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

Abstract

BackgroundTaenia 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 findingsPrimers 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.ConclusionsT3qPCR 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 deve...

The epidemiology of Taenia spp. infection and Taenia solium cysticerci exposure in humans in the Central Highlands of Vietnam

“…following DNA sequencing. The methods and results have been described and published elsewhere [ 14 ].…”

“…In Dak Lak province, there is no information available on the infection status of T. saginata in livestock, however our research data on the seroprevalence of cysticercosis in pigs was 0.95% [ 13 ]. A previous study carried out by our group using a newly developed multiplex real-time PCR found the prevalence of Taenia solium, T. saginata and T. asiatica taeniasis to be 1.2%, 5.8% and 1.5%, respectively in Dak Lak province [ 14 ]. In this study, our aims were to: i) estimate the seroprevalence of T. solium cysticerci exposure using the lentil-lectin purified glycoprotein enzyme-linked immunoelectrotransfer blot (LLGP-EITB) assay; ii) identify and quantify risk factors for human T. solium cysticerci exposure and taeniasis ( Taenia spp.…”

Spatial distribution of Taenia solium exposure in humans and pigs in the Central Highlands of Vietnam

“…Within the province, three districts namely Buon Don, Krong Nang and M’Drak were chosen as the study sites based on their diverse geographic characteristics ( Fig 1 ). The characteristics of these districts have been described in detail elsewhere [ 22 ]. A sampling frame listing the name of all villages in Dak Lak province was obtained from Sub-Department of Animal Health office within the Ministry of Agriculture and Rural Development.…”

“… The black shaded areas show the study locations in Buon Don, Krong Nang and M’Drak districts. This figure was adapted from Ng-Nguyen et al (2017) [ 22 ]. …”

“…Human stool samples were tested to determine T . solium tapeworm carriers using a real-time PCR (T3qPCR) described by Ng-Nguyen et al (2017) [ 22 ]. The assay has been reported to have a diagnostic sensitivity of 94% and a diagnostic specificity of 98%.…”