Detection Of Classical Swine Fever Research Articles

Oral Fluids for the Early Detection of Classical Swine Fever in Commercial Level Pig Pens.

The early detection of classical swine fever (CSF) remains a key challenge, especially when outbreaks are caused by moderate and low-virulent CSF virus (CSFV) strains. Oral fluid is a reliable and cost-effective sample type that is regularly surveilled for endemic diseases in commercial pig herds in North America. Here, we explored the possibility of utilizing oral fluids for the early detection of CSFV incursions in commercial-size pig pens using two independent experiments. In the first experiment, a seeder pig infected with the moderately-virulent CSFV Pinillos strain was used, and in the second experiment, a seeder pig infected with the highly-virulent CSFV Koslov strain was used. Pen-based oral fluid samples were collected daily and individual samples (whole blood, swabs) every other day. All samples were tested by a CSFV-specific real-time RT-PCR assay. CSFV genomic material was detected in oral fluids on the seventh and fourth day post-introduction of the seeder pig into the pen, in the first and second experiments, respectively. In both experiments, oral fluids tested positive before the contact pigs developed viremia, and with no apparent sick pigs in the pen. These results indicate that pen-based oral fluids are a reliable and convenient sample type for the early detection of CSF, and therefore, can be used to supplement the ongoing CSF surveillance activities in North America.

Rope-based oral fluid sampling for early detection of classical swine fever in domestic pigs at group level.

BackgroundNon-invasive sampling techniques based on the analysis of oral fluid specimen have gained substantial importance in the field of swine herd management. Methodological advances have a focus on endemic viral diseases in commercial pig production. More recently, these approaches have been adapted to non-invasive sampling of wild boar for transboundary animal disease detection for which these effective population level sampling methods have not been available. In this study, a rope-in-a-bait based oral fluid sampling technique was tested to detect classical swine fever virus nucleic acid shedding from experimentally infected domestic pigs.ResultsSeparated in two groups treated identically, the course of the infection was slightly differing in terms of onset of the clinical signs and levels of viral ribonucleic acid detection in the blood and oral fluid. The technique was capable of detecting classical swine fever virus nucleic acid as of day 7 post infection coinciding with the first detection in conventional oropharyngeal swab samples from some individual animals. Except for day 7 post infection in the “slower onset group”, the chances of classical swine fever virus nucleic acid detection in ropes were identical or higher as compared to the individual sampling.ConclusionsWith the provided evidence, non-invasive oral fluid sampling at group level can be considered as additional cost-effective detection tool in classical swine fever prevention and control strategies. The proposed methodology is of particular use in production systems with reduced access to veterinary services such as backyard or scavenging pig production where it can be integrated in feeding or baiting practices.Electronic supplementary materialThe online version of this article (doi:10.1186/s12917-016-0930-2) contains supplementary material, which is available to authorized users.

Comparative evaluation of single step real-time RT-PCR and gel based RT-PCR assay for detection of classical swine fever

To control classical swine fever (CSF), a highly contagious viral disease causing serious losses in the pig industry worldwide, rapid detection and identification of the causative agent is a crucial step. In the present study, real-time RT-PCR and gel based RT-PCR techniques were compared for detection of CSF virus nucleic acid in both clinical as well as tissue samples. Clinical and tissue samples (325) were collected from CSF suspected outbreaks from different part of north eastern region of India. In gel based RT-PCR, % positivity was 44.61% while in realtime RT-PCR it was 57.23%. Highest per cent positivity was recorded in tonsil followed by mesenteric lymph node, blood, nasal swab, spleen, kidney and ileum. The study indicated that probe based RT-PCR could specifically detect CSF virus genome and detection limit was about one log higher than a gel based PCR assay targeting the non translated region. Total time required to complete the gel based RT-PCR including extraction of viral RNA was about 6 h. On the other hand, real-time RT-PCR assay can be performed in 2 to 3 h, thus providing a rapid detection tool.

Molecular Diagnosis of CSF Virus Isolated from Indian Origin

This case study describes the detection of Classical Swine Fever (CSF) in Karnataka state, India during the year 2009-10.Three cases of acute mortality in small-scale pig farming units were investigated. In all the cases, affected pigs were not vaccinated against CSF. Constant pyrexia, dyspnoea, anorexia, lethargy, conjunctivitis, hyperaemia of the ears and convulsions were the primary clinical signs observed in all the cases. Clinical signs and necropsy findings suggested an early presumptive diagnosis of CSF. Presumptive diagnosis was further strengthened by molecular diagnosis by specific amplification of the 5' UTR region of the virus by subjecting pooled tissue materials to one-step RT-PCR. Strengthening the presumptive diagnosis by the aid of molecular diagnostic techniques is important not only to prevent the spread of the disease but also to evolve suitable strategy to control the disease.

