title: Differentiating Schistosoma haematobium from Schistosoma magrebowiei and other closely related schistosomes by polymerase chain reaction amplification of a species specific mitochondrial gene.
abstract: BACKGROUND
  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.
  
  
  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).
  
  
  CONCLUSIONS
  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.
authors.affiliation: Public Health Division, Molecular Parasitology Research Laboratory, Nigerian Institute of Medical Research, Yaba, Lagos State, Nigeria.
authors.forename: Olaoluwa P
authors.initials: OP
authors.lastname: Akinwale
authors.affiliation: Infectious Disease Cluster, Advanced Medical and Dental Institute, Universiti Sains Malaysia, Bandar Putra Bertam, 13200 Kepala Batas, Pulau Pinang, Malaysia.
authors.forename: Tang T
authors.initials: TT
authors.lastname: Hock
authors.affiliation: Department of Parasitology, College of Medicine, Taipei Medical University, Taipei 1101, Taiwan.
authors.forename: Fan
authors.initials: F
authors.lastname: Chia-Kwung
authors.affiliation: National Institute of Parasitic Diseases, Chinese Center for Disease Control and Prevention, Shanghai 20025, PR, China.
authors.forename: Qi
authors.initials: Q
authors.lastname: Zheng
authors.affiliation: National Institute of Parasitic Diseases, Chinese Center for Disease Control and Prevention, Shanghai 20025, PR, China.
authors.forename: Shen
authors.initials: S
authors.lastname: Haimo
authors.affiliation: National Institute of Parasitic Diseases, Chinese Center for Disease Control and Prevention, Shanghai 20025, PR, China.
authors.forename: Charles
authors.initials: C
authors.lastname: Ezeh
authors.affiliation: Public Health Division, Molecular Parasitology Research Laboratory, Nigerian Institute of Medical Research, Yaba, Lagos State, Nigeria ; Department of Parasitology, College of Medicine, Taipei Medical University, Taipei 1101, Taiwan.
authors.forename: Pam V
authors.initials: PV
authors.lastname: Gyang
keywords: Differential identification; Schistosoma haematobium; Schistosoma magrebowiei; nicotinamide adenine dinucleotide subunit 3 gene; polymerase chain reaction assay
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