Differentiating Schistosoma haematobium from Schistosoma magrebowiei and other closely related schistosomes by polymerase chain reaction amplification of a species specific mitochondrial gene

Abstract

Introduction:Schistosoma haematobium infection afflicts about 150 million people in 53 countries in Africa and the Middle East. In many endemic areas, S. haematobium is sympatric with Schistosoma bovis, Schistosoma mattheei, Schistosoma curassoni, Schistosoma intercalatum and Schistosoma magrebowiei, its closely related species. In addition, they also develop in the same intermediate snail hosts. Since these schistosome species often infect snails inhabiting the same bodies of water, examining cercariae or infected snails for estimating transmission of S. haematobium is always confounded by the need to differentially identify S. haematobium from these other species. Recently, differentiating S. haematobium by polymerase chain reaction (PCR) from S. bovis, S. mattheei, S. curassoni and S. intercalatum, but not from S. magrebowiei was reported. However, to be able to evaluate residual S. haematobium transmission after control interventions in areas where S. haematobium may be sympatric with S. magrebowiei, a differential tool for accurate monitoring of infected snails is needed.Materials and Methods:Thus in this study, we developed a new PCR assay using a pair of primers, ShND-1/ShND-2, to amplify a target sequence of 1117 bp (GenBank accession number KF834975) from S. haematobium mitochondrion complete genome (GenBank accession number DQ157222). Sensitivity of the assay was determined by PCR amplification of different concentrations of S. haematobium gDNA serially diluted from 10ng to 0.1pg. For assay specificity, different concentrations of gDNA from S. haematobium and the other schistosome species, 20 positive urine samples and five controls as well as 20 infected snails were subjected to PCR amplification, while some of the PCR products were sequenced.Results:The assay detected up to 1pg of S. haematobium gDNA, while a differential identification of S. haematobium DNA content from other closely related species was achieved when applied to urine and naturally infected snails. When a protein-protein blast search was carried out using Blastp, the amplified sequence was found to encode a protein that shows a 100% similarity with S. haematobium nicotinamide adenine dinucleotide dehydrogenase subunit 3 (GenBank accession number YP_626524.1).Conclusion:The PCR assay was sensitive, specific and was able to successfully differentiate S. haematobium from S. magrebowiei, in addition to its other closely related animal infective schistosome species.

Isothermal Recombinase Polymerase amplification (RPA) of Schistosoma haematobium DNA and oligochromatographic lateral flow detection

“…The Dra1 LF-RPA specificity to schistosome DNA was high with no cross-reaction with Enterobacteria or human gDNA. As found previously with RT-PCR [ 24 ], RPA Dra1 cross-amplification with S. mansoni was 1000 fold less sensitive than for S. haematobium but as predicted cross-amplification was successful with S. curassoni and S. bovis, both members of the S. haematobium group, known to possess the Dra1 repeat sequence [ 25 ]. Thus there is potential to utilise this assay and / or develop alternative species-specific assays for the identification of other S. haematobium -group species infections in livestock and humans.…”

A review of bovine fasciolosis and other trematode infections in Nigeria

“…Molecular identification of paramphistome species from Asia and elsewhere in Africa has been carried out (Lotfy et al, 2010); however, at time of writing this review, there are no records of such studies having been carried out in Nigeria. However, PCR to amplify species-specific genes has been used in differentiating human species of Schistosoma in Nigeria (Akinwale et al, 2014) and also to identify S. bovis in Kenya (Kamanja et al, 2011).…”

Nucleic acid detection in the diagnosis and prevention of schistosomiasis

“…This method was successfully applied to detect S. japonicum DNA in human stool samples [ 28 , 52 , 53 ] with low-intensity infections, and in pig feces [ 54 ]. Recently, a specific and efficient PCR assay based on NADH-3 [GenBank accession no: KF834975] was described and successfully used to detect snails and urine samples of patients infected with schistosome [ 55 ]. This approach can accurately differentiate S. haematobium from S. bovis , S. mattheei , S. curassoni , S. intercalatum , and S. magrebowiei , indicating a potential application in the identification of a coinfection.…”

“…This method may enable the accurate evaluation of infected snails where S. haematobium is coexistent with other prevalent schistosome species. Recently, a simple and efficient PCR technique based on the mitochondrial gene NADH-3 [GenBank accession no: KF834975] was described [ 55 ]. Its detection limit was shown to be 1 pg and no cross-reactions with other related schistosomes were observed.…”