West Nile fever

OIE Reference Laboratory Reports

Activities in 2010

Name of disease (or topic) for which you are a designated OIE Reference Laboratory: / West Nile fever
Address of laboratory: / Istituto Zooprofilattico Sperimentale dell’Abruzzo e del Molise “G. Caporale”
Via Campo Boario, 64100 Teramo - ITALY
Tel.: / (+39-0861) 33.22.05
Fax: / (+39-0861) 33.22.51
e-mail address: /
website: / www.izs.it
Name of Head of Laboratory (Responsible Official): / Vincenzo Caporale
Name of OIE Reference Expert: / Rossella Lelli
Name of writer of this report
(if different from above):


Part I: Summary of general activities related to the disease

1. Test(s) in use/or available for the specified disease/topic at your laboratory

Test / For / Specificity / Total
ELISA IgG / Antibody /

Flavivirus Group

/ 6,952
ELISA IgM / Antibody /

Type

/ 1,524

VERO cell culture

/ Virus isolation / Type / 2,436
RT-PCR Real time / Antigen /
Type
/ 39
RT-PCR Real time / Antigen /
WNV lineage 1
/ 7,178

IF

/ Antigen / Flavivirus Group / 2,436
Serum Neutralization (microtitre) /
Antibody
/

Type

/ 5,777

Plaque Reduction neutralization test (PRNT)

/ Antibody / Type / 400

Genome sequencing

/ Antigen / Type / 4
Morphologic examination / Mosquitoes identification / Species / 983

2. Production and distribution of diagnostic reagents

Production and testing of West Nile (WN) virus (WNV) (strain Eg101) antigen, positive antisera and monoclonal antibodies, to be used in serological and virological tests, have continued throughout the year.

In the 2010, WNV i-ELISA reagents sufficient for 6,952 tests have been produced and used for the activities of the National Surveillance program.

Isolates of WNV collected during the 2008 and 2009 Italian outbreaks were aliquoted and stocked. The genome of the most representative strains were fully sequenced to obtain molecular epidemiological understanding of the WNV origin.

Type of reagent / Amount supplied nationally
(including for own use) / Amount supplied to other countries
Control positive serum / 31 ml / 1 ml (Bulgaria)
Control negative serum / 31 ml / 1 ml (Bulgaria)
Positive antigen for ELISA test (Eg101 strain) / 25 ml / -
Negative antigen for ELISA test / 25 ml / -

WNV strain Eg101

/ 200 ml / -

WNV strain NY99

/ - / 1 ml (Bulgaria)


Part II: 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 laboratory participated at the proficiency test on serological (IgG ELISA, IgM ELISA, PRNT) and virological (real-time RT-PCR) methods, organized by the EU Reference laboratory (EU-RL) for equine diseases.

4. Preparation and supply of international reference standards for diagnostic tests or vaccines

WN virus strain NY99 and specific antiserum have been sent to the National Diagnostic and Research Veterinary Institute “Pencho Slavceikov” di Sofia, Bulgaria.

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 2010 particularly focussed on:

·  a program for the automatization and validation of the real time RT-PCR for detecting WNV strains from insects, organs and blood samples has been put in place. The system is now under testing and the validation phase is in progress;

·  a new competitive ELISA for detecting WNV antibodies from serum samples is under standardization and will be validated according to the OIE standards.

6. Collection, analysis and dissemination of epizootiological data relevant to international disease control

·  In the framework of a research project titled “A multidisciplinary approach to the epidemiology of the West Nile virus in Italy” funded by the Italian Ministry of Health, OIE-laboratory personnel is involved in the definition of epidemiological models on WN distribution in Italy with specific regards to resident bird species. In particular, the role of some species of resident birds, like magpies and carrion crows, is under study, with the determination of the ecological and behavioral aspects as well as the length of viraemia in these species.

·  In the effort to characterize the WNV strain circulating in Italy, the genome of the representative isolates of different years has been fully sequenced and phylogenetic analysis performed in the attempt to investigate the possible origin. Furthermore an experimental infection on mouse model has been carried out. Mortality rate, pathogenicity and neuroinvasivity were assessed and compared to the prototype strain isolated in USA (New York 99).

·  According to the Italian WN surveillance program, insects collected from nearly 700 catches originating from different traps (CO2 CDC light-trap, BG sentinel and Gravid trap) located in all Italian regions, were coordinated and examined. These, together with all relevant virological, serological and epidemiological data on west Nile in Italy, were published through a web-based information system on a weekly basis (website: www.izs.it). Furthermore continuous advices on WN entomological surveillance and laboratory diagnosis have been given to Italian institutions involved in the WN Surveillance program.

