Spiroplasma strain PUP-1was isolated from the gut fluids of a firefly beetle () collected in Maryland. Cells of the strain were shown by dark-field microscopy to be helical, motile filaments. Ultrastructural examination by electron microscopy revealed filamentous cells bounded by a single cytoplasmic membrane and no evidence of a cell wall. The cells were not sensitive to 500 U of penicillin per ml and grew under aerobic conditions in M1D, SP-4, and M-2 broth formulations, as well as in conventional mycoplasma medium. The doubling times at 15, 20, 25, and 30°C were 83.1, 32.0, 14.9, and 9.8 h, respectively. Suboptimal growth occurred at 37°C, and no growth was apparent in cultures maintained at 10 or 40°C. The organism required cholesterol for growth and produced acid from glucose, fructose, and trehalose; arginine and urea were not hydrolyzed. The results of previous serological analyses of strain PUP-1indicated that the organism was not related to the then currently established species or group representatives, and the organism was classified as the representative of group XIX. Additional testing of strain PUP-1against recently recognized species or group representatives by both metabolism inhibition and deformation tests confirmed the unique serological status of the organism. The guanine-plus-cytosine content of the DNA was 26 ± 1 mol%, and the genome size was 1,375 kbp. These values clearly differentiate strain PUP-1from group XXI strain W115, with which it cross-reacted reciprocally at a low level in deformation and metabolism inhibition tests. We propose that strain PUP-1 (= ATCC 43206) should be recognized as the type strain of a new species, .


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