TDK conference 2009

This thesis deals with occurrence and diversity of the Equine infectious anemia virus (EIAV) strains in Hungary. EIAV belongs to the Lentivirus genus of the family Retroviridae. The virus causes life-long infection and slowly developing lethal disease in equids. The virus is transmitted from viremic animals usually through blood by insect vectors; however other transmission routes (semen, milk, iatrogenic) may also occur. Symptoms in the acute phase include fever, lack of appetite, thrombocytopenia, and hemorrhage. In its chronic phase immune-mediated anemia and wasting develops. The disease occurs world-wide, but is usually sporadic. Due to its lethal character and to the lack of specific preventive methods (vaccination), control measures are based on the identification and destruction of infected animals. EIA appears on the list of communicable diseases of the World Organization for Animal Health (OIE). The Hungarian rules also order that EIAV-positive horses have to be destroyed. A reliable diagnostic system is necessary for the early detection of the infected animals, and hence for the effective eradication measures. Currently the gold-standard for EIA diagnosis is an agar-gel immunodiffusion test (the Coggins-test), but ELISA can also be used in surveys. These assays detect antibodies against EIAV, but they differ in sensitivity and specificity. In this study a reverse transcription polymerase chain reaction (RT-PCR) based diagnostic assay was developed for detection of EIAV RNA. Primers were selected to amplify a 313bp fragment of the partial 5’ untranslated region, and partial the gag gene region of EIAV, which was found to be a relatively conserved region, according to the comparison of the available EIAV nucleotide sequences. The assay was tested on samples from ELISA positive horses from Hungary, and sample from France was used as positive control. EIAV was detected in 7 out of 10 samples. The RT-PCR was capable to demonstrate the presence of the virus RNA in both spleen samples and from peripheral blood mononuclear cells. The nucleotide sequences of the amplification products were determined and subjected to phylogenetic analysis. Although one of the most conserved regions of the virus genome was investigated, the Hungarian strains showed considerable sequence diversity. This diversity indicates that several strains of EIAV circulate in the Hungarian horse populations; however the infections are often not diagnosed. The results of this study emphasize the importance of the regular, comprehensive and accurate testing of the Hungarian horse populations to EIAV. The RT-PCR assay, which was developed in this study, might be a useful tool for the validation of the results of serological tests, and for the genetic characterization of the strains.