scholarly journals A Comparison of Polymerase Chain Reaction with and without RNA Extraction and Virus Isolation for Detection of Bovine Viral Diarrhea Virus in Young Calves

2002 ◽  
Vol 14 (5) ◽  
pp. 433-437 ◽  
Author(s):  
D. Deregt ◽  
P. S. Carman ◽  
R. M. Clark ◽  
K. M. Burton ◽  
W. O. Olson ◽  
...  
Keyword(s):  
Virus Isolation ◽  
Rna Extraction ◽  
Bovine Viral Diarrhea ◽  
Chain Reaction ◽  
Pcr Assay ◽  
Diarrhea Virus ◽  
Viral Diarrhea ◽  
Polymerase Chain ◽  
Pcr Assays ◽  
Viral Diarrhea Virus

Previously, the authors described a multiplex reverse transcriptase–polymerase chain reaction (PCR) assay for detection and typing of bovine viral diarrhea virus (BVDV) from blood of persistently infected (PI) cattle that could be used with or without RNA extraction. In the present study, the PCR assay was evaluated for its ability to detect BVDV in young calves as a screening tool for detection of persistent infections. Both methods, PCR after RNA extraction (rPCR) and the direct method without RNA extraction (dPCR) were applied and compared with virus isolation (VI) with diagnostic specimens. From 450 whole blood samples from Ontario calves, 47 and 39 samples were positive by rPCR and VI, respectively. From the 47 samples positive by rPCR, 45 (96%) also were positive by dPCR when samples were tested both undiluted and diluted 1:10. In comparison to VI, the relative sensitivities of both PCR assays were 100%. Examination of the results indicates that both PCR assays can be used for screening calves for persistent infection with BVDV.

scholarly journals Comparison of Tests for Detection of Bovine Viral Diarrhea Virus in Diagnostic Samples

2007 ◽  
Vol 19 (4) ◽  
pp. 376-381 ◽  
Author(s):  
Misty A. Edmondson ◽  
M. Daniel Givens ◽  
Paul H. Walz ◽  
Julie A. Gard ◽  
David A. Stringfellow ◽  
...  
Keyword(s):  
Virus Isolation ◽  
Bovine Viral Diarrhea ◽  
Infected Cattle ◽  
Rt Pcr ◽  
Chain Reaction ◽  
Diarrhea Virus ◽  
Viral Diarrhea ◽  
Polymerase Chain ◽  
Viral Diarrhea Virus ◽  
Level Of Agreement

Currently, a variety of tests are used to detect bovine viral diarrhea virus (BVDV) in persistently infected (PI) cattle. These tests include immunohistochemical staining (IHC), antigen capture enzyme-linked immunosorbent assay (ACE), virus isolation (VI), and reverse transcription-polymerase chain reaction (RT-PCR). However, a lack of methods standardization could compromise the ability to consistently identify animals infected with BVDV. This study evaluated the diagnostic proficiency of current methods for detecting BVDV in infected cattle using intra- and interlaboratory comparisons. Samples were collected from 4 animals more than 7 months of age (2 BVDV negative animals, a PI animal, and a PI animal that previously lacked detectable virus in serum as determined by VI). Samples were submitted to 23 participating diagnostic laboratories using the respective laboratory's standard submission protocol. Samples collected for submission included: 1) serum for ACE, RT-PCR, and VI; 2) whole blood for RT-PCR and VI; and 3) skin biopsies for ACE and IHC. The ACE performed on skin provided the greatest consistency in detecting positive samples and a perfect level of agreement among laboratories. Reverse transcription-polymerase chain reaction and IHC performed well by correctly identifying ≤85% of samples positive for BVDV. Virus isolation performed on serum yielded the lowest consistency in detecting positive samples and the lowest level of agreement. The level of agreement between laboratories for detecting BVDV in persistently infected cattle ranged from perfect to less than expected by chance. The variation between laboratories suggests a need for training opportunities in standardized laboratory protocols and proficiency testing.

scholarly journals Comparison of Virus Isolation and Reverse Transcription Polymerase Chain Reaction Assay for Detection of Bovine Viral Diarrhea Virus in Bulk Milk Tank Samples

