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PubMedDate not supplied

Immunomodulatory effect of plasmids co-expressing cytokines in classical swine fever virus subunit gp55/E2-DNA vaccination.

The aim of this study was to determine the immunomodulatory effects of IL-12, IL-18 and CD154 (CD40 ligand, CD40L) in DNA-vaccination against the classical swine fever virus. Four recombinant plasmids were constructed including the CSFV coding region for the glycoprotein gp55/E2 alone or together with porcine IL-12, IL-18 or CD154 genes. Five groups of four pigs each were immunized intramuscularly (i.m.) three times with the respective constructs. The control group was inoculated with empty plasmid DNA. Eighteen days after the final immunization, the pigs were challenged with a lethal dose of CSFV strain Eystrup and monitored for a further 16 days. This study showed that co-delivery of IL-18 and CD154 induced an earlier appearance of serum antibodies, reduced B-cell deficiency after infection and protected pigs against a lethal CSFV infection. In contrast, co-delivery of IL-12 led to a reduced titer of neutralizing antibodies and protection against a lethal CSFV challenge in comparison to the other pigs and to pigs that were immunized with a gp55/E2 plasmid alone.

Adjuvants, Immunologic↗
PubMed2000

An experimental infection with classical swine fever in E2 sub-unit marker-vaccine vaccinated and in non-vaccinated pigs.

The clinical and virological protection induced by an E2 sub-unit marker-vaccine against Classical Swine Fever (CSF) was examined during an experimental infection in vaccinated and non-vaccinated pigs. Forty-five pigs were equally distributed over three adjacent pens of an isolation unit, there was only indirect (airborne) contact between pigs in the different pens. In pen 3 all pigs were vaccinated twice with 4 weeks interval. Pigs in pens 1 and 2 were not vaccinated. Two weeks after booster vaccination, one randomly selected pig in the middle pen was experimentally inoculated with CSF virus. After the initial virus spread in the infected pen, all pigs in the non-vaccinated adjacent pen were infected. In the vaccinated pen, seven out of 14 pigs became infected during the experiment. Survival analysis showed that virus transmission by direct and indirect contact was significantly (p<0.001) delayed in vaccinated pigs as compared to non-vaccinated pigs. In the non-vaccinated pens over 40% of the pigs died and typical clinical signs were noticed. In the vaccinated pen no mortality and no clinical symptoms were observed. Although double vaccination with an E2 sub-unit marker-vaccine was able to prevent the clinical course of the disease it was unable to prevent infection through indirect contact. This finding combined with the slow serological response after vaccination will complicate the possible use of the vaccine in emergency vaccination programmes.

Animals↗
PubMedDate not supplied

An RT-PCR assay for the specific detection of classical swine fever virus in clinical samples.

A simple reverse transcriptase-polymerase chain reaction (RT-PCR) assay has been developed for the specific amplification of DNA after reverse transcription of RNA from the classical swine fever virus (CSFV). A pair of oligonucleotides was selected from an area of high homology in the genome of CSFV strains, but which differed from the corresponding sequences in the genome of bovine viral diarrhea virus (BVDV) strains. Using these primers (CSFV1-CSFV2), a CSFV specific DNA band of 174 bp was amplified from the CSFV RNA extracted from four reference strains and 14 field isolates, as well as from 25 organ extracts and eight buffy coats and serum samples of experimentally infected animals. No amplification was observed with the RNA from four BVDV reference and vaccine strains and seven field isolates. This RT-PCR assay made it possible, in a one-step reaction, to detect CSFV rapidly, sensitively and specifically in cell culture supernatants and in clinical specimens.

Animals↗
PubMed2004

Development of an optimized protocol for the detection of classical swine fever virus in formalin-fixed, paraffin-embedded tissues by seminested reverse transcription-polymerase chain reaction and comparison with in situ hybridization.

An optimized protocol was developed for the detection of classical swine fever virus (CSFV) in formalin-fixed, paraffin-embedded tissues obtained from experimentally and naturally infected pigs by seminested reverse transcription-polymerase chain reaction (RT-PCR). The results for seminested RT-PCR were compared with those determined by in situ hybridization. The results obtained show that the use of deparaffinization with xylene, digestion with proteinase K, extraction with Trizol LS, followed by seminested RT-PCR is a reliable detection method. An increase in sensitivity was observed as amplicon size decreased. The highest sensitivity for RT-PCR on formalin-fixed, paraffin-embedded tissues RNA was obtained with amplicon sizes less than approximately 200 base pairs. An hybridization signal for CSFV was detected in lymph nodes from 12 experimentally and 12 naturally infected pigs. The sensitivity of seminested RT-PCR compared with in situ hybridization was 100% for CSFV. When only formalin-fixed tissues are available, seminested RT-PCR and in situ hybridization would be useful diagnostic methods for the detection of CSFV nucleic acid.

