MOLECULAR IDENTIFICATION OF SCHISTOSOMA MATTHEEI FROM FECES OF KINDA (PAPIO CYNOCEPHALUS KINDAE) AND GRAYFOOT BABOONS (PAPIO URSINUS GRISEIPES) IN ZAMBIA Anna H. Weyher, Jane E. Phillips-Conroy*, Kerstin FischerÀ, Gary J. WeilÀ, Wilbroad Chansa`, and Peter U. FischerÀ Department of Anthropology, Washington University in St. Louis, One Brookings Drive, St. Louis, Missouri 63130. e-mail: baboon@pcg.wustl.edu ABSTRACT: Terminal-spined Schistosoma sp. eggs were detected in several groups of baboons living in Kafue National Park in central Zambia. A total of 166 fecal samples was screened; egg prevalence overall ranged between 7% and 10%, while infection intensities were low. Formalin-fixed eggs had an average length of 144.5 mm and a breadth of 48.3 mm, but the schistosome species could not be unambiguously identified by size or morphology. We used molecular methods to definitively identify the parasite species. Parasite DNA was amplified from stools by polymerase chain reaction (PCR). Sequence analysis of fragments of the first internal transcribed spacer (ITS-1), mitochondrial 12S rDNA, NADH dehydrogenase subunit 6 (nad6), and cytochrome C oxidase subunit 1 (cox1) from 3 egg-positive samples revealed the presence of S. mattheei in these samples. This is the first molecular identification of S. mattheei from free-ranging baboons. Schistosoma mattheei is typically a parasite of bovids, but it can also infect humans. Schistosoma mattheei in baboons in Zambia may affect other wildlife species and humans that live in close proximity to baboons. Schistosoma species are blood fluke parasites that are a major cause of morbidity and mortality in humans. It is estimated that 200 million people are infected and about 10% of them show severe disease (Chitsulo et al., 2000). Schistosomes have an indirect life cycle with snail intermediate hosts. Once eggs are introduced to suitable snail habitats by feces, massive asexual reproduction within the intermediate hosts leads to contamina- tion of water resources with cercariae that may be infective for human and other animal hosts. Schistosome species that commonly reside in humans as definitive hosts include Schistosoma mansoni, S. haematobium, S. japonicum, and S. intercalatum. A number of schistosome species related to the S. haematobium group infect domestic and wild animals. Schistosoma bovis, S. mattheei, and S. curassoni are common in domesticated ruminants and wild antelopes, while Schistosoma margrebowiei and S. leiperi prefer the latter. A recent revision of Schistosoma based on molecular phylogenetic data showed that what was formerly known as S. intercalatum actually comprises 2 taxa, S. intercalatum and S. guineensis (Pages et al., 2001; Webster et al., 2006). Although most species are relatively host specific, animal schistosomes sometimes infect humans and vice versa (Hira, 1975; Muller-Graf et al., 1997; Murray et al., 2000). A stool survey of Kinda (Papio cynocephalus kindae) and grayfoot (Papio ursinus griseipes) baboons revealed schistosome eggs with terminal spines from a number of localities in Kafue National Park, Zambia. As the size and shape of eggs of terminal- spined schistosome species overlap considerably, we were unable to identify the species of these eggs by microscopy. In addition, naturally occurring hybrids have been observed within the S. haematobium group, further complicating species identification using egg morphology (Kruger et al., 1986; Pages et al., 2002; Lockyer et al., 2003). Baboons (Papio spp.) are a widespread and very successful primate taxon in Africa (Jolly, 2007). In many areas, baboons live close to humans and their domesticated animals; in areas where they live in the wild, baboons often form mixed-group (‘‘poly- specific’’) associations with ungulates. Schistosomes have been recorded in baboons in a number of localities in Africa, including South Africa (S. mattheei and S. haematobium), Kenya (S. mattheei), Senegal (S. haematobium), and Tanzania (S. haemato- bium). In all of these studies, the parasite species was identified morphologically, but its specific identity was never confirmed using molecular markers (Taylor et al., 1972; Cheever et al., 1974; McConnell et al., 1974; Pitchford and Visser, 1975; Appleton et al., 1991). Schistosoma mansoni has been found in several studies of baboon parasites (Muller-Graf et al., 1997; Murray et al., 2000; Weyher et al., 2006). Baboons have been postulated to be a zoonotic reservoir of S. mansoni (Fenwick, 1969; Muller-Graf et al., 1997), but their reservoir capacity is not clear and it is likely that they may be a reservoir for other human schistosomes. The objective of the present study was to identify Schistosoma eggs as to species in Zambian baboons. The molecular phylogeny of schistosomes has been intensively studied over the last decade and a multi-marker approach, which also includes the cox1 gene, allows detailed molecular barcoding. Unfortunately, most studies have characterized markers from adult worms collected from natural or laboratory hosts (Lockyer et al., 2003; Webster et al., 2006). In contrast, reports of molecular barcoding of animal schistosome DNA from fecal samples that contain only small numbers of eggs are scarce. The relatively small amount of target DNA and the diversity of plant, animal, bacterial, and fungal DNA in feces can cause problems when polymerase chain reaction (PCR) primers based on conserved DNA sequences are selected, or when low stringency conditions are applied, or both. In the present study, we evaluated several primer sets to amplify nuclear and mitochondrial DNA markers of schistosomes found in baboon feces. The PCR products were sequenced and the schistosome species was determined by comparison with nucleo- tide sequences archived in GenBank. MATERIALS AND METHODS Animals and study site The fecal samples came from baboons living in Kafue National Park, the largest national park in Zambia and the second largest in Africa. It is Received 2 June 2009; revised 30 June 2009; accepted 20 August 2009. * To whom correspondence should be addressed: Department of Anatomy and Neurobiology, Washington University School of Medicine, 660 S. Euclid Ave., St Louis, Missouri 63110. { Infectious Disease Division, Department of Internal Medicine, Washington University School of Medicine, 660 S. Euclid Ave., St. Louis, Missouri 63110. { Department of Research, Zambian Wildlife Authority, Chilanga, Zambia. DOI: 10.1645/GE-2186.1 J. Parasitol., 96(1), 2010, pp. 184–190 F American Society of Parasitologists 2010 184