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PubMed2003

Molecular epidemiology of classical swine fever in the Russian Federation.

The ability to discriminate between various classical swine fever virus (CSFV) strains and isolates is a prerequisite for following the spread of the virus after an outbreak. To determine the relatedness between Russian CSFV isolates from different geographical regions, three fragments of the viral genome (5' NTR, the variable region of the E2 gene and a fragment of the NS5B gene) were sequenced and used for genetic typing. Thirty-one field isolates were obtained from CSF outbreaks which occurred between 1994 and 1999. In addition, three attenuated strains were included in the study, namely the LK and CS vaccine strains, and the moderately virulent 238H isolate. The vaccine strains have been used in Russia for more than 30 years. Our results showed that all field isolates are in subgroup 1.1 together with Alfort 187 and with the highly virulent strain Shimen. In contrast, the CS and LK vaccine strains belong to subgroup 1.2. While there is no evidence for the reversion of the two vaccine strains to wild type, it is feasible that the highly virulent Shimen strain, which has been used as a challenge strain for many years, contributed to field strain generation. The Russian field isolates from the 1990s can be distinguished from the CSF virus isolates which occurred in the EU Member States in the same decade, as here all outbreaks were caused by CSF viruses belonging to subgroup 2.

Animals↗
PubMed2002

Pig chromosome aberrations after vaccination against classical swine fever in field trials.

Chromosomal aberrations were observed after vaccination against classical swine fever (CSF) in a previous study done on experimental pigs. To determine if the same effect occurs in farm animals, field trials were done with immunized pigs. The cytogenetic analysis was made from lymphocyte cultures of pigs sampled in three farms of Río Cuarto region on six different periods: one, pre-vaccination (control) and in five post-vaccination (days 3, 7, 10, 15 and 22). Vaccine inoculation induced significant increase of cell frequency with chromosomal aberrations from days 3 to 10, when maximal frequencies of 6.78, 10.36 and 7.21% were observed in farms A, B and C, respectively. Afterwards, a decrease was recorded, reaching values not significantly different from controls. Mean frequencies of cells with chromosomal aberrations were 2.8+/-0.3, 5.9+/-0.4, 3.1+/-0.3%, for A, B and C farms, respectively. Cytogenetic effect was clastogenic, being chromosome breaks the first type of alteration to increase from the day 3 post-vaccination onwards, followed by chromatid exchanges and cells with multiple abnormalities. Chromosome pulverization was the last type to show increment, reaching a top value on day 10, after that it started to diminish gradually. There was no difference in the frequency of polyploid cells among sampling dates meaning that this type of alteration would not be induced by vaccination. Results of this field study confirm the mutagenic capacity of live virus vaccines against CSF and indicate that the evolution of chromosome alterations on the five evaluated post-vaccination periods was similar at different environments.

Animals↗
PubMed1994

Epidemiology of classical swine fever in Sardinia: a serological survey of wild boar and comparison with African swine fever.

A serological survey was carried out to establish the distribution of classical swine fever among wild boar in Sardinia, where that disease and African swine fever have been endemic in free-ranging domestic pigs and wild boar living in the mountainous areas of the province of Nuoro for several years. Blood samples were collected from 4752 wild boar shot during the period December 1988 to January 1992. An overall prevalence of 11 per cent was observed and the almost constant rate of about 9.8 per cent detected in the past three years indicates that the infection is well established. Wild boar seropositive to classical swine fever were found not only in the areas of the province of Nuoro where they share their habitat with free-ranging domestic pigs but also in other areas of the island where contacts between wild and domestic pigs are unlikely to occur. Therefore, transmission from wild boar to wild boar seems to play an important role in the spread and persistence of classical swine fever virus. In contrast, African swine fever virus is probably unable to persist in the wild boar population in the absence of the risk factor represented by their cohabitation with domestic free-ranging pigs infected with African swine fever.

African Swine Fever↗
PubMed1998

Molecular cloning and nucleotide sequence of 3'-terminal region of classical swine fever virus LPC vaccine strain.

