Molecular characterisation of b-tubulin gene of fasciola hepatica isolates obtained from different hosts

Fasciolosis causes many economical problems in livestock such as loss of meat, milk and wool production. In recent era use of anti-helminthic drugs is common way for controlling Fasciolasis.Triclabendasole (TCBZ) is one of the most frequently used between thoose drugs. Careless and intensive use of that drug cause developing resistance against drug.In this study molecular characterisation of β-tubulin gene from Fasciolahepatica isolates of different hosts and geographic areas have been investigated for possible polyphormism indicating triclabendasole resistance.For this purpose adult Fasciolahepatica isolates from Erzurum and Kayseri have been collected from 20 sheep and 20 herds and they transported to laboratory in %70 ethanol. Then genomic DNA isolation have been done from each parasites separetely and with using spesific primers PCR have been designed for β-tubulin gene. The primers β-tub295f and β-tub1230r have been used for PCR ampilification and SSCP and RFLP analyses have been done for further investigation.β-tubulin gene of all samples succesfully amplified and spesific bant of 935 bp have been observed.Two bants of 350 and 390 bp have been observed with a result of cutting PCR products with MboIIenzyme. Three bants of 210, 340 and 540 bp have been obtained as a result of use of HphI enzyme.HindIIused as last ezyme cut all sampled and spesific bants as 380 and 450 bp have been obtained. Similar bant profiles have been observed of SSCP analyses of samples.Six samples have randomly selected and DNA sequence analysis have been performed. Samples have been obtained from sheep showed more nucleotide polymorphism than the samples from cows. Some nucleotide changes have been observed of first and the last part of protein sequences but the mutation 200th nucloetide which could be associated with triclebendasole resistant couldnt be identified. With this studyβ-tubulin gene polymorphism of F.hepaticaisolates from sheep and cattles of two distinct geographical ares of Turkey have been researched for first time. Despite of lack of datas which could be indicating drug resistance, using experimental studies with molecular studies can be a way of getting more accurate and clear data.