A Novel qPCR Assay for the Detection of African Animal Trypanosomosis in Trypanotolerant and Trypanosusceptible Cattle Breeds

Abstract

This study was conducted to (i) determine the prevalence of African Animal Trypanosomosis (AAT) in tsetse challenged areas, (ii) compare conventional with qPCR detection systems and (iii) evaluate the host genetic background and biology as risk factors. AAT prevalence studies are often confronted with low levels of parasitaemia. Hence, we designed a novel qPCR assay using primers and species specific probes amplifying the Internal Transcribed Spacer 1 (ITS1) gene. Thereby all three AAT species could be detected simultaneously. 368 individuals from three cattle types (Baoulé, Zebu and hybrids) originating from 72 farms in Burkina Faso were analysed. Farmers were interviewed and morphometric measurements of the cattle taken. A chi-squared test and a logistic regression model were calculated to detect associations with infection. In our study, the overall rate of prevalence detected with the novel qPCR assay was 11.14%. Compared to conventional PCR we identified a concordance of 91.30%. We tested 41 animals positive for trypanosome DNA, five animals showed multiple infections. Zebus were twice as often infected (21.74%) compared to Baoulé (9.70%) and hybrids (9.57%). Trypanosoma vivax is the dominant species (9.24%), as compared to T. congolense (2.44%) and T. brucei (0.82%). The chi-squared tests linking the infection events to the breeds (Zebu vs. Baoulé and Zebu vs. hybrids) were on the border of significance. No significant association with other tested parameters could be detected. We introduce a novel qPCR technique for the fast, sensitive and simultaneous detection of the three AAT species. Our results suggest that associations with breed and infection exist since Zebu cattle are more likely to be infected compared to Baoulé and hybrids. Indigenous taurine cattle breeds, like the Baoulé, therefore provide a unique and valuable genetic resource.

Comparison of conventional and molecular techniques for Trypanosoma vivax diagnosis in experimentally infected cattle

“…South American isolates of T. vivax do present some genetic differences despite being phylogenetically related to African isolates (GARCIA et al, 2014). The present study was the first to use the qPCR protocol, developed for African isolates (SILBERMAYR et al, 2013), for one South American isolate, detecting the T. vivax DNA, "Lins" isolate. Clearly it will be possible to develop a rapid and accurate measurement of parasite numbers in whole blood through target DNA quantification.…”

“…It was also observed that the PCR assay possessed an identical capacity of detection for T. vivax DNA as the qPCR assay (61.1%). This similarity in detection capacity could be explained by the fact that the qPCR assay described by Silbermayr et al (2013) was directed to ITS1 region that has a high CG content, which can interfere with the diagnostic accuracy of qPCR (FIKRU et al, 2016). Fluctuations in parasitaemia and aparasitemic intervals make the diagnosis of animal trypanosomiasis challenging since it is difficult to directly detect the parasite, especially in the subpatent phase of infection (CADIOLI et al, 2015); thus, the use of more sensitive diagnostic tools such as PCR and qPCR is necessary.…”

“…TaqMan qPCR was performed as described by Silbermayr et al (2013). Primers for concurrent detection of the ITS1 regions of T. congolense, T. brucei and T. vivax and for the bovine toll-like-receptor 8 (TLR-8) (endogenous gene) were initially used with the addition of probes labelled with FAM fluorophore and BHQ2 quencher for T. vivax detection and HEX fluorophore and BHQ2 quencher for TLR-8 detection.…”

Trypanosomosis: potential driver of selection in African cattle

“…Silbermayr et al ( 2013 ) developed a novel qPCR assay for indication of infection status of animals with the three trypanosome species involved in AAT ( T. vivax, T. congolense , and T. brucei ) from blood samples of most of the animals involved in this study. Zebus were twice as often infected (21.74%) compared to Baoule (9.70%) and composites (9.57%).…”

Hematocrit alterations and its effects in naturally infected indigenous cattle breeds due to Trypanosoma spp. on the Adamawa Plateau - Cameroon

“…The pathological consequences become severe depending on the trypanosome species and sub-species, within a livestock, between livestock species and among breeds depending on the challenge and virulence of the strains [ 9 , 10 ]. Due to ignorance of the symptoms, signs and pathology of the disease, farmers tend to treat these animals with trypanocides without prior laboratory diagnosis and confirmation.…”