Classical swine fever: genetic detection and analysis of differences between virus isolates

Lowings, J. P.; Paton, David; Sands, J. J.; Mia, Gian Mario De; Rutili, D.

doi:10.1099/0022-1317-75-12-3461

Cited by 62 publications

(20 citation statements)

References 29 publications

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“…In the last decade, various PCR assays have been developed for the detection of ASFV [1,18,35,37] and of CSFV [16,17,19,23,30,38]. These assays provide novel diagnostic tools, but a disadvantage is the restricted ability to detect only a single virus, ASFV or CSFV.…”

Section: Discussionmentioning

confidence: 99%

A highly sensitive and specific gel-based multiplex RT-PCR assay for the simultaneous and differential diagnosis of African swine fever and Classical swine fever in clinical samples

Agüero

Fernández²,

Romero

et al. 2004

Vet. Res.

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-The development and standardisation of a novel, highly sensitive and specific one-step hot start multiplex RT-PCR assay is presented for the simultaneous and differential diagnosis of African swine fever (ASF) and Classical swine fever (CSF). The method uses two primer sets, each one specific for the corresponding virus, amplifying DNA fragments different in length, allowing a gel-based differential detection of the PCR products. Universal detection of ASF and CSF virus strains was achieved through selection of primers in conserved viral genome regions. The detection range was confirmed by analysis of a large collection of isolates of the two viruses. The high specificity of the assay was proven by testing related viruses, uninfected cell line cultures and healthy pig tissues. Additional confirmatory tests of the ASF and CSF virus amplicon specificity, based on restriction endonuclease analysis with BsmA I or Ban II, respectively, are also described. The analysis of whole blood and serum samples from experimentally infected animals proved the usefulness of the method for an early diagnosis of both diseases, even before the appearance of the first clinical signs. A study of 150 positive field samples from several ASF and CSF outbreaks showed the suitability of this method for a rapid (less than five hours), sensitive and specific differential diagnosis in clinical samples. In addition, a highly sensitive and specific uniplex RT-PCR for CSFV was also developed and standardised as a powerful tool for fast and early diagnosis of the disease.

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Section: Discussionmentioning

confidence: 99%

A highly sensitive and specific gel-based multiplex RT-PCR assay for the simultaneous and differential diagnosis of African swine fever and Classical swine fever in clinical samples

Agüero

Fernández²,

Romero

et al. 2004

Vet. Res.

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“…In addition, the sequence of PCR and sequencing. The molecular cloning and sequencing of some of the Italian isolates have been described (Lowings et al, 1994). The same PCR primers were used to amplify the remaining virus gene segments which were sequenced directly without molecular cloning.…”

Section: Methodsmentioning

confidence: 99%

“…The PHYLIP (Felsenstein, 1989) and GCG (Devereux et al, 1984) software used for the analysis of molecular sequences and MAb data were as described (Lowings et al, 1994). In addition, the PHYLIP program RestML was used to analyse RE derived data (Harding et al, 1994).…”

Section: Methodsmentioning

confidence: 99%

“…The proposed rate of E2 evolution was calculated from three sources: (a) the rate of divergence of the central Italian viruses clW and c4D (Lowings et al, 1994) which were strongly suspected of being from a closed virus population: (b) by averaging the rates obtained from all suspected terminal branch progenitor/progeny relationships produced by E2 phylogenetic analysis; and (c) by assuming that all of the current isolates originated from viruses similar to our oldest isolates (1940s).…”

Section: Estimation Of Virus Divergence Timesmentioning

confidence: 99%

“…This region of the protein is encoded by one of the most variable parts of the pestivirus genome. Typing with monoclonal antibodies (MAbs) (Edwards & Sands, 1990) and some limited molecular data (Harding et al, 1994;Hofmann et al, 1994;Lowings et al, 1994) have suggested that CSFV, although probably the least variable member of the pestivirus genus, can be subdivided into distinct groups, the most obvious division being between old and recent isolates. The identification of virus subtypes should improve our understanding of CSFV evolution and epidemiology and might provide markers for biological differences, such as in virulence.…”

Section: Introductionmentioning

confidence: 99%

See 2 more Smart Citations

Classical swine fever virus diversity and evolution

Lowings

Ibata

Needham

et al. 1996

Journal of General Virology

181

114

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By analysing the nucleotide sequence data generated from both the E2 (gp55) and the NS5B genes of classical swine fever virus (CSFV), in addition to previously published data from the 5'NCR, we were able to divide 115 CSFV isolates into two major groups, five subgroups and two disparate isolates. Further discrimination was possible by analysis of sequence data from the E2 region. The three sequencing based methods were compared to monoclonal antibody (MAb) typing and to limited restriction enzyme (RE) mapping. Although both MAb and RE methods confirmed the previous classification the resolution was inferior. We estimated an approximate evolution rate for CSFV from an analysis of the virus variation observed in a single geographical area over a 6 year period. Applying this proposed rate to each of our deduced CSFV subgroups enabled us to calculate the approximate dates of divergence for each subgroup.

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