Establishment and validation of two duplex one-step real-time RT-PCR assays for diagnosis of foot-and-mouth disease

Górna, Kamila; Relmy, Anthony; Romey, Aurore; Zientara, Stéphan; Blaise‐Boisseau, Sandra; Bakkali‐Kassimi, Labib

doi:10.1016/j.jviromet.2016.03.020

Cited by 11 publications

(8 citation statements)

References 26 publications

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“…A one‐step duplex pan‐FMDV real‐time RT‐PCR (rtRT‐PCR) assay was used as previously described (Gorna et al, ). The assay targeted the FMDV 3D coding region and the cellular β‐actin gene as an internal control.…”

Section: Methodsmentioning

confidence: 99%

Model of persistent foot‐and‐mouth disease virus infection in multilayered cells derived from bovine dorsal soft palate

Hägglund

Laloy

Näslund

et al. 2019

Transbound Emerg Dis

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Foot‐and‐mouth disease virus (FMDV) causes a highly contagious vesicular disease in livestock, with serious consequences for international trade. The virus persists in the nasopharynx of cattle and this slows down the process to obtain an FMDV‐free status after an outbreak. To study biological mechanisms, or to identify molecules that can be targeted to diagnose or interfere with persistence, we developed a model of persistent FMDV infection in bovine dorsal soft palate (DSP). Primary DSP cells were isolated after commercial slaughter and were cultured in multilayers at the air‐liquid interface. After 5 weeks of culture without further passage, the cells were infected with FMDV strain O/FRA/1/2001. Approximately, 20% of cells still had a polygonal morphology and displayed tight junctions as in stratified squamous epithelia. Subsets of cells expressed cytokeratin and most or all cells expressed vimentin. In contrast to monolayers in medium, multilayers in air demonstrated only a limited cytopathic effect. Integrin αVβ6 expression was observed in mono‐ but not in multilayers. FMDV antigen, FMDV RNA and live virus were detected from day 1 to 28, with peaks at day 1 and 2. The proportion of infected cells was highest at 24 hr (3% and 36% of cells at an MOI of 0.01 and 1, respectively). At day 28 after infection, at a time when animals that still harbour FMDV are considered carriers, FMDV antigen was detected in 0.2%–2.1% of cells, in all layers, and live virus was isolated from supernatants of 6/8 cultures. On the consensus level, the viral genome did not change within the first 24 hr after infection. Only a few minor single nucleotide variants were detected, giving no indication of the presence of a viral quasispecies. The air‐liquid interface model of DSP brings new possibilities to investigate FMDV persistence in a controlled manner.

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Section: Methodsmentioning

confidence: 99%

Model of persistent foot‐and‐mouth disease virus infection in multilayered cells derived from bovine dorsal soft palate

Hägglund

Laloy

Näslund

et al. 2019

Transbound Emerg Dis

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“…Classical methods for the food-and-mouth diagnostics involve virus isolation, ELISA and RT-PCR tests [ 109 ]. Recently an improved duplex one-step RT-PCR assays was validated [ 110 ]. The main advantage of this test is in the co-amplification of foot-and-mouth virus RNA and host β-actin mRNA.…”

Section: Detection Of Foot-and-mouth Disease Virusesmentioning

confidence: 99%

Advanced biosensors for detection of pathogens related to livestock and poultry

Vidić

Manzano

Chang

et al. 2017

Vet Res

208

124

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Infectious animal diseases caused by pathogenic microorganisms such as bacteria and viruses threaten the health and well-being of wildlife, livestock, and human populations, limit productivity and increase significantly economic losses to each sector. The pathogen detection is an important step for the diagnostics, successful treatment of animal infection diseases and control management in farms and field conditions. Current techniques employed to diagnose pathogens in livestock and poultry include classical plate-based methods and conventional biochemical methods as enzyme-linked immunosorbent assays (ELISA). These methods are time-consuming and frequently incapable to distinguish between low and highly pathogenic strains. Molecular techniques such as polymerase chain reaction (PCR) and real time PCR (RT-PCR) have also been proposed to be used to diagnose and identify relevant infectious disease in animals. However these DNA-based methodologies need isolated genetic materials and sophisticated instruments, being not suitable for in field analysis. Consequently, there is strong interest for developing new swift point-of-care biosensing systems for early detection of animal diseases with high sensitivity and specificity. In this review, we provide an overview of the innovative biosensing systems that can be applied for livestock pathogen detection. Different sensing strategies based on DNA receptors, glycan, aptamers and antibodies are presented. Besides devices still at development level some are validated according to standards of the World Organization for Animal Health and are commercially available. Especially, paper-based platforms proposed as an affordable, rapid and easy to perform sensing systems for implementation in field condition are included in this review.

