Authors

Discussion

Urogenital schistosomiasis, a snail-borne disease caused by infection with Schistosoma haematobium, accounts for more than 110 million cases of human schistosomiasis in sub-Saharan Africa alone. Zanzibar (Unguja and Pemba Island) is endemic for urogenital schistosomiasis but is now targeted for elimination through the ‘Zanzibar Elimination of Schistosomiasis Transmission’ (ZEST). However new measures are required to evaluate ‘true’ elimination (cessation of Schistosoma transmission), when it is finally reached. Snail xenomonitoring has been suggested as an alternative method to assess levels of transmission/freshwater-schistosome-contamination and certify eventual elimination. Two morphologically similar species of freshwater snail, Bulinus globosus and B. nasutus, are linked with S. haematobium transmission on Pemba Island, however the latter is widely regarded as naturally refractory to schistosome infection in this setting. The development of sensitive and specific xenomonitoring molecular markers may therefore offer a more efficient approach to evaluate these two Bulinus species individual roles in transmitting Schistosoma spp. in Pemba Island. Here we report on the development of protocols for the xenomonitoring of Schistosoma infection in Bulinus in Pemba Island. Snails have been collected over 3 times points in up to 11 shehias (smallest administrative region) of 4 differing transmission statuses. Transmission status categories were developed from results of the annual ZEST S. haematobium prevalence surveys between 2012 and 2016 with shehias grouped in either 1 of 4 categories: persisting hot-spots (high prevalence areas with persisting high prevalence of >10%), declining hot-spots (high prevalence areas with decline to <10% by 2016), low non-responders (prevalence persisting at <10%) and low decliners (starting <10% prevalence and declining to <1%). In each shehia, human freshwater contact sites were mapped, and snail surveys conducted at each site involving searching and collecting Bulinus and recording ecological characteristics. Snails were checked for shedding schistosomes (patent infections) and any cercariae were molecularly identified by amplifying and sequencing the mitochondrial cytochrome oxidase subunit 1 (cox1) and internal transcribed spacer 1 (ITS 1+2) DNA regions. All snails were then fixed in absolute ethanol for morphological and molecular identification to differentiate B. globosus and B. nasutus and for xenomonitoring. Different primer combinations are currently being developed to amplify snail and schistosome DNA in a single reaction, to confirm Bulinus species and pre-patent schistosome infections. Initial results have been surprising, in that although S. haematobium cercariae have been identified shedding from B. globosus, the new emergence of S. bovis, a parasite of cattle, also identified from multiple shedding B. globosus complicates the development of the xenomonitoring tool that now requires an increased specificity to differentiate these two species of schistosome. Not only this, but the confirmed presence of B. nasutus in identified transmission areas, questions their individual involvement in transmission. Our data therefore shows the necessity for the molecular identification and the development of species-specific molecular xenomonitoring tools of the snails and their schistosomes to truly evaluate the levels of human schistosome transmission/contamination. Without such specificity the transmission of S. bovis will be misidentified and assumed to be S. haematobium.