Comparison of a Multiplex Real-Time PCR Assay with a Multiplex Luminex Assay for Influenza Virus Detection

Munro, Sandra B.; Kuypers, Jane; Jerome, Keith R.

doi:10.1128/jcm.03113-12

Cited by 43 publications

(36 citation statements)

References 28 publications

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“…Air was sampled weekly from a university campus in Hong Kong from October 2016 to June 2018 except during the summer holidays in 2017 (weeks [19][20][21][22][23][24][25][26][27][28][29][30][31][32][33][34][35] and on public holidays. Samples were collected from canteens, lecture halls, shuttle buses, and the University Health Service (UHS) during peak hours of human flow, using 6-13 air samplers per week.…”

Section: Air Sample Collectionmentioning

confidence: 99%

“…Viral RNA (vRNA) was extracted from 140 μL of resuspended media using the QIAamp Viral RNA Mini Kit (Qiagen) and eluted into 60 μL water. Respiratory viruses were detected using quantitative real-time reverse-transcription polymerase chain reaction (rRT-qPCR) by the ViiA 7 Real-time PCR System (ThermoFisher) using 5 μL eluent, AgPath-IDTM One-step RT-PCR Reagents (Life Technology), and specific primers and probes (Supplementary Table 1) [24][25][26][27][28]. IAV M gene-positive samples were further subtyped (H1 and H3) by rRT-qPCR [25].…”

Section: Detection and Quantification Of Viral Rna Genomes From Airbomentioning

confidence: 99%

See 1 more Smart Citation

Detection of Influenza and Other Respiratory Viruses in Air Sampled From a University Campus: A Longitudinal Study

Xie

Lau

Yoshida

et al. 2019

Clinical Infectious Diseases

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Background. Respiratory virus-laden particles are commonly detected in the exhaled breath of symptomatic patients or in air sampled from healthcare settings. However, the temporal relationship of detecting virus-laden particles at nonhealthcare locations vs surveillance data obtained by conventional means has not been fully assessed.Methods. From October 2016 to June 2018, air was sampled weekly from a university campus in Hong Kong. Viral genomes were detected and quantified by real-time reverse-transcription polymerase chain reaction. Logistic regression models were fitted to examine the adjusted odds ratios (aORs) of ecological and environmental factors associated with the detection of virus-laden airborne particles.Results. Influenza A (16.9% [117/694]) and influenza B (4.5% [31/694]) viruses were detected at higher frequencies in air than rhinovirus (2.2% [6/270]), respiratory syncytial virus (0.4% [1/270]), or human coronaviruses (0% [0/270]). Multivariate analyses showed that increased crowdedness (aOR, 2.3 [95% confidence interval {CI}, 1.5-3.8]; P < .001) and higher indoor temperature (aOR, 1.2 [95% CI, 1.1-1.3]; P < .001) were associated with detection of influenza airborne particles, but absolute humidity was not (aOR, 0.9 [95% CI, .7-1.1]; P = .213). Higher copies of influenza viral genome were detected from airborne particles >4 μm in spring and <1 μm in autumn. Influenza A(H3N2) and influenza B viruses that caused epidemics during the study period were detected in air prior to observing increased influenza activities in the community.Conclusions. Air sampling as a surveillance tool for monitoring influenza activity at public locations may provide early detection signals on influenza viruses that circulate in the community.

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Section: Air Sample Collectionmentioning

confidence: 99%

Section: Detection and Quantification Of Viral Rna Genomes From Airbomentioning

confidence: 99%

Detection of Influenza and Other Respiratory Viruses in Air Sampled From a University Campus: A Longitudinal Study

Xie

Lau

Yoshida

et al. 2019

Clinical Infectious Diseases

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“…The use of PCR has grown increasingly in different applications ranging from microorganisms detection to diagnosis of complex genetic diseases [1–3]. The simple implementation and the possibility of post-PCR analysis automation make PCR a great tool for high throughput analysis [3].…”

