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PubMed2006

[Genetic typing of classical swine fever viruses--a review].

Classical swine fever (CSF) is a notifiable disease of domestic pigs and wild boar. It is caused by the highly contagious CSF virus and in its acute form the disease generally results in high morbidity and mortality. Due to the great economical impact an outbreak can cause to the pig industry it is one of the most important swine diseases worldwide. To limit the damage in the case of a new outbreak it is necessary to identify the virus as fast as possible. This information helps epidemiologists to trace the origin of the virus and to follow the virus spread. Genetic typing revealed that CSF virus genotypes, subgroups and types show a regional distribution making it an important tool for epidemiologists. Meanwhile, besides epidemiological data and nucleotide sequences from European isolates, information from isolates from South- and Central America, the Caribbean, Asia and recently from South Africa have become available. The data are stored in a database in the EU Reference Laboratory for CSF, accessible by the WWW (http://viro08.tiho-hanno ver.de). A new module was implemented that allows efficient automated genotyping.

Animals↗

PubMed2004

Transmission of classical swine fever. A review.

Classical swine fever (CSF) is one of the most important viral pig diseases. Basic measures to control epidemics of CSF comprise eradication of infected herds and preventive emptying of herds at risk. Identification of these herds at risk is based on knowledge of the different transmission routes of the virus. Direct transmission of CSF is undoubtedly the most efficient way of CSF virus transmission. Data on indirect transmission are variable and often equivocal. Various indirect transmission routes like swill feeding, wild boar and artificial insemination are well described, and the importance is beyond discussion. Mechanical transmission via vehicles and persons are categorized as very important based upon epidemiological research, whereas in experiments they can only be reproduced under worst case conditions. The role of arthropods, birds, rodents, and other animals in the spread of CSF virus remains doubtful. Active transmission by these has never been demonstrated and also very sparse indications for mechanical transmission are available. Also the role of airborne transmission remains debated. However epidemiological as well as experimental data indicate that airborne spread over short distances is probable.

Animals↗

PubMed2006

A sequence database allowing automated genotyping of Classical swine fever virus isolates.

Classical swine fever (CSF) is a highly contagious viral disease of pigs. According to the OIE classification of diseases it is classified as a notifiable (previously List A) disease, thus having the potential for causing severe socio-economic problems and affecting severely the international trade of pigs and pig products. Effective control measures are compulsory, and to expose weaknesses a reliable tracing of the spread of the virus is necessary. Genetic typing has proved to be the method of choice. However, genotyping involves the use of multiple software applications, which is laborious and complex. The implementation of a sequence database, which is accessible by the World Wide Web with the option to type automatically new CSF virus isolates once the sequence is available is described. The sequence to be typed is tested for correct orientation and, if necessary, adjusted to the right length. The alignment and the neighbor-joining phylogenetic analysis with a standard set of sequences can then be calculated. The results are displayed as a graph. As an example, the determination is shown of the genetic subgroup of the isolate obtained from the outbreaks registered in Russia, in 2005. After registration (Irene.greiser-wilke@tiho-hannover.de) the database including the module for genotyping are accessible under http://viro08.tiho-hannover.de/eg/eurl_virus_db.htm.

Algorithms↗

PubMedDate not supplied

[Sequence, necessary for initiating RNA synthesis, in the 3'-noncoding region of the classical swine fever virus genome].

Classical swine fever virus (CSFV) is the causative agent of swine fever, which represents an economically important disease in hogs. We previously made a prediction about the recognition sites of replication initiation of CSFV by using the information content method, and it was predicted that the 21 nucleotides located at 3' end of the CSFV genome 3'UTR were essential to CSFV replication. In this paper, we experimentally studied these 21 nucleotides by site-directed mutagenesis. It was found that the 3'UTRs with the 21 nucleotides had the function of initiating RNA synthesis, while the 3'UTRs without the 21 nucleotides did not. The 21 nucleotides alone, without the rest of 3'UTR, were able to initiate RNA synthesis, though with a slump. It was demonstrated that the 21 nucleotides were essential to the replication of CSFV genome. The other part of 3'UTR was also required for sufficient RNA synthesis. It is highly likely that the 21 nucleotides were the necessary site for the CSFV genome replication initiation, and that the elements required for sufficient RNA synthesis were in the other part of 3'UTR. It was assumed that the CSFV replicase bound to the site and initiated the replication of the CSFV genome. In the 21 nucleotides, it was found that the mutation of position 216 and destruction of the 3' terminus in the 3'UTR precluded initiation of RNA synthesis, that the mutation of position 212 did not affect the capacity for initiation of RNA synthesis but attenuated the synthesis of RNA. Among the four mutants of 3'UTR at position 219, three produced the 3'UTR without initiation of RNA synthesis, and the other one produced the 3'UTR with initiation of less RNA synthesis. Therefore, it could be concluded that T216 was the most important while T212 was the least important, and that G219 and C228 were also important for RNA synthesis. The normal base component within the 21 nucleotides was essential to sufficient RNA synthesis.