A cDNA of the 3'-terminus of classical swine fever virus (LPC vaccine strain) was cloned and sequenced. The 3431 nucleotides and deduced amino acid sequences were compared with those of other pestiviruses, and the similarity of nucleotide sequences and deduced amino acid sequences were found to be 84-95% and 95-98%, respectively. Similar to other isolates of classical swine fever virus, the sequenced region included the non-structural gene p58 (NS5A) and part of p76 (NS5B) gene. The p76 gene of LPC vaccine strain also contained a highly conserved motif G-D-D (Gly-Asp-Asp) that is present in the RNA replicase of positive-stranded RNA viruses. With the sequence data currently available, we carried out a phylogenetic analysis and obtained a genealogical relationship among members of the classical swine fever virus.

Amino Acid Sequence↗
PubMed2003

Virulence of recent and former classical swine fever virus isolates evaluated by their clinical and pathological signs.

The clinical diagnosis of classical swine fever (CSF) still caused problems to the veterinarians during the last decade. The primary CSF outbreak was often detected too late and, meanwhile, the virus had spread. Consequently, the recent classical swine fever virus isolates (CSFV) were suspected to be of low virulence. The purpose of the study was to quantify the virulence of four recent CSFV by evaluating the clinical and pathological signs caused by different CSFV. Pigs of the same breed and age group were inoculated intranasally with CSFV from recent epidemics in European Union (EU) member states. The CSFV used are registered in the data base of the EU Reference Laboratory for CSF and belong to different genotypes: 2.1, 2.2 and 2.3 respectively. Clinical signs of CSF were evaluated by using a score system suggested previously (Mittelholzer et al., 2000: Vet. Microbiol. 74, 293). For the evaluation of pathological lesions, a new pathological score was introduced. The four CSFV tested here were classified as moderately virulent in general, although one CSFV may cause different clinical courses, ranging from highly virulent to avirulent. This indicates the importance of additional factors in the host animal for virulence. Differences in the clinical and pathological signs between these four recent CSFV were rather minor, emphasizing that the genetic typing of CSFV is absolutely essential. Differences towards former CSFV (e.g. reference virus strain Alfort 187) were more pronounced, especially regarding the onset and duration of the disease, the occurrence of skin haemorrhages and pathological lesions of kidney, subcutis and serosae. It is concluded that clinical diagnosis of CSF is rather difficult in pigs up to 14 days post-CSFV infection using these four CSFV, emphasizing the need for careful differential diagnosis and the laboratory investigation for CSF at an early stage.

Animals↗
PubMed2000

Chimeric classical swine fever viruses containing envelope protein E(RNS) or E2 of bovine viral diarrhoea virus protect pigs against challenge with CSFV and induce a distinguishable antibody response.

Three chimeric classical swine fever virus (CSFV)/bovine viral diarrhoea virus (BVDV) full-length DNA copies were constructed, based on the infectious DNA copy of the CSFV vaccine strain C. The antigenic region of E2 and/or the complete E(RNS) gene were replaced by the analogous sequence of BVDV II strain 5250. Viable chimeric virus Flc11, in which E(RNS) was replaced, was directly recovered from supernatant of SK6.T7 cells transfected with full-length DNA. Viable chimeric virus Flc9, in which E2 was replaced, resulted in recovery of virus only when SK6.T7 transfected cells were maintained for several passages. However, no virus could be recovered after replacement of both E(RNS) and E2, even after 10 cell passages. Both Flc9 and Flc11 grow in swine kidney cells (SK6), stably maintain their heterologous BVDV sequences and, as assessed by monoclonal antibody typing and radio-immunoprecipitation assays, express their heterologous proteins. Flc9 showed a slower growth rate on SK6 cells than Flc11 and wild-type Flc2 virus. Replacement of E(RNS) or E2 of C-strain-based chimeric viruses did not alter cell tropism compared to wild-type C-strain virus for SK6 and FBE cells. Both Flc9 and Flc11 induced E2 or E(RNS) antibodies, which could be discriminated from those induced after wild-type virus infection, even after repeated vaccination. Furthermore, pigs were completely protected against a lethal CSFV challenge. These results indicate the feasibility of introduction of marker antigens in a live-attenuated marker C-strain vaccine for CSFV.

Amino Acid Sequence↗
PubMed1999

[Classical swine fever in wild boars in Switzerland].

In May 1998, wild boars with classical swine fever (CSF) symptoms were detected in the southern part (Canton Ticino) of Switzerland. CSF virus was isolated from the submitted samples and RT-PCR followed by direct nucleotide sequencing of the 5' non-translated region showed that this virus was identical to the isolate previously recognized in wild boars from the area of Varese (Italy). In most animals, antibodies to CSF virus were detected as well. An immediate measurement was taken by limiting the movement of pigs and identifying both risk and surveillance zones. In order not to disturb potentially infected wild boars within their habitat a complete hunting prohibition for 2 months was enforced. The different possibilities of the control of CSF outbreaks in wild boars are discussed.

