Strong-LAMP Assay Based on a Strongyloides spp.-Derived Partial Sequence in the 18S rRNA as Potential Biomarker for Strongyloidiasis Diagnosis in Human Urine Samples

Abstract

Human strongyloidiasis a soil-transmitted infection caused by Strongyloides stercoralis is one of the most neglected amongst the so-called Neglected Tropical Diseases (NTDs). S. stercoralis is a nematode, which is distributed worldwide; it has been estimated that it could affect millions of people, mainly in tropical and subtropical endemic regions. The difficulties of diagnosis lead to infection rates being underreported. Asymptomatic patients have chronic infections that can lead to severe hyperinfection syndrome or disseminated strongyloidiasis in immunocompromised patients. Strongyloidiasis can easily be misdiagnosed because conventional faecal-based techniques lack of sensitivity for the morphological identification of infective larvae in faeces. None of the currently used molecular methods have used urine samples as an alternative to faecal samples for diagnosing strongyloidiasis. This study was thus aimed at comparing, for the first time, the use of a new loop-mediated isothermal amplification (LAMP) molecular assay (Strong-LAMP) to traditional methods on patients’ urine samples. Twenty-four urine samples were taken from patients included in a study involving two Spanish hospitals for strongyloidiasis screening using parasitological and serological tests. Strongyloides larvae were found in 11 patients’ faecal samples, thereby ascertaining that they had the disease. Other patients had high antibody titres but no larvae were found in their faeces. All urine samples were analysed by PCR and Strong-LAMP assay. No amplification occurred when using PCR. Strong-LAMP led to detecting S. stercoralis DNA in urine samples from patients having previously confirmed strongyloidiasis by parasitological tests and/or a suspicion of being infected by serological ones. The Strong-LAMP assay is a useful molecular tool for research regarding strongyloidiasis in human urine samples. After further validation, the Strong-LAMP assay could also be used for complementary and effective diagnosis of strongyloidiasis in a clinical setting.

Loop-Mediated Isothermal Amplification in Schistosomiasis

“…This has allowed the use of alternative specimens, such as urine samples, for the molecular diagnosis of intestinal schistosomiasis [ 44 , 49 , 51 ]. As handling and storage of large numbers of stool samples for field surveys is very demanding, urine has been proposed as more suitable specimen for large-scale field studies [ 44 , 62 ]. Nevertheless, further evidence is needed to prove the usefulness of urine for intestinal schistosomiasis diagnosis.…”

Loop-Mediated Isothermal Amplification as Point-of-Care Diagnosis for Neglected Parasitic Infections

“…Regarding Strogyloides spp., LAMP sensitivity (10 −3 ng) was greater than PCR sensitivity (0.1 ng) when amplifying the 18S rRNA sequence from DNA extracted from S. venezuelensis [175]. No positive PCR results were obtained when using the same primers on urine samples [173]. Concerning T. trichiura, the set of primers which amplified the ITS-2 region designed by Ngari et al [172] was able to detect very low levels of infection, having greater sensitivity than the Kato-Katz technique.…”

Development of a Duplex LAMP Assay with Probe-Based Readout for Simultaneous Real-Time Detection of Schistosoma mansoni and Strongyloides spp. -A Laboratory Approach to Point-Of-Care

“…infection, based on a modified DARQ probe method [25]. The duplex DARQ-LAMP was based on two well-stablished conventional LAMP assays previously described by our group and successfully tested on samples from an experimental model of schistosomiasis [38] and strongyloidiasis [39] as well as on clinical samples, including application in field conditions [40][41][42].…”