Abstract
BackgroundSchistosoma mansoni is the main species causing hepatic and intestinal schistosomiasis in Sub-Saharan Africa, and it is the only species in South America. Adult stages of the parasite reside in the mesenteric venous plexus of infected hosts, and eggs are shed in feces. Collecting patient stool samples for S. mansoni diagnostic purposes is difficult in large-scale field trials. Urine samples would be an alternative approach for molecular S. mansoni detection since they have several advantages over stool samples, including better handling, management and storage. Additionally, loop-mediated isothermal amplification (LAMP) technology is a powerful molecular diagnostic tool for infectious diseases, particularly under field conditions in developing countries. The present study aimed to assess the effectiveness of our previously developed LAMP assay (SmMIT-LAMP) for S. mansoni-specific detection in clinical urine samples.Methodology/Principal findingsThe sensitivity of SmMIT-LAMP in urine was established in simulated fresh human urine samples artificially spiked with genomic DNA from S. mansoni. LAMP for 120 min instead of 60 min improved the sensitivity, reaching values of 0.01 fg/μL. A set of well-defined frozen stored human urine samples collected from Sub-Saharan immigrant patients was selected from a biobank to evaluate the diagnostic validity of SmMIT-LAMP. The set included urine samples from patients with microscopy-confirmed infections with S. mansoni, S. haematobium and other nonschistosome parasites, as well as urine samples from patients with microscopy-negative eosinophilia without a confirmed diagnosis. The SmMIT-LAMP was incubated for 60 and 120 min. A longer incubation time was shown to increase the LAMP-positive results in patient urine samples. We also tested urine samples from mice experimentally infected with S. mansoni, and LAMP-positive results were obtained from the third week after infection. A real-time LAMP assay was also performed with three individual urine samples.Conclusions/SignificanceThe SmMIT-LAMP could effectively detect S. mansoni DNA in mouse urine samples and produced promising results for human clinical samples. The detection of S. mansoni DNA in mouse urine samples from the third week after infection indicates that early diagnosis of active S. mansoni infection is possible using urine as a source of DNA. Further studies are still needed, but our method could be used as a promising molecular tool applicable to urine samples to diagnose human intestinal schistosomiasis caused by S. mansoni.
Highlights
Human schistosomiasis, a parasitic disease caused by several species of trematode flatworms of the genus Schistosoma, is an endemic disease in 74 countries, infecting more than 200 million people worldwide [1]
We examine the application of SmMIT-loop-mediated isothermal amplification (LAMP) for urine in a mouse model of S. mansoni and in human urine samples, both spiked and naturally infected
The detection limit of LAMP was 100 fg/μL when incubating the tubes for 60 min (Fig 1A), whereas the detection limit was established at 0.01 fg/μL when incubating for 120 min (Fig 1B)
Summary
A parasitic disease caused by several species of trematode flatworms of the genus Schistosoma, is an endemic disease in 74 countries, infecting more than 200 million people worldwide [1]. The traditional Kato-Katz (KK) fecal microscopic examination for counting schistosome eggs and immunology-based analyses detecting schistosome-derived circulating anodic (CAA) and cathodic (CCA) antigens mainly lack sensitivity in low-intensity infections and posttreatment conditions [8, 9]. Antibody detection lacks specificity, causing a high level of cross-reactivity [9]. The detection of CAA and CCA has a number of potential advantages over specific antibody detection because both can be detected in urine [9, 10]. New technologies and better diagnostic tests for schistosomiasis in field conditions in endemic areas, as well as for routine diagnosis in developed countries, are still needed
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