Cloning and expression of Rift Valley fever virus nucleocapsid (N) protein and evaluation of a N-protein based indirect ELISA for the detection of specific IgG and IgM antibodies in domestic ruminants

Fafetine, José; Tijhaar, Edwin; Pawęska, Janusz T.; Neves, Luís; Hendriks, Judith; Swanepoel, Robert; Coetzer, Jacobus A.W.; Egberink, Herman; Rutten, Victor P.M.G.

doi:10.1016/j.vetmic.2006.11.008

Cited by 72 publications

(60 citation statements)

References 31 publications

(29 reference statements)

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“…In addition, it was also reported that N protein is the main immunodominant viral protein in other members of the Bunyaviridae family [22,25]. Since rNp lacks infectivity with considerable stability, it is one of the suitable candidates for use as a diagnostic antigen in an ELISA [7,8,15,18].…”

Section: Discussionmentioning

confidence: 99%

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Competitive ELISA for the Detection of Antibodies to Rift Valley Fever Virus in Goats and Cattle

Kim

Nah

Moon

et al. 2012

The Journal of Veterinary Medical Science

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ABSTRACT. Rift Valley fever virus (RVFV) is one of the important emerging viral diseases of serious impact in public health and animal hygiene both in human and animal industries. In this study, we developed a monoclonal antibody-based competitive ELISA for the detection of antibodies to RVFV in goats and cattle. The recombinant N protein of RVFV was expressed in E. coli with a six-histidine tag, and the purified N protein was used for detecting antigen with a competitive monoclonal antibody against RVFV antibodies. The competitive ELISA (C-ELISA) could detect antibodies at 9-11 days after inoculation in goats and cattle with a sensitivity of 94.7% (virus neutralization titer >32) and specificity of 99.7%, respectively. In addition, the C-ELISA did not show any cross-reactivity with positive sera against arboviruses such as Akabane, Aino, Chuzan, Ibaraki and bovine ephemeral fever virus, which are prevalent viral agents in ruminant animals throughout Southeast Asia. The results of the present study indicate that the C-ELISA is a simple, rapid and convenient serodiagnostic method for RVFV in goats and cattle.

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Section: Discussionmentioning

confidence: 99%

“…Accurate diagnosis of RVF can be achieved by serological tests in combination with clinical observation and epidemiological history [7]. Classical methods for the detection of antibodies to RVFV are hemagglutination inhibition, complement fixation, virus neutralization (VN) test, and immunofluorescence assay (IFA) [24].…”

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confidence: 99%

Competitive ELISA for the Detection of Antibodies to Rift Valley Fever Virus in Goats and Cattle

Kim

Nah

Moon

et al. 2012

The Journal of Veterinary Medical Science

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“…In recent years, various ELISA formats with high diagnostic accuracy and specificity have been developed for the specific detection of IgG, IgM, and total antibodies; in particular, for example, recombinant antigens have been used for accurate, specific detection of antibodies to a number of viruses in the family Bunyaviridae (3,12,18). In this study, we took another phlebovirus, specifically, Rift Valley fever virus, as a reference, and we developed a unique double-antigen sandwich ELISA for detection of SFTSV-specific total antibodies in sera from humans and a variety of animals using SFTSV recombinant N (rN) protein.…”

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confidence: 99%

Preparation and Evaluation of Recombinant Severe Fever with Thrombocytopenia Syndrome Virus Nucleocapsid Protein for Detection of Total Antibodies in Human and Animal Sera by Double-Antigen Sandwich Enzyme-Linked Immunosorbent Assay

