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PubMed1999

The effectiveness of routine serological surveillance: case study of the 1997 epidemic of classical swine fever in The Netherlands.

The authors describe the value of routine serological surveillance in detecting the introduction of classical swine fever virus into a disease-free population. The first investigation concerned the question of whether the epidemic of classical swine fever (CSF), which occurred from 1997 to 1998 in the Netherlands, could have been detected using the existing monitoring system for notifiable diseases. The investigation used data from the CSF epidemic of 1997/1998 and from the existing monitoring system. Secondly, the probability of detecting a case of CSF using routine serological surveillance was modelled both for multiplier herds and for finishing herds, and then for different herd size categories. The first investigation concluded that the probability of detecting the epidemic at the current level of routine serological surveillance is very low. The second investigation concluded that even employing a sampling scheme of sixty blood samples per month, the probability of detecting an outbreak of CSF within forty days of the introduction of the virus, is less than 40%.

Animals↗
PubMed2003

Comparison of antibody values in sera of pigs vaccinated with a subunit or an attenuated vaccine against classical swine fever.

Ten pigs, aged 85 days, were vaccinated with a subunit vaccine containing 32 microg of classical swine fever virus glycoprotein E2 (gp E2) (group 1), and a further 10 pigs were vaccinated with a C strain vaccine (10(4+/-0.15) TCID50/ml), produced by amplification in minipig kidney (MPK) cell culture (group 2). Nine non-vaccinated pigs served as a control group (group 3). Serum samples were collected before (day 0) and at 4, 10, 21 and 28 days after vaccination and were analysed by two commercially available enzyme immunoassays and by a neutralizing peroxidase-linked assay (NPLA). At the same times, peripheral blood was taken for determining the total leukocyte count and the body temperature was taken daily. Antibodies were not detected in serum samples collected before vaccination (day 0), and no side-effects that could be connected with vaccination were observed during the trial. Ten days after vaccination 6/10 pigs vaccinated with the subunit vaccine were seropositive. On days 21 and 28, the ratios of serologically positive to vaccinated pigs were 9/10 and 10/10, respectively. Four of the ten pigs that were vaccinated with the C strain vaccine were positive on day 21 and 9/10 on day 28. However, the results of the NPLA showed that only 4/10 pigs had an antibody titre > 1:32 at the end of the trial in both the vaccinated groups, even though the subunit vaccine initiated an earlier and higher level of neutralizing antibodies than the vaccine produced from the C strain. Challenge was performed 28 days after vaccination on four randomly selected pigs from both vaccinated groups. The pigs survived the challenge without showing any clinical signs of classical swine fever (CSF), while two nonvaccinated control pigs died on the 10th and 12th days after infection.

Animals↗
PubMed1992

A review on classical swine fever infections in pigs: epizootiology, clinical disease and pathology.

A review is given on classical swine fever (CSF) including epizootiology, clinical disease and pathology. Under the item of epizootiology the history of CSF is briefly summarized. Ways of transmission are described with special reference to CSF in wild boars. The chapter about clinical disease includes the description of different courses of CSF such as peracute, acute, subacute form and chronic disease with reference to the course of transplacental infection and fate of the progeny associated with the "carrier sow syndrome". The most typical lesions in CSF are summarized in the chapter of pathology.

Animals↗
PubMed2002

Lipopolysaccharide-induced impairment of classical swine fever virus infection in monocytic cells is sensitive to 2-aminopurine.

Lipopolysaccharide (LPS) impairs classical swine fever virus (CSFV) replication in monocytic cells, which are primary targets for CSFV and mediators of virus-induced immunomodulation. Although soluble antiviral factors including interferons (IFN) were not detected, IFN-alpha and IFN-beta mRNA were induced. The serine threonine protein kinase inhibitor 2-aminopurine, impeded this antiviral activity. These results indicate that the LPS-induced antiviral state employs signaling pathways, in which the double-stranded RNA-dependent protein kinase (PKR) is actively involved.

2-Aminopurine↗
PubMed2002

Modeling and real-time prediction of classical swine fever epidemics.

We propose a new method to analyze outbreak data of an infectious disease such as classical swine fever. The underlying model is a two-type branching process. It is used to deduce information concerning the epidemic from detected cases. In particular, the method leads to prediction of the future course of the epidemic and hence can be used as a basis for control policy decisions. We test the model with data from the large 1997-1998 classical swine fever epidemic in The Netherlands. It turns out that our results are in good agreement with the data.

Animals↗
PubMed2000

Apoptosis in the lymphatic organs of piglets inoculated with classical swine fever virus.