3' Untranslated Regions↗

PubMed2006

Sensitivity analysis to evaluate the impact of uncertain factors in a scenario tree model for classical swine fever introduction.

Introduction of classical swine fever virus (CSFV) is a continuing threat to the pig production sector in the European Union. A scenario tree model was developed to obtain more insight into the main risk factors determining the probability of CSFV introduction (P(CSFV)). As this model contains many uncertain input parameters, sensitivity analysis was used to indicate which of these parameters influence model results most. Group screening combined with the statistical techniques of design of experiments and meta-modeling was applied to detect the most important uncertain input parameters among a total of 257 parameters. The response variable chosen was the annual P(CSFV) into the Netherlands. Only 128 scenario calculations were needed to specify the final meta-model. A consecutive one-at-a-time sensitivity analysis was performed with the main effects of this meta-model to explore their impact on the ranking of risk factors contributing most to the annual P(CSFV). The results indicated that model outcome is most sensitive to the uncertain input parameters concerning the expected number of classical swine fever epidemics in Germany, Belgium, and the United Kingdom and the probability that CSFV survives in an empty livestock truck traveling over a distance of 0-900 km.

Animals↗

PubMed1979

[Viral carrier state and viral excretion in classical swine fever].

Studies on swine pest virus carrying and elimination on swine vaccinated by lapinizated vaccine strain "K" from rabbits were performed. Vaccinated swine were injected with the pathogenic virus on the 60th day post vaccination and to them were added not immunized swine with the aim to discover virus elimination. The experimental swine were decapitated on the 3, 5, 6 and 7th day post infection. Alternating passages were made by emulsion of the inner organs of these swine on not immunized swine as well as pathohistological investigation of the parenchymal organs and the brain. Presence of the virus and of pathohistological changes were not detected 7 days post injection of the virulent virus. Contact having swine which served as a indicator for virus elimination were observed for 21 days, but remained healthy. These swine, when infected with the virulent virus, suffered from pest.

Animals↗

PubMed1993

Application of a blocking enzyme-linked immunosorbent assay for serological monitoring of hog cholera (classical swine fever) in Poland.

Between 1990 and 1992, serum samples from 55,478 domestic swine were tested by enzyme-linked immunosorbent assay (ELISA) for the presence of hog cholera virus (HCV) antibodies. The amount of antibody in the sera was expressed as the mean percentage inhibition (PI). For diagnosis, the tested sera were diluted 1:2 and considered positive if the PI was less than 25%. Sera giving PI values in the range of 25-50% were retested against HCV and bovine virus diarrhoea virus (BVDV), by neutralising peroxidase-linked assay. Comparison of the serum titres obtained was used for serological diagnosis of hog cholera; the tested sera were considered negative for hog cholera if the titre for BVDV was higher than that obtained for HCV. All sera with a PI higher than 50% were considered negative for HCV and BVDV. All sera were found to be free of antibodies to HCV. BVDV antibodies were demonstrated in 0.40% of the sera tested in 1990, in 1.80% in 1991 and 1.06% in 1992.

Animals↗

PubMed1998

Isolation and characterization of cytopathogenic classical swine fever virus (CSFV).

Two new classical swine fever virus (CSFV) isolates obtained from naturally infected swine were found to exhibit a cytopathogenic (cp) phenotype. According to their reactivity with monoclonal antibodies (mabs) the isolates cpBW1 and cpMVP1 were classified as antigenic types "Lothringen'92" and "Flandern'90", respectively. In Northern blot analyses and PCR assays CSFV RNA of subgenomic length was detected in infected cells indicating the presence of defective interfering particles. Nucleotide sequencing of cDNA fragments obtained by reverse transcription (RT)-PCR allowed to localize in the genomes of the cpMVP1 and cpBW1 DIs deletions of 4,764 and 4,746, respectively. From the latter isolate a noncytopathogenic (noncp) biotype was obtained by cloning in cell culture. CSFV cpBW1 and a noncp isolate were used separately for infection of pigs. Viremia occurred much quicker after infection with cp virus and clinical signs were more prominent compared to animals infected with noncp virus.

