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To shed first light on caprine schistosomiasis and its zoonotic potential in Malawi, we conducted a molecular epidemiological investigation, sampling goats ( n  = 230) across three districts, using faecal miracidia hatching test. Molecular genotyping of miracidia later revealed the prevalence of Schistosoma mattheei to be 0.0 % in Nsanje District ( n  = 30), 16.7 % in Chikwawa District (n = 30) and 25.3 % in Mangochi District ( n  = 170). Notably, a miracidium of Schistosoma haematobium was observed from a single goat in Chikwawa. Inspection of carcasses ( n  = 51) at two local abattoirs in Mangochi District did not find any evidence of caprine schistosomiasis where only a single herd, at Mangochi 3, was infected. Here, despite sampling several other herds nearby, the prevalence was 87.7 % ( n  = 49), with an animal found excreting 1000 miracidia per 5 g of faeces. At this location, our praziquantel treatment ( n  = 14) and GPS animal tracking ( n  = 2) pilot sub-study compared two local goat herds over a three-month period. The daily foraging ranges across a 10 km 2 area were recorded, alongside targeted schistosome surveillance within local freshwater intermediate snail hosts. Analysis of GPS data revealed only one herd (infected) to have regular daily water contact with Lake Malawi whereas the other herd (not infected) totally avoided the lake. One week after praziquantel treatment administered at 40 mg/kg, anthelminthic cure rate was 92.3 % while at three months approximately a third of treated animals were shedding schistosome miracidia. Cercariae from several field-caught snails locally were genotyped, inclusive of finding a Schistosoma haematobium - mattheei hybrid. Our findings reveal the focalized nature of caprine schistosomiasis, signposting a novel alert for S. haematobium transmission, and highlight where zoonotic transmission can be intense. To better address zoonotic spill-over from S. mattheei (and/or S. haematobium ), the national control programme for schistosomiasis should formally develop targeted surveillance of caprine schistosomiasis and where appropriate, attempt an integrated One Health intervention in future., Competing Interests: The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (© 2024 The Author(s). Published by Elsevier B.V.)

Background: In areas of low disease endemicity, highly sensitive diagnostic tools to identify, diagnose, and monitor intestinal schistosomiasis transmission are needed to reliably measure the burden and risk of infection. Here, we used highly sensitive molecular diagnostic methods to investigate Schistosoma mansoni prevalence and transmission along the southern shoreline of Lake Malawi, five years post-disease outbreak., Methodology and Principal Findings: Faecal and urine samples were provided by school-aged children situated along the southern shoreline of Lake Malawi. Kato-Katz faecal-egg microscopy and point-of-care circulating cathodic antigen (POC-CCA) rapid diagnostic tests were then performed to diagnose infection with S. mansoni. Urine-egg microscopy was also used to diagnose infection with Schistosoma haematobium. In addition, Schistosoma miracidia were isolated from faecal material using a standard miracidium hatching technique. A two-step real-time PCR approach was then used to diagnose infection with S. mansoni using DNA isolated from faecal samples. Furthermore, isolated miracidia were genotyped to species level through PCR and Sanger sequencing. Phylogenetic analyses were then carried out to identify which previously defined S. mansoni cox1 lineage group S. mansoni miracidia were most closely related to. The measured prevalence of S. mansoni infection varied considerably depending on which diagnostic assay was used. When compared to real-time PCR, faecal-egg microscopy had a sensitivity of 9% and a specificity of 100%. When POC-CCA 'trace' results were considered positive, POC-CCA had a sensitivity of 73% and a specificity of 81% when compared to real-time PCR. However, when considered negative, POC-CCA sensitivity was reduced to 56%, whereas specificity was increased to 90%. In addition, a high degree of S. haematobium DNA was detected in DNA isolated from faecal samples and motile S. haematobium miracidia were recovered from faecal samples. Schistosoma mansoni miracidia were closely related to two independent cox1 lineage groups, suggesting multiple recent introduction and colonisation events originating from surrounding east African countries., Conclusions and Significance: Intestinal schistosomiasis is now highly prevalent along the southern shoreline of Lake Malawi just five years post-disease outbreak. In addition, a high prevalence of urogenital schistosomiasis persists. The revision of ongoing schistosomiasis control programmes in this area is therefore recommended. Our study also highlights the need for reliable diagnostic assays capable of distinguishing between Schistosoma species in multispecies co-endemic areas., Competing Interests: The authors have declared that no competing interests exist., (Copyright: © 2024 Archer et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

In Malawi, the putative origin of a newly described Schistosoma haematobium - mattheei hybrid human schistosome was assessed upon a seminal molecular parasitological survey of cattle. Using miracidia hatch test (MHT) and carcass inspection at slaughter, mean prevalence of bovine schistosomiasis was 49.1% (95% CI: 43.7-54.6%) and 10.3% (95% CI: 6.0-16.2%) respectively, though significant spatial heterogeneity was noted. Approximately 2.0% of infected cattle, and only those from Mangochi District, shed S. haematobium - mattheei and/or S. haematobium in faeces. To quantify schistosome (re)infection dynamics, where a S. haematobium - mattheei hybrid was present, we undertook a novel pilot GPS-datalogging sub-study within a specific herd of cattle ( n  = 8) on the Lake Malawi shoreline, alongside a praziquantel (40 mg/kg) treatment efficacy spot check. At sub-study baseline, all GPS-tagged cattle had proven daily water contact with the lake. Each animal was patently infected upon MHT, with older animals shedding less miracidia. At one month review, whilst parasitological cure was 100.0%, from six weeks onwards, (re)infection was first noted in the youngest animal. By three-month review, all animals were patently (re)infected though only miracidia of S. mattheei were recovered, albeit in much lower numbers. To conclude, infection with S. mattheei is particularly common in cattle and demonstrates a previously cryptic burden of bovine schistosomiasis. Within Mangochi District, bovine transmission of both S. haematobium - mattheei hybrids and S. haematobium are now incriminated, with unequivocal evidence of contemporary zoonotic spill-over. Future control of urogenital schistosomiasis here in the southern region needs to develop, then successfully integrate, a One Health approach with appropriate mitigating strategies to reduce and/or contain bovine schistosomiasis transmission., Competing Interests: The authors report no conflicts of interest., (© 2024 The Authors.)

Schistosomiasis is a neglected tropical disease (NTD) caused by infection with parasitic trematodes of the genus Schistosoma that can lead to debilitating morbidity and mortality. The World Health Organization recommend molecular xenomonitoring of Biomphalaria spp. freshwater snail intermediate hosts of Schistosoma mansoni to identify highly focal intestinal schistosomiasis transmission sites and monitor disease transmission, particularly in low-endemicity areas. A standardised protocol to do this, however, is needed. Here, two previously published primer sets were selected to develop and validate a multiplex molecular xenomonitoring end-point PCR assay capable of detecting S. mansoni infections within individual Biomphalaria spp. missed by cercarial shedding. The assay proved highly sensitive and highly specific in detecting and amplifying S. mansoni DNA and also proved highly sensitive in detecting and amplifying non- S. mansoni trematode DNA. The optimised assay was then used to screen Biomphalaria spp. collected from a S. mansoni- endemic area for infection and successfully detected S. mansoni infections missed by cercarial shedding as well as infections with non- S. mansoni trematodes. The continued development and use of molecular xenomonitoring assays such as this will aid in improving disease control efforts, significantly reducing disease-related morbidities experienced by those in schistosomiasis-endemic areas., Competing Interests: The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (© 2024 Published by Elsevier B.V.)