The involvement of apoptosis in the lymphatic organs of piglets infected with classical swine fever (CSF) virus was investigated. Piglets were inoculated with CSF virus and 3, 5, 7 and 10 days post inoculation (DPI), the thymus, spleen and lymph node were examined. In the thymus cortex, macrophages phagocytizing the nuclear remnants or apoptotic bodies increased after 3 DPI. Thymus atrophy due to the loss of the cortex increased markedly during the observation period. Compact and shrunken nuclei indicating apoptosis were observed in the spleen and lymph node. DNA fragmentation was detected in the nuclei of lymphocytes in the thymus, spleen and lymph nodes, as well as at sites of focal necrosis. Using the terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling method, ultrastructural characteristics of apoptosis, i.e. margination of condensed nuclear chromatin, were observed in the lymphatic organs from 2 DPI onward. These results suggest that apoptosis is involved in the pathology of CSF.

Animals↗
PubMed2001

Phylogenetic comparison of classical swine fever virus in China.

An N-terminal fragment of the E2 gene of classical swine fever (CSF) virus encoding major immunogenic sites was amplified by RT-PCR directly from 110 clinical specimens representing 109 epizootic sites during the last decade in China. Phylogenetic relationships between these viruses as well as 20 reference strains were determined by comparison of their nucleotide sequences. A phylogenetic tree showed that 103 of the 110 field viruses (93.6%) were clustered within group 2 and subdivided into three subgroups, while the remaining seven viruses (6.4%), along with two Chinese reference strains, Shimen and HCLV (attenuated vaccine strain), were clustered into subgroup 1.1 within group 1. However, none of the Chinese CSF viruses were members of subgroup 1.2 (represented by reference strain Brescia). This is the first report on the distribution of CSF virus genotypes in China. Results indicated that CSF viruses predominating in recent epizootics within China are genetically divergent from the reference strain Shimen and the vaccine strain HCLV.

Animals↗
PubMed2001

Development of a competitive ELISA using a truncated E2 recombinant protein as antigen for detection of antibodies to classical swine fever virus.

The sequence encoding a truncated E2 glycoprotein of the Alfort/187 strain of classical swine fever virus (CSFV) was expressed in Escherichia coli using the pET expression system and the recombinant product purified by Ni-NTA agarose affinity chromatography. The antigenicity of this recombinant protein was demonstrated by immunoblot using anti- CSFV-specific antibodies. A monoclonal antibody was produced against the truncated E2 protein and used as competitor in an ELISA for the detection of antibodies to CSFV. Specific antibodies were demonstrated by competitive ELISA (C-ELISA) as early as 21 days post-infection (dpi) in experimentally infected pigs. Seroconversion was demonstrated by C-ELISA and neutralising peroxidase-linked assay (NPLA) in all infected animals by 4 weeks. No cross-reaction with antibodies to bovine viral diarrhoea virus (BVDV) was seen in the C-ELISA using sera from experimentally infected pigs. The C-ELISA is not intended as a substitute for the NPLA. However, it is expected it will be useful for monitoring and prevalence studies. It will also assist in testing a large number of samples in the event of an outbreak.

Animals↗
PubMed1999

Differentiation between vaccine strain and field isolates of classical swine fever virus using polymerase chain reaction and restriction test.

The CS vaccine strain of Classical Swine Fever Virus is a derivative from the LK parental strain that has been used in Russia for more than 30 years. A 10697 nucleotide fragment of the CS strain's genome has been sequenced. Sixteen unique restriction markers have been found in the CS genome comparing to the following strains: Alfort187, Alfort Tubingen, Brescia, CAP, Glentorf, ALD, GPE-, Chinese, C-strain, Riems, P97. Fourteen of these sites (AflII, AvaI, CfoI, Eco47II, HaeII, KpnI, MunI, NspI, PstI, ScaI, SmaI, SpeI, StyI, VspI) are only present in the CS strain genome. The 2 sites (BgII, NdeI) are present in all other genomes except for the genome of vaccine strain. A PCR/restriction test has been developed based on these findings in order to distinguish the vaccine strain from field isolates. Two pairs of nested primers and a criteria of analysis have been designed for each restriction marker site. The tests have been conducted first on the reference strains resulting in predicted restriction patterns. Finally, the tests have been applied to a number of field isolates obtained at different locations in Russia in different years. These results give further evidence that PCR/restriction tests can identify the LK and CS vaccines helping to avoid confusion with field strains.

Animals↗
PubMed1997

An improved ELISA for the detection of serum antibodies directed against classical swine fever virus.

