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Detection of feline norovirus using commercial real-time RT-PCR kit for the diagnosis of human norovirus infection.

ABSTRACT: Feline noroviruses (FNoVs) are potential clinical pathogens in cats. To perform an epidemiological study of FNoV infection, it is necessary to develop a simple and effective method for virus detection. We investigated whether a commercial human NoV quantitative RT-PCR kit for the detection of human NoVs used in medical practice can be applied for FNoV detection. This kit was capable of detecting the FNoV gene regardless of the genogroup (GIV and GVI) in experimental and field samples. Based on the above findings, it is possible to detect FNoVs using human NoV tests. The relationship between FNoV infection and gastroenteritis in cats may be clarified by applying these methods to an epidemiological survey of FNoVs.

SUBMITTER: Takano T

PROVIDER: S-EPMC8182310 | biostudies-literature | 2021 May

REPOSITORIES: biostudies-literature

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Detection of feline norovirus using commercial real-time RT-PCR kit for the diagnosis of human norovirus infection.

Takano Tomomi T Watanabe Haruna H Doki Tomoyoshi T Kusuhara Hajime H

The Journal of veterinary medical science 20210311 5

Feline noroviruses (FNoVs) are potential clinical pathogens in cats. To perform an epidemiological study of FNoV infection, it is necessary to develop a simple and effective method for virus detection. We investigated whether a commercial human NoV quantitative RT-PCR kit for the detection of human NoVs used in medical practice can be applied for FNoV detection. This kit was capable of detecting the FNoV gene regardless of the genogroup (GIV and GVI) in experimental and field samples. Based on t ...[more]

PMID: 33692232

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Project description:IntroductionQuery (Q) fever is an important zoonosis and a cause of concern for humans, due to the potential bioterrorism threat posed by the causative agent, Coxiella burnetii. Because of the danger of contracting the illness, isolation attempts are seldom made. Serological and molecular diagnostic tests are the main option.AimTo study the prevalence of acute Q fever in Puducherry and surrounding districts of Tamil Nadu, India, employing a new commercial Real-Time Polymerase Chain Reaction (RT-PCR) kit and confirming it by the gold standard Immunofluorescence Assay (IFA).Materials and methodsAcute phase blood samples from 72 consecutive febrile patients and 24 healthy individuals were included in this prospective study. DNA was extracted from the buffy coats and preserved at -80°C. Detection of C. burnetii was carried out employing a commercial Real-Time PCR kit. Serum samples were tested for IgM (Phase I+II) and IgG (Phase I+II) by QM-120 and QG-120, Coxiella burnetii IFA Fuller Laboratories, California, USA. Sensitivity, Specificity, Positive Predictive Value (PPV) and Negative Predictive Value (NPV) were calculated keeping IFA as the reference.ResultsPresumptive diagnosis of acute Q fever was made in two febrile patients by the Genesig Easy kit (2.78%). In addition to these two PCR positive cases, one more patient was positive for both Phase II IgM and Phase II IgG antibodies by the gold standard IFA. All 24 healthy controls were negative for Q fever by both PCR and IFA. The sensitivity, specificity, NPV and PPV for Genesig Easy kit PCR were: 66.67%, 100%, 100% and 98.57 % respectively against IFA as the reference.ConclusionThe true prevalence of Q fever in India and other developing countries is poorly understood, owing to the difficulties in the diagnosis of this infection. Since molecular diagnostic tests have good specificity and are mandated for confirmation of single acute samples, validation of commercial Q fever PCR kits is the need of the hour. Genesig Easy kit in our hands was found to be reliable with the moderate sensitivity and high specificity. Performing both PCR (with acute specimens) and IFA (with paired sera) would be ideal for Q fever diagnosis.

