ANNUAL REPORT on surveillance for avian influenza in wild birds in the EU in 2008

Transcription

1 EUROPEAN COMMISSION ANNUAL REPORT on surveillance for avian influenza in wild birds in the EU in 2008 Community Reference Laboratory for Avian Influenza

2 Neither the European Commission nor any person acting on behalf of the Commission is responsible for the use which might be made of the following information. Online information about the European Union is available at: Further information on the Health and Consumers Directorate-General is available on the internet at: ISBN-13: DOI: /87920 European Union, 2010 Reproduction is authorised provided the source is acknowledged.

3 About the report Firstly, the resources and efforts invested by Chief Veterinary Officers, veterinary services, national reference laboratories, ornithologists, bird watching organisations, hunters and all the people who have contributed to the collection of samples in the conduct of the surveillance and compilation of this data are hereby specifically acknowledged. This Report was prepared by the Community Reference Laboratory for Avian Influenza, Veterinary Laboratory Agency (VLA), Weybridge, Addlestone, Surrey, KT15 3NB according to its work programme. It was prepared by Andrew Breed, George Gould, David Rae, Uta Hesterberg, Kate Harris, Ruth Moir, Bhudipa Choudhury, Adnan Younas and Alasdair Cook under the main responsibility of: Andrew Breed Tel: and the Director of Community Reference Laboratory for avian influenza: Dr. Ian H. Brown Tel: The European Commission provides for the collection of surveillance data submitted by the Member States via an online reporting system operated by the Commission services of Directorate General for Health and Consumers (DG SANCO). The technical support provided by Caroline Fabre from the SANCO Information systems unit A4 is specifically acknowledged. For more information on the Commission's online reporting system please contact: Valentina Piazza, Unit 04 - Veterinary control programmes Tel: valentina.piazza@ec.europa.eu. For more general information on the Annual Report on surveillance for avian influenza in wild birds in the European Union and its publication on the Commission's website: please contact: Maria Pittman, Unit D1-Animal Health and Standing Committees Tel: maria.pittman@ec.europa.eu Reproduction is authorised, provided the source is acknowledged, save where otherwise stated. The views or positions expressed in this report do not necessarily represent in legal terms the official position of the European Commission. The European Commission assumes no responsibility or liability for any errors or inaccuracies that may appear

4 EXECUTIVE SUMMARY Avian Influenza (AI) is a highly contagious viral infection, which can affect all species of birds. Highly Pathogenic Avian Influenza (HPAI) can spread rapidly, causing serious disease with high mortality in many bird species and has so far comprised only H5 or H7 subtypes. The current H5N1 Highly Pathogenic Avian Influenza panzootic has affected over 60 countries across Asia, Africa and Europe, resulting in the loss of hundreds of millions of birds. Low Pathogenicity Avian Influenza (LPAI) can comprise subtypes H1 to H16 and usually causes a mild disease in poultry. LPAI strains of the H5 and H7 subtypes have the potential to mutate into HPAI following introduction to poultry populations. Birds of the Orders Anseriformes and Charadriiformes are thought to be the major reservoirs for LPAI viruses. Although historically HPAI infection has been rarely observed in wild birds and only in connection with poultry outbreaks, since the continuing outbreaks of H5N1 HPAI in Eurasia and Africa, wild birds have been implicated in the long distance spread of that virus. Therefore wild bird surveillance and the reporting of the results have become compulsory since 2005 in the European Union. The three main objectives of the surveillance are: The early detection of H5N1 HPAI in wild birds; the investigation of possible carrier or bridge species following an incident of H5N1 HPAI; and baseline monitoring of circulation of LPAI H5 and H7 strains in wild birds. The surveillance results reported here were collected between January and December 2008 according to EU harmonised guidelines. A total of 49,597 wild birds, from 26 Member States of the European Union were tested during the 2008 survey. In contrast to 2007, where cases in wild birds were reported from four Member States, in 2008 H5N1 HPAI was reported from only one Member State. The incident was detected through testing of samples from Mute Swans and a Canada Goose that were found dead or would have died if not euthanized on welfare grounds, and was limited in extent with H5N1 HPAI virus detected in a total of 11 birds over approximately six weeks in January and February. H5N1 HPAI infection was also identified in Mute Swans and Canada Geese in 2006 and However, the 2008 data suggest that the occurrence of the H5N1 HPAI incident in 2008 was the result of a new introduction rather than a continuous undetected circulation of the virus. Phylogenetic analysis showed that the HPAI virus found in 2008 could be clearly differentiated from those associated with previous incidents in poultry and wild birds in the EU. It appears possible that the 2008 HPAI virus arose from independent spill-over potentially from infected poultry populations in Turkey, Ukraine or the wider region into wild birds. LPAI of subtypes H5 or H7 was detected in 98 sampled birds from 11 Member States: Austria (3), Czech Republic (5), Germany (34), Denmark (20), France (8), Italy (5), Lithuania (2), Netherlands (6), Portugal (10), Sweden (1) and the United Kingdom (4). Consistent with previous years, the large majority of these infections were identified through active surveillance of key species, especially dabbling ducks (Anas spp.) and swans (Cygnus spp.). Evidence has accumulated for the ability of wild birds to transfer H5N1 HPAI from one area to another over hundreds of kilometres. However the exact species involved and the role of wild birds in maintaining the virus is not clear. H5N1 HPAI is currently circulating in poultry in Asia and Africa and was declared to be endemic in Egypt in mid The areas where H5N1 HPAI is currently circulating include wetlands on major water-bird migratory flyways where large numbers of birds will spend time before moving into the EU. The EU survey provides detection of AI incidents in wild birds, often not associated with outbreaks in poultry, illustrating the value and role of wild bird surveillance as a potential early detection and monitoring system for the presence of H5N1 HPAI and other AI viruses in the EU

