Primena lančane reakcije polimeraze u dokazivanju prisustva goveđeg herpesvirusa 1 u kliničkim uzorcima materijala

The objective of this study was the application of polymerase chain reaction in detection of bovine herpesvirus 1 in clinical samples. Twenty samples of bovine nasal swabs were separately inoculated in Vero cells. Laboratory strain TN 41 of bovine herpesvirus 1 served as a control in the experiment. The cytopathic effects were recovered in the cell line previously inoculated with a sample of bovine nasal swab, as well as in cells inoculated with the laboratory strain TN 41 of the bovine herpesvirus 1 after a period of incubation of 24h, 36h and 48h. Identification of the isolated strains of the virus was done by the virus - neutralization test with specific immune sera against BHV 1, with a titre of 1:16. Comparative analysis of DNA fragments of the laboratory strain of BHV1 and isolated strain obtained by polymerase chain reaction with primers for viral gB glycoprotein and thymidine-kinase coding region, confirmed that the isolated strain of the virus belongs to bovine herpesvirus 1 (BHV 1). Cilj nàših istraživanja je bio primena lančane reakcije polimeraze u dokazivanju prisustva goveđeg herpesvirusa 1 u kliničkim uzorcima materijala. Dvadeset uzoraka nosnih briseva goveda pojedinačno je inokulisano u ćelijsku liniju Vero. Referentni soj TN 41 virusa BHV 1 služio je kao pozitivna kontrola u ogledu. Posle 24h, 36h i 48h od inokulacije, utvrđena je pojava citopatogenog efekta u ćelijskoj liniji inokulisanoj uzorkom poreklom od jednog nosnog brisa goveda. Iste promene su ustanovljene i posle inokulacije referentnog soja TN 41 virusa BHV 1 u ćelijsku liniju Vero. Identifikacija izolovanog soja virusa poreklom iz uzorka nosnog brisa goveda, vršena je primenom testa virus - neutralizacije uz korišćenje specifičnog imunog seruma protiv virusa BHV 1, titra od 1:16. Uporednom analizom DNK fragmenata referentnog soja virusa BHV 1 i izolovanog soja virusa, dobijenih primenom lančane reakcije polimeraze uz korišćenje dva para prajmera koji su amplifikovali gene na molekulu DNK koji kodiraju sintezu glikoproteina B spoljašnjeg omotača virusa BHV 1 i timidin - kinaze, potvrđena je pripadnost izolovanog soja virusa goveđem herpesvirusu 1.