Bluetongue
Activities in 2009
Bluetongue
Dr Giovanni Savini
Istituto Zooprofilattico Sperimentale dell’ Abruzzo e Molise “G. Caporale”,
Via Campo Boario, 64100 Teramo, Italy
Tel.: (39-0861) 33.24.40, Fax: 39-0861) 33.22.51
e-mail: bsite:
Summary of general activities related to the disease
1.Test(s) in use/or available for the specified disease at your laboratory
Test / For / Specificity / TotalC-ELISA / Antibody / Group / 33,999
AGID / Antibody / Group / 40
VNT / Antibody / Type / 141,840
KC + VERO cell culture
/ Virus isolation / Group / 46ECE+VERO cell culture
/ Virus isolation / Group / 144VERO cell culture
/ Virus isolation / Group / 162VERO cell culture
/ Virus titration / Group / 631Serotype specific PCR real time
/ Virus identification / Type / 39PCR real time
/ Agent identification / Group / 5,356Virus typing
/ Virus identification / Type / 24Microscopic examination
/ Culicoides imicola identification / Species / 5,110PCR
/ Culicoides obsoletus and Culicoides scoticus identification / Obsoletus complex species / 179Microscopic examination
/ Culicoides spp. /Species
/ 5,1092.Production and distribution of diagnostic reagents
Production and testing of bluetongue (BT) virus (BTV) antigen and sheep antisera, to be used in serological tests and diagnostic kits, have continued throughout the year.
In the 2009, BTV C-ELISA reagents sufficient for 146,475 tests have been produced and 77,400 were supplied to Italian laboratories.
Isolates of BTV-8, BTV-4 and BTV-1 collected during the 2009 Italian outbreaks and the trace back activity from animals coming from Northern Europe, were aliquoted and stocked. The most representative strains were cloned and sequenced to obtain molecular epidemiological understanding of the BTV origin.
From the 2009 National Surveillance Program 4,123 new sera were collected, aliquoted, lyophilised and stored.
Activities specifically related to the mandate
of OIE Reference Laboratories
3.International harmonisation and standardisation of methods for diagnostic testing or the production and testing of vaccines
The eighth inter-laboratory proficiency testing for BT ELISA including 37 Italian laboratories and the Croatian Veterinary Institute of Split, Croatia, was organised. All ring trial records (subscriptions, data submission, final report) were handled through an on-line internet interface.
The third inter-laboratory proficiency testing for RT-PCR including 12 Italian laboratories was also organised. As for the serology, all ring trial records were handled through an on-line internet interface.
The laboratory also participated to the “fourth European Union (EU) ring trial for BT ELISAs and RT-PCR” organised by the Institute for Animal Health, Pirbright Laboratory, Pirbright, UK, (European Community Reference Laboratory for BT).
The laboratory participated at the second ring trial specific for the molecular identification of Culicoides spp. organised by CIRAD (Cetre-Sossa) within the MEDREONET (EU Sixth Framework Programme) activities.
4.Preparation and supply of international reference standards for diagnostic tests or vaccines
Antiserum and virus stocks were continuously refilled and supplied to other Member Countries when requested.
BTV-1, BTV-6, BTV-8 and BTV-11 reference strains have been sent to the Institute of Diagnosis and Animal Health, Bucharest, Romania.
BTV-1, BTV-4 and BTV-9 vaccine and reference strains have been sent to the Faculty of Veterinary Medicine, Institute of Comparative Medicine, University of Glasgow, Scotland, UK.
During the 2009, 68 light traps for catching Culicoides spp. were built and sold to Norway, France, Poland, Romania and Spain.
5.Research and development of new procedures for diagnosis and control
Works on the Italian Administration-funded projects coordinated by personnel of the OIE reference laboratory continued throughout 2009 as follows:
- The program for validating the manually executed real time RT-PCR for detecting all BTV serotypes from insects, organs and blood samples was completed. The system is now under automatization and the validation program for this new step is in progress.
- A new competitive ELISA for detecting BTV antibodies from serum samples using new antigens and monoclonal antibodies was standardised and validated according to the OIE standards.
- The use of ovine and human monocytes and other cell lines to grow BTV is under study.
- A trial funded by the Italian Ministry of Health which aims to study the transplacental transmission of different strains of BTV-8 in cows started.
- A trial which aimed to study the transplacental transmission of different strains of BTV-8 in ewes in the last month of pregnancy was completed.
- The laboratory part of the Project on “New expressing vector for the bluetongue virus” has been completed. In this project an ORF virus has been used to express the VP2 and VP5 gene segments of BTV in order to immunize susceptible animals.
