Commercial Simplex and Multiplex PCR Assays for the Detection of Intestinal Parasites Giardia intestinalis, Entamoeba spp., and Cryptosporidium spp.: Comparative Evaluation of Seven Commercial PCR Kits with Routine In-House Simplex PCR Assays

Abstract

Nowadays, many commercial kits allowing the detection of digestive parasites by DNA amplification methods have been developed, including simplex PCR assays (SimpPCRa) allowing the identification of a single parasite, and multiplex PCR assays (MultPCRa) allowing the identification of several parasites at once. Thus, aimed at improving the diagnosis of intestinal protozoal infections, it is essential to evaluate the performances of these new tools. A total of 174 DNA samples collected between 2007 and 2017 were retrospectively included in this study. Performances of four commercial SimpPCRa (i.e., CerTest-VIASURETM) and three MultPCRa (i.e., CerTest-VIASURETM, FAST-TRACK-Diagnostics-FTD-Stool-ParasiteTM and DIAGENODE-Gastroenteritis/Parasite-panel-ITM) were evaluated for the detection of Cryptosporidium spp., Entamoeba spp., and Giardia intestinalis in stool samples compared to our routinely used in-house SimpPCRa. Globally, the SimpPCRa showed better sensitivity/specificity for the detection of G. intestinalis, E. histolytica, E. dispar, and Cryptosporidium spp. (i.e., 96.9/93.6%; 100/100%; 95.5/100%; and 100/99.3%, respectively), compared to the three commercial MultPCRa tested. All in all, we showed that MultPCRa offer an interesting alternative for the detection of protozoans in stool samples depending on the clinical context.

Evaluation of a Novel Commercial Real-Time PCR Assay for the Simultaneous Detection of Cryptosporidium spp., Giardia duodenalis, and Entamoeba histolytica

“…Finally, the VIASURE assay achieved good diagnostic performance for the detection of E. histolytica , with sensitivity and specificity values of 0.96 and 1, respectively. This is in agreement with previous results obtained using the same assay and its singleplex variant ( 30 ). Similar results were also obtained by most commercial kits evaluated to date ( Table 1 ).…”

“…of 0.96 and 0.99, respectively. These values were in line with those (1 and 0.99, respectively) estimated in a recent French study with the same multiplex assay and its singleplex version ( 30 ). Large differences in the diagnostic performance for the detection of Cryptosporidium spp.…”

“…Regarding G. duodenalis , the VIASURE assay achieved diagnostic sensitivity and specificity of 0.94 and 1, respectively. Sensitivity values of 0.81 and 0.96 were previously obtained with the multiplex and singleplex versions of this kit, respectively ( 30 ). Other commercially available assays have been demonstrated to be particularly suited for the detection of G. duodenalis in clinical samples; these include the BD MAX ( 22 , 23 ), FilmArray ( 25 ), and NanoCHIP gastrointestinal panel (Savyon Diagnostics, Ashdod, IL, USA) ( 27 , 28 ) assays, all of which consistently achieve diagnostic sensitivities of >0.97.…”

“…have been reported for other commercially available multiplex qPCR assays (summarized in Table 1 ). Whereas sensitivity values of 1 have been achieved with the EasyScreen enteric parasite detection kit (Genetic Signatures, Sydney, Australia) ( 24 ) and the FilmArray gastrointestinal panel (BioFire Diagnostics, Salt Lake City, UT, USA) ( 25 ), lower values of 0.96 to 0.97 have been reported with the BD MAX enteric parasite panel (Becton, Dickinson and Company) ( 22 , 23 ), 0.87 with the RIDAGENE parasitic stool panel II (R-Biopharm AG, Germany) ( 29 ), 0.74 to 0.75 with the gastroenteritis/parasite panel I (Diagenode, Liège, Belgium) ( 4 , 29 , 30 ), and 0.53 to 0.64 with the FTD stool parasite assay (FAST-Track Diagnostics, Esch-sur-Alzette, Luxembourg) ( 29 , 30 ). As discussed before, these differences can be attributed, at least partially, to the inability of some assays to detect Cryptosporidium species other than C. hominis or C. parvum .…”

“…A major methodological contribution is the use of a large reference panel of well-characterized DNA samples. Most evaluation studies conducted previously were based on prospectively collected stool samples submitted to clinical laboratories for parasite investigation ( 4 , 12 , 22 – 28 ), whereas studies based on selected DNA panels were far less common ( 29 , 30 ). The latter approach allows additional benefits, including the inclusion of DNA samples from less prevalent or rare species/genotypes and animal-adapted genetic variants with zoonotic potential.…”