: a macrophage subpopulation contains peroxisomes and lacks bacilli.

Lepromatous tissue from armadillos inoculated 24--36 months earlier with Mycobacterium leprae was obtained for electron microscopic studies. Cytochemically stained lepromas revealed a subpopulation of macrophages containing peroxisomes. These peroxidase reactive macrophages were not infected with bacilli. Acid phosphatase was present in macrophages and many of these were infected with bacilli and contained vacuoles and lipid globules. Within the membrane-bound vacuoles, acid phosphatase surrounded bacilli. However, the reaction product ended abruptly at a 15--40 millimicron thick zone of low electron density surrounding intact bacilli. Acid phosphatase was more intensely reactive and localized less precisely in heavily infected and vacuolated macrophages than in lightly and non-infected cells. The effectiveness of this bacillary barrier and the numerous infected macrophages with substantial acid phosphatase argue against the ability of acid phosphatase to protect host cells from leprosy bacilli. Evidence suggests a protective action of peroxidase or the rapid turnover of macrophages within lepromas. Granular and membranous debris were commonly seen within vacuoles of infected macrophages. A portion of the debris was ultrastructurally similar to bacillary matrix and was nonreactive for peroxidase and acid phosphatase. Following homogenization and centrifugation, similar materials banded with bacilli above 60% sucrose. Another portion of the debris was ultrastructurally similar to host lysosomal matrix and was reactive for acid phosphatase. Results support the concept of dual host and parasitic origins of the debris found in phagolysosomes of infected macrophages. Transparent, oval Epon defects remained eccentric to the majority of intact bacilli in centrifuged fractions. Apparently, an intrinsic property of leprosy produced these Epon defects.

The immune system of the nine-banded armadillo (Dasypus novemcinctus, Linn).

The armadillo is rapidly emerging as an animal for biomedical research. A morphological study of the immune system of the armadillo was undertaken to try to demonstrate a possible defect in the morphogenesis of the lymphoid organs that could account for their susceptibility to leprosy. Blood from 20 domesticated adult armadillos revealed lymphocytes predominated among circulating leukocytes in the majority of animals, whereas in wild animals neutrophils predominated. Necropsies revealed well-developed thymus glands and the thymus-dependent areas in lymph nodes and spleens contained abundant lymphoid cells. Hence, we conclude that the susceptibility of the armadillo to leprosy cannot be explained on a morphological basis.