Project description:Rapid and accurate detection of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is essential for the successful control of the current global COVID-19 pandemic. The real-time reverse transcription polymerase chain reaction (Real-time RT-PCR) is the most widely used detection technique. This research describes the development of two novel multiplex real-time RT-PCR kits, AccuPower® COVID-19 Multiplex Real-Time RT-PCR Kit (NCVM) specifically designed for use with the ExiStation™48 system (comprised of ExiPrep™48 Dx and Exicycler™96 by BIONEER, Korea) for sample RNA extraction and PCR detection, and AccuPower® SARS-CoV-2 Multiplex Real-Time RT-PCR Kit (SCVM) designed to be compatible with manufacturers' on-market PCR instruments. The limit of detection (LoD) of NCVM was 120 copies/mL and the LoD of the SCVM was 2 copies/μL for both the Pan-sarbecovirus gene and the SARS-CoV-2 gene. The AccuPower® kits demonstrated high precision with no cross reactivity to other respiratory-related microorganisms. The clinical performance of AccuPower® kits was evaluated using the following clinical samples: sputum and nasopharyngeal/oropharyngeal swab (NPS/OPS) samples. Overall agreement of the AccuPower® kits with a Food and Drug Administration (FDA) approved emergency use authorized commercial kit (STANDARD™ M nCoV Real-Time Detection kit, SD BIOSENSOR, Korea) was above 95% (Cohen's kappa coefficient ≥ 0.95), with a sensitivity of over 95%. The NPS/OPS specimen pooling experiment was conducted to verify the usability of AccuPower® kits on pooled samples and the results showed greater than 90% agreement with individual NPS/OPS samples. The clinical performance of AccuPower® kits with saliva samples was also compared with NPS/OPS samples and demonstrated over 95% agreement (Cohen's kappa coefficient > 0.95). This study shows the BIONEER NCVM and SCVM assays are comparable with the current standard confirmation assay and are suitable for effective clinical management and control of SARS-CoV-2.

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Project description:Mucormycosis is a severe and emerging invasive fungal infection associated with high mortality rates. Early diagnosis is crucial for initiating specific antifungal treatment, with molecular tools currently representing the most efficient diagnostic approach. Presently, a standardized in-house real-time PCR method is widely employed for diagnosing mucormycosis. Our study aimed to evaluate the agreement for the Mucorales DNA detection between two commercial real-time PCR assays-the Fungiplex Mucorales Real-Time PCR Kit and the MycoGENIE Aspergillus-Mucorales spp. Real-Time PCR Kit-in comparison with the in-house PCR. We retrospectively analyzed 58 samples previously identified as positive for Mucorales using the in-house PCR. These samples, obtained from 22 patients with proven or probable mucormycosis, were tested with both commercial kits. Additionally, samples from 40 patients without mucormycosis served as negative controls. Our findings revealed that the MycoGENIE Kit demonstrated superior performance in detecting Mucorales DNA in samples identified as positive by the in-house PCR. Notably, we observed minimal variability in cycle threshold (CT) values when comparing the results of the MycoGENIE Kit with those of the in-house PCR, with an average difference of 1.8 cycles. In contrast, the Fungiplex Kit exhibited a larger discrepancy in CT values compared to the in-house PCR, with an average difference of 4.1 cycles. The MycoGENIE Kit exhibited very good agreement (kappa of 0.82) with the in-house PCR for detecting Mucorales DNA across various sample types. These findings are important for the choice of kits that could be used to diagnose mucormycosis in clinical microbiology laboratories.ImportanceEarly diagnosis of mucormycosis is crucial for initiating effective treatment. The detection of Mucorales DNA by PCR in serum has revolutionized the diagnosis of this infection. However, the use of in-house methods can be time consuming. The availability of a commercial kit eliminates the need for in-house assay development, reducing laboratory workload and ensuring consistent performance across different healthcare settings. Currently, there are several commercial assays available, but many have limited evaluation. In this study, we compared two commercial kits and found that the MycoGENIE Kit offers a promising alternative to the in-house method.

Dataset Information

Detection of feline norovirus using commercial real-time RT-PCR kit for the diagnosis of human norovirus infection.

Publications

Detection of feline norovirus using commercial real-time RT-PCR kit for the diagnosis of human norovirus infection.

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