Animals↗
PubMed2003

Localization of classical swine fever virus from chronically infected pigs by in situ hybridization and immunohistochemistry.

Classical swine fever (CSF) virus (CSFV) nucleic acid and antigen were detected in 15 pigs with naturally occurring chronic CSF by in situ hybridization and immunohistochemistry. The most consistent and prominent microscopic lesions were perivascular mononuclear cell infiltration and gliosis in the central nervous system of pigs with chronic CSF. Positive cells typically exhibited a dark brown (in situ hybridization) or red (immunohistochemistry) reaction product in the cytoplasm without background staining. A positive signal for both in situ hybridization and immunohistochemistry was detected in mononuclear cells and lymphocytes of lymphoid tissues. Viral nucleic acid was detected in some tissue sections in the absence of viral antigen. The in situ hybridization technique developed in this study was useful for the detection of CSFV RNA in tissues taken from chronically infected pigs and may be a valuable technique for studying the pathogenesis of chronic CSFV infection.

Animals↗
PubMed2005

Antibody responses of pigs to defined Erns fragments after infection with classical swine fever virus.

Antibody responses of pigs to defined Erns fragments, after classical swine fever virus (CSFV) infection, were studied by using an enzyme-linked immunosorbent assay (ELISA). Selection of various E(rns) fragments was based on an immunodominant Erns region encompassing three overlapping antigenic regions, amino acids 65 to 145 (Erns(aa)65-145) (AR1), 84 to 160 (Erns(aa)84-160) (AR2), and 109 to 220 (Erns(aa)109-220) (AR3), identified earlier by our group (M. Lin, E. Trottier, J. Pasick, and M. Sabara, J. Biochem., in press). Defined Erns fragments, including AR1, AR2, AR3, Erns(aa)65-160 (AR12), Erns(aa)84-220 (AR23), Erns(aa)65-220 (AR123), Erns(aa)109-145 (the consensus region defined by the three overlapping regions), and Erns(aa)109-160 (a fragment 15 amino acids larger than the consensus region), were expressed in Escherichia coli, purified by nickel chelate affinity chromatography, and used to measure antibody responses in 20 sera serially collected from pigs experimentally infected with CSFV. Based on the optimum cutoffs determined by receiver operating characteristic analysis after testing 238 negative field sera from Canadian sources, all the Erns fragments were capable of distinguishing positive from negative antibody responses with sensitivities ranging between 75 and 90% and specificities ranging between 83.2 and 100%. Detection of antibody responses to refolded Erns(aa)109-145 and Erns(aa)109-160 by ELISA (this study) but not by Western blots (Lin et al., in press) indicated that the epitopes within the consensus region are conformational. When cutoff values were raised to give a specificity of 100%, four Erns fragments (AR2, AR23, Erns(aa)109-145, and Erns(aa)109-160) offered much higher sensitivities (75 to 90%) than those obtained with other fragments (20 to 65%). Erns(aa)109-145 and Erns(aa)109-160 were capable of detecting antibody responses in infected pigs as early as 7 days postinfection. Demonstration of antibody responses to either one of the four fragments can thus be an alternative to use of the full-length protein in ELISA for serological diagnosis of CSFV infection. An advantage of such a test would be its utilization for serological survey in a classical swine fever-free country (e.g., Canada) in biocontainment level 2 laboratories.

Animals↗
PubMed2002

An experimental infection to investigate the indirect transmission of classical swine fever virus by excretions of infected pigs.

In this experiment transmission of classical swine fever (CSF) virus via excretions of infected pigs was investigated under experimental conditions. Five pairs of pigs were experimentally infected with CSF virus. Eight days after experimental infection, when all pigs were viraemic for at least 3 days, the pens were depopulated and 20 h later, restocked with five pairs of susceptible pigs which stayed in these pens for 35 days. During the first 3 weeks of the experiment, the pens were neither cleaned nor disinfected. During the observation period, none of the susceptible pigs became infected. This result indicates that CSF virus spread via excretions is of minor importance in the early stages of infection. For extrapolation of these findings to the field situation and to increase the validity of the conclusions further research is needed to evaluate the effect of factors like virus strain, interval, ..., that may influence the outcome of the experiment.

