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Background   Parafilaria bovicola  (Nematoda: Filariidae) causes cutaneous bleedings in bovine species. Flies serve as intermediate hosts. In recent years, reports on bovine parafilariosis have become more frequent, corroborating the necessity of reliable diagnostic interventions especially since no molecular or serological test has been available. We aimed to establish a polymerase chain reaction assay to detect DNA of  P. bovicola  in flies, skin biopsies and serohemorraghic exudates of bleeding spots.    Methods  PCRs targeting the cytochrome  c  oxidase subunit 1 ( cox 1) gene and the internal transcribed spacer region (ITS) of the ribosomal RNA gene cluster were evaluated for their diagnostic sensitivity as well as performance and specificity on biopsy and serohemorrhagic exudate samples from  P. bovicola- infected cattle.    Results  Using serohemorrhagic exudates ( n  = 6), biopsies ( n  = 2) and flies ( n  = 1), the PCR targeting the  cox 1 gene resulted in a gel band of almost 700 bp. Cloning, sequencing, and removal of primer sequences yielded a 649-bp fragment of the  P. bovicola cox 1 gene. The PCR targeting the ITS region showed a band of about 1100 bp. Cloning, sequencing, and removal of primer sequences resulted in a 1083 bp stretch of the  P. bovicola  ITS region. Testing samples from presumably affected animals, the  cox 1-PCR resulted in bands with the expected size and they were all confirmed as  P. bovicola  by sequencing. In contrast, the ITS-PCR proved to be less sensitive and less specific and additionally amplified the ITS region of  Musca domestica  or buttercup DNA. When analysing for sensitivity, the  cox 1-PCR yielded visible bands up to 2 ng of genomic DNA, whereas the ITS-PCR produced bands up to 3 ng. In a plasmid dilution series, the minimum number of target DNA copies was 10 2  for the  cox 1-PCR and 10 1  in the ITS-PCR.    Conclusions  The evaluated  cox 1-PCR enables reliable detection of  P. bovicola  DNA in skin biopsies and serohemorrhagic exudates. This PCR and, to a limited extent, the ITS-PCR, may help evaluate different therapeutic approaches. Furthermore, the  cox 1-PCR may be useful for epidemiological studies on the geographical distribution of  P. bovicola . Further understanding of the epidemiology of this parasite will help develop and implement effective control strategies.

parasites and vectors

Diagnosing bovine parafilariosis: utility of the cytochrome c oxidase subunit 1 gene and internal transcribed spacer region for PCR detection of Parafilaria bovicola in skin biopsies and serohemorrhagic exudates of cattle