Molecular methods.

<div><p>Background</p><p>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 <i>Schistosoma mansoni</i> prevalence and transmission along the southern shoreline of Lake Malawi, five years post-disease outbreak.</p><p>Methodology and principal findings</p><p>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 <i>S</i>. <i>mansoni</i>. Urine-egg microscopy was also used to diagnose infection with <i>Schistosoma haematobium</i>. In addition, <i>Schistosoma</i> 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 <i>S</i>. <i>mansoni</i> 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 <i>S</i>. <i>mansoni cox</i>1 lineage group <i>S</i>. <i>mansoni</i> miracidia were most closely related to.</p><p>The measured prevalence of <i>S</i>. <i>mansoni</i> 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 <i>S</i>. <i>haematobium</i> DNA was detected in DNA isolated from faecal samples and motile <i>S</i>. <i>haematobium</i> miracidia were recovered from faecal samples. <i>Schistosoma mansoni</i> miracidia were closely related to two independent <i>cox</i>1 lineage groups, suggesting multiple recent introduction and colonisation events originating from surrounding east African countries.</p><p>Conclusions and significance</p><p>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 <i>Schistosoma</i> species in multispecies co-endemic areas.</p></div>