5 GLOSSARY AND ABBREVIATIONS Table 1: Key to Member State abbreviations Abbreviation Country AT Austria BE Belgium BG Bulgaria CY Cyprus CZ Czech Republic DE Germany DK Denmark EE Estonia ES Spain FI Finland FR France GR Greece HU Hungary IE Ireland IT Italy LT Lithuania LU Luxembourg LV Latvia NL Netherlands PL Poland PT Portugal RO Romania SE Sweden SI Slovenia SK Slovakia UK United Kingdom Active surveillance: ADNS: AI: Bird Origin: Bridge Species: CRL: DG SANCO: EFSA: For the purpose of this report, active surveillance will be used as an equivalent to the surveillance birds that were live without clinical signs, hunted without clinical signs and hunted with clinical signs. Animal Disease Notification System Avian Influenza Relates to the collected information on the origin of the bird when sampled. The six categories are: live without clinical signs, live with clinical signs, injured, hunted without clinical signs, hunted with clinical signs and found dead. Birds sampled with no information on their origin were reported in the first quarter (Jan-Mar) prior to the onset of online submission. Species listed as those that may provide contact between risk species and poultry through sharing wetlands or farmlands with poultry (EFSA, 2006). Community Reference Laboratory Directorate General for Health and Consumers. European Food Safety Authority - 3 -

