Development of a Blocking ELISA for Screening Antibodies to Porcine Rubulavirus, La Piedad Michoacan Virus

Nordengrahn, Ann; Svenda, M.; Moreno‐López, J.; Bergvall, Ann-Christin; Hernández, Pablo E.; McNeilly, F.; Allan, Gordon; Mérza, M.

doi:10.1177/104063879901100404

Cited by 18 publications

(7 citation statements)

References 15 publications

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“…Blocking ELISA, a specific method for antibody detection, has been used widely to diagnose human diseases [18], monitor animal infectious diseases [19], [20], [21], [22], [23], and detect viral antibodies in clinics or laboratories [24], [25], [26], [27], [28], [29]. The distinct advantages of blocking ELISA include high-volume sample testing, applicability across multiple species, and greater objectivity than some traditional techniques.…”

Section: Discussionmentioning

confidence: 99%

Development of a Blocking ELISA for Detection of Serum Neutralizing Antibodies against Newly Emerged Duck Tembusu Virus

Teng

et al. 2012

PLoS ONE

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BackgroundSince April 2010, domesticated ducks in China have been suffering from an emerging infectious disease characterized by retarded growth, high fever, loss of appetite, decline in egg production, and death. The causative agent was identified as a duck Tembusu virus (DTMUV), a member of the Ntaya virus (NTAV) group within the genus Flavivirus, family Flaviviridae. DTMUV is highly contagious and spreads rapidly in many species of ducks. More than 10 million shelducks have been infected and approximately 1 million died in 2010. The disease remains a constant threat to the duck industry; however, it is not known whether DTMUV can infect humans or other mammalians, despite the fact that the virus has spread widely in southeast China, one of the most densely populated areas in the world. The lack of reliable methods to detect the serum antibodies against DTMUV has limited our ability to conduct epidemiological investigations in various natural hosts and to evaluate the efficiency of vaccines to DTMUV.Methodology/Principal FindingsA neutralizing monoclonal antibody (mAb) 1F5 binding specifically to the E protein was developed. Based on the mAb, a blocking enzyme-linked immunosorbent assay (ELISA) was developed for the detection of neutralizing antibodies against DTMUV. The average value of percent inhibition (PI) of 350 duck serum samples obtained from DTMUV-free farms was 1.0% ±5.8% (mean ± SD). The selected cut-off PI values for negative and positive sera were 12.6% (mean +2SD) and 18.4% (mean +3SD), respectively. When compared with a serum neutralizing antibody test (SNT) using chicken embryonated eggs, the rate of coincidence was 70.6% between the blocking ELISA and SNT, based on the titration of 20 duck DTMUV-positive serum samples.Conclusions/SignificanceThe blocking ELISA based on a neutralizing mAb allowed rapid, sensitive, and specific detection of neutralization-related antibodies against DTMUV.

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Section: Discussionmentioning

confidence: 99%

Development of a Blocking ELISA for Detection of Serum Neutralizing Antibodies against Newly Emerged Duck Tembusu Virus

Teng

et al. 2012

PLoS ONE

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“…Clinical symptoms, necropsy findings, and histopathological changes can often provide an insight into the aetiology of the disease. Currently different methods have been used for the diagnosis of PorPV infection, including serological tests and virus isolation ( 40 , 41 , 46 , 56 , 57 ). The virus has been shown to be able to grow in many different cells including pig kidney cells with typical syncytial formation, demonstrating fusion activity ( 1 ).…”

Section: Diagnosismentioning

confidence: 99%

“…The most common serological method for diagnosis is hemagglutination inhibition test. Other serological techniques frequently used are indirect fluorescent antibody, serum-virus neutralisation ( 40 , 41 , 46 , 57 ) and a blocking ELISA ( 46 , 54 , 57 ). Classical PCR technologies have been used for different research purposes 16 , 41 , 46 ), and a real-time PCR (qRT-PCR) method specific to PorPV has been developed for PorPV to study epidemiology aspects of the disease ( 58 , 59 ).…”

Section: Diagnosismentioning

confidence: 99%

Molecular and epidemiological studies ofPorcine rubulavirusinfection – an overview

Cuevas-Romero

Blomström

Berg

2015

Infection Ecology & Epidemiology

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Porcine rubulavirus-La Piedad-Michoacan-Mexico virus (PorPV-LPMV) was identified as the causative agent of a viral disease that emerged spontaneously in Mexican swine in the 1980s. Since the report of the initial outbreak of the disease, only one full-length genome from a strain isolated in 1984 (PorPV-LPMV/1984) has been sequenced; sequence data are scarce from other isolates. The genetic variation of this virus that has spread throughout the main endemic region of Mexico is almost a complete mystery. The development of molecular techniques for improved diagnostics and to investigate the persistence, molecular epidemiology, and the possible reservoirs of PorPV are needed. Together, this will provide greater knowledge regarding the molecular genetic changes and useful data to establish new strategies in the control of this virus in Mexico.

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“…Concerning possible diagnostic tests, at present there are only serological tests and classical virus isolation techniques available (McNeilly et al, 1997;Nordengrahn et al, 1999). In addition, classical PCR technology has been used for various research purposes Wiman et al, 1998;Cuevas et al, 2009).…”

Section: Introductionmentioning

confidence: 99%

Development of a real-time RT-PCR method for detection of porcine rubulavirus (PoRV-LPMV)

Cuevas-Romero

Blomström

Alvarado

et al. 2013

Journal of Virological Methods

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In order to provide a rapid and sensitive method for detection of the Porcine rubulavirus La Piedad-Michoacan-Mexico Virus (PoRV-LPMV), we have developed a specific real-time reverse transcriptase polymerase chain reaction assay. The detection of PoRV-LPMV, represents a diagnostic challenge due to the viral RNA being present in very small amounts in tissue samples. In this study, a TaqMan(®) real-time PCR assay was designed based on the phosphoprotein gene of PoRV-LPMV, to allow specific amplification and detection of viral RNA in clinical samples. Assay conditions for the primers and probe were optimized using infected PK15 cells and ten-fold serial dilutions of a plasmid containing the whole P-gene. The sensitivity of the developed TaqMan(®) assay was approximately 10 plasmid copies per reaction, and was shown to be 1000 fold better than a conventional nested RT-PCR. The performance of this real-time RT-PCR method enables studies of various aspects of PoRV-LPMV infection. Finally, the assay detects all current known variants of the virus.

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Development of a Blocking ELISA for Screening Antibodies to Porcine Rubulavirus, La Piedad Michoacan Virus

Cited by 18 publications

References 15 publications

Development of a Blocking ELISA for Detection of Serum Neutralizing Antibodies against Newly Emerged Duck Tembusu Virus

Development of a Blocking ELISA for Detection of Serum Neutralizing Antibodies against Newly Emerged Duck Tembusu Virus

Molecular and epidemiological studies ofPorcine rubulavirusinfection – an overview

Development of a real-time RT-PCR method for detection of porcine rubulavirus (PoRV-LPMV)

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