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The Cultivation in Vitro of B. Leprae with Thiamin (Vitamin B1) Culture Medium
Walker L. Loving
Walker L. Loving
Department of Pathology and Bacteriology, Oglethorpe University, School of Medicine, Oglethorpe University, Georgia
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Summary
It is well known that several members of the vitamin B complex possess growth promoting properties (7). Riboflavin, pantothenic acid, biotin, nicotinic acid, thiamin, and folic acid excite certain groups of bacteria and yeast to proliferate beyond their ordinary limits. The researches of Sure (8) have contributed considerably in clarifying vitamin B1 potency relative to its growth accelerating factors in the lower animal.
In these leprosy experiments vitamin B1 (thiamin) was employed as a growth stimulating substance. Cultures were prepared directly from tissue emulsion that was rich in the bacillus of Hansen. Organ culture preparations were made from the affected tissues of rabbits which had previously been injected with emulsion (tissue juice) of a leprosy nodule containing the Hansen bacillus in great numbers.
Direct cultivation upon the artificial medium of a chromogenic acid-fast bacillus from the human leprous nodule was obtained. The culture secured directly from the human lesion was identical with the chromogenic culture isolated from the lesions of the rabbit induced with human leprous tissue.
Growth was not observed in the cultured material before the intervention of two and one-half months for the indirect (leprous tissue → animal passage → in vitro cultivation) and two months had elapsed before visible growth was noted for the direct (leprous tissue → in vitro culture) cultivation. This feature is quite constant as regards B. leprae culture.
The author (9) has demonstrated that acid-fast saprophytes (B. phlei and B. smegmatis) can be cultivated readily on an ordinary medium in twenty-four hours as contrasted to the slow growing tendency of the chromogenic acid-fast bacilli isolated and reported herein. Duval and Harris (10) observed that B. leprae is extremely slow to grow even under the most favorable conditions requiring eight to ten weeks to attain maximum growth. Initial growth is apparently dependent upon a vital “activator” or stimulatory agent.
There have been reports in the literature regarding the production of infection in the experimental animal with cultures obtained from human leprosy tissue; however, there has been no mention made of an experimental procedure where animal passage and infection was fulfilled, resulting in the cultivation of a chromogenic acid-fast bacillus. Furthermore, to establish authenticity of this culture a tissue emulsion of a fresh leprosy nodule was cultured directly on the artificial pabulum (vitamin B1). This preparation also yielded an acid-fast chromogenic bacillus. The special medium “vitalized” with thiamin or vitamin B1 was employed in these isolations.
These microbial isolations were identical from the cultural, morphological, and tinctorial points of view to the acid-fast chromogenic leprosy culture of Duval. These premises herein reported are significantly correlated with Duval's (2) in vitro cultivation of the acid-fast chromogenic bacillus from the uncontaminated leprous nodule. This representation gives strong supporting evidence for the authenticity of Duval's leprosy culture regarding its etiological relationship to the disease of human leprosy.
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