Animals↗
PubMed2006

Validation of a real-time RT-PCR assay for rapid and specific diagnosis of Classical Swine Fever virus.

Two new real-time RT-PCR kits developed by LSI (TaqVet CSF) and ADIAGENE (Adiavet CSF) obtained a manufacturing agreement in France during the past year. For that purpose, the Classical Swine Fever (CSF) National Reference Laboratory (NRL) planned a schedule of conditions to be fulfilled by commercial real-time RT-PCR assays. The producers were asked to introduce an internal control to check the RNA extraction efficacy. The different criteria assessed were: sensitivity, specificity, especially "pestivirus specificity", reproducibility and easy handling, using 187 different samples distributed in four different panels. These samples were either CSFV inactivated strains or organs collected from CSF experimental SPF infected pigs, or naturally infected wild boars. All these samples were previously tested for genome detection using an RT-nested PCR assay and for virus isolation on cell culture. The LSI TaqVet kit was used for the CSF surveillance of wild boars in an area known to be infected, during the winter of 2004-2005. This field evaluation was carried out on 4710 spleen samples. In summary, the new CSF real-time RT-PCR assays have a higher predictive value than the current diagnostic standard, Virus Isolation.

Animals↗
PubMed2006

Protection of gruntlings against classical swine fever virus-infection after oral vaccination of sows with C-strain vaccine.

The objective of this study was to investigate the maternal protection of gruntlings derived from wild sows vaccinated orally against classical swine fever (CSF) using C-strain vaccine. Three vaccinated sows and one unvaccinated control sow were included. Challenge infection of the progeny was carried out either intranasally or by contact at the beginning of the third month of life (61-65 days post-natum). Whereas, two of three litters had maternal antibodies, the progeny of one vaccinated sow was seronegative at challenge. The progeny of the control sow, which was challenged by contact infection, developed moderate clinical signs except for one animal which became ill and died. Two gruntlings derived from the vaccinated sows also died of CSF, although one of them had a relatively high maternal antibody titre (128 ND(50)). The transient infection and partial virus shedding observed in a small number of gruntlings with maternal antibodies and the fact that one animal with maternal antibodies became ill and died confirm the incomplete maternal protection at this age. The reason for this incomplete protection is discussed. As none of the surviving gruntlings could be shown to carry CSFV or viral RNA at the end of the experiment (36 or 70 d.p.i.), it may be concluded that these animals do not represent a potential CSFV reservoir.

Animals↗
PubMed1999

Spatial and stochastic simulation to evaluate the impact of events and control measures on the 1997-1998 classical swine fever epidemic in The Netherlands. II. Comparison of control strategies.

Using the spatial, temporal and stochastic simulation model InterCSF, several alternative pre-emptive slaughter strategies that could have been applied in the Dutch Classical Swine Fever (CSF) epidemic of 1997-1998 were evaluated. Furthermore, effects of changes in some disease-spread and disease-control parameters were studied. InterCSF simulates the spread of CSF between farms through local spread and contacts (animals, transport and persons). Disease spread is affected by control measures implemented through different mechanisms (e.g. depopulation of infected farms, pre-emptive slaughter, movement control). The starting point for the evaluation of strategies was a simulated basic scenario, which mimicked the real epidemic. Strategies were compared using epidemiological as well as economic results. Economic results were generated by a separate model (EpiLoss) that calculated the direct losses and consequential losses for farmers and related industries. The comparison of the different alternatives to the basic scenario led to some general conclusions on the Dutch CSF-epidemic. Pre-emptive slaughter seemed to be an effective strategy to reduce the size of an epidemic, if started at an early stage. Economically, pre-emptive slaughter was not as expensive as expected; the resulting smaller size of the epidemic, combined with less welfare slaughter, led to much lower overall losses. Furthermore, although large movement control areas seemed effective in reducing the size of the epidemic, the total losses were relatively high because of subsequent welfare slaughter. If infection probabilities could be reduced, for example by improved biosecurity, the resulting epidemics would be much smaller.

Animal Husbandry↗
PubMed2005

A DNA vaccine expressing the E2 protein of classical swine fever virus elicits T cell responses that can prime for rapid antibody production and confer total protection upon viral challenge.