Animal Husbandry↗
PubMedDate not supplied

Evaluation of the oral immunisation of wild boar against classical swine fever in Baden-Württemberg.

The oral immunisation of wild boar against classical swine fever (CSF) in Baden-Württemberg is described and evaluated. The bait vaccine based on the CSF virus (CSFV) strain "C" proved to be safe in wild boar of all age classes. The modified immunisation procedure consisting of three double vaccinations per year was very effective. CSFV was not detected beyond the second immunisation campaign. The average rate of seropositive wild boar diagnosed over all immunisation periods was 49.2%. The seroprevalence rate increased significantly during the first year of immunisation and reached its maximum after the third vaccination period with 72% antibody positive animals. The higher percentage of seropositive young boars in this field trial compared to the seroprevalence rates in this age class in other field trials in Germany may be attributed to the new vaccination scheme. Factors that may be responsible for the decreased herd immunity after the fourth or sixth immunisation period are discussed.

Administration, Oral↗
PubMed2004

Efficacy of E2-sub-unit marker and C-strain vaccines in reducing horizontal transmission of classical swine fever virus in weaner pigs.

At present, two types of vaccines against classical swine fever (CSF) virus are commercially available: E2 sub-unit marker vaccines and the conventional attenuated live C-strain vaccines. To evaluate the reduction of the horizontal virus transmission, three comparable experiments were carried out in which groups of weaner pigs (vaccinated with a marker vaccine or a C-strain vaccine) were challenged with CSF virus at 0, 7, and 14 days post-vaccination (dpv). Virus transmission was prevented totally when the challenge occurred at 14 dpv with an E2-marker vaccine (0/12 contact pigs positive in virus isolation (VI); R = 0 (0; 1.5)). At 7 dpv, transmission was reduced slightly (5/12 contact pigs positive in VI; R = 1.0 (0.3; 3.0)), whereas at 0dpv, vaccination had no effect on transmission (10/12 contact pigs positive in VI; R = 2.9 (1.5; 10.8)). In the C-strain-vaccinated pigs, no virus transmission was detected even when the challenge was performed at the same day as the vaccination (0/12 contact pigs positive in VI; R = 0 (0; 1.5)).

Animals↗
PubMed1999

An experimental marker vaccine and accompanying serological diagnostic test both based on envelope glycoprotein E2 of classical swine fever virus (CSFV).

Envelope glycoprotein E2 is the most immunogenic protein of classical swine fever virus (CSFV). In a proposed model of the antigenic structure of E2, the N-terminal half of E2 forms two independent structural antigenic units, A and BC. E2 without transmembrane region (E2-TMR) is expressed and secreted into the medium of insect cells by use of the baculovirus expression system. The immune response induced by E2 protects pigs against CSFV. Recently, we showed that the protective immune response to a homologous CSFV challenge can be induced by a single unit, A or BC, of E2. An indirect blocking ELISA, or complex trapping blocking assay (CTB) based on both units is routinely used worldwide for serological diagnosis of CSFV infections. Here we show that E2-TMR is secreted into the medium as a homodimer. This E2 homodimer was used to develop a CTB detecting antibodies directed against one immunogenic unit of E2. Thus, the protective immune response induced by E2 containing one unit was not detected with a modified CTB based on the other unit, whereas immune responses induced by a variety of low virulent CSFV strains were detected with such a modified CTB. These results indicate that a deletion E2 protein in combination with a modified CTB are feasible as CSF marker vaccine and accompanying differentiating diagnostic test.

Animals↗
PubMed2003

Rapid detection of classical swine fever virus by a portable real-time reverse transcriptase PCR assay.

A fluorogenic-probe hydrolysis (TaqMan)-reverse transcriptase PCR assay for classical swine fever virus (CSFV) was developed and evaluated in experimentally infected swine. The assay detected CSFV, representing different phylogenetic groupings, but did not amplify viral RNA from related pestiviruses. The assay met or exceeded the sensitivity (1 to 100 50% tissue culture infective doses per ml) of viral cultures of samples from experimentally infected animals. Viral RNA was detected in nasal and tonsil scraping samples 2 to 4 days prior to the onset of clinical disease. The assay can be performed in 2 h or less, thus providing a rapid method for the diagnosis of classical swine fever.

