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Detection of rodent parvoviruses by use of fluorogenic nuclease polymerase chain reaction assays
1University of Arizona, Central Animal Facility, Tucson 85721-0101, USA.
Abstract:
Polymerase chain reaction (PCR) assays have proven useful for detection of rodent parvoviruses in animals and contaminated biological materials. Fluorogenic nuclease PCR assays combine PCR with an internal fluorogenic hybridization probe, eliminating post-PCR processing and potentially enhancing specificity. Consequently, three fluorogenic nuclease PCR assays were developed, one that detects all rodent parvoviruses, one that specifically detects minute virus of mice (MVM), and one that specifically detects mouse parvovirus 1 (MPV) and hamster parvovirus (HaPV). When rodent parvoviruses and other rodent DNA viruses were evaluated, the rodent parvovirus assay detected only rodent parvovirus isolates, whereas the MVM and MPV/HaPV assays detected only the MVM or MPV/ HaPV isolates, respectively. Each assay detected the equivalent of 10 or fewer copies of target template, and all fluorogenic nuclease PCR assays exceeded the sensitivities associated with previously reported PCR assays and mouse antibody production testing. In addition, each fluorogenic nuclease PCR assay detected the targeted parvovirus DNA in tissues obtained from mice experimentally infected with MVM or MPV. Results of these studies indicate that fluorogenic nuclease PCR assays provide a potentially high-throughput, PCR-based method to detect rodent parvoviruses in infected mice and contaminated biological materials.
Insights
New fluorogenic nuclease PCR assays offer sensitive and specific detection of rodent parvoviruses. These methods improve detection in infected mice and contaminated materials, aiding in disease control.
Area of Science:
- Molecular Biology
- Virology
- Biotechnology
Background:
- Polymerase chain reaction (PCR) is valuable for detecting rodent parvoviruses.
- Fluorogenic nuclease PCR assays enhance specificity and eliminate post-PCR processing.
Purpose of the Study:
- Develop and evaluate fluorogenic nuclease PCR assays for rodent parvovirus detection.
- Establish assays for broad rodent parvovirus detection, specific minute virus of mice (MVM) detection, and specific mouse parvovirus 1 (MPV)/hamster parvovirus (HaPV) detection.
Main Methods:
- Developed three distinct fluorogenic nuclease PCR assays.
- Tested assay specificity against various rodent parvoviruses and other DNA viruses.
- Assessed assay sensitivity by detecting low copies of target DNA.
- Validated assays using tissues from experimentally infected mice.
Main Results:
- All developed assays demonstrated high specificity for their target rodent parvoviruses.
- Assays detected as few as 10 copies of target DNA, surpassing previous PCR methods and mouse antibody production testing.
- Target parvovirus DNA was successfully detected in tissues from MVM- or MPV-infected mice.
Conclusions:
- Fluorogenic nuclease PCR assays provide a sensitive and specific method for rodent parvovirus detection.
- These assays are effective in detecting parvoviruses in infected animals and contaminated biological materials.
- The developed assays offer a potentially high-throughput solution for rodent parvovirus surveillance and diagnostics.