et al. 2012

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The recent emergence of the human infection confirmed to be caused by severe fever with thrombocytopenia syndrome virus (SFTSV) in China is of global concern. Safe diagnostic immunoreagents for determination of human and animal seroprevalence in epidemiological investigations are urgently needed. This paper describes the cloning and expression of the nucleocapsid (N) protein of SFTSV. An N-protein-based double-antigen sandwich enzyme-linked immunosorbent assay (ELISA) system was set up to detect the total antibodies in human and animal sera. We reasoned that as the double-antigen sandwich ELISA detected total antibodies with a higher sensitivity than traditional indirect ELISA, it could be used to detect SFTSV-specific antibodies from different animal species. The serum neutralization test was used to validate the performance of this ELISA system. All human and animal sera that tested positive in the neutralization test were also positive in the sandwich ELISA, and there was a high correlation between serum neutralizing titers and ELISA readings. Cross-reactivity was evaluated, and the system was found to be highly specific to SFTSV; all hantavirus-and dengue virus-confirmed patient samples were negative. SFTSV-confirmed human and animal sera from both Anhui and Hubei Provinces in China reacted with N protein in this ELISA, suggesting no major antigenic variation between geographically disparate virus isolates and the suitability of this assay in nationwide application. ELISA results showed that 3.6% of the human serum samples and 47.7% of the animal field serum samples were positive for SFTSV antibodies, indicating that SFTSV has circulated widely in China. This assay, which is simple to operate, poses no biohazard risk, does not require sophisticated equipment, and can be used in disease surveillance programs, particularly in the screening of large numbers of samples from various animal species.

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“…The nucleocapsid (N) protein is the most abundant and highly immunogenic component of the RVF virion and has been used for development of diagnostic assays for detection of RVFV-specific antibodies in human and animal sera (Fafetine et al 2007, Paweska et al 2008. Although the N protein is shown to be highly conserved among members of the Bunyaviridae family (Schwarz, et al 1996, Magurano and Nicoletti 1999), a previous indirect ELISA based on the recombinant protein did not show cross-reactivity with other African phleboviruses that could hamper the reliability of using this protein in assays for serodiagnosis of RVFV infection ).…”

Section: Figmentioning

confidence: 99%

Rift Valley Fever Virus Structural and Nonstructural Proteins: Recombinant Protein Expression and Immunoreactivity Against Antisera from Sheep

Faburay

Wilson²,

McVey³

et al. 2013

Vector-Borne and Zoonotic Diseases

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The Rift Valley fever virus (RVFV) encodes the structural proteins nucleoprotein (N), aminoterminal glycoprotein (Gn), carboxyterminal glycoprotein (Gc), and L protein, 78-kD, and the nonstructural proteins NSm and NSs. Using the baculovirus system, we expressed the full-length coding sequence of N, NSs, NSm, Gc, and the ectodomain of the coding sequence of the Gn glycoprotein derived from the virulent strain of RVFV ZH548. Western blot analysis using anti-His antibodies and monoclonal antibodies against Gn and N confirmed expression of the recombinant proteins, and in vitro biochemical analysis showed that the two glycoproteins, Gn and Gc, were expressed in glycosylated form. Immunoreactivity profiles of the recombinant proteins in western blot and in indirect enzyme-linked immunosorbent assay against a panel of antisera obtained from vaccinated or wild type (RVFV)-challenged sheep confirmed the results obtained with anti-His antibodies and demonstrated the suitability of the baculo-expressed antigens for diagnostic assays. In addition, these recombinant proteins could be valuable for the development of diagnostic methods that differentiate infected from vaccinated animals (DIVA).

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Cloning and expression of Rift Valley fever virus nucleocapsid (N) protein and evaluation of a N-protein based indirect ELISA for the detection of specific IgG and IgM antibodies in domestic ruminants

Cited by 72 publications

References 31 publications

Competitive ELISA for the Detection of Antibodies to Rift Valley Fever Virus in Goats and Cattle

Competitive ELISA for the Detection of Antibodies to Rift Valley Fever Virus in Goats and Cattle

Preparation and Evaluation of Recombinant Severe Fever with Thrombocytopenia Syndrome Virus Nucleocapsid Protein for Detection of Total Antibodies in Human and Animal Sera by Double-Antigen Sandwich Enzyme-Linked Immunosorbent Assay

Rift Valley Fever Virus Structural and Nonstructural Proteins: Recombinant Protein Expression and Immunoreactivity Against Antisera from Sheep

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