6 EU: EU 27: EURING code: HP: HPAI: Higher-Risk (HRS): Incident: LPAI: LPAI H5: LPAI H7: LPAI other: MS: NAI: NRL: NUTS: NUTS 3: Species European Union Refers to the 27 Member States of the European Union. European Union for Bird Ringing; A 5-digit number allocated to a species or subspecies of bird. Highly Pathogenic Highly Pathogenic Avian Influenza Species listed as those with an increased probability to contribute to the transmission of the Asian-lineage H5N1 HPAI viruses within Europe as defined in the scientific report by EFSA (EFSA, 2006) and the EC guidelines (EC, 2007) (in total 29 species). The EURING codes corresponding to these species can be found at: For the purpose of this report H5N1 HPAI cases were grouped into incidents based on proximity of 10km radius, which is equivalent to the size of monitoring areas (EC, 2006a). Low Pathogenic Avian Influenza birds positive for LPAI of subtype H5 birds positive for LPAI of subtype H7 birds reported as LPAI of other subtypes Member State(s) Notifiable Avian Influenza. Influenza A virus of subtypes H5 or H7 according to the OIE definition. National Reference Laboratory Nomenclature of Units for Territorial Statistics Nomenclature of Units for Territorial Statistics. At NUTS 3 level this refers to, for example, a region, district, county, municipal or unitary authority (depending on the MS). Other positives: Passive surveillance: PCR: Positive/ Infected: VI: birds positive for Avian Influenza but that were not clearly reported as either LPAI or HPAI. For the purpose of this report, passive surveillance will be used as an equivalent to the surveillance of birds that were live with clinical signs, injured and found dead. Polymerase chain reaction is a generic term for laboratory methodology that acts through the amplification of specific viral nucleic acid from clinical specimens. For the purpose of this report, a positive/ infected case of avian influenza is defined as a wild bird, from which at least one sample tested positive on either PCR or virus isolation. Virus isolation is a laboratory methodology that enables the propagation of infectious virus directly from clinical specimens

7 Table of contents Executive Summary... 2 Glossary and Abbreviations... 3 Introduction... 7 Objectives...7 Surveillance Programmes...7 Results... 8 Sampling...8 Overview...8 Geographical targeting...11 Seasonal targeting...13 Targeting of bird species...14 H5N1 HPAI Positives...15 Descriptive overview of H5N1 HPAI incidents in wild birds...15 Temporal pattern of H5N1 wild bird incidents...17 Origin of the HPAI H5N1 infected birds...18 Order and species of birds affected by HPAI H5N1 HPAI infections...18 Phylogenetic analysis of HPAI H5N1 viruses relevant to this surveillance...18 LPAI...20 Overview of LPAI...20 Timing of LPAI H5/H7 detections...22 Origin of LPAI H5/H7 positive birds...23 Order and species of birds positive for LPAI H5 H Discussion Materials and methods The testing of samples...27 Data processing...27 Data completeness (EU 27)...27 Species of birds...27 Bird origin...27 Subtype / pathotype information...27 Date of sampling...28 Spatial information...28 Assumptions...28 Phylogenetic analysis...28 References Annex I - Diagnosis H5N1 positives...32 Other positives (LPAI, including pathotype not reported, unidentifiable and pending)...32 Annex II Type of surveillance by quarter Annex III Proportion of higher-risk species sampled by MS and quarter Annex IV Overview of results for higher-risk and other species Annex V Scientific and English names of bird species

8 List of tables Table 1: Key to Member State abbreviations 3 Table 2: Birds sampled in 2008 by surveillance type and MS 9 Table 3: Bird Orders most frequently sampled in Table 4: Bird Species most frequently sampled in Table 5: Number and proportion of wild birds positive for H5N1 HPAI in Table 6: Details of H5N1 HPAI incidents in Table 7: Number tested and percentage positive for H5N1 HPAI in 2008 by Order 18 Table 8: Number of birds tested and number positive for H5N1 HPAI in 2008 by species 18 Table 9: Total number and proportion of birds testing positive for LPAI H5, H7 and other subtypes for MS in Table 10: Number and percentage of Birds positive for LPAI H5 or H7 by surveillance type in Table 11: Apparent prevalence of LPAI by order in EU MS in Table 12: Apparent prevalence of LPAI H5 by species in EU MS in Annex I - Table 1: Number and proportion of samples collected by Status of bird Annex I - Table 2: Test-results and samples taken of dead birds positive to H5N1 HPAI Annex I - Table 3: Test-results and samples taken for live birds without clinical signs positive to subtypes other than H5N1 HPAI (data April September) Annex I - Table 4: Test results and samples taken for hunted birds with and without clinical signs positive to subtypes other than H5N1 HPAI Annex I - Table 5: Test results and samples taken dead birds positive to subtypes other than H5N1 HPAI Annex II - Table 1: Number of birds tested in active surveillance by MS (live and hunted, healthy birds) Annex II - Table 2: Number of birds tested in passive surveillance by MS (injured, diseased and dead birds) Annex IV - Table 1: Number of high risk species birds sampled and number of positive results in each MS Annex IV - Table 2: Number sampled and positive by MS for birds not listed as high risk species Annex IV - Table 3: LPAI Other Positive Birds in Annex V - Table 1: High risk species as well as all other birds testing positive for AI in 2008 (English/Latin names) List of figures Figure 1: Total number of birds sampled in 2008 by EU Member State... 8 Figure 2: Proportion of birds sampled by surveillance type for all EU Member States Figure 3: Intensity of sampling by active surveillance (live and hunted birds) in EU MS in Figure 4: Intensity of sampling by passive surveillance (birds found dead, injured or live with clinical signs) in EU MS in Figure 5: Proportion of all birds sampled in 2008 by quarter and MS Figure 6: Proportion of birds sampled by quarter for the 26 MS during Figure 7: Total number of birds sampled by surveillance type during 2008 showing the incidents of H5N1 HPAI Figure 8: Sampling intensity and distribution of H5N1 HPAI incidents and LPAI detections (H5 and H7) in wild birds in EU MS in 2008 (Q1 data randomized at NUTS 0 level) Figure 9: Number of H5N1 HPAI incidents in wild birds and number of wild birds sampled in the EU by week in Figure 10: Neighbor-Joining phylogenetic tree of a 959 base pair HA1 fragment of H5 gene Figure 11: Number and week of detection of LPAI H5/ H7 positive birds by MS Figure 12: Number of LPAI H5 and H7 detections in wild birds and number of wild birds sampled in the EU by week in

