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Spiroplasma chrysopicola sp. nov., Spiroplasma gladiatoris sp. nov., Spiroplasma helicoides sp. nov., and Spiroplasma tabanidicola sp. nov., from Tabanid (Diptera: Tabanidae) Flies

Robert F. Whitcomb^1,*, Frank E. French², Joseph G. Tully³, Gail E. Gasparich⁴, David L. Rose³, Patricia Carle⁵, Joseph M. Bove⁵, Roberta B. Henegar¹, Meghnad Konai⁴, Kevin J. Hackett⁴, Jean R. Adams⁴, Truman B. Clark^4, and David L. Williamson⁶

¹Vegetable Laboratory, U. S. Department of Agriculture, Beltsville, Maryland 20705
⁴Insect Biocontrol Laboratory, U. S. Department of Agriculture, Beltsville, Maryland 20705
²Department of Biology, Georgia Southern University, Statesboro, Georgia 30460
³Mycoplasma Section, Laboratory of Molecular Microbiology, National Institute of Allergy and Infectious Diseases, Frederick Cancer Research Facility, Frederick, Maryland 21702
⁵Laboratoire de Biologie Cellulaire et Moléculaire, Institut Nationale de Recherche Agronomique, 33883 Villenave d'Ornon Cedex, France
⁶Department of Anatomical Sciences, State University of New York, Stony Brook, New York 11794

^* Corresponding author. Mailing address: Vegetable Laboratory, Range 2, HH3, BARCW, Beltsville, MD 20705. Phone: (301) 504–8339. Fax: (301) 504–6017.

ABSTRACT

Four spiroplasma strains, DF-1^T, TG-1^T, TABS-2^T, and TAUS-1^T, all of which were isolated from deerflies or horseflies (Diptera: Tabanidae), were serologically distinct from previously described spiroplasma species, groups, and subgroups. Strain DF-1^Toriginated from a Maryland deerfly (Chrysops sp.); strain TG-1^Twas isolated from a Maryland horsefly (Tabanus gladiator); strain TAUS-1^Toriginated from a member of the Tabanus abdominalis-limbatinevris complex of horseflies collected in Maryland; and strain TABS-2^Twas isolated from a horsefly (Tabanus abactor) collected in Oklahoma. Cells of all of the strains appeared to be helical and motile when they were examined by dark-field microscopy. Cells of strain DF-1^Tgrowing in M1D medium were short helices with less than six turns; the helical cells of the other strains were long and usually had six or more turns. The short cells of strain DF-1^Tpassed through 450- and 300-nm filter pores with no reduction in titer, but the longer cells of the other strains were partially retained by 450-nm-pore-size filters. Electron microscopic examination of all of the strains revealed wall-less cells surrounded only by a single cytoplasmic membrane. All of the strains grew well in SP-4 liquid media and in conventional mycoplasma or M1D media supplemented with horse or fetal bovine serum. Strains TABS-2^T, TAUS-1^T, and DF-1^Trequired serum or sterol for growth, but strain TG-1^Twas able to grow in the absence of serum or sterol. The optimum temperatures for growth of the four strains varied from 30 to 32°C, and growth occurred at 10 to 37°C. All of the strains catabolized glucose but did not hydrolyze urea. Only strain DF-1^Thydrolyzed arginine. The guanine-plus-cytosine contents of the DNAs of the strains were: DF-1^T, 29 ± 1 mol%; TG-1^T, 26 ± 1 mol%; TABS-2^T, 27 ± 1 mol%; and TAUS-1^T, 26 ± mol%. The genome sizes of strains DF-1^Tand TAUS-1^Twere 1,270 and 1,375 kbp, respectively. Strain DF-1 (= ATCC 43209), the representative of spiroplasma subgroup VIII-2, is designated the type strain of a new species, Spiroplasma chrysopicola. We also propose that strain TG-1^T(= ATCC 43525^T), the designated representative of group XXIII, should be placed in a new species, Spiroplasma gladiatoris. In addition, group XXXII spiroplasma strain TABS-2 (= ATCC 51746) is designated the type strain of Spiroplasma helicoides sp. nov., and group XXXIII representative strain TAUS-1 (= ATCC 51747) is designated the type strain of another new species, Spiroplasma tabanidicola.

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