Animals↗
PubMed2003

Risk analysis of the spread of classical swine fever virus through "neighbourhood infections" for different regions in Belgium.

Risk factors associated with the occurrence of "neighbourhood infections" [Epidemiology of classical swine fever. In: Truszczynski, M. (Ed.), Proceedings of the Workshop on Diagnostic Procedures and Measures to Control Classical Swine Fever in Domestic Pigs and the European Wild Boar. Pulaway, Poland, pp. 119-130] during classical swine fever (CSF) outbreaks were examined based on information collected during a CSF-epidemic, which occurred in the East Flanders Province of Belgium in 1994. The only risk factor that was associated with the occurrence of "neighbourhood infections" was a kernel estimation of the intensity of neighbouring herds (P=0.055) [Interactive spatial data analysis. Pearson Education Limited, Harlow, Essex], i.e. the higher the kernel estimation, the higher the risk for the occurrence of neighbourhood infections. In a second part of the study, the likelihood for the occurrence of neighbourhood infections within an area with a 1 km radius was predicted for every Belgian pig herd, assuming that the herd was infected with CSF-virus. For the prediction of these likelihoods, the model resulting from the risk assessment was used. Finally, the predicted likelihoods were transformed into a raster map after applying a smoothing technique. As a result, different areas in Belgium of higher or lower risk for CSF-virus spread through "neighbourhood infections" could be identified on the map. The areas in Belgium where CSF-outbreaks including "neighbourhood infections" occurred in the past decades were all predicted by the model to be of high risk.

Animals↗
PubMed2001

Highly sensitive one-tube RT-PCR and microplate hybridisation assay for the detection and for the discrimination of classical swine fever virus from other pestiviruses.

Rapid, sensitive and specific laboratory diagnostic methods are necessary to confirm outbreaks of classical swine fever. The detection of classical swine fever virus (CSFV) and its discrimination from other pestiviruses can be achieved by virus isolation on cell culture, antigen detection, or molecular methods. To reduce the time and the number of steps in the diagnostic procedure a sensitive and rapid detection method based on specific amplification of the pestiviral RNA by one-step reverse transcription-polymerase chain reaction (RT-PCR) followed by detection and differentiation of the amplification products by pestivirus-, bovine viral diarrhoea virus- (BVDV-) and CSFV-specific capture probe hybridisation and colorimetric assay in microwell plates (enzyme liked immunosorbent assay (ELISA)) was developed. Two different methods using two gene regions for pestivirus RT-PCR amplification were carried out. One pair of primers was selected from the 5'-UTR region and the second one from the gene region coding for N(pro), C and E0 proteins. The designed oligonucleotide primers were used for several pestivirus reference strains as well as for some field isolates detection in cell culture supernatants and in clinical specimens. The specificity and sensitivity of both methods were compared using EZ rTth RNA PCR kit and ACCESS RT-PCR system for combined RT-PCR assay. The use of one-step RT-PCR eliminates the additional manipulations that are generally required for a two reaction system and limits the risk of carry-over contamination. Labelling of PCR products with digoxigenin (DIG) during the amplification reaction enables colorimetric assessment of hybridisation reactions. For solution hybridisation pestivirus-, BVDV- and CSFV-specific biotin-labelled capture probes were used. By serial dilutions of DIG-labelled PCR products the RT-PCR-ELISA was found to be 100-times more sensitive than the conventional agarose gel electrophoresis. Higher sensitivity of RT-PCR-ELISA detection using specific biotin-labelled probes offers the opportunity to eliminate strain specific nested PCR and to overcome the problems with contamination and false positive results.

Animals↗
PubMed2002

Classical swine fever (hog cholera) in wild boar in Europe.

Classical swine fever (CSF) is of increasing concern in Europe where wild boar appear to play an important epidemiological role. In most parts of the continent, demographic trends are on the increase, due to improvement in game management. As a result of higher densities, populations become more susceptible to various infectious diseases, among which CSF is cause for particular concern. Wild boar do not appear to be a classic reservoir in most cases, but nevertheless may perpetuate foci of infection over the long term, constituting a real threat for the pig farming industry. Since the infection does not appear to spread easily in natural populations of free-ranging wild boars, control of the disease may be feasible. However, most of the appropriate measures, such as banning hunting, are not considered acceptable. Consequently, the expertise of wildlife disease specialists is required to help solve the problem when it occurs.

Animals↗