9 INTRODUCTION Wild bird surveillance in the EU is aimed at identifying the risk of introduction of AI viruses (LPAI and HPAI) into domestic poultry (EC, 2007a). Voluntary surveillance for AI in wild birds in EU MS was first carried out in under Commission Decision 2002/649/EC (EC, 2002), although several MS had already been conducting wild bird surveillance prior to this. In response to the cases in wild birds and outbreaks in poultry and the evolving epidemiological situation of H5N1 HPAI principally in Asia, activities with regard to wild bird surveillance were increased. Wild bird surveillance became compulsory in 2005 for all MS and information collection on wild birds was extended and harmonised. In 2006, EFSA completed a scientific opinion on migratory birds and their possible role in the spread of HPAI (EFSA, 2006). This included an assessment of birds of predominantly the orders Anseriformes and Charadriiformes regarding their likelihood to introduce H5N1 HPAI following the criteria of gregariousness during migration/ wintering periods (group size and group density), degree of mixing during migration wintering periods, main habitat during migration/ wintering and degree of mixing with other species. This opinion has lead to inclusion of a higher risk species (HRS) list into the guidelines for targeting of surveillance. Objectives The main objective of this report is to present the surveillance results of 2008 and to discuss the main findings. The objectives of the EU wild bird AI surveillance are: Ensuring early detection of H5N1 HPAI by investigating increased incidence of morbidity and mortality in wild birds, in particular in selected higher risk species. In the event that H5N1 HPAI is detected in wild birds, then surveillance of live and dead wild birds shall be enhanced to determine whether wild birds of other species can act as asymptomatic carriers or bridge species. Continuing a baseline surveillance of different species of free-living migratory birds as part of continuous monitoring of LPAI viruses. Anseriformes (waterfowl) and Charadriiformes (shorebirds and gulls) shall be the main sampling targets to assess if they carry LPAI viruses of H5 and H7 subtypes (which would in any case also detect H5N1 HPAI and other HPAI, if present). Higher risk species must be targeted in particular (EC, 2007a). Surveillance Programmes In 2008 the surveillance programmes were performed according to the guidelines laid down in Commission Decision 2007/268/EC (EC, 2007a) which are available at the DG SANCO website under: &Submit22=GO For 2008, the surveillance programmes of the MS were evaluated and approved through Decisions 2007/782/EC (EC, 2007b) and 2008/920/EC (EC, 2008). Details of the survey programmes for each Member State are available at: Samples were tested in accordance with the diagnostic Manual for avian influenza (EC, 2006b)

