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Spiroplasma poulsonii sp. nov., a new species associated with male-lethality in Drosophila willistoni, a neotropical species of fruit fly

David L. Williamson^1,, Bungo Sakaguchi², Kevin J. Hackett³, Robert F. Whitcomb^4,, Joseph G. Tully⁵, Patricia Carle⁶, Joseph M. Bové, Jean R. Adams³, Meghnad Konai^3, and Roberta B. Henegar³

¹Department of Anatomical Sciences, State University of New York, Stony Brook, NY 11794, USA
²Faculty of Agriculture, Kyushu University, Fukuoka, Japan
³Insect Biocontrol Laboratory, US Department of Agriculture, Beltsville, MD 20705, USA
⁴Vegetable Laboratory, US Department of Agriculture, Beltsville, MD 20705, USA
⁵Mycoplasma Section, Laboratory of Molecular Microbiology, National Institute of Allergy and Infectious Diseases, Frederick Cancer Research Facility, Frederick, MD 21702, USA
⁶Laboratoire de Biologie Cellulaire et Moléculaire, Institut National de la Recherche Agronomique and Université de Bordeaux II, 33883 Villenave d'Ornon Cedex, France

Author for correspondence: David L. Williamson. Tel: +1 516 724 0897. Fax: +1 516 444 3947. e-mail: dwmson{at}physiology.pnb.sunysb.edu

ABSTRACT

Progenies from some wild-caught females of Drosophila willistoni and three other sibling species are entirely female. The proclivity for production of unisexual female progeny by these flies was named the sex ratio (SR) trait and was originally thought to be genetic. However, experiments in the laboratory of Donald F. Poulson in the early 1960s demonstrated that this ‘trait’ was vertically transmitted and infectious, in that it could be artificially transferrd by injection from infected females to non-infected females. Motile, helical micro-organisms were observed in females showing the trait. In 1979, the SR organisms were designated as group II in the informal spiroplasma classification system. The organisms proved to be extremely fastidious, but were eventually cultivated in a very complex cell-free medium (H-2) after initial co-cultivation with insect cells. Cultivation in the H-2 medium and the subsequent availability of a triply cloned strain (DW-1^T) permitted comparaltiv studies. Cells of strain DW-1^Twere helical, motile filaments 200--250 nm in diameter and were bound by a single trilaminar membrane. Cells plated on 1.8% Noble agar formed small satellite-free colonies 60--70 µm in diameter with dense centres and uneven edges. The temperature range for growth was 26--30 °C; optimum growth occurred at 30 °C, with a doubling time in H-2 medium of 15.8 h. The strain passed through filters with 220 nm, but not 100 nm, pores. Reciprocal serological comparisons of strain DW-1^Twith representatives of other spiroplasma groups showed an extensive pattern of one-way crossing when strain DW-1^Twas used as antigen. However, variable, usually low-level reciprocal cross-reactions were observed between strain DW 1^Tand representatives of group I sub-groups. The genome size of strain DW-1^Twas 2040 kbp, as determined by PFGE. The G+C content was 26·1 mol%, as determined by buoyant density and melting point methods. The serological and molecular data indicate that strain DW-1^Tis separated from group I representative strains sufficiently to justify retention of its group status. Continued group designation is also indicated by the ability of SR spiroplasma to induce male lethality in Drosophila, their vertical transmissibility and their extremely fastidious growth requirements. Group II spiroplasmas, represented by strain DW-1^T(ATCC 43153^T), are designated Spiroplasma poulsonii.

Key Words: Spiroplasma poulsonii sp. nov. • mollicute • sex ratio • Drosophila willistoni

Present address: 9125 Tymat Court, Laurel, MD 20723, USA.

Present address: Carl Hayden Bee Research Center, USDA/ARS, 2000 E Allen Road, Tucson, AZ 85719, USA.

resent address: 4 Galahad Lane, Nesconset, NY 11767--2220, USA.

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