Contents lists available at ScienceDirect Acta Tropica journal homepage: www.elsevier.com/locate/actatropica PCR-based verification of positive rapid diagnostic tests for intestinal protozoa infections with variable test band intensity Sören L. Becker a,b,c, ⁎ , Ivan Müller a,b,d , Pascal Mertens e , Mathias Herrmann f , Leyli Zondie g , Lindsey Beyleveld g , Markus Gerber d , Rosa du Randt h , Uwe Pühse d , Cheryl Walter h , Jürg Utzinger a,b a Swiss Tropical and Public Health Institute, P.O. Box, CH-4002 Basel, Switzerland b University of Basel, P.O. Box, CH-4003 Basel, Switzerland c Institute of Medical Microbiology and Hygiene, Saarland University, Kirrberger Straße, Building 43, D–66421 Homburg/Saar, Germany d Department of Sport, Exercise and Health, University of Basel, Birsstrasse 320B, CH-4056 Basel, Switzerland e Coris BioConcept, Rue Jean Sonet 4A, B-5032 Gembloux, Belgium f Medical Faculty, University of Münster, P.O. Box, D-48149 Münster, Germany g Department of Medical Laboratory Sciences, Nelson Mandela Metropolitan University, P.O. Box 77000, Port Elizabeth, 6031, South Africa h Department of Human Movement Science, Nelson Mandela Metropolitan University, P.O. Box 77000, Port Elizabeth, 6031, South Africa ARTICLE INFO Keywords: Cryptosporidium spp. Giardia intestinalis Intestinal protozoa Multiplex PCR Rapid diagnostic test South Africa ABSTRACT Stool-based rapid diagnostic tests (RDTs) for pathogenic intestinal protozoa (e.g. Cryptosporidium spp. and Giardia intestinalis) allow for prompt diagnosis and treatment in resource-constrained settings. Such RDTs can improve individual patient management and facilitate population-based screening programmes in areas without microbiological laboratories for confirmatory testing. However, RDTs are difficult to interpret in case of ‘trace’ results with faint test band intensities and little is known about whether such ambiguous results might indicate ‘true’ infections. In a longitudinal study conducted in poor neighbourhoods of Port Elizabeth, South Africa, a total of 1428 stool samples from two cohorts of schoolchildren were examined on the spot for Cryptosporidium spp. and G. intestinalis using an RDT (Crypto/Giardia DuoStrip; Coris BioConcept). Overall, 121 samples were positive for G. intestinalis and the RDT suggested presence of cryptosporidiosis in 22 samples. After a storage period of 9–10 months in cohort 1 and 2–3 months in cohort 2, samples were subjected to multiplex PCR (BD Max™ Enteric Parasite Panel, Becton Dickinson). Ninety-three percent (112/121) of RDT-positive samples for G. intestinalis were confirmed by PCR, with a correlation between RDT test band intensity and quantitative pa- thogen load present in the sample. For Cryptosporidium spp., all positive RDTs had faintly visible lines and these were negative on PCR. The performance of the BD Max™ PCR was nearly identical in both cohorts, despite the prolonged storage at disrupted cold chain conditions in cohort 1. The Crypto/Giardia DuoStrip warrants further validation in communities with a high incidence of diarrhoea. 1. Introduction According to the 2010 Global Burden of Disease (GBD) study, in- testinal protozoa infections were responsible for more than 10 million disability-adjusted life years (DALYs) (Hotez et al., 2014; Murray et al., 2012). Intestinal protozoa are mainly acquired through ingestion of contaminated water or food and they affect the gastrointestinal tract (Shirley et al., 2012; Speich et al., 2016). Cryptosporidium spp. and Giardia intestinalis (syn.: G. lamblia and G. duodenalis), for instance, cause a wide variety of clinical manifestations, ranging from asymp- tomatic carriage to severe diarrhoea with subsequent exsiccosis (DuPont, 2016; Sow et al., 2016). Immunocompromised individuals (e.g. with HIV infection or alpha1-antitrypsin deficiency) are particu- larly vulnerable to intestinal protozoa infections (Becker et al., 2014; Stark et al., 2009). The long-term effects of chronic or multiple infec- tions, particularly among children in areas of poor sanitation, remain controversial. While some studies from high-endemicity settings sug- gest a causal role of both cryptosporidiosis and giardiasis to the de- velopment of malnutrition and persistent diarrhoea (Muhsen and Levine, 2012), other studies found no clear associations (Donowitz et al., 2016). Hence, a recent review highlighted the need for addi- tional, high-quality epidemiological and clinical data on giardiasis and http://dx.doi.org/10.1016/j.actatropica.2017.06.012 Received 8 April 2017; Received in revised form 7 June 2017; Accepted 10 June 2017 ⁎ Corresponding author at: Institute of Medical Microbiology and Hygiene, Saarland University, Kirrberger Straße, Building 43, D-66421, Homburg/Saar, Germany. E-mail address: soeren.becker@uks.eu (S.L. Becker). Acta Tropica 174 (2017) 49–55 Available online 17 June 2017 0001-706X/ © 2017 Elsevier B.V. All rights reserved. MARK