Revista Colombiana de Ciencias Pecuarias
Print version ISSN 0120-0690
CORDERO CAMACHO, Claudia P et al. Detection of seven viruses and Mycoplasma in fetal bovine serum by real time PCR. Rev Colom Cienc Pecua [online]. 2011, vol.24, n.4, pp. 585-597. ISSN 0120-0690.
Objective: real time PCR analysis for the detection of seven bovine pathogenic viruses: Bovine Adenovirus (BAdV), Bovine Viral Diarrhea Virus Types 1 and 2 (BVDV-1 and BVDV-2), Bovine Respiratory Syncytial Virus (BRSV), Vesicular Stomatitis Virus (VSV), Bovine Parainfluenza Virus 3 (BPIV-3), Bovine Herpes Virus-1 (BoHV-1), and Mycoplasma was conducted using fetal bovine serum (FBS, MICROGEN®) obtained in Colombia, aiming to include it as part of the serum quality control. Methods: bovine derived MDBK and human derived HEp-2 cell lines were cultured with the test serum for 21 days, collecting supernatant and cellular samples every 7-days. Once DNA and RNA were extracted, the later was converted into cDNA and both samples were subjected to real time PCR using specific primers and Resolight® (DNA-binding fluorescent dye). Standard curves were generated using serial dilutions of cloned specific viral sequences. Accurate amplification and high efficiency was demonstrated in these reactions. Results: realtime PCR amplification did not show a persistent increase of viral counts in cultures during the 21-day follow-up. However, for vesicular stomatitis virus, a transient increase was observed at 7 and 14 days in both cell lines, but considered as not conclusive for viral presence. Conclusions: real time PCR analysis showed to be a suitable method for viral detection in fetal bovine serum samples and through this method no consistent viral or mycoplasma presence was detected in the MICROGEN® fetal bovine serum.
Keywords : bovine virus; fetal bovine serum; real time PCR; viral detection.