·  A collaboration with the Biopharma Laboratory of Rabat, Morocco, and Moroccan Veterinary Authorities has been put in place in the fields of diagnostic tests standardisation, vaccine researches and in the epidemiological analysis of West Nile Fever (WNF) outbreaks occurred in Morocco in 2010 (environmental data from satellite resources has been collected, processed and preliminary analysed). On the 30th of September and the 1st of October 2010, OIE-laboratory personnel visited Morocco to make a preliminary assessment of the epidemiological situation regarding WNF and other emerging diseases. The analysis of the epidemiological relevant data is carried out in collaboration with the OIE Collaborating Centre for Veterinary Training, Epidemiology, Food Safety and Animal Welfare of Institute “G. Caporale”.

7. Provision of consultant expertise to OIE or to OIE Members

·  In the framework of a research project titled “Network for vector-borne infections surveillance” funded by the Italian Ministry of Health, OIE laboratory personnel is collaborating with experts of the Central Veterinary Laboratory of Etlik, Ankara, Turkey, on the main aspects of the West Nile epidemiology and diagnosis. In particular, from 27 September to 1 October a visit to Turkey was performed by OIE-laboratory personnel with the aim of exploring a possible twinning project.

·  In the framework of the collaboration with the Biopharma Laboratory of Rabat, Morocco, OIE-laboratory personnel, jointly with Biopharma experts, is carrying out a spatial analysis of WNF outbreaks occurred in 2010 in Morocco. The environmental conditions of the areas where outbreaks occurred are analysed to better define the geographical and climatic patterns playing a possible role in WNF spread in Northern Africa.

·  The laboratory also contributed to the 2nd EU Workshops on “Annual meeting of the National Reference laboratories for equine diseases”, 19th October 2010, ANSES, Maisons-Alfort, France, presenting the 2008, 2009 and 2010 WN circulation in Italy.

·  Experts from the Laboratory also attended:

·  Meeting of the OIE ad hoc Group on Scientific Partnerships of OIE Laboratories, Paris, France, 6-7 May 2010;

·  Second Global Conference of OIE Reference Laboratories and Collaborating Centres, Paris, France, 21-23 June 2010;

·  OIE Global Conference on Veterinary Legislation, Djerba, Tunisia, 7-9 December 2010.

8. Provision of scientific and technical training to personnel from other OIE Members

In the framework of a research project titled “International Network for Capacity Building for the Control of Emerging Viral Vector Borne Zoonotic Diseases, in particular West Nile Fever, Rift Valley Fever and Crimean-Congo Haemorrhagic Fever ARBOZOONET” (KBBE–2007–1–3–06) (a project which aimed to create a Surveillance Network for vector borne zoonosis) a workshop on WND diagnostic techniques was held in OIE laboratories (27-29 April, Teramo, Italy). This workshop was jointly organised by the OIE laboratory and the Veterinary Laboratories Agency (VLA) (UK). It was hosted in Teramo with the aim of giving participants the ability to perform molecular and serological methods for the diagnosis of West Nile disease (WND) in accordance with the official and/or most commonly used techniques. The workshop was attended by 20 participants, representing 12 different countries, and included presentations, instructional videos, discussions and practical sessions given by both IZS AM and VLA staff.

9. Provision of diagnostic testing facilities to other OIE Members

The following table summarises the official diagnostic services provided and the Member Countries where the samples originated.

Country / Test / No. of samples
Bulgaria
primary diagnostic / IgG ELISA / 11
IgM ELISA / 11
SN / 11
PRNT / 11

VERO cell culture

/ 8
RT-PCR Real time / 8
Morocco
confirmatory testing / IgG ELISA / 18
IgM ELISA / 18
SN / 18
PRNT / 18
RT-PCR Real time / 6
VERO cell culture / 6
Country / Test / No. of samples
Turkeyconfirmatory testing / IgM ELISA / 382
SN / 382
PRNT / 37
RT-PCR / 382
Croatia
confirmatory testing / IgG ELISA / 34
IgM ELISA / 34
SN / 34
PRNT / 34
Greece
confirmatory testing / IgG ELISA / 5
IgM ELISA / 5
SN / 5
PRNT / 5

10. Organisation of international scientific meetings on behalf of OIE or other international bodies

–

11. Participation in international scientific collaborative studies

The OIE lab is one of the participants of the research project “Emerging Diseases in a changing European eNvironment (EDEN)” 6th FP (2005-2010) funded by the European Commission. It is an Integrated Project of the European Commission that aims to identify and catalogue those European ecosystems and environmental conditions which can influence the spatial and temporal distribution and dynamics of human pathogenic agents. OIE lab personnel has been involved in the WN WP providing data related to vertebrate hosts and vectors in Italian study area (Padule di Fucecchio marshes, Tuscany Region). A database was build to collect data related to:

·  Birds: species (54 caught, 48 bleeded, 1405 blood samples tested serologically and virologically)

distributions in ecotopes

abundance index

·  Horses: 420 blood samples of resident horses were collected and processed

·  Mosquitoes: 16 species identified.