Modelling the spread of Swine Vesicular Disease Virus and Vesicular Stomatitis Virus in an area of livestock units without any control measures and measurement of consequential impact resulting from an assumed introduction into one livestock farm

EFSA Supporting PublicationsVolume 9, Issue 5 285E External scientific reportOpen Access Modelling the spread of Swine Vesicular Disease Virus and Vesicular Stomatitis Virus in an area of livestock units without any control measures and measurement of consequential impact resulting from an assumed introduction into one livestock farm Dr. Dirk Eisinger, Dr. Dirk Eisinger Dr. Dirk EisingerSearch for more papers by this author Dr. Dirk Eisinger, Dr. Dirk Eisinger Dr. Dirk EisingerSearch for more papers by this author First published: 21 May 2012 https://doi.org/10.2903/sp.efsa.2012.EN-285 This external report is not produced by EFSA. It is published to help keep the public informed of developments related to EFSA's scientific work. EFSA reserves its rights, view and position as regards the issues addressed and conclusions reached in the present document, without prejudice to the rights of the authors. Published date: 21 May 2012 Question number: EFSA-Q-2011-01135 AboutPDF ToolsExport citationAdd to favoritesTrack citation ShareShare Give accessShare full text accessShare full-text accessPlease review our Terms and Conditions of Use and check box below to share full-text version of article.I have read and accept the Wiley Online Library Terms and Conditions of UseShareable LinkUse the link below to share a full-text version of this article with your friends and colleagues. Learn more.Copy URL Share a linkShare onFacebookTwitterLinkedInRedditWechat References Backer, J.A., Brouwer, H., van Schaik, G., van Roermind, H.J.W., 2011. Using mortality data for early detection of Classical Swine Fever in The Netherlands. Prev. Vet. Med. 99, 38– 47. Backer, J.A., Hagenaars, T.J., van Roermund, H.J.W., de Jong, M.C.M., 2009. Modelling the effectiveness and risks of vaccination strategies to control classical swine fever epidemics. Journal of the Royal Society Interface 6, 849– 861. Bates, T.W., Thurmond, M.C., Carpenter, T.E., 2003. Description of an epidemic simulation model for use in evaluating strategies to control an outbreak of foot-and-mouth disease. Am. J. Vet. Res. 64, 195– 204. EFSA, 2012. Scientific Opinion on Swine Vesicular Disease and Vesicular Stomatitis. The EFSA Journal 10, 2631. Elbers, A.R.W., Stegeman, A., Moser, H., Ekker, H.M., Smak, J.A., Pluimers, F.H., 1999. The classical swine fever epidemic 1997–1998 in the Netherlands: descriptive epidemiology. Prev. Vet. Med. 42, 157– 184. Garner, M.G., Beckett, S.D., 2005. Modelling the spread of foot-and-mouth disease in Australia. Aust Vet J 83, 758– 766. Grimm, V., Berger, U., Bastiansen, F., Eliassen, S., Ginot, V., Giske, J., Goss-Custard, J., Grand, T., Heinz, S., Huse, G., Huth, A., Jepsen, J.U., Jørgensen, C., Mooij, W.M., Müller, B., Pe'er, G., Piou, C., Railsback, S.F., Robbins, A.M., Robbins, M.M., Rossmanith, E., Rüger, N., Strand, E., Souissi, S., Stillman, R.A., Vabø, R., Visser, U., DeAngelis, D.L., 2006. A standard protocol for describing individual-based and agent-based models. Ecol. Modell. 198, 115– 126. Grimm, V., Berger, U., DeAngelis, D.L., Polhill, J.G., Giske, J., Railsback, S.F., 2010. The ODD protocol: a review and first update. Ecol. Modell. 221, 2760– 2768. Harvey, N., Reeves, A., Schoenbaum, M.A., Zagmutt-Vergara, F.J., Dubé, C., Hill, A.E., Corso, B.A., McNab, W.B., Cartwright, C.I., Salman, M.D., 2007. The North American Animal Disease Spread Model: A simulation model to assist decision making in evaluating animal disease incursions. Prev. Vet. Med. 82, 176– 197. Ribbens, S., Dewulf, J., Koenen, F., Laevens, H., de Kruif, A., 2004. Transmission of classical swine fever. A review. Veterinary Quarterly 26, 146– 155. Staubach, C., Teuffert, J., Thulke, H.-H., 1997. Risk analysis and local spread mechanisms of classical swine fever. Epidémiol. santé anim. 31–32, 6.12.1– 3. Stegeman, A.J., Elbers, A.R.W., Bouma, A., de Jong, M.C.M., 2002. Rate of inter-herd transmission of classical swine fever virus by different types of contact during the 1997–8 epidemic in The Netherlands. Epidemiol. Infect. 128, 285– 291. Thulke, H.-H., Eisinger, D., Beer, M., 2011. The role of movement restrictions and pre-emptive destruction in the emergency control strategy against CSF outbreaks in domestic pigs. Prev. Vet. Med. 99, 28– 37. Volume9, Issue5May 2012285E ReferencesRelatedInformation

