Single step multiplex real-time RT-PCR for H5N1 influenza A virus detection

Payungporn, Sunchai; Chutinimitkul, Salin; Chaisingh, Arunee; Damrongwantanapokin, Sudarat; Buranathai, Chantanee; Amonsin, Alongkorn; Theamboonlers, Apiradee; Poovorawan, Yong

doi:10.1016/j.jviromet.2005.08.004

Cited by 165 publications

(106 citation statements)

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“…According to the type of detection target, these methods could be categorized into, for example, virus isolation and identification, nucleic acid-based detection, antigen detection, and antibody detection [3][4][5][6][7][8]. Most of these methods need demanding conditions and professional operations, and some of the detection processes is time-consuming.…”

Section: Open Access Research Articlementioning

confidence: 99%

Ultra Sensitive Detection of Influenza A Virus Based on Cdse/Zns Quantum Dots Immunoassay

Wu¹,

Mao²,

Liu³

et al. 2016

SOJB

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Open Access Research articlesevere outbreaks disease, and they can cause serious infections among people.To date, numerous analytical methods have being used to detect influenza A virus. According to the type of detection target, these methods could be categorized into, for example, virus isolation and identification, nucleic acid-based detection, antigen detection, and antibody detection [3][4][5][6][7][8]. Most of these methods need demanding conditions and professional operations, and some of the detection processes is time-consuming. For these reasons, some new biosensors based on nano materials such as gold nanoparticles, magnetic nano beads and quantum dots (QDs) have been coming to the forefront [9][10][11][12].Developing a sensitive, specific and fast biosensor for diagnosing the influenza virus at the early stages of infection is a challenge. Lateral flow immune assay method has been widely used in detection of infectious diseases with the advantages of rapid, easy to operate, and low cost [13]. However, traditional method such as colloidal gold lateral flow tests has limitations when high sensitivity is needed [14].QDs have been developed to replace colloidal gold owing to their excellent optical properties such as high fluorescence efficiency, wide range of excitation wavelength, narrow and symmetric emission spectra, and QDs based lateral flow tests have been proved higher sensitivity and reproducibility for detection of pathogen, characteristic protein and even nucleic acid [15][16][17][18].In this paper, we present an ultrasensitive lateral flow immunoassay (LFIA) for detecting influenza A virus. This immune sensor is based on site-specific covalent binding with the Fc end of influenza A virus antibodies to CdSe/ ZnS QDs. The carboxylfunctionalized CdSe/ ZnS QDs conjugated with antibodies via an amide bond often result in random links of the antibody structure in the QD conjugates and block certain antigenbinding sites. In order to obtain site-specific linking, we modified carboxyl-functionalized QDs with adipicdihydrazide (ADH) and oxidized the carbohydrate groups on the antibody's Fc region AbstractAn ultrasensitive lateral flow immunoassay system (LFIAS) was established for the detection of influenza A virus. In this LFIAS, hydrophilic dihydrazide-modified CdSe/ZnSquantom dots (QDs) were conjugated with specific antibodies and used as fluorescent labels, and a pair of matched anti-nucleoprotein of influenza A virus antibodies were used to form a sandwich immunoassay. The QDs were in conjugation with the fragment crystallizable region (Fc region) of specific influenza A virus antibodies through aldehydehydrazide covalent chemistry, conferring high sensitivity. The antibodies used for detection are specific for the most conserved and popular nucleoprotein of influenza A virus and ensure the accuracy and specificity. The QDs-LFIAS can analyze the nasal-pharyngeal swab samples through simple steps and get results within 15 min. Detection of nasal-pharyngeal swab samples makes it more rapid and conve...

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Section: Open Access Research Articlementioning

confidence: 99%

Ultra Sensitive Detection of Influenza A Virus Based on Cdse/Zns Quantum Dots Immunoassay

Wu¹,

Mao²,

Liu³

et al. 2016

SOJB

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“…N1 RRT-PCR used the primers (Sigma Genosys) and minor groove binding probe originally designed by Payungporn et al (2006), but the NED fluorophore was replaced by VIC (Applied Biosystems). Addition of RNA, the thermocycling conditions and the core chemistry plus supplements were as described above for H5 CS RRT-PCR with the ROX reference dye included.…”

Section: Virus Isolationmentioning

confidence: 99%

“…M-gene, H5 and N1 RRT-PCRs are validated methods (Spackman et al, 2002;Payungporn et al, 2006;Slomka et al, 2007a) that have been recommended in the European Union (EU) Diagnostic Manual for AI (EU, 2006). Suspect AI investigations in the EU normally utilize the M-gene RRT-PCR as a generic first-line screening test, and positive AI results are promptly followed by H5 and H7-specific RRT-PCRs to identify NAI (EU, 2006).…”

Section: Introductionmentioning

confidence: 99%

Challenges for accurate and prompt molecular diagnosis of clades of highly pathogenic avian influenza H5N1 viruses emerging in Vietnam

