Validation of alphavirus chimeras for diagnosis of Eastern and Venezuelan equine encephalitis
Journal of Equine Veterinary Science
cases. In a previous study, real-time (RT) PCRs for each parasite, targeting the rRNA 18s gene (1, 2), were adopted for their optimal performance. This study presents and discusses the results of the use of RT-PCRs in conjunction with ELISA (VMRD, USA) and IFAT (Fuller Laboratories, USA) that were used to analyse 274 horse sera. For comparison of results, PCR was used as reference method since sequencing of derived amplicons confirmed them as specific. The B. caballi RT-PCR detected 14 positive
... samples, none confirmed in ELISA, while the IFAT had a sensitivity (Se) of 50% (7/ 14) and a specificity (Sp) of 87.7 % (228/260). The T. equi detected 137 positive samples with the ELISA showing a Se of 67.2% (92/ 137) and an Sp of 83.2% (114/137) and the IFAT, a Se of 86.1% (118/ 137) and an Sp of 81% (111/137). For both parasites, the IFAT presented a limited major sensitivity when compared to the ELISA. Results for B. caballi are preliminary as number of samples recruited is limited. In particular, for T. equi the serological tests showed a high agreement and a relatively high specificity. An initial evaluation of the correlation between the PCR result and clinical status of the animal was carried out defining as cases those presenting an EP acute form based on temperature >38 C and at least one of the following signs, jaundice, anaemia, and petechial haemorrhages together with a PCR positive result. Of the PCR positive horses for B. caballi and T. equi, only 28.7% and 19.7% respectively were defined as cases, possibly due to parasite persistence beyond the acute form. Seropositve results of PCR negative samples could be due to sterilization of infected horses, occurring spontaneously or following treatment with a longer persistence of antibodies as usually occurs in most infections. An additional value of the use of quantitative PCR is that treatment efficacy may be monitored especially in view of the side effects this possess. This study demonstrates that the simultaneous use of PCR with serological tests increases the diagnostic probability to define the sanitary state for EP for the purposes stated above and underlines the need for revision of the prescribed diagnostic tests for trade that should include more sensitive methods.