The complex-trapping-blocking (CTB) ELISA for detection of antibodies against classical swine fever virus (CSFV) using two monoclonal antibodies (mAbs) directed against envelope glycoprotein E2, has been improved using recombinant CSFV E2-antigen. The newly developed Ceditest ELISA for CSFV-Ab is a modification of the CTB-ELISA as described by Wensvoort et al. (1988) and Bloemraad et al. (1993). The old CTB-ELISA format comprised of a two-step, single-dilution test which had to be performed by hand at 37 degrees C. The Ceditest ELISA is a one-step, single-dilution test which can be performed by hand, or automated at room temperature. A set of 505 pig sera was tested for CSFV antibodies in the CTB-ELISA, in the Ceditest ELISA and in the neutralizing peroxidase-linked assay (NPLA). Concordance between the two ELISAs was 96.7%. Sensitivity and specificity of the Ceditest ELISA were 99% and 99% compared to the CTB-ELISA. The Ceditest ELISA discriminates between antibodies directed against bovine viral diarrhea virus (BVDV) and CSFV. The discrimination between antibodies directed against some Border disease virus (BDV) strains and CSFV is as inconsistent as with the NPLA. We conclude that the Ceditest ELISA for CSFV-Ab is a promising tool for the fast and simple screening of large numbers of pig sera for detection of antibodies directed against CSFV.

Animals↗
PubMed2003

Assessment of the use of gross lesions at post-mortem to detect outbreaks of classical swine fever.

The performance of pathological findings as a diagnostic tool for the detection of classical swine fever (CSF) outbreaks during the 1997/1998 CSF-epidemic in The Netherlands was evaluated by constructing and analysing receiver operating characteristic (ROC) curves. This was done at the individual pig level and at the submission level (a group of pigs from the same herd submitted together for post-mortem investigation). At post-mortem examination, the tonsils, spleen, ileo-caecal valve and renal pelvis were sampled, sent to the reference laboratory, and tested by means of a CSF-specific fluorescent antibody test in combination with a confirmatory test. This resulted in an infection status at the individual pig level. The infection status and pathological findings of 1072 individual pigs from a total of 230 infected herds were included in this analysis. We also included submissions of pigs from herds that were sent to post-mortem examination because of a clinically CSF-suspect situation but afterwards were concluded to be from non-infected herds. Infection status and pathological findings of 1224 individual pigs from a total of 241 non-infected herds were included in the analysis. Pneumonia, pleuritis, chronic bronchitis, pulmonary oedema, chronic gastric ulceration, dry faecal contents in the colon, conjunctivitis, haemorrhages in the renal pelvis, renal haemorrhages, splenic enlargement, haemorrhages in the urinary bladder, haemorrhagic and enlarged lymph nodes were the most frequently recorded pathological findings during a post-mortem examination of pigs submitted in a CSF-suspect clinical situation. However, some of these pathological findings (e.g. pneumonia, pleuritis) were almost evenly distributed in infected and in non-infected pigs, resulting in a high sensitivity combined with a low specificity. The area under the ROC curve of pathological findings at the individual pig level and at the submission level was 0.720 and 0.782, respectively, which was significantly (P<0.0001) larger than the area under the random ROC curve. It was concluded that, although gross pathology is a legitimate test, its quantitative contribution to the detection of CSF is limited.

Animals↗
PubMed2005

Origin and evolution of viruses causing classical swine fever in Cuba.

We have analyzed the origin and evolution of viruses from the classical swine fever (CSF) epidemic that affects Cuba since 2001 by nucleotide sequencing of regions within the E2 glycoprotein and the NS5B (polymerase) genes. The sequence of 190 nucleotides from E2 gene was determined for 10 CSF viruses isolated at different locations of the island, and used for phylogenetic analyses, including sequences from viruses of the 1993--1997 epizootic, previously determined, as well as those from representatives of the different CSFV genotypes. The phylogenetic tree obtained indicates that viruses circulating at present belong to the subgroup 1.2 and are closely related to those isolated during the 1993--1997 epizootic, including the strain Margarita used for vaccine potency tests in Cuba. However, the pattern of evolution revealed by these analyses was different than that observed previously, in which western isolates were almost identical to Margarita strain, while eastern isolates showed a higher level of genetic diversification. In this case, all the viruses analyzed grouped in an independent, define cluster that is closely related, albeit distinguishable, from that of Margarita-related viruses that previously circulated in the western part of Cuba. In addition, the 2001--2003 viruses showed a branched pattern with a level of sequence diversification similar to that observed in the eastern 1993--1997 viruses. Interestingly, a significant fraction (about 54%) of the mutations found in the E2 sequence led to amino acid replacements. This high rate of non-synonymous mutations was not found in the previous Cuban epizootic and has not been reported for other CSF outbreaks. In spite of these amino acid replacements, no antigenic changes were observed in the reactivity of different isolates with CSFV-specific MAbs and polyclonal sera. The phylogenetic tree derived from 409 nucleotides of NS5B gene of seven isolates and Margarita strain, was consistent with that obtained from E2 sequences. In this region, encoding a non-structural protein, a low level of fixation of non-synonymous mutations was observed. The results obtained suggests that epidemiological factors affecting CSFV spread during the current epizootic in Cuba can favour the fixation of non-synonymous mutation in the E2 gene, which could be associated with a lower severity in the clinical signs developed by most of the affected animals.

