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Background  Only a few studies have examined the presence of  Anaplasma marginale  and  Anaplasma centrale  in South Africa, and no studies have comprehensively examined these species across the whole country. To undertake this country-wide study we adapted a duplex quantitative real-time PCR (qPCR) assay for use in South Africa but found that one of the genes on which the assay was based was variable. Therefore, we sequenced a variety of field samples and tested the assay on the variants detected. We used the assay to screen 517 cattle samples sourced from all nine provinces of South Africa, and subsequently examined  A. marginale  positive samples for  msp1α  genotype to gauge strain diversity.    Results  Although the  A. marginale msp1β  gene is variable, the qPCR functions at an acceptable efficiency. The  A. centrale groEL  gene was not variable within the qPCR assay region. Of the cattle samples screened using the assay, 57% and 17% were found to be positive for  A. marginale  and  A. centrale , respectively. Approximately 15% of the cattle were co-infected.  Msp1α  genotyping revealed 36 novel repeat sequences. Together with data from previous studies, we analysed the Msp1a repeats from South Africa where a total of 99 repeats have been described that can be attributed to 190  msp1α  genotypes. While 22% of these repeats are also found in other countries, only two South African genotypes are also found in other countries; otherwise, the genotypes are unique to South Africa.    Conclusions   Anaplasma marginale  was prevalent in the Western Cape, KwaZulu-Natal and Mpumalanga and absent in the Northern Cape.  Anaplasma centrale  was prevalent in the Western Cape and KwaZulu-Natal and absent in the Northern Cape and Eastern Cape. None of the cattle in the study were known to be vaccinated with  A. centrale , so finding positive cattle indicates that this organism appears to be naturally circulating in cattle. A diverse population of  A. marginale  strains are found in South Africa, with some  msp1α  genotypes widely distributed across the country, and others appearing only once in one province. This diversity should be taken into account in future vaccine development studies.

Co-infections with multiple genotypes of Anaplasma marginale in cattle indicate pathogen diversity