- The validation of the commercial kits “VIROTYPE BTV" of the Labor Diagnostik Leipzig and “ADIAVET BTV REAL TIME KIT” of the Adiagene to detect BTV from blood and tissue samples, has been completed. Validation program of the commercial kit “VetMAX bluetongue virus reagents” of the Applied Biosystem for detecting BTV RNA from blood and tissue samples is in progress.
- A project on “the use of “landscape analysis and Remote Sensing” techniques to investigate the microclimatic and environmental factors associated to the C. obsoletus s.s., C. scoticus, C. dewulfi and C. montanus distribution, started. The project aims to identify the most important factors which determine the presence of C. obsoletus s.s., C. scoticus, C. dewulfi and C. montanus in the Italian environment.
6.Collection, analysis and dissemination of epizootiological data relevant to international disease control
Relevant data on BTV control were collected, analysed and disseminated through the use of a web-GIS system and the establishment an OIE Bluetongue reference laboratories network (OIE-BTNet). More information on these activities are given in the Report of the IZS A&M OIE Collaborating Centre for ‘Veterinary Training, Epidemiology, Food Safety and Animal Welfare’ and in the details of the 2nd meeting listed in paragraph 10.
EU-BTNet – a web-based information system aiming to collect and spread data on bluetongue surveillance in Europe has been developed and set up. The system, sponsored by the European Union (Decision 2007/367/CE), has been designed for collecting all information on bluetongue gathered in the course of the implementation of the bluetongue monitoring and/or surveillance programmes and on the mass vaccination campaigns that are taking place in the Member States.
According to the Italian BT surveillance program, insects collected from nearly 6,600 catches originating from permanent (249) and mobile light traps (24) in a total of 320 municipalities located in all Italian regions, were coordinated and examined. These, together with all relevant virological, serological and epidemiological data on bluetongue in Italy, were published through a web-based information system on a weekly basis (website:
7.Provision of consultant expertise to OIE or to OIE Members
Personnel of the OIE reference lab:
participated to the European Union working group on BTV surveillance to discuss on the BTV-6 and BTV-11 inCentral Europe and on possible amendments on the Bluetongue regulation (EU) 1266/2007. Brussels, Belgium, 3 March 2009;
participated as expert to the OIE meeting on “Amendments to terrestrial code chapter on Bluetongue: infective period, BT vaccines, the risk of maternal transmission”. Paris, France, 14 April 2009;
participated to the European Union working group on BTV to discuss on 2009 and 2010 vaccination plans, trade of vaccinated animals and vaccination report. Brussels, Belgium, 26 October 2009;
completed the Scientific reviews on Crimean Congo Hemorrhagic Fever (CCHF) and Epizootic Hemorrhagic Disease (EHD) (Call for Proposal CFP/EFSA/AHAW/2008/04);
participated to the European Food Safety Authority (EFSA) Working Group on “Epizootic Haemorragic Disease” and contributed to the document “EFSA - Opinion of the Scientific Panel on Animal Health and Welfare on request from the Commission on “Epizootic Haemorragic Disease” which is available on line: The panel met in Parma the 30th June-01st of July and the 07th-09th of September, the 14th-16th of October in Barcelona and the 18th-19th of November in Brussels;
participated as invited speaker to the Veterinary Information Day, organised by the Central Veterinary Laboratory, lecturing on “Bluetongue: introduction to the disease, aetiology, laboratory activities and research, disease control”. Windhoek, Namibia, 3 April 2009;
participated as invited speakers to the 2nd Leipziger Labor Diagnostik, Symposium, 25-26 June Leipzig, Germany;
reviewed the Bluetongue chapter of the “Manual of Diagnostic Tests and Vaccines for Terrestrial Animals (Terrestrial Manual)”.
The laboratory also contributed to the EU Workshop on “Annual meeting of National Reference Laboratories for bluetongue”, Brussels, Belgium, 30 November 2009, presenting the 2009 BT situation in Italy and the results on the “BTV-8 vaccination and infection”.
8.Provision of scientific and technical training to personnel from other OIE Members
Within the framework of the MIUR (Italian Ministry of Universities and Research) project “Innovative eLearning system for the agricultural sector in the Mediterranean countries: recognition, diagnosis, control and prevention of Arthropod-borne diseases”, material for the course on BT was prepared.
Training on BTV diagnostic methods was provided to veterinary officers from State Veterinary Institute, Split, Croatia.
Within the agreement with the Ministry of Agriculture, Water and Forestry and the Department of Veterinary Services of Namibia, the reference laboratory supported the Central Veterinary Laboratory of Windhoek in the application of bluetongue diagnostic protocols.