Amino Acid Sequence↗
PubMed2000

[Oral immunization against classical swine fever (CSF) in the Federal State of Brandenburg from 1995 to 1997].

The first findings of classical swine fever in wild boar occurred in the rural district Ostprignitz-Ruppin of Brandenburg in March 1995. Up to this time the area was considered to be free of swine fever in domestic and wild boars. The swine fever cases were marked particularly by the occurence of dead games and clinically decreased individuals as well as by the involvement of young wild boars. (< 10 kg carcass weight). In Brandenburg, the eradication programme of classical swine fever was based on oral immunization with an attenuated type C vaccine (so called Dessauer wild boar lure) carried out within the frame of a field trial since 1995. The success of oral immunization was examined by the development of swine fever positive-findings within the swine fever endangered district and by the seroconversion in the entire vaccination area. Due to the oral immunization the classical swine fever prevalence decreased from 4.65% in March 1995 to 0.58% in December 1997. After the third immunization the seroconversion reached on average rates between 30% and 35%. The seroconversion increased with carcass weight and with number of subsequent immunization. During the investigation period the rate of seroconversion in wild boars up to 17 kg carcass weight was 13.2%, in wild boars from 18 to 28 kg carcass weight 23.1%, sub-adult wild boars from 29 to 56 kg carcass weight 33.2%, and the adult wild boars above 57 kg carcass weight 41.5%.

Administration, Oral↗
PubMed1998

Lymphocyte apoptosis during classical swine fever: implication of activation-induced cell death.

Infection of pigs with classical swine fever virus (CSFV), a member of the Flaviviridae family, causes a severe leukopenia, particularly notable with the lymphocytes. The goal of this study was to analyze mechanisms behind this CSFV-induced lymphopenia. To this end, the kinetics of leukocyte depletion, the appearance of apoptotic cells, and virus infection of leukocytes after infection of pigs with the virulent CSFV strain Brescia were analyzed. Depletion of B and T lymphocytes was noted as early as 1 day postinfection (p.i.). Circulating viable lymphocytes with reduced mitochondrial transmembrane potential--a particular early marker for apoptosis--were also detectable as early as 1 day p.i. When isolated peripheral blood mononuclear cells were cultured for 6 h, significantly more sub-G1 cells with reduced DNA content were detected among the lymphocytes from CSFV-infected animals, again as early as 1 to 3 days p.i. The first time virus was first found in the plasma, as well as infection of leukocytes, was 3 days p.i. However, throughout the observation time of 1 week, <3% of the circulating leukocytes and no lymphocytes contained virus or viral antigen. Further analysis of the T lymphocytes from infected animals demonstrated an increase in CD49d, major histocompatibility complex class II, and Fas expression. An increased susceptibility to apoptosis in vitro was also observed, particularly after addition of concanavalin A as well as apoptosis-inducing anti-Fas antibody to the cultures. Taken together, these results imply that activation-induced programmed cell death was the mechanism behind lymphopenia during classical swine fever.

Animals↗
PubMed2005

Comparison of six RNA extraction methods for the detection of classical swine fever virus by real-time and conventional reverse transcription-PCR.

Six RNA extraction methods, i.e., RNAqueous kit, Micro-to-midi total RNA purification system, NucleoSpin RNA II, GenElute mammalian total RNA kit, RNeasy mini kit, and TRIzol LS reagent, were evaluated on blood and 7 tissues from pig infected with classical swine fever virus (CSFV). Each of the 6 extraction methods yielded sufficient RNA for positive results in a real-time reverse transcription-PCR (RT-PCR) for CSFV, and all RNA, except the one extracted from blood by TRIzol LS reagent, yielded positive results in both a conventional RT-PCR for CSFV and a conventional RT-PCR for an endogenous gene encoding beta-actin. The RNA extracted from blood by TRIzol LS reagent became positive in both conventional RT-PCR assays when it was diluted to 1:2, 1:4, or up to 1:64 in nuclease-free water. It is concluded that all 6 methods are more or less useful for the detection of CSFV by real-time and conventional RT-PCR in swine blood and tissues. However, some of the 6 reagents offer certain advantages not common to all 6 extraction procedures. For example, RNA extracted by the TRIzol LS reagent constantly had the highest yield; that by the RNAqueous kit had the highest A260/A280 ratio for almost all samples; and that by the NucleoSpin RNA II and the GenElute mammalian total RNA kit was most likely to be free of contaminations with genomic DNA.

Animals↗