Slomka

Tong

et al. 2012

Avian Pathology

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Forty-six chickens and 48 ducks were sampled from four Vietnamese poultry premises in 2009 infected with H5N1 highly pathogenic avian influenza (HPAI) clade 2.3.2 and 2.3.4 viruses, which also differed by cleavage site (CS) sequences in their haemagglutinin (HA) genes. All clinical specimens (n0282), namely tracheal and cloacal swabs plus feathers, were tested by five Eurasian reverse-transcriptase AI RealTime polymerase chain reaction (RRT-PCR) methods. Bayesian modelling showed similar high sensitivity for the validated H5 HA2 RRT-PCR and a new modified M-gene RRT-PCR that utilizes lyophilized reagents. Both were more sensitive than the validated ''wet'' M-gene RRT-PCR. Another RRT-PCR, which targeted the H5-gene CS region, was effective for clade 2.3.4 detection, but severely compromised for clade 2.3.2 viruses. Reduced sensitivity of the H5 CS and ''wet'' M-gene RRT-PCRs correlated with mismatches between the target and the primer and/or probe sequences. However, the H5 HA2 RRT-PCR sensitively detected both clade 2.3.2 and 2.3.4 viruses, and agreed with N1 RRT-PCR results. Feather testing from diseased chicken and duck flocks by AI RRT-PCRs resulted in the most sensitive identification of H5N1 HPAI-infected birds. Evolution of new H5N1 HPAI clades remains a concern for currently affected Asian countries, but also for more distant regions where it is important to be prepared for new incursions of H5N1 HPAI viruses. Genetic evidence for adamantane resistance and sensitivity was also observed in isolates from both clades.

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Section: Introductionmentioning

confidence: 99%

“…So it is very important to develop a detecting method for SRBSDV with simple operation, strong specificity, high sensitivity, good reproducibility and quick speed. The real-time RT-PCR is suitable and gradually become an important technique for virus detecting [10][11][12].This study aimed to design specific primers and develop an absolute quantification real-time RT-PCR method for detecting SRBSDV, and this method was also applied to detect SRBSDV infection in field samples of rice. For demonstrating the excellent performance of developed real-time RT-PCR, the detection efficiency was also compared with conventional RT-PCR.…”

mentioning

confidence: 99%

Development of a Real-time RT-PCR Method for Rapid Detection and Quantification of Southern Rice Black-streaked Dwarf Virus in Rice

Zhang¹

2013

J Plant Pathol Microbiol

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IntroductionSouthern rice black-streaked dwarf virus (SRBSDV), a latest member of the genus Fijivirus within the family Reoviridae, was simultaneously isolated and identified from the rice in southern of China in 2008 [1,2]. In the following three years (2009)(2010), SRBSDV caused the most important rice disease in South of China, there were over ten millions hectare of rice infected, and resulted in massive devastation of rice yield [3].SRBSDV was most closely related to rice black-streaked dwarf virus (RBSDV); symptoms of infected rice by SRBSDV were similar to RBSDV, such as stunting, dark leaf and small enations on stem and leaf back. Its genome consisted of 10 linear segments of double-stranded RNAs (dsRNA) called S1-S10 in an increasing order of electrophoretic mobility in polyacrylamide gels [1]. The homology of nucleotide sequence of genome segments of SRBSDV was 70.6%-80.0% similar to the genome segments of RBSDV. Nevertheless, the SRBSDV was transmitted to rice by white-backed planthoppers, but not by small brown planthopper (RBSDV transmitted vector), and whole genome comparisons and phylogenetic analyses also supported suggestions that SRBSDV represented a new species within the genus Fijivirus, family Reoviridae [4]. So it is exceeding to develop an accurate detecting method for identifying SRBSDV from RBSDV.Currently, the identification of SRBSDV and its infection is mainly by typical symptoms and RT-PCR [5]. For typical symptoms, it is time consuming, false positive in early diagnosis and severe yield loss by the time it is identified in the infected rice [6]. For RT-PCR, thought it is specific to SRBSDV, false positive still constantly occur, especially in early infected stage in rice because of low quantity of virion in the host [7]. The serological methods are more fast and economical than other methods [8] including in rice and transmitted vector, nevertheless, they are under development. These methods are difficult to precisely quantify SRBSDV, and difficult to rapidly detect the disease for early warning [9]. So it is very important to develop a detecting method for SRBSDV with simple operation, strong specificity, high sensitivity, good reproducibility and quick speed. The real-time RT-PCR is suitable and gradually become an important technique for virus detecting [10][11][12].This study aimed to design specific primers and develop an absolute quantification real-time RT-PCR method for detecting SRBSDV, and this method was also applied to detect SRBSDV infection in field samples of rice. For demonstrating the excellent performance of developed real-time RT-PCR, the detection efficiency was also compared with conventional RT-PCR. Materials and Methods Leaf samples of riceLeaf samples of rice were collected from heading stage of rice in different districts of Hunan province, China. The leaf samples were gathered in period from 2009 to 2011, there into, some of samples had typical or suspected symptoms of SRBSDV infected, such as stunting, dark leaf and/or small enations on stem and leaf back....

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Single step multiplex real-time RT-PCR for H5N1 influenza A virus detection

Cited by 165 publications

References 9 publications

Ultra Sensitive Detection of Influenza A Virus Based on Cdse/Zns Quantum Dots Immunoassay

Ultra Sensitive Detection of Influenza A Virus Based on Cdse/Zns Quantum Dots Immunoassay

Challenges for accurate and prompt molecular diagnosis of clades of highly pathogenic avian influenza H5N1 viruses emerging in Vietnam

Development of a Real-time RT-PCR Method for Rapid Detection and Quantification of Southern Rice Black-streaked Dwarf Virus in Rice

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