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“…Indeed, real-time RT-PCR has been used for the detection of footand-mouth disease targeting the 3D and IRES regions of the viral genome (Laor, Torgersen, Yadin, & Becker, 1992;Reid, Ferris, Hutchings, Samuel, & Knowles, 2000). Multiplex RT-PCR has been used for typing FMD virus (Callens & De Clercq, 1997;Giridharan, Hemadri, Tosh, Sanyal, & Bandyopadhyay, 2005;Gorna et al, 2016). However, this test does not allow the sequencing because the amplified part is very short (250 nt) which is insufficient for complete sequencing of the VP1 gene (Gorna et al, 2016).…”

Section: Laboratory Analysesmentioning

confidence: 99%

“…Multiplex RT-PCR has been used for typing FMD virus (Callens & De Clercq, 1997;Giridharan, Hemadri, Tosh, Sanyal, & Bandyopadhyay, 2005;Gorna et al, 2016). However, this test does not allow the sequencing because the amplified part is very short (250 nt) which is insufficient for complete sequencing of the VP1 gene (Gorna et al, 2016). Finally, conventional RT-PCR was used to amplify the complete gene encoding the VP1 protein (Relmy et al, 2017).…”

Section: Laboratory Analysesmentioning

confidence: 99%

Seroprevalence and molecular characterization of foot‐and‐mouth disease virus in Chad

Abdel-Aziz

Romey

Relmy

et al. 2019

Veterinary Medicine & Sci

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This study aimed at determining the seroprevalence of foot‐and‐mouth disease (FMD) in domestic ruminants and at characterizing the virus strains circulating in four areas of Chad (East Batha, West Batha, Wadi Fira and West Ennedi). The study was carried out between October and November 2016. A total of 1,520 sera samples (928 cattle, 216 goats, 254 sheep and 122 dromedaries) were collected randomly for FMD serological analyses. Nine epithelial tissue samples were also collected from cattle showing clinical signs, for FMDV isolation and characterization. Serological results showed an overall NSP seroprevalence of 40% (375/928) in cattle in our sample (95% CrI [19–63]). However, seroprevalences of 84% (27/32), 78% (35/45) and 84% (21/25) were estimated in cattle over 5 years of age in East Batha, West Batha and Wadi Fira, respectively. In cattle under 1 year of age, 67% (18/27) seroprevalence was estimated in Wadi Fira, 64% (14/22) in East Batha and 59% (13/22) in West Batha. It was found that the high seroprevalences have been obtained in areas where pastures are shared by several different herds but also in farms where two to three species (bovine, caprine and ovine) are raised together. ELISA PrioCHECK®FMDV types O and A and in‐house solid phase competition ELISA serotyping results showed that the four O, A, SAT1 and SAT2 serotypes have circulated in Chad in 2016. However, the type SAT2 dominated with an overall seroprevalence of 43% (29/67) and was present in the four areas investigated. The phylogenetic analyses of the VP1 coding sequence allowed determining the serotype SAT2 topotype VII, close to viral strains found in Cameroon in 2015 with a similarity of 98.60%.

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Establishment and validation of two duplex one-step real-time RT-PCR assays for diagnosis of foot-and-mouth disease

Cited by 11 publications

References 26 publications

Model of persistent foot‐and‐mouth disease virus infection in multilayered cells derived from bovine dorsal soft palate

Model of persistent foot‐and‐mouth disease virus infection in multilayered cells derived from bovine dorsal soft palate

Advanced biosensors for detection of pathogens related to livestock and poultry

Seroprevalence and molecular characterization of foot‐and‐mouth disease virus in Chad

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