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Detection of phocine distemper virus using the polymerase chain reaction
Abstract:
During the fatal seal epizootics in the North and Baltic Seas in summer 1988 a virus was isolated which was shown to be the causal agent. It was subsequently classified as morbillivirus by neutralization assays, reaction with monoclonal antibodies and nucleic acid hybridization studies. The virus (tentatively called Phocine Distemper Virus, PDV) is difficult to grow in culture making rapid diagnosis difficult. We have used the Polymerase Chain Reaction (PCR) as an alternative and fast method to detect the presence of virus-specific nucleic acid and we describe here the amplification of cell culture derived PDV RNA in a "one-tube" reaction using heterologous (Rinderpest Virus cDNA derived) F gene primers. The resulting 370 bp DNA fragment was shown to be morbillivirus derived by Southern blot hybridization using cloned RPV F gene as probe.
Insights
Researchers developed a fast Polymerase Chain Reaction (PCR) method for detecting Phocine Distemper Virus (PDV). This technique aids in diagnosing the morbillivirus responsible for fatal seal epizootics, overcoming challenges with traditional culture methods.
Area of Science:
- Virology
- Marine Mammal Health
- Molecular Diagnostics
Background:
- Fatal seal epizootics in the North and Baltic Seas in 1988 were caused by an unidentified virus.
- The causative agent was classified as a morbillivirus, tentatively named Phocine Distemper Virus (PDV).
- Traditional methods for PDV diagnosis, such as virus culture, are slow and challenging.
Purpose of the Study:
- To develop a rapid and reliable method for detecting virus-specific nucleic acid.
- To utilize the Polymerase Chain Reaction (PCR) as an alternative diagnostic tool for PDV.
- To amplify PDV RNA using heterologous primers for efficient detection.
Main Methods:
- Isolation and classification of the causative virus from affected seals.
- Application of Polymerase Chain Reaction (PCR) for detecting viral RNA.
- Amplification of cell culture-derived PDV RNA in a one-tube reaction using Rinderpest Virus F gene primers.
- Southern blot hybridization with cloned Rinderpest Virus F gene probe to confirm morbillivirus origin.
Main Results:
- A 370 bp DNA fragment was successfully amplified from PDV RNA.
- The amplified DNA fragment was confirmed to be of morbillivirus origin.
- The PCR method proved to be a fast and effective alternative to conventional virus culture for PDV detection.
Conclusions:
- The developed PCR assay provides a rapid diagnostic tool for Phocine Distemper Virus.
- This molecular method overcomes the limitations of slow and difficult virus culture.
- The findings contribute to improved surveillance and understanding of morbillivirus in marine mammals.