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Background   Taenia solium , the cause of neurocysticercosis (NCC), has significant socioeconomic impacts on communities in developing countries. This disease, along with taeniasis is estimated to infect 2.5 to 5 million people globally. Control of  T .  solium  NCC necessitates accurate diagnosis and treatment of  T .  solium  taeniasis carriers. In areas where all three species of  Taenia  tapeworms ( T .  solium ,  Taenia saginata  and  Taenia asiatica ) occur sympatrically, conventional microscope- and copro-antigen based diagnostic methods are unable to distinguish between these three  Taenia  species. Molecular diagnostic tools have been developed to overcome this limitation; however, conventional PCR-based techniques remain unsuitable for large-scale deployment in community-based surveys. Moreover, a real-time PCR (qPCR) for the discrimination of all three species of  Taenia  in human stool does not exist. This study describes the development and validation of a new triplex Taq-Man probe-based qPCR for the detection and discrimination of all three  Taenia  human tapeworms in human stools collected from communities in the Central Highlands of Vietnam. The diagnostic characteristics of the test are compared with conventional Kato Katz (KK) thick smear and copro-antigen ELISA (cAgELISA) method utilizing fecal samples from a community based cross-sectional study. Using this new multiplex real-time PCR we provide an estimate of the true prevalence of taeniasis in the source population for the community based cross-sectional study.    Methodology/Principal findings  Primers and TaqMan probes for the specific amplification of  T .  solium ,  T .  saginata  and  T .  asiatica  were designed and successfully optimized to target the internal transcribed spacer I (ITS-1) gene of  T .  solium  and the cytochrome oxidase subunit I (COX-1) gene of  T .  saginata  and  T .  asiatica . The newly designed triplex qPCR (T3qPCR) was compared to KK and cAgELISA for the detection of  Taenia  eggs in stool samples collected from 342 individuals in Dak Lak province, Central Highlands of Vietnam. The overall apparent prevalence of taeniasis in Dak Lak province was 6.72% (95% confidence interval (CI) [3.94–9.50]) in which  T .  solium  accounted for 1.17% (95% CI [0.37–3.17]), according to the T3qPCR. There was sympatric presence of  T .  solium ,  T .  saginata  and  T .  asiatica . The T3qPCR proved superior to KK and cAgELISA for the detection and differentiation of  Taenia  species in human feces. Diagnostic sensitivities of 0.94 (95% credible interval (CrI) [0.88–0.98]), 0.82 (95% CrI [0.58–0.95]) and 0.52 (95% CrI [0.07–0.94]), and diagnostic specificities of 0.98 (95% CrI [0.94–1.00]), 0.91 (95% CrI [0.85–0.96]) and 0.99 (95% CrI [0.96–1.00]) were estimated for the diagnosis of taeniasis for the T3qPCR, cAgELISA and KK thick smear in this study, respectively.    Conclusions  T3qPCR is not only superior to the KK thick smear and cAgELISA in terms of diagnostic sensitivity and specificity, but it also has the advantage of discriminating between species of  Taenia  eggs in stools. Application of this newly developed T3qPCR has identified the existence of all three human  Taenia  tapeworms in Dak Lak province and proves for the first time, the existence of  T .  asiatica  in the Central Highlands and the south of Vietnam.

plos neglected tropical diseases

Comparison of a new multiplex real-time PCR with the Kato Katz thick smear and copro-antigen ELISA for the detection and differentiation of Taenia spp. in human stools