Immunization of domestic pigs with a DNA vaccine expressing the complete E2 protein of classical swine fever virus (CSFV) conferred total protection against a severe viral challenge. Immunization with three doses of plasmid pcDNA3.1/E2 elicited a consistent and specific, MHC class II restricted T cell response in the three domestic pigs analyzed, in the absence of detectable anti-CSFV antibodies in serum. Upon challenge specific T cell responses were boosted in the three vaccinated pigs, and a rapid rise in the titers of CSFV neutralizing antibodies was noticed in two of them, which correlated with a total protection. In these two pigs, neither disease symptoms were observed nor was virus detected at any time after CSFV infection. Neutralizing antibody titers were lower in the third vaccine, which developed a mild and transient peak of pyrexia. As expected, similar analyses in three control pigs (injected with the empty vector or PBS) did not reveal the induction of specific T cells or viral antibodies and, upon challenge, animals developed severe symptoms of the disease, including high titers of viremia, hyperthermia and virus spread to different organs. Control pigs developed, also, a marked leucopenia, resulting in SWC3+ (myelomonocytic cells) being the major PBMC population, and a drastic decrease CD3+ T cells. This T cell depletion was prevented in animals immunized with pcDNA3.1/E2. The total protection achieved, in the absence of CSFV antibodies before challenge, supports the relevance in the antiviral response observed of specific T cell responses primed by pcDNA3.1/E2 vaccine, which, upon challenge, led to a rapid induction of neutralizing antibodies. The observation that CSFV antibodies could only be detected in protected animals after viral challenge opens the possibility of exploring the potential of the DNA vaccine approach used to develop marker vaccines against CSF.

Amino Acid Sequence↗
PubMed2002

Within- and between-pen transmission of Classical Swine Fever Virus: a new method to estimate the basic reproduction ratio from transmission experiments.

We present a method to estimate basic reproduction ratio R0 from transmission experiments. By using previously published data of experiments with Classical Swine Fever Virus more extensively, we obtained smaller confidence intervals than the martingale method used in the original papers. Moreover, our method allows simultaneous estimation of a reproduction ratio within pens R0w and a modified reproduction ratio between pens R'0b. Resulting estimates of R0w and R'0b for weaner pigs were 100 (95% CI 54.4-186) and 7.77 (4.68-12.9), respectively. For slaughter pigs they were 15.5 (6.20-38.7) and 3.39 (1.54-7.45), respectively. We believe, because of the smaller confidence intervals we were able to obtain, that the method presented here is better suited for use in future experiments.

Animals↗
PubMed2004

Ability of veterinary pathologists to diagnose classical swine fever from clinical signs and gross pathological findings.

Clinical signs recorded in a classical swine fever (CSF)-suspect situation and the results of the subsequent post-mortem examination (PME) from swine submitted to post-mortem during the 1997-1998 CSF epidemic in the Netherlands, were presented in an experiment as anonymous cases (without knowledge of the actual infection status of the submission) to five veterinary pathologists for their judgment: CSF-suspect or non-suspect. It was presented to them in two hypothetical situations: country was free of CSF for 5 years and CSF was detected in the country 2 weeks ago. Subsequently, their judgment was compared to the gold standard (infection status of the submission on the basis of an immunofluoresence assay on tissue samples) and the sensitivity (Se) and specificity (Sp) of clinical diagnosis was estimated. Furthermore, intra- and inter-pathologist agreement on pathological diagnosis was measured. Adding information on clinical signs to information on PME resulted in a significant increase in median Se and a significant decrease in median Sp for a clinical judgment by pathologists. Also, median Se was significantly higher-and Sp significantly lower-for a clinical judgment by pathologists in a situation in which CSF was detected 2 weeks ago in the country compared to the situation in which the country had been free of CSF for 5 years. Apparently, the internal threshold of pathologists is severely adjusted depending on the actual disease situation in a country. Intra-rater agreement ranged from fair to almost perfect for a clinical diagnosis on the basis of PME. There was considerable variation between pathologists, especially if clinical diagnosis was based on the combined information on PME and clinical signs observed in the herd. Inter-rater agreement was substantial for the diagnosis based on information on PME. Inter-rater agreement was considerably lower for the diagnosis based on the combined information on PME and clinical signs observed in the herd.

Animals↗
PubMed2003

Simulated epidemiological and economic effects of measures to reduce piglet supply during a classical swine fever epidemic in The Netherlands.

