Twenty-Third Annual Meeting and Symposium of the
Desert Tortoise Council, April 3-5, 1998
Abstracts

Mollicutes are a group of about nine distinct genera of wall-less bacteria occurring in a variety of plant, insect and animal hosts. Two species in the genus Mycoplasma have been identified as etiologic agents in respiratory disease of tortoises. Mycoplasma agassizii is pathogenic for both the desert (Gopherus agassizii) and gopher (Gopherus polyphemus) tortoise in Southwestern and Southeastern United States, and in tortoises in France. A currently unnamed Mycoplasma species (strain HB29) has been isolated from desert tortoises with respiratory disease in California, and identified in a single Florida gopher tortoise with a respiratory infection. Experimental transmission of Mycoplasma agassizii from tortoise to tortoise through an infectious nasal discharge has been documented.

Ticks are an important vector for transmission of numerous infectious bacterial agents, including spirochete infections (Lyme disease and relapsing fever due to Borrella infections), tularemia, and various rickettsial infections. In addition, special mollicutes of the genus Spiroplasma, a group of helical wall-less prokaryotes occurring widely in insects have been found in two tick hosts (Ixodes pacificus, Haemaphysalis leporis-palustris) in the US. Previous studies have documented the frequent occurrence of ticks, specifically Ornithodoros parkeri, as ectoparasites of tortoises. Also, tick transmission of mycoplasmas involved in bovine pleuropneumonia disease in Africa has been demonstrated in some earlier experimental studies but not confirmed under natural conditions. In view of this association between ticks and bacterial/mycoplasmal agents, it was thought worthwhile to attempt cultivation of the two Mycoplasma species from ticks infesting tortoises.

Four hundred fifty nine ticks (all Ornithodoros parkeri, J. Oliver, pers. comm.) were collected from Gopherus agassizii tortoises in the Mojave, Colorado, and Sonoran deserts in California, Arizona, and Nevada from March 18 to November 14, 1997. Eighty three percent of the ticks (382) came from the Desert Tortoise Conservation Center in Las Vegas. Upon arrival by express air mail with information on tortoise number and health profile forms and other collection data, the ticks were assigned pool numbers and immediately placed in a sterile mortar with a small piece of dry ice. After carbon dioxide anaesthesia, the ticks were ground with a pestle in a small amount of sterile sand and about 2 ml of SP-4 mycoplasma medium. This formulation contains 17% sterile fetal bovine serum and 500 units/ml of penicillin G to suppress bacterial growth. The tick triturate was then drawn into a sterile syringe and the contents passed through a sterile, 25mm bacteriologic filter (450 nm) into a 4ml screw cap glass vial containing about 1.5 ml SP-4 broth. Vials were incubated at 30^oC and examined periodically for turbidity or pH changes in the medium. Each fresh lot of the SP-4 broth was tested to confirm the ability to grow the two Mycoplasma species identified in tortoises.

No Mycoplasma or Spiroplasma species were isolated in cultivation attempts from the ticks examined in this study. Although a preponderance of the ticks originated from one location, the disease is apparent in many of the tortoises at this location. These findings would seem to suggest that tick transmission of the mycoplasma infection is an unlikely event.

(The authors wish to thank M. Berkowitz, S. Boland, P. Frank, G. Goodlett, T. Goodlett, S. Hart, L. Stockton, M. Vaughn, and A. P. Woodman, for their special efforts in providing ticks from the various field sites).

References

Brown, M. B., I. M. Schumacher, P.A. Klein, K. Harris, T. Correll, and E.R. Jacobson. Mycoplasma agassizii causes upper respiratory tract disease in the desert tortoise. Infect. Immun. 62:4580-4586, 1994.

Tully, J. G., R. F. Whitcomb, D. L. Rose, D. L. Williamson, and J. M. Bove. Characterization and taxonomic status of tick spiroplasmas: a review. Yale J. Biol. Med., 56 :599-603, 1983.

Tully, J. G., D. L. Rose, C. E. Yunker, P. Carle, J. M. Bove, D. L. Williamson, and R. F. Whitcomb. Spiroplasma ixodetis sp. nov., a new species from Ixodes pacificus ticks collected in Oregon. Int. J. Syst. Bacteriol. 45:23-28, 1995.