Continuous advices on BT entomological surveillance and laboratory diagnosis have been given to the Croatian Veterinary Institute, Zagreb and Split, Croatia; Ministry of Rural Affairs and the Environment (MRAE), Valletta, Malta; Kosovo Central Veterinary Laboratory, Prishtina, Kosovo; Veterinary Faculty, University of Slovenia, Ljubljana, Slovenia; Institute for Diagnosis and Animal Health, Bucharest, Romania, State Veterinary Institute, Zvolen, Slovakia, State Veterinary Institute, Jihlava, Czech Republic. Kimron Veterinary Institute, Beit Dagan. Israel
.9.Provision of diagnostic testing facilities to other OIE Members
Within the framework of the agreement for the co-operation between the Istituto Zooprofilattico Sperimentale dell’Abruzzo e del Molise “G. Caporale” and the Department of Veterinary Services of the Republic of San Marino for assistance in Bluetongue surveillance signed in 2007, the surveillance activities continued.
Within the agreement with the SIVtro- Vétérinaires sans Frontières Italy, the laboratory assisted the Saharawi population in problems related to animal health including BT.
An agreement between the “Istituto Zooprofilattico Sperimentale dell’Abruzzo e delMolise” and the Veterinary Authorities of The Netherlands and Germany for assistance in bluetongue virus serotype 6 surveillance was settled.
The following table summarises the official diagnostic services provided and the Member Countries where the samples originated.
Country / Test / No. of samplesIsrael / BTV Typing RT- PCR / 17
BTV sequencing / 17
Virus neutralisation / 17
Virus isolation / 17
EHDV RT-PCR / 17
Netherlands / VNT / 274
San Marino / C-ELISA / 350
VNT / 350
Germany / VNT / 1515
Croatia / VNT / 2
10.Organisation of international scientific meetings on behalf of OIE or other international bodies
The second STEREO2 - EPISTIS project Annual Meeting was organised and held in Teramo, Italy, 18-19February 2009
The 2nd “OIE Bluetongue Reference Laboratory Network Meeting Workshop”, was organised and held in Cerrano, Pineto (TE), Italy 1-2 June, 2009. The Conference assembled bluetongue specialists and OIE bluetongue reference laboratories. The main purposes of the conference were to update and strengthen a reference laboratory network in order to handle, under the auspices of the OIE headquarters, epidemiological information on BT global occurrence and spread, on vector presence and abundance and on BTV typing and genetic characterization. As in the previous meeting, it was a good chance to enforce the necessity of making available and exchanging diagnostic material, the importance of harmonizing, standardizing and validating diagnostic tests between the ref labs, of assisting national reference laboratories in relation to BT diagnosis, BTV and vectors identification and of providing expertise and training to OIE and OIE Member Countries in relation to BT diagnosis, surveillance and control.
11.Participation in international scientific collaborative studies
Within the European call “KBBE-2009-1-3-02: Diseases caused by Orbiviruses: African horse sickness, bluetongue and epizootic haemorrhagic disease: development of new generation vaccines and accompanying tests” the IZS A&M led a group including the Agricultural Research Council - Onderstepoort Veterinary Institute, Onderstepoort, SA; the Kimron Veterinary Institute, Beit Dagan, Israel; Pfizer Animal Health, UK; Svanova Biotech AB, Uppsala, Sweden; Animal Health Service (GD), Deventer, the Netherlands; the Friedrich-Loeffler-Institute, Federal Research Institute for Animal Health, Tuebingen, Germany. A project proposal was prepared and submitted to the European Union.
Activities were carried out on the project “Oral susceptibility of European and South African Culicoides species for European and South African bluetongue virus isolates”, in collaboration with the Agricultural Research Council – Onderstepoort Veterinary Institute in South Africa, within the framework of the “Executive programme of scientific and technological cooperation between Italy and South Africa”, signed in 2007.
Within the Med_Reo_Net Project (a project which aimed to create a Surveillance Network of Reoviruses, Bluetongue and African Horse Sickness, in the Mediterranean basin and Europe) serological surveillance data have been analysed in order to evaluate their capability of detecting the infection in population of vaccinated and unvaccinated animals. Simulation models to assess the sensitivity and the costs of the surveillance system have been developed. The models will be validated on the Italian and Catalonia surveillance data. For this last target, a collaboration with the Animal Health Research Centre (CRESA), Barcelona (Spain) has been signed.
STEREO2 - EPISTIS – Within the project on remote sensing tools to study the epidemiology and space/time dynamics of diseases sponsored by Ministry for Science Policy of Belgium (STERO II Project), spread models able to characterize and define the vector distribution based on genetic characteristics were developed. In collaboration with “l’ Universitè Libre de Bruxelles”, models which investigate the space/time dynamics of C. imicola through the analysis of 7 years of entomological surveillance were set up. Beside l’ Universitè Libre, the project also involves “AVIAGIS”, the Agriculture and Veterinary Information and Analysis. With this partner the soil will be classified through the satellite images and the optimal habitat of the C. imicola will be determined through the landscape analysis.