The effects of additional measures adopted during a classical swine fever (CSF) epidemic to reduce piglet supply, namely, an insemination ban, abortion of sows and killing of young piglets, are studied using a stochastic, spatial, dynamic epidemiological simulation model of the pig sector in The Netherlands. The piglet supply derived from the epidemiological model is used as input for a sector-level market and trade model that simulates the pig market in The Netherlands. Changes in the economic welfare of different stakeholders are measured, as is the net welfare effect for the economy in The Netherlands. Sensitivity analysis is performed on parameters such as destruction capacity constraints and the duration of the high-risk period. Additional measures to reduce piglet supply are found to have no epidemiological impact, but they do involve larger economic welfare changes for stakeholders and a larger net welfare loss for the economy in The Netherlands. These findings do not support the use of the additional measures. Moreover, sensitivity analysis shows that such measures do not solve the problem of a shortage in rendering capacities.

Abortion, Induced↗
PubMed2003

The risk of the introduction of classical swine fever virus at regional level in the European Union: a conceptual framework.

Recent classical swine fever (CSF) epidemics in the European Union (EU) have clearly shown that preventing the introduction of CSF virus (CSFV) deserves high priority. Insight into all the factors contributing to the risk of CSFV introduction is a prerequisite for deciding which preventive actions are cost-effective. The relations between virus introduction and spread, prevention and control, and economic losses have been described using the conceptual framework presented in this paper. A pathway diagram provides insight into all the pathways contributing to the likelihood of CSFV introduction (LVI_CSF) into regions of the EU. A qualitative assessment based on this pathway diagram shows that regions with high pig densities generally have a higher LVI_CSF, although this cannot be attributed to pig density only. The pathway diagram was also used to qualitatively assess the reduction in LVI_CSF achieved by restructuring the pig production sector. Especially integrated chains of industrialised pig farming reduce the LVI_CSF considerably, but are also difficult and costly to implement. Quantitative assessment of the LVI_CSF on the basis of the pathway diagram is needed to support the results of the qualitative assessments described.

Animal Husbandry↗
PubMed2004

Specific interaction between the classical swine fever virus NS5B protein and the viral genome.

The NS5B protein of the classical swine fever virus (CSFV) is the RNA-dependent RNA polymerase of the virus and is able to catalyze the viral genome replication. The 3' untranslated region is most likely involved in regulation of the Pestivirus genome replication. However, little is known about the interaction between the CSFV NS5B protein and the viral genome. We used different RNA templates derived from the plus-strand viral genome, or the minus-strand viral genome and the CSFV NS5B protein obtained from the Escherichia coli expression system to address this problem. We first showed that the viral NS5B protein formed a complex with the plus-strand genome through the genomic 3' UTR and that the NS5B protein was also able to bind the minus-strand 3' UTR. Moreover, it was found that viral NS5B protein bound the minus-strand 3' UTR more efficiently than the plus-strand 3' UTR. Further, we observed that the plus-strand 3' UTR with deletion of CCCGG or 21 continuous nucleotides at its 3' terminal had no binding activity and also lost the activity for initiation of minus-strand RNA synthesis, which similarly occurred in the minus-strand 3' UTR with CATATGCTC or the 21 nucleotide fragment deleted from the 3' terminal. Therefore, it is indicated that the 3' CCCGG sequence of the plus-strand 3' UTR, and the 3' CATATGCTC fragment of the minus-strand are essential to in vitro synthesis of the minus-strand RNA and the plus-strand RNA, respectively. The same conclusion is also appropriate for the 3' 21 nucleotide terminal site of both the 3' UTRs.

3' Untranslated Regions↗
PubMed1998

In vivo effect on pig chromosomes of high dosage vaccine against classic swine fever.

Hog cholera virus (HCV) can induce chromosome abnormalities in diseased pigs as well as in those vaccinated with attenuated virus vaccine against classic swine fever. An experiment was made using animals from potency and safety control tests of commercial vaccines in Argentina. The different types of chromosomal alterations observed were chromatid and chromosome breaks, chromatid exchanges, polyploid, multiple aberrations cells, and chromosome pulverization. In this study the occurrence of chromosome alterations in pigs receiving either 1 or 10 vaccine doses was evaluated by means of blood sampling at different periods after vaccination. An essay comparing prolificity between treated and non-treated sows was also made. Significant differences in the amount of damaged chromosomes as well as differences in the type of predominant alterations between the two treatments were observed. Aberration frequencies increased from the 5-day postvaccination period reaching the highest value of 4.14% at the 10th, for the one-dose treatment; and highest value of 42.7% including 33.96% of cell with chromosome pulverization which was found in the 7th day interval when applying 10 doses. From then on, the proportion of affected cells dropped until the 20th day interval, which was the last recorded. The prolificity trial did not show any difference between treated and control sows, indicating that chromosome alteration might be limited to lymphocytes. It is concluded that HCV maintains its mutagenic potentiality in the attenuated vaccine, being able to induce chromosomal damage as it does in classic swine fever diseased animals.

Animals↗
PubMed2006

Development of a reverse-transcription polymerase chain reaction assay with fluorogenic probes to discriminate Korean wild-type and vaccine isolates of Classical swine fever virus.

A 1-step reverse-transcription polymerase chain reaction (RT-PCR) assay using TaqMan minor-groove-binding (MGB) probes was developed to distinguish between vaccine-type and wild-type strains of Classical swine fever virus (CSFV) in Korea. Because attenuated Korean LOM strains have been used in animal vaccination in Korea for some time but CSF remains a serious problem, there was a need for a practical approach to differentiating vaccine and field strains. We examined the fluorescence of 5 vaccine strains, 10 field strains, and 5 mixed samples. Three clusters of the samples could be distinguished: those with only fluorescence of the vaccine-type-specific probe, VIC; those with only fluorescence of the wild-type-specific probe, FAM; and those with both VIC and FAM fluorescence. The RT-PCR assay with fluorogenic probes is sensitive and accurate and is therefore useful for differentiating vaccine and field strains of CSFV in Korea.

Animals↗
PubMed2004

An avirulent chimeric Pestivirus with altered cell tropism protects pigs against lethal infection with classical swine fever virus.

A chimeric Pestivirus was constructed using an infectious cDNA clone of bovine viral diarrhea virus (BVDV) [J. Virol. 70 (1996) 8606]. After deletion of the envelope protein E2-encoding region, the respective sequence of classical swine fever virus (CSFV) strain Alfort 187 was inserted in-frame resulting in plasmid pA/CP7_E2alf. After transfection of in vitro-transcribed CP7_E2alf RNA, autonomous replication of chimeric RNA in bovine and porcine cell cultures was observed. Efficient growth of chimeric CP7_E2alf virus, however, could only be demonstrated on porcine cells, and in contrast to the parental BVDV strain CP7, CP7_E2alf only inefficiently infected and propagated in bovine cells. The virulence, immunogenicity, and "marker vaccine" properties of the generated chimeric CP7_E2alf virus were determined in an animal experiment using 27 pigs. After intramuscular inoculation of 1 x 10(7) TCID(50), CP7_E2alf proved to be completely avirulent, and neither viremia nor virus transmission to contact animals was observed; however, CSFV-specific neutralizing antibodies were detected from day 11 after inoculation. In addition, sera from all animals reacted positive in an E2-specific CSFV-antibody ELISA, but were negative for CSFV-E(RNS)-specific antibodies as determined with a CSFV marker ELISA. After challenge infection with highly virulent CSFV strain Eystrup, pigs immunized with CP7_E2alf were fully protected against clinical signs of CSFV infection, viremia, and shedding of challenge virus, and almost all animals scored positive in a CSFV marker ELISA. From our results, we conclude that chimeric CP7_E2alf may not only serve as a tool for a better understanding of Pestivirus attachment, entry, and assembly, but also represents an innocuous and efficacious modified live CSFV "marker vaccine".

Animals↗
PubMed2005

Estimating the probability of freedom of classical swine fever virus of the East-Belgium wild-boar population.

A report of the Scientific Committee on Animal Health and Animal Welfare of the European Commission (CEC, 1999.) includes recommendations for setting up monitoring programmes for classical swine fever (CSF) infection in a wild-boar population, based on the assumption that one would detect at least 5% prevalence in a CSF-infected wild-boar population. This assumption, however, is not science based. We propose an alternative method to provide evidence for a wild-boar population being free of CSF and evaluate the efficiency of a surveillance programme that was implemented in Belgium in 1998. In our study, the probability of freedom of CSF-virus was estimated based on 789 samples; these were collected from wild-boars within the surveillance programme (within the three provinces which include 95% of the Belgian wild-boar population) and examined by three diagnostics methods (antibody detection, virus detection and virus RNA detection). A Bayesian framework was used for the estimation, accounting for the diagnostic test characteristics without the assumption of the presence of a gold standard. The median probability of freedom of CSF-virus was estimated at 0.970, with a 95% credibility interval of 0.149-1.000. Independent on the choice of the prior information, the posterior distributions for the probability of freedom of CSF-virus were always skewed close to the upper boundary of 1. This represents a big gain of knowledge since we did not use any prior information for the probability of freedom of CSF-virus and took the uncertainty about the accuracy of the diagnostic methods into account.

Animals↗