Differentiation between porcine epidemic diarrhea virus and transmissible gastroenteritis virus in formalin-fixed paraffin-embedded tissues by multiplex RT-nested PCR and comparison with in situ hybridization

Kwonil Jung; Junghyum Kim; Okjin Kim; Bongtae Kim; Chanhee Chae

doi:10.1016/s0166-0934(02)00253-7

Differentiation between porcine epidemic diarrhea virus and transmissible gastroenteritis virus in formalin-fixed paraffin-embedded tissues by multiplex RT-nested PCR and comparison with in situ hybridization

J Virol Methods. 2003 Mar;108(1):41-7. doi: 10.1016/s0166-0934(02)00253-7.

Authors

Kwonil Jung¹, Junghyum Kim, Okjin Kim, Bongtae Kim, Chanhee Chae

Affiliation

¹ Department of Veterinary Pathology, College of Veterinary Medicine and School of Agricultural Biotechnology, Seoul National University, Seoul, Kwanak-Gu 151-742, South Korea.

PMID: 12565152
DOI: 10.1016/s0166-0934(02)00253-7

Abstract

Porcine epidemic diarrhea virus (PEDV) and transmissible gastroenteritis virus (TGEV) were detected and differentiated in formalin-fixed, paraffin-embedded tissues from experimentally and naturally infected pigs by multiplex reverse transcription-nested polymerase chain reaction (RT-nPCR). The results of this new method were compared with in situ hybridization. A method based on xylene deparaffinization followed by proteinase K digestion yielded RNA of a suitable quality for reliable and consistent multiplex RT-nPCR analyses. PEDV and TGEV cDNAs were detected in jejunal tissues from experimentally and naturally infected pigs by multiplex RT-nPCR. Distinct positive signals for PEDV and TGEV were also detected in the same jejunal tissues by in situ hybridization. The rate of conformity between multiplex RT-nPCR and in situ hybridization was 100% for the detection of PEDV and TGEV in formalin-fixed paraffin-embedded jejunal tissues.

Publication types

Comparative Study
Research Support, Non-U.S. Gov't

MeSH terms

Animals
Base Sequence
Coronavirus / classification
Coronavirus / genetics*
Coronavirus / isolation & purification*
Coronavirus M Proteins
DNA Primers / genetics
DNA, Viral / genetics
DNA, Viral / isolation & purification
Formaldehyde
Gastroenteritis, Transmissible, of Swine / epidemiology
Gastroenteritis, Transmissible, of Swine / virology
In Situ Hybridization
Jejunum / virology
Molecular Epidemiology
Paraffin Embedding
Reverse Transcriptase Polymerase Chain Reaction
Sensitivity and Specificity
Sus scrofa
Tissue Fixation
Transmissible gastroenteritis virus / classification
Transmissible gastroenteritis virus / genetics*
Transmissible gastroenteritis virus / isolation & purification*
Viral Matrix Proteins / genetics
Virology / methods*
Virology / statistics & numerical data

Substances

Coronavirus M Proteins
DNA Primers
DNA, Viral
M protein, Porcine epidemic diarrhea virus
M protein, Transmissible gastroenteritis virus
Viral Matrix Proteins
Formaldehyde