Isolation of Coccidioides Immitis (Stiles) from the Soil

Abstract

The epidemiological studies on coccidioidal granuloma, recently summarized by Beck, 1 suggest the soil and the vegetation as the probable source of the fungus infection. The peculiar geographic distribution of the disease both in man and animals to the central and southern sections of the State of California with a predominance of the cases in Kern and Los Angeles Counties supports this contention. Although repeated attempts have been made to find the organism in nature the examinations, made with the aid of the customary bacteriologic technique, have thus far yielded negative results. The relatively slow growing atypical fungi are, as a rule, rapidly overgrown by the concomitant bacteria present in the soil and vegetations. Little progress could be made until a procedure had been developed which would concentrate, possibly enrich the stage of the mold (chlamydospore) supposed to exist in nature. In the course of detailed metabolism studies it was found that Coccidioides immitis satisfies its nitrogen and carbon requirements from relatively simple compounds. The medium, which gives selective cultures, has the following composition: Ammonium chloride 1 part, sodium acetate 1 part, equal mixture of KH₂PO₄ and K₂HPO₄ 0.4 part and magnesium sulfate 0.01 part; distilled water 100 parts. While the majority of bacteria grow slowly or very poorly, Coccidioides, certain Blastomyces and Aspergillus varieties develop exceedingly well in the nutrient solution. Bacterial multiplication may be entirely suppressed and the purification of the fungus greatly facilitated by the addition of acriflavine in a concentration of 1:25,000. For several years the medium has been employed in the examination of clinical specimens (sputum, pus, blood, autopsy specimens, etc.) suspected to contain the mold. Samples of sputum, which contained very few typical double contoured fungoidal cells yet teeming with the usual respiratory and buccal flora, gave pure cultures of Coccidioides immitis in the dye culture medium.