Detection of antibodies against porcine epidemic diarrhea virus in serum and colostrum by indirect ELISA

Porcine epidemic diarrhea virus (PEDV) is the highly contagious, causative agent of an economically important acute enteric disease in pigs of all ages. The disease is characterized by diarrhea and dehydration causing mortality and growth retardation. In the last few decades, only classical PEDV was reported sporadically in Europe, but in 2014 outbreaks of PEDV were described in Germany. Phylogenetic analysis showed a very high nucleotide similarity with a variant of PEDV that was isolated in the US in January 2014. The epidemiological situation of PEDV infections in the Netherlands in 2014 was unknown and a seroprevalence study in swine was performed. In total, 838 blood samples from sows from 267 farms and 101 samples from wild boars were collected from May till November 2014 and tested for antibodies against PEDV by ELISA. The apparent herd prevalence of 0.75% suggests that PEDV was not circulating on a large scale in the Netherlands at this time. However, in November 2014 a clinical outbreak of PEDV was diagnosed in a fattener farm by PCR testing. This was the first confirmed PEDV outbreak since the early nineties. Sequence analyses showed that the viruses isolated in 2014 and 2015 in the Netherlands cluster with recently found European G1b strains. This suggests a one event introduction of PEDV G1b strains in Europe in 2014, which made the Netherlands and other European countries endemic for this type of strains since then.

Section: Discussionmentioning

confidence: 97%

Section: Indirect Elisamentioning

confidence: 99%

Porcine epidemic diarrhea virus (PEDV) introduction into a naive Dutch pig population in 2014

Dortmans

Wolf

et al. 2018

“…Recombinant PEDV S1 protein was obtained from PK-rS1-Ig cells (Oh et al, 2014) and purified as described previously (Oh et al, 2014). Anti-PEDV immunoglobulin (Ig) A antibodies in serum and colostrum were detected using an in-house PEDV G2b S1-based indirect enzymelinked immunosorbent assays (ELISA) as described previously (Gerber et al, 2014;Gerber and Opriessnig, 2015), with minor modifications. Briefly, microtiter plates (Nunc, Naperville, IL) were coated with 1.6 ng per well of the S1 antigen diluted in coating buffer (50 mM bicarbonate buffer, pH 9.6) and incubated overnight at 4°C.…”

Section: Enzyme-linked Immunosorbent Assaysmentioning

confidence: 99%

Assessment of the safety and efficacy of an attenuated live vaccine based on highly virulent genotype 2b porcine epidemic diarrhea virus in nursing piglets

Jang

Won

Lee³

et al. 2019

We have previously reported the generation of the attenuated KNU-141112-S DEL5/ORF3 virus by continuous propagation of highly virulent G2b porcine epidemic diarrhea virus (PEDV) in Vero cells. The present study aimed to assess the safety of S DEL5/ORF3 and to evaluate its effectiveness as a live vaccine for prime-booster vaccinations. Reversion to virulence experiments revealed that the S DEL5/ORF3 strain retains its attenuated phenotype and genetic stability after five successive passages in susceptible piglets. Pregnant sows were primed orally with an S DEL5/ORF3 live vaccine and boosted intramuscularly twice with a commercial killed vaccine at 2-week intervals prior to parturition. This sow vaccination regimen completely protected nursing piglets against virulent G2b challenge, as evidenced by the increase in survival rate from 0% to 100% and the significant reduction in diarrhea intensity, including the amount and duration of PEDV fecal shedding. In addition, despite a 2-3 day period of weight loss in piglets from vaccinated sows after challenge, their daily weight gain was recovered at 7 days post-challenge and became similar to that of unchallenged pigs from unvaccinated sows over the course of the experiment. Furthermore, strong antibody responses to PEDV were verified in the sera and colostrum of immunized sows with the prime-boost treatment and their offspring. Altogether, our data demonstrated that the attenuated S DEL5/ORF3 strain guarantees the safety to host animals with no reversion to virulence and is suitable as an effective primary live vaccine providing durable maternal lactogenic immunity for passive piglet protection.

“…The constructs used for this study are listed in Table 1. For transient expression in mammalian cells, the pcDNA-3.1 vector was used, and the expression plasmid pcDNA-PEDV-S1-Fc has been described in the previous study (Gerber et al, 2014).…”

Section: Expression and Purification Of Recombinant Pedv Proteinsmentioning

confidence: 99%

“…Another common diagnostic method is serological testing for the presence of specific antibodies against viral proteins, which is also fast and convenient for epidemiologic investigations. Many tools have been developed for the detection of anti-PEDV antibodies based upon the major structural proteins (such as S, M or N proteins) in serum, colostrum, milk, feces and oral fluid, including indirect immunofluorescence assays (IFA), virus neutralization assays (SN), enzyme-linked immunosorbent assays (ELISA), and fluorescent microsphere immunoassays (FMIA) (Diel et al, 2016;Gerber et al, 2014;Gerber and Opriessnig, 2015;Gimenez-Lirola et al, 2017;Okda et al, 2015). But comparative studies of the above assays using different PEDV structural and nonstructural proteins as antigens are rarely conducted.…”

Section: Introductionmentioning

confidence: 99%

Specific recombinant proteins of porcine epidemic diarrhea virus are immunogenic, revealing their potential use as diagnostic markers

Lei

Yang

et al. 2019

Self Cite

A B S T R A C TGiven the highly contagious and acute nature of porcine epidemic diarrhea (PED), especially in piglets, there is an urgent need for the development of rapid and sensitive diagnostic assays. The diagnostic potentials of specific porcine epidemic diarrhea virus (PEDV) accessory and nonstructural proteins, if any, have not yet been investigated. In order to determine and compare which of the viral proteins may be useful as diagnostic antigens, whole virus (WV) particles and a panel of structural and nonstructural PEDV proteins [spike subunit 1 (S1), the C-terminal part of ORF3 (ORF3C), envelope (E), nonstructural protein 1 (Nsp1), Nsp2, Ac (acidic domain of Nsp3), and ADRP (ADP-ribose-1-monophosphatase domain of Nsp3), expressed individually in bacterial and/or mammalian cells] were tested for reactivity with sera from PEDV-infected pigs by ELISA and/or western blot analysis. According to western blots, serum antibody interactions with the S1 protein were relatively more sensitive and specific than ORF3C, E and Ac. Furthermore, a total of 851 serum samples from diarrheal pigs of different ages were analyzed by ELISA, with most showing immune-reactivity towards the WV, S1, ORF3C, and E proteins. The earliest IgG antibody response was observed in the one-week-old piglets, with similar antibody ontogeny and patterns of seroconversion for S1, ORF3C, E, and WV antigens. In addition, the pattern of neutralizing antibody was more similar to that of IgA in weaning piglets after PEDV infection. Collectively, these data provide more reliable information on the host immune response to different viral proteins, which will be useful for development of novel serological assays and for design of vaccines that better stimulate protective immunity.