Highly specific confirmatory Western blot test for African swine fever virus antibody detection using the recombinant virus protein p54

Alcaraz, C.; Rodrı́guez, Fernando; Oviedo, J.M.; Eiras, A.; Diego, M. De; Alonso, Covadonga; Escribano, José M.

doi:10.1016/0166-0934(94)00150-f

Cited by 32 publications

(21 citation statements)

References 9 publications

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“…In conclusion, this work extends the recombinant antigens (2,9,19,22) with the potential for use for the diagnosis of ASFV infections, has confirmed the usefulness of p54, and has identified pB602L as a novel diagnostic tool for use in areas where ASF is endemic. Given that the variability of ASFV isolates in Africa is greater than that in Europe, further studies are needed in African countries in order to adequately validate the ELISAs.…”

Section: Discussionmentioning

confidence: 91%

“…A particular disadvantage of the conventional ELISA (which uses semipurified virions as the antigen) is the lack of sensitivity and the unacceptable level of false-negative results obtained with poorly preserved samples, hence demanding the routine confirmation of the result by immunoblotting (2,3,20). In order to evaluate the stability of the new ELISAs, 39 ASFVpositive field serum samples were incubated for 1 month at 37°C in order to simulate deterioration in the field and were then retested by the ELISA with the recombinant proteins.…”

Section: Discussionmentioning

confidence: 99%

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Recombinant Antigen Targets for Serodiagnosis of African Swine Fever

Gallardo

Reis

Kalema‐Zikusoka

et al. 2009

Clin Vaccine Immunol

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African swine fever (ASF) is an infectious and economically important disease of domestic pigs. There is no vaccine, and so reliable diagnosis is essential for control strategies. The performance of four recombinant ASF virus (ASFV) protein (pK205R, pB602L, p104R, and p54)-based enzyme-linked immunosorbent assays (ELISAs) was evaluated with European porcine field sera that had been established by Office International des Epizooties (OIE)-approved tests to be ASFV negative (n ‫؍‬ 119) and ASFV positive (n ‫؍‬ 80). The values showed that there was almost perfect agreement between the results of the "gold standard" test (immunoblotting) and the results obtained by the p54-specific ELISA ( ‫؍‬ 0.95; 95% confidence interval [CI], 0.90 to 0.99) and the pK205R-specific ELISA or the pB602L-specific ELISA ( ‫؍‬ 0.92; 95% CI, 0.86 to 0.97). For the pA104R-specific ELISA, there was substantial to almost perfect agreement ( ‫؍‬ 0.81; 95% CI, 0.72 to 0.89). Similar results were observed by the OIE-approved ELISA ( ‫؍‬ 0.89; 95% CI, 0.82 to 0.95). Importantly, antibodies against these proteins were detectable early after infection of domestic pigs. Preliminary testing of 9 positive and 17 negative serum samples from pigs from West Africa showed identical results by the recombinant protein-based ELISA and the OIE-approved tests. In contrast, there was a high degree of specificity but a surprisingly a low level of sensitivity with 7 positive and 342 negative serum samples from pigs from East Africa. With poorly preserved sera, only the p104R-specific ELISA showed a significant reduction in sensitivity compared to that of the OIE-approved ELISA. Finally, these recombinant proteins also detected antibodies in the sera of the majority of infected warthogs. Thus, recombinant ASFV proteins p54, pB602L, and pK205R provide sensitive and specific targets for the detection of antibodies in European and West African domestic pigs and warthogs.