Section: Introductionmentioning

confidence: 99%

In silicosingle strand melting curve: a new approach to identify nucleic acid polymorphisms in Totiviridae

et al. 2014

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BackgroundThe PCR technique and its variations have been increasingly used in the clinical laboratory and recent advances in this field generated new higher resolution techniques based on nucleic acid denaturation dynamics. The principle of these new molecular tools is based on the comparison of melting profiles, after denaturation of a DNA double strand. Until now, the secondary structure of single-stranded nucleic acids has not been exploited to develop identification systems based on PCR. To test the potential of single-strand RNA denaturation as a new alternative to detect specific nucleic acid variations, sequences from viruses of the Totiviridae family were compared using a new in silico melting curve approach. This family comprises double-stranded RNA virus, with a genome constituted by two ORFs, ORF1 and ORF2, which encodes the capsid/RNA binding proteins and an RNA-dependent RNA polymerase (RdRp), respectively.ResultsA phylogenetic tree based on RdRp amino acid sequences was constructed, and eight monophyletic groups were defined. Alignments of RdRp RNA sequences from each group were screened to identify RNA regions with conserved secondary structure. One region in the second half of ORF2 was identified and individually modeled using the RNAfold tool. Afterwards, each DNA or RNA sequence was denatured in silico using the softwares MELTSIM and RNAheat that generate melting curves considering the denaturation of a double stranded DNA and single stranded RNA, respectively. The same groups identified in the RdRp phylogenetic tree were retrieved by a clustering analysis of the melting curves data obtained from RNAheat. Moreover, the same approach was used to successfully discriminate different variants of Trichomonas vaginalis virus, which was not possible by the visual comparison of the double stranded melting curves generated by MELTSIM.ConclusionIn silico analysis indicate that ssRNA melting curves are more informative than dsDNA melting curves. Furthermore, conserved RNA structures may be determined from analysis of individuals that are phylogenetically related, and these regions may be used to support the reconstitution of their phylogenetic groups. These findings are a robust basis for the development of in vitro systems to ssRNA melting curves detection.Electronic supplementary materialThe online version of this article (doi:10.1186/1471-2105-15-243) contains supplementary material, which is available to authorized users.

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“…At present, real-time reverse transcription polymerase chain reaction (RT-PCR) is a powerful molecular diagnostic method for influenza Dovepress Dovepress 2646 ge et al virus infections. [6][7][8][9] In contrast to PCR-based target gene detection, which requires bulky and expensive equipments and highly skilled technicians, recent research has focused on the development of isothermal amplification methods for simple and user-friendly pathogen detection. As the reaction is conducted under isothermal conditions, the thermal cycler is not required, which makes these methods more suitable for use in resource-limited regions or for field use.…”

Section: Introductionmentioning

confidence: 99%

Detection of influenza viruses by coupling multiplex reverse-transcription loop-mediated isothermal amplification with cascade invasive reaction using nanoparticles as a sensor

Ge¹,

Zhou²,

Zhao³

et al. 2017

IJN

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Influenza virus infections represent a worldwide public health and economic problem due to the significant morbidity and mortality caused by seasonal epidemics and pandemics. Sensitive and convenient methodologies for detection of influenza viruses are essential for further disease control. Loop-mediated isothermal amplification (LAMP) is the most commonly used method of nucleic acid isothermal amplification. However, with regard to multiplex LAMP, differentiating the ladder-like LAMP products derived from multiple targets is still challenging today. The requirement of specialized instruments has further hindered the on-site application of multiplex LAMP. We have developed an integrated assay coupling multiplex reverse transcription LAMP with cascade invasive reaction using nanoparticles (mRT-LAMP-CIRN) as a sensor for the detection of three subtypes of influenza viruses: A/H1N1pdm09, A/H3 and influenza B. The analytic sensitivities of the mRT-LAMP-CIRN assay were 10 1 copies of RNA for both A/H1N1pdm09 and A/H3, and 10 2 copies of RNA for influenza B. This assay demonstrated highly specific detection of target viruses and could differentiate them from other genetically or clinically related viruses. Clinical specimen analysis showed the mRT-LAMP-CIRN assay had an overall sensitivity and specificity of 98.3% and 100%, respectively. In summary, the mRT-LAMP-CIRN assay is highly sensitive and specific, and can be used as a cost-saving and instrument-free method for the detection of influenza viruses, especially for on-site use.

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Comparison of a Multiplex Real-Time PCR Assay with a Multiplex Luminex Assay for Influenza Virus Detection

Cited by 43 publications

References 28 publications

Detection of Influenza and Other Respiratory Viruses in Air Sampled From a University Campus: A Longitudinal Study

Detection of Influenza and Other Respiratory Viruses in Air Sampled From a University Campus: A Longitudinal Study

In silicosingle strand melting curve: a new approach to identify nucleic acid polymorphisms in Totiviridae

Detection of influenza viruses by coupling multiplex reverse-transcription loop-mediated isothermal amplification with cascade invasive reaction using nanoparticles as a sensor

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