Evaluation of three extraction methods for molecular detection of Schistosoma mansoni infection in human urine and serum samples

Abstract

The diagnostic techniques based on polymerase chain reaction (PCR) for the detection of Schistosoma spp. DNA in stool, serum, plasma and urine has shown high sensitivity and specificity solving the problems for the low worm burdens and low transmission rates facing the routine microscopic diagnosis. Since PCR assays require efficient unbiased procedures of extraction and purification of nucleic acids. This study compared the efficiencies of simple, manual and feasible DNA extraction methods; a salting out and resin method, phenol/chloroform method to a commercial extraction kit through PCR analysis of human urine and serum samples spiked with known amounts of adult Schistosoma mansoni DNA confirmed by the application on real samples from patients. In artificially spiked urine gradient, the best mean diagnostic performance was that of salting out and resin then phenol/chloroform and last for the commercial kit. All three methods gave positive results in all tested urine samples which insures comparable high efficiency for DNA detection. In artificially spiked serum gradient, the highest mean diagnostic performance was obtained by the kit then salting out and resin and last by phenol chloroform. In patients' urine samples the phenol/chloroform method showed the highest mean diagnostic performance followed by the resin and then the kit. Using patients' serum samples the resin method showed equal mean diagnostic performance with the phenol/chloroform method which was higher compared to the kit. As regards sensitivity from urine samples the resin and phenol/chloroform showed equal results using artificial gradients and patients' samples. In serum samples the resin and phenol/chloroform showed equal results using artificial gradients while the resin showed better results in patients' samples. It is recommended to extract DNA from urine samples and to use the salting out and resin as a manual DNA extraction method from patients' samples for the molecular diagnosis of Schistosoma mansoni infection.

Recent advances in nucleic acid-based methods for detection of helminth infections and the perspective of biosensors for future development

“…The PCR is based on using the ability of DNA polymerase to synthesize a new strand of DNA complementary to the template strand. The technique has been successfully used for detecting schistosome DNA from host serum, urine and faeces as well as water containing cercaria and infected snails (Enk et al ., 2012; Sarhan et al ., 2015; Sengupta et al ., 2019). A new PCR-based approach has been shown to identify different Schistosoma species ( S. mansoni , S. japonicum , S. haematobium , S. intercalatum and S. bovis ) and is applicable to human urine samples with high specificity and sensitivity (Sandoval et al ., 2006).…”

Tools for Detection of Schistosomiasis in Resource Limited Settings

“…Recently, the use of molecular methods in the detection and characterisation of schistosomes have been perfected [ 50 , 51 ]. PCR has been used in detecting schistosomes in human serum, urine and faeces, water, snails, and many other sample types [ 47 , 52 , 53 , 54 , 55 ]. Amplification of cell-free DNA has higher detection sensitivity and specificity than KK or serum CCA detection [ 56 ].…”

Detection of Schistosoma mansoni DNA using polymerase chain reaction from serum and dried blood spot card samples of an adult population in North-western Tanzania

“…In the reviews of Weerakoon et al [ 8 , 21 ] all the different diagnostic methods were presented in great detail. Considering that the result of molecular diagnostic methods depends directly on the quantity and quality of the DNA used, the type of sample, sample conservation, sample storage as well as DNA extraction from these samples play a central role [ 16 , 22 – 24 ]. The samples usually have to be transported to a central laboratory for processing and cannot be examined directly in the field [ 8 ].…”