Analysis of clinical samples for early detection of classical swine fever during infection with low, moderate, and highly virulent strains in relation to the onset of clinical signs

Classical swine fever (CSF) is a transboundary viral disease affecting swine. The clinical course of disease and the best diagnostic samples for early detection were examined using low, moderate, and highly virulent strains of CSFV inoculated into 8-12 week old domestic pigs. Clinical signs were monitored and recorded. Nasal swabs, tonsil scrapings, blood and tonsils were tested using virus isolation, immunohistochemistry, and real-time reverse transcriptase PCR (rRT-PCR).Severe clinical signs appear 3 days post infection (dpi) with the highly virulent strain, correlating with positive tonsil scrapings, tonsil and blood by virus isolation and rRT-PCR (83-100%), whereas nasal swabs become comparable by 5dpi (89-100%). The moderate strain caused less severe clinical signs between 5 and 7dpi, with tonsil scrapings, tonsil and blood positive by 7dpi (83-100%), and nasal swabs were comparable at 10dpi (67-90%). The low virulent strain showed mild clinical signs at 7dpi, with blood, tonsil and tonsil scrapings positive by virus isolation and rRT-PCR. Except for one sample at 10dpi, nasal swabs remained negative throughout the course of infection. This study indicates that irrespective of virulence, whole blood and tonsil scrapings are the sample of choice for early detection of CSFV in live pigs.

Using mortality data for early detection of Classical Swine Fever in The Netherlands

Early detection of the introduction of an infectious livestock disease is of great importance to limit the potential extent of an outbreak. Classical Swine Fever (CSF) often causes non-specific clinical signs, which can take considerable time to be detected. Currently, the disease can be detected by three main routes, that are all triggered by clinical signs. To improve the early detection of CSF an additional program, based on mortality data, aims to routinely perform PCR tests on ear notch samples from herds with a high(er) mortality. To assess the effectiveness of this new early detection system, we have developed a stochastic model that describes the virus transmission within a pig herd, the development of disease in infected animals and the different early detection programs. As virus transmission and mortality (by CSF and by other causes) are different for finishing pigs, piglets and sows, a distinction is made between these pig categories. The model is applied to an extensive database that contains all unique pig herds in The Netherlands, their herd sizes and their mortality reports over the CSF-free period 2001–2005. Results from the simulations suggest that the new early detection system is not effective in piglet sections, due to the high mortality from non-CSF causes, nor in sow sections, due to the low CSF-mortality. In finishing herds, the model predicts that the new early detection system can improve the detection time by two days, from 38 (27–53) days to 36 (24–51) days after virus introduction, when assuming a moderately virulent virus strain causing a 50% CSF mortality. For this result up to 5 ear notch samples per herd from 8 (0–13) finishing herds must be tested every workday. Detecting a source herd two days earlier could considerably reduce the number of initially infected herds. However, considering the variation in outcome and the uncertainty in some model assumptions, this two-day gain in detection time is too small to demonstrate a substantial effect of the new early detection system based on mortality data. But when the alertness of herd-owners and veterinarians diminishes during long CSF-free periods, the new early detection system might gain in effectiveness.