2000 ◽  
Vol 12 (2) ◽  
pp. 184-186 ◽  
Author(s):  
R. W. Renshaw ◽  
R. Ray ◽  
E. J. Dubovi
Keyword(s):  
Infected Animal ◽  
Virus Isolation ◽  
Rt Pcr ◽  
Chain Reaction ◽  
Pcr Assay ◽  
Diarrhea Virus ◽  
Bulk Milk ◽  
Polymerase Chain ◽  
Viral Diarrhea Virus ◽  
Bulk Milk Tank

The use of a reverse transcriptase polymerase chain reaction (RT-PCR) assay to screen bulk milk tank samples for bovine viral diarrhea virus (BVDV) has proven to be a sensitive and economical means to evaluate the lactating animals in a herd. The assay is capable of detecting the presence of a single persistently infected animal within a group of several hundred cows. Over a 3–year period, 144 samples from 97 farms were tested for BVDV using an RT-PCR assay in conjunction with a classical virus isolation (VI) procedure to measure the relative effectiveness of the techniques. Virus could be detected with both methods when the milk from a single persistently infected animal was diluted 1:600 with the milk from a herd of BVDV-negative animals. Based on individual farms, there was an overall prevalence of 12.4% BVDV infection, and the correlation between the 2 assays was 95.9%. In terms of sensitivity, specificity, and turnaround time, RT-PCR was superior to VI. However, of the 17 samples that were VI positive, 4 were RT-PCR negative. RT-PCR may not detect all naturally occurring BVDV isolates because they may contain minor sequence variations in the primer regions. VI and RT-PCR are both suitable for detection of BVDV in bulk milk samples when used independently, but to increase the probability of successful detection and to provide cross-checks against assay contamination, it is desirable to utilize both methods in parallel.

scholarly journals Deteksi Kontaminasi Bovine Viral Diarrhea Virus Pada Fetal Bovine Serum Yang Tersedia Secara Komersial

2021 ◽  
Vol 22 (2) ◽  
pp. 229-236
Author(s):  
Hastari Waryastuty ◽  
Sri Handayani Irianingsih ◽  
Raden Wasito
Keyword(s):  
Bovine Viral Diarrhea Virus ◽  
Gold Standard ◽  
Fetal Bovine Serum ◽  
Bovine Viral Diarrhea ◽  
Chain Reaction ◽  
Diarrhea Virus ◽  
Viral Diarrhea ◽  
Fetal Bovine ◽  
Polymerase Chain ◽  
Viral Diarrhea Virus

Teknik diagnostik menggunakan kultur sel merupakan gold standard untuk menentukan penyebab utama suatu penyakit terutama yang disebabkan oleh virus seperti bovine viral diarrhea virus (BVDV). Prosedur kultur sel memerlukan suplementasi serum pada media pertumbuhan. Serum yang digunakan sebagai suplementasi umumnya berasal dari hewan spesies yang sama agar sel dapat tumbuh sesuai dengan habitatnya. Serum yang paling umum digunakan dalam teknik kultur sel adalah fetal bovine sera (FBS) karena kandungan faktor pertumbuhannya yang tinggi. Infeksi BVDV sering terjadi pada populasi sapi. Infeksi BVDV in utero menyebabkan materi (serum, sel dan jaringan) yang berasal dari fetus sapi terkontaminasi BVDV. Penelitian telah membuktikan bahwa dua dari sepuluh sampel FBS dengan merek dagang yang berbeda, menunjukkan hasil positif terhadap virus BDV dengan uji polymerase chain reaction.. Isolasi terhadap BVDV dari FBS yang positif dilakukan pada kultur sel Madin-Darby bovine kidney (MDBK), dilanjutkan dengan pengecatan menggunakan uji immuno peroxidase monolayer assay, memberikan hasil negatif. Meskipun BVDV yang terkandung di dalam FBS sampel tidak bereplikasi, tetapi hasil ini tidak dapat diekstrapolasikan untuk setiap produk FBS yang tersedia secara komersial. Berdasarkan hasil tersebut disarankan adanya pengujian rutin terhadap kemungkinan terjadinya kontaminasi oleh BVDV yang dapat berpengaruh pada hasil diagnostik di suatu laboratorium.

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