ABSTRACT
VP2 gene coding region of a vaccinal strain [D78] of infectious bursal disease virus [IBDV] was cloned in a eukaryotic expression vector, pSec Tag2A. The gene was placed downstream of Ig kappa chain leader sequence, under the control of human cytomegalovirus [hCMV] immediate early enhancer and promoter. The construct pSec Tag2A-VP2 was transferred in COS-7 cell line and the expression and secretion of VP2 was assessed by dot blotting and antigen capture ELISA. The antibody used in the immunological assays was a neutralizing monoclonal antibody [1A6] against VP2. Positive reaction with the antibody indicated the construct was functional with respect to expression and secretion of a native VP2
ABSTRACT
To compare PCR and bacterial culture methods for diagnosis of subclinical mastitis caused by Staphylococcus aureus, 100 milk samples from cattle with subclinical mastitis and 20 samples from healthy cattle were collected and tested. The samples were cultured on selective blood agar and bacteria were identified by standard methods. DNA extracted from samples was subjected to PCR reaction with species specific primers and PCR products were analyzed by agarose gel electrophoresis. Based on the PCR results the prevalence of subclinical mastitis due to S. aureus was 25%. In the bacteriological culture of single milk sampling, S. aureus was isolated from the same samples being positive in PCR. A correlation of 100% was found between PCR and single milk sampling culture method by Mc Nemar test. All of the CMT negative samples were also negative in culture and PCR methods. The results of this study indicate that the PCR reaction is sensitive and specific for diagnosis of S. aureus in subclinical mastitis and can detect this pathogen in milk samples at species level in few hours