Rapid pre-clinical detection of classical swine fever by reverse transcription loop-mediated isothermal amplification

The usefulness of reverse transcription loop-mediated isothermal amplification (RT-LAMP) for rapid pre-clinical detection of classical swine fever virus (CSFV) infection was evaluated. The RT-LAMP reaction could be finished in 60 min under isothermal condition at 65 °C by employing a set of four primers targeting the 5′ untranslated region of CSFV. The RT-LAMP assay of CSFV showed higher sensitivities than that of RT-PCR, with a detection limit of 5 copies per reaction. No cross-reactivity was observed from the samples of other related viruses including porcine circovirus type 2, porcine parvovirus, porcine pseudorabies virus, Japanese encephalitis virus, and porcine reproductive and respiratory syndrome virus. The detection rates of CSFV RT-LAMP, RT-PCR and virus isolation for samples including blood, tonsil, nasal and rectal swabs from uninoculated pigs without any clear clinical symptom were 89%, 78% and 71%, respectively. Furthermore, all of the assays showed higher sensitivity for blood and tonsil swabs samples than nasal and rectal swabs. These results indicate that the CSFV RT-LAMP assay is a valuable tool for its rapid, cost-effective detection and has potential usefulness for rapid pre-clinical detection and surveillance of classical swine fever in developing countries.

Evaluation of real-time RT-PCR assay for the routine intra vitam diagnosis of classical swine fever

The aim of the study was to evaluate the diagnostic quality of the real-time RT-PCR assay described by Hoffmann et al. [Hoffmann, B., Beer, M., Schelp, C., Schirrmeier, H., Depner, K., 2005. Validation of a real-time RT-PCR assay for sensitive and specific detection of classical swine fever. J. Virol. Methods 130, 36–44] for the routine intra vitam diagnosis of classical swine fever (CSF). We compared the assay with conventional diagnostic methods by using defined diagnostic material from an animal experiment with pigs showing different clinical forms of CSF. Compared to virus isolation and antigen ELISA an enhanced sensitivity of the real-time RT-PCR could be shown. We were able to detect all infected pigs regardless of the clinical course of CSF. CSF infection was detected already during the incubation period, during the entire clinical phase as well as at the beginning of convalescence when the first antibodies were detected and no virus could be isolated any more. In most cases, positive PCR results were obtained 2 days earlier than with virus isolation and 2–4 days earlier than with the antigen ELISA.

Validation of a real-time RT-PCR assay for sensitive and specific detection of classical swine fever

A fully validated, ready-to-use, real-time reverse transcription-polymerase chain reaction (RT-PCR) assay, multiplexed for simultaneous detection of an internal control, for the simple and rapid diagnosis of classical swine fever (CSF) was developed. Primers and FAM-labeled TaqMan-probes specific for classical swine fever virus (CSFV) were selected from the consensus sequence of the 5′ non-translated region (5′ NTR) of 78 different CSFV strains. For determining analytical sensitivity, an in vitro transcript (T7-PC3alf) of the 5′ NTR was constructed and tested. In addition, the T7-PC3alf transcript was further used as a positive control and a standard for quantitation of CSFV genome copies. A second heterologous in vitro transcript based on a specific primer–probe HEX-system was designed as an internal positive control for the RNA isolation step and RT-PCR. By using limited primer concentrations for the internal control, no adverse effects on the sensitivity of the CSF-system could be observed, and the newly designed duplex real-time RT-PCR proved to have a sensitivity of approximately eight copies. The primer–probe combination selected was strictly CSFV-specific and no amplification was observed in all non-CSFV pestiviruses tested.

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.

Quantitative assessment of clinical signs for the detection of classical swine fever outbreaks during an epidemic

The performance of clinical signs as a diagnostic test for the detection of classical swine fever (CSF) outbreaks during the 1997–1998 CSF epidemic in The Netherlands was evaluated by constructing and analysing a receiver operating characteristic (ROC) curve. This curve assesses the discriminating ability of a diagnostic test over a range of test signals. The cut-off values for a defined diagnostic test to detect CSF outbreaks were set by different combinations of clinical signs observed. The area under the ROC curve, which is a quantitative measure of test performance, was significantly ( P<0.001) larger than the area under the random ROC curve. This indicates that clinical signs have a significantly higher performance as a diagnostic test for the detection of CSF than for flipping a coin. However, the gain in diagnostic performance compared to a random process is not as much as we would wish it to be. The optimal efficient diagnostic test combined a sensitivity of 72.7% with a specificity of 52.7%, with a combination of the following clinical signs: unsteady gait/ataxia, not eating, not reacting to antibiotic treatment, conjunctivitis, hard faecal pellets.