In collaboration with the Faculty of Veterinary Medicine, Institute of Comparative Medicine, University of Glasgow, Scotland, UK within the project on the “identification of the Molecular determinants of BTV virulence”, studies on the pathogenicity of some field and vaccine BTV strains using mouse model were completed.
Within the framework of the ERA-Net on Emerging and Major Infectious Diseases of Livestock EMIDA, the laboratory, together with other 9 dedicated laboratories of 9 different countries, participated to the European call presenting a proposal on “Combating orbivirus infections of livestock: understanding of the molecular basis for protein function / virus phenotype, molecular epidemiology and improving diagnostic assays.”
12.Publication and dissemination of information relevant to the work of OIE (including list of scientific publications, internet publishing activities, presentations at international conferences)
Presentations at international conferences and meetings
Savini G., Paladini C., Leone A., Teodori L., Migliaccio P., Lelli R., Monaco F., 2009. Infezione transplacentare di alcuni ceppi del virus della bluetongue. III Workshop Nazionale di Virologia Veterinaria, Bari 11 e 12 Giugno 2009.
Savini G., Bonelli P., Nardini R., Leone A., Di Ventura M., Ruggeri F.M., Pasquali P., Cannas E.A., Re R., Nicolussi P., 2009. Clinical signs and viraemia in sheep vaccinated against bluetongue virus serotype 1 and challenged with bluetongue virus serotype 2. World Association of Veterinary Laboratory Diagnosticians-14th International Symposium, Madrid, Spain, 17-20 June 2009
Napp S., Calistri P., Giovannini A., Gubbins S., García-Bocanegra I., Alba A., Allepuz A., Casal J.. Quantitative Assessment of the risk of bluetongue by Culicoides introduced via transport and trade networks. Third Annual Meeting of MEDREONET, Lisbona, 2-4 December 2009.
Napp S., Gubbins S., Calistri P., Allepuz A., Alba A., García-Bocanegra I., Giovannini A., Casal J. Quantitative Assessment of the probability of Bluetongue virus overwintering by horizontal transmission in vectors, ruminants or in both: application to Germany. Third Annual Meeting of MEDREONET, Lisbona, 2-4 December 2009.
Caporale M., Chiam R., Savini G., Franchi P., Di Gialleonardo L., Golder M., Armillotta G., Di Francesco G., Lelli R., Pini A., Kellam P., Palmarini M.. Molecular determinants of BTV virulence Third Annual Meeting of MEDREONET, Lisbona, 2-4 December 2009
Savini G. “RT-PCR and diagnosis of Bluetongue” 2nd Leipziger Labor Diagnostik, Symposium, 25-26 June Leipzig, Germany
Di Gialleonardo L. “Accuracy of the “VIROTYPE BTV” kit in detecting bluetongue virus serotypes circulating in Europe”. 2nd Leipziger Labor Diagnostik, Symposium, 25-26 June Leipzig, Germany
Savini G.. “Cross placental transmission of some Bluetongue isolates other than BTV-8”. 2nd OIE Bluetongue Reference Laboratory Network Meeting Workshop. Cerrano, Pineto (TE), Italy 1-2 June, 2009
Scientific publications in peer-reviewed journals
Savini G., Hamers C., Conte A., Migliaccio P., Bonfini B., Teodori L., Di Ventura M. Hudelet P., Schumacher C. Caporale V. 2009. Assessment of efficacy of a bivalent BTV-2 and BTV-4 inactivated vaccine by vaccination and challenge in cattle. Vet. Microbiol. 133, 1-8.
Listeš E., Labrović A., Monaco F., Paladini C., Leone A., Di Gialleonardo L., Camma’ C., Savini G. 2009. First evidence of bluetongue virus serotype 16 inCroatia. Vet. Microbiol doi:10.1016/j.vetmic.2009.03.011
Listeš E., Bosnić S., Benić M., Madić J., Cvetnić Ž., Lojkić M., Šeparović S., Labrović A., Meiswinkel R., Savini G., 2009. An outbreak of bluetongue and entomology study in Southern Croatia, Archiv Vet., submitted
Eschbaumer M., Hoffmann B., Moss A., Savini G., Leone A., König P., Zemke J., Conraths F. and M. Beer. 2009. Emergence of bluetongue virus serotype 6 inEurope – German field data and experimental infection of cattle. Vet. Microbiol. doi:10.1016/j.vetmic.2009.11.040
Conte A., Gilbert M., and Goffredo M. (2009). Eight years of entomological surveillance in Italy show no evidence of Culicoides imicola geographical range expansion. Journal of Applied Ecology 2009, 46, 1332–1339. doi: 10.1111/j.1365-2664.2009.01723.x
Other communications
The public web site ( disseminating information and data on bluetongue has been continuously updated in order to have: