Abstract

Atoxoplasma are coccidian parasites that infect passerine birds causing significant morbidity and mortality, especially in young animals. While disease caused by Atoxoplasma or Atoxoplasma-like organisms has been reported in a number of species, including the highly endangered Bali mynah (Leucopsar rothschildi), no definitive diagnostic test for identifying Atoxoplasma exists. In addition, the lack of information about the host-species specificity of the organism and the number of Atoxoplasma sp. that exist causes significant problems with regard to preventive medicine and captive population management. In the past, the most reliable method for detecting Atoxoplasma infection has been demonstration of sporozoites in mononuclear leukocytes on blood impressions. Finding typical isosporoid oocysts after sporulation of fecal samples and histology are additional techniques for identifying suspected cases of atoxoplasmosis. The purpose of this study was to develop a polymerase chain reaction (PCR) based molecular diagnostic test for atoxoplasmosis, to begin molecular characterization of the organism, and to begin compiling data on the host-range of different species of Atoxoplasma. Since 1988 we have identified 85 cases of suspected atoxoplasmosis in 32 passerine species based on microscopic evaluation of blood impressions, fecal flotation analyses, and/or histologic examination. Eighty-two of these 85 animals died, and atoxoplasmosis was cited as the cause of death or directly implicated in the death of 50 (61%) of these animals. In order to provide a more definitive diagnosis of atoxoplasmosis and to facilitate a molecular epidemiologic investigation of these cases, we have recently implemented a PCR method that amplifies conserved small subunit rRNA gene regions of apicomplexan protozoa. PCR amplicons from frozen or formalin-fixed, paraffin-embedded tissues of nine species of passerine birds from the Zoological Society of San Diego with suspected Atoxoplasma infection have now been generated, cloned, and sequenced. These genetic sequences are distinct from other coccidia for which sequence information is available but are most similar to Isospora and Caryospora. In each animal, a homogenous PCR product was obtained, indicating that infection in the individual was likely limited to a single species of Atoxoplasma. Our findings also suggest that at least two species of Atoxoplasma probably exist in passerines, and that transmission of the same Atoxoplasma sp. between different avian taxa may occur.