This is complementary information to the poster :
Putting research into practice: Identification of Haemonchus contortus in routine diagnostics
Poster nr 101
Full title: IDENTIFICATION OF HAEMONCHUS CONTORTUS ON EGGS USING REAL TIME PCR IN ROUTINE DIAGNOSTICS
Authors: Sara Hägglund, Bitte Ljungström, Pia Svedberg, Mikael Juremalm. Vidilab Enköping Sweden.
Conclusions
PCR provided a higher sensitivity allowing detection of lower infection burden of H.contortus compared to culturing. The ability to diagnose samples using parasite eggs, without the need of cultivation, shortens the time needed for analyses from 7+ to 2 days allowing faster diagnosis and ability to treat the affected animals in time. The method has been implemented into the daily routine diagnostics at Vidilab.
Introduction
Haemonchus contortus is one of the most pathogenic parasites in small ruminants. In Sweden testing for parasites of sheep is routine. Today most samples are analysed using McMaster flotation and if needed followed by larval cultivation (7+ days) and morphologic identification of L3 larvae for confirmation. The goal of the project was to find a faster way to confirm infection of H.contortus providing faster diagnosis to the farmer/veterinarian.
Materials and methods
Faeces were collected from routine sampling (n = 151). Samples were analysed by McMaster flotation, larval cultivation and PCR. Eggs were purified and concentrated by flotation with saturated NaCl-solution and washed with PBS. DNA extraction were done by bead beating and automated magnetic extraction. Amplification were done by rtPCR with primers and probes described by Reslova et al. (2021).
Laboratory Flow Chart
Results
Using clustered multiple comparison the cut off was set to a Cq-value of 32. Samples with a value of ≤ 32 were considered positive.
Among 151 samples, 40 were positive using larval cultivation. Using PCR, 52 samples were found to be positive for H.contortus. Of the 40 samples found positive using larval culture, 36 were found positive using PCR. The remaining 16 positive PCR samples were not detected using larval culture.
The sensitivity of the PCR-method was 90 % compared to culturing. However, considering that all PCR positive were true positive, the sensitivity of culturing was 71 %.
References
Nikol Reslova, Lucie Skorpikova, Iveta Angela Kyrianova, Jaroslav Vadlejch, Johan Höglund, Philip Skuce, Martin Kasny.
The identification and semi-quantitative assessment of gastrointestinal nematodes in faecal samples using multiplex real-time PCR assays. Parasit Vectors. 2021 Aug 9;14(1):391.