During the study period, an intense collaboration with the Romanian EDEN team was carried out for the serological analysis of samples: 1209 sera of wild birds and 150 horse sera were analyzed.

The personnel of the OIE lab participate to the research project titled “International Network for Capacity Building for the Control of Emerging Viral Vector Borne Zoonotic Diseases, in particular West Nile Fever, Rift Valley Fever and Crimean-Congo Haemorrhagic Fever Arbo-Zoonet” (KBBE–2007–1–3–06). The Arbo-Zoonet project aims at creating common knowledge on West Nile (WN) fever, Crimean- Congo hemorrhagic fever (CCHF) and Rift Valley fever (RVF), arthropod borne diseases of domestic and wild animals that can affect humans, posing a great threat to public health because of their epidemic and zoonotic potential.

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

·  Monaco F, Savini G., Calistri P., Bruno R., Lelli R. West Nile viruses responsible for the 2008 and 2009 Italian outbreaks. Oral presentation. EDEN Annual Meeting 10-12 May 2010, Montpellier, France.

·  Lelli R., Pascucci I., Calistri P., Bruno R., Monaco F., Savini G. West Nile Transmission in resident birds in Italy. Oral presentation. EDEN Annual Meeting 10-12 May 2010, Montpellier, France.

¡  Scientific publications in peer-reviewed journals

·  Lelli R., Calistri P., Bruno R., Monaco F., Savini G., Di Sabatino D., Corsi I., Pascucci I. West Nile transmission in resident birds in Italy. Submitted to Transboundary and Emerging Diseases.

·  Lelli R. 2010. West Nile virus in Europe: understanding the present to gauge the future. Euro Surveillance 2010;15(15):pii=19538 Available online: http://www.eurosurveillance.org/ViewArticle.aspx?ArticleId=19538

·  Calistri P., Giovannini A., Savini G., Monaco F., Bonfanti L., Ceolin C., Terregino C., Tamba M., Cordioli P., Lelli R. 2010. West Nile virus transmission in 2008 in North-Eastern Italy. Zoonoses and Public Health. 57: 211-219.

·  Calistri P., Giovannini A., Hubalek Z., Ionescu A., Monaco F., Savini G., Lelli R. 2010. Epidemiology of West Nile in Europe and in the Mediterranean Basin. The Open Virology Journal. 4: 29-37.

·  Calistri P., Monaco F., Savini G., Guercio A., Purpari G., Vicari D., Cascio S., Lelli R. 2010. Further spread of West Nile infection in Italy. Vet Ital. 46 (4), 471-474.

·  Monaco F., Lelli R., Teodori L., Pinoni C., Di Gennaro A.P., Polci A., Calistri P., Savini G. 2010. Re-emergence of West Nile virus in Italy. Zoonoses and Public Health. 57: 476-486.

·  Monaco F., Savini G., Calistri P., Polci A., Pinoni C., Bruno R., Lelli R. 2009 West Nile disease epidemic in Italy: first evidence of overwintering in western Europe?. Research in Veterinary Science. In press.

·  Patregnani T., Bonfanti L., Montarsi F., Savini G., Ravagnan S., Marangon S., Capelli G. 2010 West Nile Disease (WND) outbreak in Italy and the role of dogs as potential sentinel for surveillance programs. Proceedings of the fifth Symposium of the Canine Vector Borne disease, New YorkCity, USA12-15 April 2010, pp. 16-17.

·  Busani L., Capelli G., Cecchinato M., Lorenzetto M., Savini G., Terregino C., Vio P., Bonfanti L., Marangon S. 2010. West Nile virus circulation in the Veneto Region in 2008 – 2009. Epidemiol Infect. 2010 Jul 30:1-8.

¡  Other communications

The public web site (www.izs.it) disseminating information and data on West Nile has been continuously updated in order to have:

·  the latest on the Italian and European Regulations issued by the Italian Ministry of Health;

·  the current (2010) and past (2008 and 2009) epidemiological situations in Italy;

·  maps on entomological and serological surveillance activities;

·  the current epidemiological situations in the Mediterranean Basin;

·  scientific documents on-line.

13. Inscription of diagnostic kits on the OIE Register

i) Did you participate in expert panels for the validation of candidate kits for inscription on the OIE Register? If yes, for which kits?

No

ii) Did you submit to the OIE candidate kits for inscription on the OIE Register? If yes, for which kits?

No

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Annual reports of OIE Reference Laboratories and Collaborating Centres, 2010 5