Amino Acid Sequence↗

PubMed1999

Phenotypic analysis of peripheral leukocytes in piglets infected with classical swine fever virus.

The phenotypic changes in circulating leukocytes in swine fever influenced by classical swine fever virus (CSFV) infection with different strain virulence was studied in piglets. The phenotypic differences were measured by monoclonal antibodies specific for porcine differentiation antigens. The pattern of phenotypic change varied with the virulence of CSFV. Infection with virulent, but not the attenuated strain of CSFV resulted in the dramatic early loss of CD8-bearing T lymphocytes from the circulation. A similar trend was also seen in the gammadelta T-cell compartment following infection with the highly virulent strain, Washington. The loss of circulating B-lymphocytes was consistent with the failure to generate neutralising antibody. These observations contrasted the finding that the number of leukocytes expressing the CD4 surface antigen increased in piglets infected with CSFV. These data provide preliminary information on the potential range of leukocyte changes produced in piglets following infection with CSFV.

Animals↗

PubMed2000

Laboratory diagnosis, epizootiology, and efficacy of marker vaccines in classical swine fever: a review.

Detection of classical swine fever virus (CSFV) can be achieved by a range of assays of which the most commonly used are: immunohistochemical and virus culture techniques. New developments have enabled the detection of viral proteins by enzyme-linked immunosorbent assays (ELISAs) and the detection of the viral genome by RT- PCR. So far, laboratory findings show that the latter assays may supplement or replace the conventional techniques in the near future. The detection of serum antibody against structural and non-structural proteins of CSFV has been improved by developments in recombinant DNA techniques and has lead to a range of ELISAs. Although the characteristics of these ELISAs are excellent, positive results still need to be confirmed in the virus neutralization test. The available amount of sequence data enables diagnosticians to type strains of CSFV as different by comparing several parts of the genome. In some cases, this can provide conclusive evidence if a primary or secondary outbreak has been detected. Increased efforts focused on the retrieval of relevant data on the introduction of CSFV in a pig holding and the spread of CSFV in- and between pig holding(s) has generated more insight into the epizootiology of the disease. A successful control and eradication programme for classical swine fever (CSF) can consist of zoosanitary measures and/or vaccination. The latter can compromise the export of live pigs and pig products considerably unless marker vaccines have been used. Several studies were performed to determine the efficacy of an E2 subunit vaccine and live recombinant vaccine candidates. Firstly, we determined the 95% protective dose of an E2 subunit vaccine at 32 microg E2 per dosage after a single application. Further studies with a single administration of the subunit vaccine showed that: the vaccine was stable for a prolonged period after production, was able to reduce horizontal and vertical transmission of CSFV among vaccinated pigs, and provided protection for at least 6 months. An E(rns) antibody discriminatory assay was developed for use in combination with the subunit vaccine. Evaluation of the E(rns) ELISA showed that the sensitivity of the assay was lower than but that the specificity was equal to that of existing antibody assays. Two live recombinant marker vaccines were evaluated for the induction of clinical protection and reduction of transmission of CSFV shortly after vaccination. Results showed that these vaccines provided good clinical protection 1 week after a single vaccination. Research has shown that marker vaccines can be used in the future to support the control and eradication of CSFV.

Animals↗

PubMed2006

The protective immune response induced by B cell epitope of classical swine fever virus glycoprotein E2.

Classical swine fever virus (CSFV) envelope glycoprotein E2 is a major protective immunogen responsible for eliciting neutralizing antibodies and conferring protective immunity against the virus. Based on the core sequence (TAVSPTTLR, 829-837 aa) of the B cell linear epitope of the CSFV E2 protein identified by Lin et al., two oligonucleotides MF and MR were synthesized and used to construct by PCR a gene cassette encoding a 15 amino acid polypeptide M (CTAVSPTTLRTEVVK), which spans 828-842 amino acids of E2. The gene cassette was fused in-frame to 3' terminal of glutathione S transferase gene (GST) of the prokaryotic expression vector pGEX-6p-1, resulting in the recombinant plasmid pGEX-M. After transformation into Escherichia coli BL21 a soluble fusion protein GST-M with expected size of 28 kDa was expressed after inducing with isopropyl-beta-d-thiogalactoside (IPTG). Enzyme-linked immunosorbent assay (ELISA) and Western blot analysis showed that the purified GST-M had good reactivity with swine anti-CSFV serum and rabbit anti-CSFV E2 serum. Further vaccination trials showed that the fusion protein GST-M could elicit effectively immune response protecting rabbits and pigs from virulent challenge. This study showed a possibility for developing epitope-based vaccines against CSFV.

Amino Acid Sequence↗

PubMed2005

De novo RNA synthesis and homology modeling of the classical swine fever virus RNA polymerase.

Classical swine fever virus (CSFV) non-structural protein 5B (NS5B) encodes an RNA-dependent RNA polymerase (RdRp), a key enzyme which initiates RNA replication by a de novo mechanism without a primer and is a potential target for anti-virus therapy. We expressed the NS5B protein in Escherichia coli. The rGTP can stimulate de novo initiation of RNA synthesis and mutation of the GDD motif to Gly-Asp-Asp (GAA) abolishes the RNA synthesis. To better understand the mechanism of viral RNA synthesis in CSFV, a three-dimensional model was built by homology modeling based on the alignment with several virus RdRps. The model contains 605 residues folded in the characteristic fingers, palm and thumb domains. The fingers domain contains an N-terminal region that plays an important role in conformational change. We propose that the experimentally observed promotion of polymerase efficiency by rGTP is probably due to the conformational changes of the polymerase caused by binding the rGTP. Mutation of the GDD to GAA interferes with the interaction between the residues at the polymerase active site and metal ions, and thus renders the polymerase inactive.

Amino Acid Sequence↗

PubMed2003

Role of birds in transmission of classical swine fever virus.

Active transmission of classical swine fever virus (CSFV) was studied in six birds (five ravens, one hooded crow) and two laying hens. Cloacal swabs, blood and organs of birds and hens as well as blood and organ samples of pigs which had been fed with faeces derived from CSFV infected birds or which had come in contact with faeces of infected hens were negative for CSFV. None of the animals seroconverted during the study. This result demonstrates that active virus transmission by these animals is unlikely. Dissemination of CSFV from wild boar to domestic pigs is discussed.

Animals↗

PubMed1999

Transmission of classical swine fever virus by artificial insemination.

Classical swine fever (CSF) virus was introduced into an artificial insemination centre during the CSF epizootic of 1997-1998 in the Netherlands. The risk of further spread of CSF virus via contaminated semen was recognised, but could not be assessed because scientific data on this issue were not available. An animal experiment was performed to determine whether CSF virus could be transmitted via artificial insemination with contaminated semen. Three boars were inoculated with a CSF virus field isolate and from Day 5 till Day 18 thereafter, ejaculates were collected and prepared for insemination. Ruttish sows were inseminated with the extended semen from Day 5 till Day 18 after inoculation of the boars. All the inoculated boars remained healthy throughout the experiment and developed CSF neutralising antibodies between 14 and 21 days after inoculation. Virus was isolated from several semen samples collected from 5 till 11 days after inoculation. Two out of six sows inseminated with CSF contaminated semen seroconverted after insemination. All the other sows remained seronegative. In the foetuses of both the seropositive sows, CSF virus was detected at approximately 35 days post insemination. These results demonstrate that adult boars infected with CSF virus can excrete virus with semen and can, subsequently, transmit the virus to sows and their foetuses via artificial insemination.

Animals↗

PubMed2003

Proportions and phenotypic expression of peripheral blood leucocytes in pigs vaccinated with an attenuated C strain and a subunit E2 vaccine against classical swine fever.

