@article{Saarangi_Polapally_Rana_Bahekar_Surendra_Chandrasekhar Reddy_Raichur_Muthappa_Sharma_2020, title={Development and laboratory validation of duplex real-time PCR for simultaneous detection of Brucella and bovine alphaherpesvirus from clinical specimens}, volume={56}, url={https://veterinariaitaliana.izs.it/index.php/VetIt/article/view/1728}, DOI={10.12834/VetIt.1728.9123.2}, abstractNote={<p><span class="fontstyle0">A duplex real‑time PCR was developed and validated for the simultaneous detection of </span><em><span class="fontstyle2">Brucella </span></em><span class="fontstyle0">and bovine alphaherpesvirus‑1 (BoHV‑1) from bovine clinical specimens. The </span><span class="fontstyle2">bcsp31 </span><span class="fontstyle0">gene of </span><em><span class="fontstyle2">Brucella </span></em><span class="fontstyle0">and </span><em><span class="fontstyle2">gB </span></em><span class="fontstyle0">gene of BoHV‑1 were used as targets in the assay. The limit of detection for BoHV‑1 was 0.03 TCID</span><span class="fontstyle0" style="font-size: 5pt;">50 </span><span class="fontstyle0">of virus and 10 plasmid copies containing the target gene while for </span><em><span class="fontstyle2">Brucella </span></em><span class="fontstyle0">it was 4.1 × 10</span><span class="fontstyle0" style="font-size: 5pt;">1 </span><span class="fontstyle0">CFUs. Intra‑assay and inter‑assay values showed high repeatability and reproducibility of the assay. The diagnostic sensitivity (dsn) and diagnostic specificity (dsp) of the duplex assay were determined by screening 443 clinical specimens and<br>comparing the results with the respective individual assays. The dsn and dsp for detection of </span><span class="fontstyle2">Brucella </span><span class="fontstyle0">were found to be 95.24% and 95.65%, respectively whereas for BoHV‑1, the dsn (100%) and dsp (99.47%) were slightly higher. The duplex assay had a very good degree of agreement with the respective individual real‑time PCR test {kappa value 0.97 for </span><span class="fontstyle2"><em>Brucella</em> </span><span class="fontstyle0">and 0.95 for BoHV‑1}. The results of the current study suggest that the duplex assay would be a cost‑effective and time‑saving alternative for the individual real‑time PCR assay for the detection of </span><span class="fontstyle2">Brucella </span><span class="fontstyle0">and BoHV‑1.</span></p>}, number={1}, journal={Veterinaria Italiana}, author={Saarangi, Laxmi Narayan and Polapally, Supriya and Rana, Samir Kumar and Bahekar, Vijay Shriram and Surendra, Kota Sri Naga Leela and Chandrasekhar Reddy, Rachamreddy Venkata and Raichur, Aparna Sri and Muthappa, Ponnanna Nadikerianda and Sharma, Girish Kumar}, year={2020}, month={Apr.}, pages={35–41} }