Novel Assays for Detection of Influenza Virus
- Conditions
- Influenza A Virus
- Interventions
- Diagnostic Test: PB2 gene RT-PCR; NS gene RT-PCR
- Registration Number
- NCT03924284
- Lead Sponsor
- The University of Hong Kong
- Brief Summary
Seasonal influenza virus causes an estimated 0.3-0.6 million deaths per year. Avian influenza virus H5N1, H7N9 and H5N6 has fatality rate of over 30%. Swine influenza viruses from pigs have also infected humans.
Molecular assays are now used routinely in the detection of influenza viruses. The M gene is often used as the target for all influenza A viruses because the nucleotide sequence of this gene is relatively conserved among all the influenza A viruses. The World Health Organization and the US Centers for Disease Control and Prevention (CDC) have published protocols for molecular detection of influenza A virus M gene.
However, recent studies have shown that mutations in the M gene have led to a reduced sensitivity of RT-PCR assay targeting this gene. Therefore, it is important to use alternative conserved genes as the target of RT-PCR. In this study, our aim is to evaluate two new RT-PCR assays that are based on PB2 and NS gene segment.
- Detailed Description
I. Background
* Seasonal influenza virus causes an estimated 0.3-0.6 million deaths per year. Avian influenza virus H5N1, H7N9 and H5N6 has fatality rate of over 30%. Swine influenza viruses from pigs have also infected humans.
* Molecular assays are now used routinely in the detection of influenza viruses. The M gene is often used as the target for all influenza A viruses because the nucleotide sequence of this gene is relatively conserved among all the influenza A viruses. The World Health Organization and the US Centers for Disease Control and Prevention (CDC) have published protocols for molecular detection of influenza A virus M gene.
* However, recent studies have shown that mutations in the M gene have led to a reduced sensitivity of RT-PCR assay targeting this gene. Therefore, it is important to use alternative conserved genes as the target of RT-PCR. In this study, our aim is to evaluate two new RT-PCR assays that are based on PB2 and NS gene segment
II. Study objective -To evaluate the sensitivity and specificity of 2 new RT-PCR assays
III. Overall study design
* The investigators will randomly retrieve archived nasopharyngeal and saliva specimens that were previously tested for influenza A virus using commercially available assays in our laboratory, tested for influenza A virus at the Public Health Laboratory Service Branch in Hong Kong. These specimens will be tested for influenza A virus by 4 different RT-PCR assays as listed below:
1. Our new RT-PCR assay targeting PB2 gene
2. Our new RT-PCR assay targeting NS gene
3. M gene RT-PCR published by the World Health Organization
4. M gene RT-PCR published by the US CDC
Sensitivity, specificity, positive predictive value and negative predictive value will be determined.
IV. Nucleic acid extraction and real-time reverse transcription-polymerase chain reaction (RT-PCR) for influenza A virus
* Saliva and nasopharyngeal specimens will be subjected to total nucleic acid (TNA) extraction by NucliSENS easyMAG (BioMerieux, Boxtel, Netherlands).
* Monoplex real-time RT-PCR assays for influenza A virus will be performed. The primers and probes for the M gene RT-PCR have been published by the WHO and the US CDC.
V. Sample size:
* The investigators will perform all 4 RT-PCR assays on a total of 320 specimens, including
* 80 nasopharyngeal specimens which tested positive for influenza A by commercially-available molecular assays or by testing performed at the Public Health Laboratory Services Branch in Hong Kong
* 80 nasopharyngeal specimens which tested negative for influenza A by commercially-available molecular assays or by testing performed at the Public Health Laboratory Services Branch in Hong Kong
* 80 saliva specimens which tested positive for influenza A by commercially-available molecular assays
* 80 saliva specimens which tested negative for influenza A by commercially-available molecular assays
Recruitment & Eligibility
- Status
- WITHDRAWN
- Sex
- All
- Target Recruitment
- Not specified
- Nasopharyngeal or saliva specimens of patients in Queen Mary Hospital of Hong Kong
- Tested for influenza A virus using a commercially available assay or by the Public Health Laboratory Services Branch in Hong Kong
- Insufficient specimen volume
Study & Design
- Study Type
- OBSERVATIONAL
- Study Design
- Not specified
- Arm && Interventions
Group Intervention Description Saliva negative PB2 gene RT-PCR; NS gene RT-PCR Saliva specimens that are tested negative for influenza A virus by commercially available assay or by the Public Health Laboratory Services Branch of Hong Kong NPA negative PB2 gene RT-PCR; NS gene RT-PCR NPA specimens that are tested negative for influenza A virus by commercially available assay or by the Public Health Laboratory Services Branch of Hong Kong Saliva positive PB2 gene RT-PCR; NS gene RT-PCR Saliva specimens that are tested positive for influenza A virus by commercially available assay or by the Public Health Laboratory Services Branch of Hong Kong NPA positive PB2 gene RT-PCR; NS gene RT-PCR NPA specimens that are tested positive for influenza A virus by commercially available assay or by the Public Health Laboratory Services Branch of Hong Kong
- Primary Outcome Measures
Name Time Method RT-PCR result Through study completion, an average of 2 months The result of RT-PCR can be positive or negative
- Secondary Outcome Measures
Name Time Method Cycle threshold value Through study completion, an average of 2 months The cycle threshold is a surrogate for viral load
Trial Locations
- Locations (1)
Queen Mary Hospital
ðŸ‡ðŸ‡°Hong Kong, Hong Kong