Performance of the Kato-Katz method and real time polymerase chain reaction for the diagnosis of soil-transmitted helminthiasis in the framework of a randomised controlled trial: treatment efficacy and day-to-day variation

Abstract

Background Accurate, scalable and sensitive diagnostic tools are crucial in determining prevalence of soil-transmitted helminths (STH), assessing infection intensities and monitoring treatment efficacy. However, assessments on treatment efficacy comparing traditional microscopic to newly emerging molecular approaches such as quantitative Polymerase Chain Reaction (qPCR) are scarce and hampered partly by lack of an established diagnostic gold standard. Methods We compared the performance of the copromicroscopic Kato-Katz method to qPCR in the framework of a randomized controlled trial on Pemba Island, Tanzania, evaluating treatment efficacy based on cure rates of albendazole monotherapy versus ivermectin-albendazole against Trichuris trichiura and concomitant STH infections. Day-to-day variability of both diagnostic methods was assessed to elucidate reproducibility of test results by analysing two stool samples before and two stool samples after treatment of 160 T. trichiura Kato-Katz positive participants, partially co-infected with Ascaris lumbricoides and hookworm, per treatment arm (n = 320). As negative controls, two faecal samples of 180 Kato-Katz helminth negative participants were analysed. Results Fair to moderate correlation between microscopic egg count and DNA copy number for the different STH species was observed at baseline and follow-up. Results indicated higher sensitivity of qPCR for all three STH species across all time points; however, we found lower test result reproducibility compared to Kato-Katz. When assessed with two samples from consecutive days by qPCR, cure rates were significantly lower for T. trichiura (23.2 vs 46.8%), A. lumbricoides (75.3 vs 100%) and hookworm (52.4 vs 78.3%) in the ivermectin-albendazole treatment arm, when compared to Kato-Katz. Conclusions qPCR diagnosis showed lower reproducibility of test results compared to Kato-Katz, hence multiple samples per participant should be analysed to achieve a reliable diagnosis of STH infection. Our study confirms that cure rates are overestimated using Kato-Katz alone. Our findings emphasize that standardized and accurate molecular diagnostic tools are urgently needed for future monitoring within STH control and/or elimination programmes.

Efficacy and safety of moxidectin and albendazole compared to ivermectin and albendazole co-administration in adolescents infected with Trichuris trichiura: a randomized controlled trial protocol

“…Since its last assessment for human helminthiases 16,17 , the tool has undergone substantial development and we aim to provide data on its applicability within the framework of this trial. Diagnostic assessment by qPCR has been recently shown by our group to be more sensitive compared to standard Kato-Katz microscopy 28 .…”

“…An in-house method based on previous work from our group will be used. Details on procedures, reagents, kits and conditions have been described in a recent publication by Keller et al 28 .…”

Towards soil-transmitted helminths transmission interruption: The impact of diagnostic tools on infection prediction in a low intensity setting in Southern Mozambique

“…It is inferred that qPCR could be detecting DNA from other worm specimens than the egg [ 8 , 36 , 45 ], and thus, detecting prepatent infections (immature worms not releasing eggs) [ 44 ]. Other studies found better correlation between eggs per gram and PCR results, but they were using other PCR outcomes such as DNA copies [ 38 , 43 , 46 , 47 ].…”

“…Multiparallel qPCR showed cost diminution [ 46 ] and some studies are attempting to optimize a sample pooling protocol to make qPCR surveillance more cost-effective [ 52 ]. Moreover, sample storage, DNA extraction and qPCR protocols should be harmonized [ 45 , 47 , 53 ]. In addition, an external quality assurance for reference labs should be established [ 49 , 54 ].…”

Long-term outcomes of ivermectin-albendazole versus albendazole alone against soil-transmitted helminths: Results from randomized controlled trials in Lao PDR and Pemba Island, Tanzania

“…For this reason, the Kato-Katz method may lead to an underestimation of the true prevalence, which in turn could result in an artificial inflation of CRs from undetected residual low-egg count infections post-treatment. Indeed, a comparison of Kato-Katz and qPCR efficacy results 14–21 days post-treatment on Pemba Island revealed significantly lower CRs for ivermectin-albendazole, when two fecal samples before and after treatment were analysed by qPCR [ 42 ]. Since the majority of participants had light infections at follow-up, our resulting apparent reinfection rates and new infections should therefore be interpreted with caution.…”