Ocorrência Do Herpesvirus Bovino 1 (Bohv-1) No Líquido Folicular E Células Epiteliais De Oviduto Bovino

Bovine viral diarrhea virus (BVDV), bovine herpesvirus type 1 (BoHV-1), and bovine herpesvirus type 5 (BoHV-5) are major cattle pathogens that can be present in biological materials used in assisted reproduction biotechnologies. The aim of the present study was to increase the sensitivity of the polymerase chain reaction (PCR) for detection of BVDV, BoHV-1, and BoHV-5 in bovine follicular fluid (FF) collected during oocyte retrieval for in vitro embryo production. Ovaries were collected immediately after slaughter at a commercial abattoir, aspirated, and the 7336 samples of FF were pooled in 84 samples. Before testing the FF field samples, sensitivity of the protocol was determined using a prenucleic acid extraction procedure that was directly compared with standard RNA or DNA extraction protocols. The prenucleic acid extraction procedure increased sensitivity of reverse transcription (RT)-PCR for BVDV and nested PCR for BoHV-1 and BoHV-5 by 100 and 10 times, respectively. The 84 FF pools were assayed for BVDV, BoHV-1, and BoHV-5 using virus isolation and RT-PCR or nested PCR. Fourteen (16.7%) FF pools were positive for BVDV RNA, and one (1.2%) was positive for BoHV-1 DNA. Two of the BVDV RT-PCR positive samples and the one BoHV-1 PCR positive sample were also positive in cell culture, demonstrating that FF contained infectious viruses. In this study, the prenucleic acid extraction procedure increased the sensitivity of RT-PCR and PCR detection. This study highlighted the importance of assuring biosecurity by detecting the presence of viral pathogens in biological materials used during in vitro embryo production.

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Evaluation of prenucleic acid extraction for increasing sensitivity of detection of virus in bovine follicular fluid pools

Weber

Galuppo

Budaszewski

et al. 2013

Theriogenology

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Comparative Study of the Clastogenic Action of Bovine Herpesvirus 1 (Bohv-1) in Primary and Established Cell Cultures

Rocha¹,

Belloti²,

Pavão³

et al. 2010

VR&R

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The concern about the transmission of infectious diseases by the use of biotechnologies, like in vitro fertilization (IVF), has been increasing. The aim of this study was to verify the clastogenic alterations in the DNA of Madin-Darby Bovine Kidney (MDBK) and Bovine Oviduct Epithelial Cells (BOEC) infected or not with bovine herpesvirus 1 (BoHV-1), for 24 hours, by the use of the Comet Assay or SingleCell Gel Electrophoresis (SCGE). The nuclear change was evaluated by measuring the extent of DNA damage, where it was possible to observe 1.7% of level II comet and 2.1% of level IV in the BOEC group "in situ". However, in BOEC infected by the virus the damages observed were 13.8% comet level I, 0.6% level III and 1.2% level IV. The MDBK cells not infected group showed displayed 1.9% level II and the infected group 26 % level I, 12.3 % level II and 6.1 % level III. All groups, regardless if they were infected or not by the virus, were considered positive by the comet assay, maybe because there has not been virus association with the cell DNA. The number of comets detected in the BOEC "in situ" corresponds to spontaneous mutations that use to be present in organisms with failure of DNA repair. In vitro established and primary cells show greater failure frequency in DNA repair mechanisms, and may present more mutations. Therefore, in vitro models provide better conditions to mimic in vivo conditions. These data have demonstrated that not always the IVF problems are due to pathogens infection.

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Ocorrência Do Herpesvirus Bovino 1 (Bohv-1) No Líquido Folicular E Células Epiteliais De Oviduto Bovino

Cited by 2 publications

References 7 publications

Evaluation of prenucleic acid extraction for increasing sensitivity of detection of virus in bovine follicular fluid pools

Evaluation of prenucleic acid extraction for increasing sensitivity of detection of virus in bovine follicular fluid pools

Comparative Study of the Clastogenic Action of Bovine Herpesvirus 1 (Bohv-1) in Primary and Established Cell Cultures

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