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Section: Discussionmentioning

confidence: 91%

Section: Discussionmentioning

confidence: 99%

Recombinant Antigen Targets for Serodiagnosis of African Swine Fever

Gallardo

Reis

Kalema‐Zikusoka

et al. 2009

Clin Vaccine Immunol

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“…In warthogs, virus is usually only isolated from warthog lymphatic tissue, with the exception of viremic neonate warthogs (Plowright, 1981;Thomson, 1985). Exposure to ASFV may be detected serologically (Plowright et al, 1994) using any one of a number of available tests, such as conventional indirect ELISA (Alcaraz et al, 1995), recombinant p30 protein ELISA (Perez-Filgueira et al, 2006), or Western blot (Wilkinson, 1984). It has, however, been shown that the percentage of positive warthog samples detected by recombinant and indirect ELISA can be low (Perez-Filgueira et al, 2006), and that confounding results, such as only one of three sibs being seropositive, can occur during warthog seroprevalence studies (Thomson et al, 1983).…”

Section: Diagnosis Of Asf In Wild Suidsmentioning

confidence: 99%

Role of Wild Suids in the Epidemiology of African Swine Fever

2009

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There is presently no vaccine to combat African swine fever (ASF), a viral hemorrhagic fever of domestic pigs that causes up to 100% morbidity and mortality in naive, commercial pig populations. In its endemic setting, ASF virus cycles between asymptomatic warthogs and soft ticks, with persistence in exotic locations being ascribed to the almost global distribution of susceptible soft tick and suid hosts. An understanding of the role played by diverse hosts in the epidemiology of this multi-host disease is crucial for effective disease control. Unlike the intensively studied Ornithodoros tick vector, the role of many wild suids remains obscure, despite growing recognition for suid-exclusive virus cycling, without the agency of the argasid tick, at some localities. Because the four wild suid genera, Phacochoerus, Potamochoerus, Hylochoerus, and Sus differ from each other in taxonomy, distribution, ecology, reservoir host potential, virus shedding, ASF symptomology, and domestic-pig contact potential, their role in disease epidemiology is also varied. This first consolidated summary of ASF epidemiology in relation to wild suids summarizes current knowledge and identifies information gaps and future research priorities crucial for formulating effective disease control strategies.

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“…(), recombinant proteins (e.g. p54) immobilized on nitrocellulose membranes can be stored at room temperature without the loss of antigenic activity for a long period of time (more than 1 year) (Alcaraz et al., ). In our case, the immunostrips have been stored at 4–8°C without the loss of activity for at least 1 year (O.…”

Section: Discussionmentioning

confidence: 99%

Recombinant Protein p30 for Serological Diagnosis of African Swine Fever by Immunoblotting Assay

Kazakova

Imatdinov

Dubrovskaya

et al. 2016

Transbound Emerg Dis

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This article is devoted to the development and evaluation of the immunoblotting test system for serological diagnosis of African swine fever (ASF), based on the highly purified recombinant p30 of ASF virus (ASFV) strain Stavropol 01/08 (Stavropol 2008), representative of the ASFV currently circulating in the Russian Federation. The main project stages are as follows: (i) cloning of the central hydrophilic region of the ASFV gene CP204L (p30) into a prokaryotic vector; (ii) expression and chromatographic purification of the recombinant product p30 with thioredoxin and poly-histidine site (p30e1_TrxA_6xHis); (iii) development of the immunoblotting test system (Rec p30-IB) using the highly purified recombinant p30; and (iv) evaluation of Rec p30-IB using sera and organ samples from domestic pigs and wild boars experimentally or naturally infected by ASFV. Testing of the Rec p30-IB showed the diagnostic specificity and sensitivity of the assay to be 98.75% and 100.00%, respectively. High sensitivity of the Rec p30-IB allowed the detection of ASFV-specific antibodies in samples of organs of the immune system and blood sera, collected from domestic pigs and wild boars, starting from 6 to 8 days post-infection, regardless of virus virulence, seroimmunotype and geographic origin of the samples (East Europe, South Europe, West Europe, Central and south-east Africa).

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Highly specific confirmatory Western blot test for African swine fever virus antibody detection using the recombinant virus protein p54

Cited by 32 publications

References 9 publications

Recombinant Antigen Targets for Serodiagnosis of African Swine Fever

Recombinant Antigen Targets for Serodiagnosis of African Swine Fever

Role of Wild Suids in the Epidemiology of African Swine Fever

Recombinant Protein p30 for Serological Diagnosis of African Swine Fever by Immunoblotting Assay

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