The influence of an attenuated classical swine fever virus C strain vaccine and a subunit E2 vaccine against classical swine fever on the peripheral blood leucocyte proportion and phenotypic expression in 12-week-old pigs was studied. The C strain was amplified in minipig kidney cell culture and final product contained 10(4 +/- 0.15) TCID50/ml, while the subunit vaccine contained 32 microg per dose of gp E2. Haematological findings showed that the vaccines did not cause leucopenia or lymphocytopenia and the number of neutrophils and eosinophils during the observation period was within physiological range. The results of the proportion of CD4a+, CD5a+, CD8a+, wCD21+, CD45RA+, CD45RC+ , non-T non-B, SWC3a+ and CD11b+ cells were gained by single-colour flow cytometry. At the end of the trial a significantly increase of percentage of CD4+, CD5a+, CD8+, wCD21+ cells has been found in pigs that received the subunit vaccine and the percentage of CD4+, CD5a+, CD8+, CD45RA+ and CD45RC+ cells was higher in pigs that received the attenuated vaccine. Twenty-eight days after vaccination the percentage of CD4+, CD45RA+ and CD45RC+ was significantly higher in pigs vaccinated with the C strain than in pigs vaccinated with the subunit vaccine. In contrary, the percentage of the wCD21- cells was higher in pigs that received the subunit vaccine. Statistically higher values of SWC3a+ and lower values of CD11b+ cells was observed in pigs that received the attenuated vaccine than in pigs vaccinated with the subunit vaccine. Taken altogether, our results showed that the subunit vaccine produced a better stimulation of B cells and CD11b+ monocytes/macrophages /granulocytes/NK cells, whereas the attenuated vaccine induced a higher response of Th cells, naive/memory cells and macrophages/neutrophils. Thus, both vaccines were able to influence the porcine immune system, by activating different subsets of the immune effector/accessory cells.

Animals↗

PubMed1988

Outbreaks of classical swine fever in Great Britain in 1986.

Classical swine fever was confirmed in 10 herds in Britain between April 10 and June 25, 1986 and typical acute disease was seen in nine of them. Serological evidence of exposure to classical swine fever virus was found in a further seven herds which, together with another nine, were slaughtered as dangerous contacts. Altogether 7781 pigs in 26 herds were slaughtered at a cost of 450,101 pounds for compensation alone. In order to detect subclinical disease, the majority of traced herds were blood sampled as well as inspected. A total of 119,169 pigs were inspected in 506 herds and 8302 blood samples were collected. Three primary outbreaks were identified, all attributed to the feeding of unprocessed waste food containing imported pig meat products. There was no spread of disease from two of these primary outbreaks.

Acute Disease↗

PubMed2000

Introduction to classical swine fever: virus, disease and control policy.

Classical swine fever virus is a spherical enveloped particle of about 40-60 nm in diameter with a single stranded RNA genome of about 12,300 bases with positive polarity, classified as a pestivirus within the family Flaviviridae. Natural hosts are domestic and wild pigs. The virus causes one of the most severe diseases in pigs world wide with grave economic consequences. The clinical picture of classical swine fever is variable, depending on the age of the affected animals and viral virulence. The virus is well characterised and reliable laboratory diagnostic procedures are available. In many parts of the world live attenuated vaccines are being used as a safe and efficient prophylactic tool. However, in EU Member States and several other countries vaccination is prohibited and CSF is controlled by a strict stamping out policy. In order to overcome the disadvantages of conventional vaccination inactivated marker vaccines have been developed that enable the distinction between vaccinated and infected animals. Whether these vaccines will be accepted as an additional tool in the framework of the stamping out policy is not yet decided.

Animals↗

PubMed2004

The effect of sample degradation and RNA stabilization on classical swine fever virus RT-PCR and ELISA methods.

Classical swine fever (CSF), also known as hog cholera, is a highly contagious viral infection of swine caused by a member of the genus pestivirus of the family, Flaviviridae. The need for accurate laboratory diagnosis of CSF is particularly important as it is more reliable than clinical diagnosis. CSF is endemic in many tropical countries where the climate is characterized by high ambient temperature and humidity. This study details the effect of sample quality on CSF antigen-capture ELISA (AC-ELISA) and reverse transcriptase-polymerase chain reaction (RT-PCR) methods. RT-PCR assessment of AC-ELISA-positive spleen samples stored in a conventional glycerol/saline buffer demonstrated that the RT-PCR was detrimentally affected by poor sample quality. To provide a more accurate representation of this effect, a 14 days study was performed to determine the effect of tropical ambient conditions on CSF virus-positive spleen samples stored in two transport media; glycerol/saline and a proprietary RNA preservation solution (RNAlater). A protective effect was demonstrated in both assays with RNAlater as samples were positive in both assays until day 14 post-exposure. Samples stored in glycerol/saline were negative at RT-PCR at day 3 post-exposure although AC-ELISA was still positive at day 14 post-exposure.

Animals↗