10 RESULTS Sampling Overview During 2008, birds were sampled in the EU in 26 MS (Figure 1). Detailed figures regarding the number of birds sampled by MS in each quarter are displayed in Annex II. The MS with the highest number of birds tested in 2008 was DE (n=17810), which approximates to 2.5 times the number of birds sampled in the next most intensive programme (NL n=6769). Seventeen MS sampled fewer than 1000 birds throughout the year Birds sampled DE NL UK IT ES PL AT DK FR SE CZ LT GR PT LV BE SI RO IE BG CY FI SK LU HU EE Member State Figure 1: Total number of birds sampled in 2008 by EU Member State - 8 -

11 Table 2 displays the number of birds sampled in each MS during 2008 by type of surveillance. All MS conducted both active and passive surveillance, although the proportion of each varied highly between MS, reflecting results of the 2006 and 2007 surveillance. The percentage of each surveillance type is shown in brackets. Table 2: Birds sampled in 2008 by surveillance type and MS Number of Birds Member State Sampled Active surveillance Passive surveillance AT 2013 (4.1%) 1008 (50.1%) 1005 (49.9%) BE 481 (1.0%) 448 (93.1%) 33 (6.9%) BG 308 (0.6%) 143 (46.4%) 165 (53.6%) CY 268 (0.5%) 94 (35.1%) 174 (64.9%) CZ 814 (1.6%) 555 (68.2%) 259 (31.8%) DE (35.9%) (79.2%) 3707 (20.8%) DK 1416 (2.9%) 1342 (94.8%) 74 (5.2%) EE 36 (0.1%) 33 (91.7%) 3 (8.3%) ES 2334 (4.7%) 1771 (75.9%) 563 (24.1%) FI 168 (0.3%) 117 (69.6%) 51 (30.4%) FR 1361 (2.7%) 968 (71.1%) 393 (28.9%) GR 620 (1.3%) 408 (65.8%) 212 (34.2%) HU 38 (0.1%) 3 (7.9%) 35 (92.1%) IE 394 (0.8%) 289 (73.4%) 105 (26.6%) IT 4018 (8.1%) 3662 (91.1%) 356 (8.9%) LT 718 (1.4%) 630 (87.7%) 88 (12.3%) LU 68 (0.1%) 49 (72.1%) 19 (27.9%) LV 487 (1.0%) 478 (98.2%) 9 (1.8%) NL 6769 (13.6%) 6262 (92.5%) 507 (7.5%) PL 2052 (4.1%) 1934 (94.2%) 118 (5.8%) PT 585 (1.2%) 433 (74.0%) 152 (26.0%) RO 421 (0.8%) 339 (80.5%) 82 (19.5%) SE 1048 (2.1%) 941 (89.8%) 107 (10.2%) SI 480 (1.0%) 348 (72.5%) 132 (27.5%) SK 127 (0.3%) 83 (65.4%) 44 (34.6%) UK 4763 (9.6%) 3344 (70.2%) 1419 (29.8%) EU TOTAL (100%) (80.2%) 9812 (19.8%) - 9 -

12 Figure 2 displays the proportion of each surveillance type for the all 26 MS. More than 80% of the birds across the EU were sampled by active surveillance (either hunted or live without clinical signs). Passive Surveillance 19.8% Active Surveillance 80.2% Figure 2: Proportion of birds sampled by surveillance type for all EU Member States

13 Geographical targeting Figures 3 and 4 illustrate the distribution of active and passive surveillance respectively on a spatial scale by displaying the number of birds sampled. For all maps in the report the classification of sampling intensity is grouped by number of samples per 100 square Kilometres: Low <0.1, Medium 0.1-1, High 1-10, Intense >10. Figure 3: Intensity of sampling by active surveillance (live and hunted birds) in EU MS in

14 Figure 4: Intensity of sampling by passive surveillance (birds found dead, injured or live with clinical signs) in EU MS in 2008 All sampling is displayed at NUTS 3 level unless less than 80% of submissions gave valid spatial information (SE, IE). In this case sample numbers were displayed at randomised national level

15 Seasonal targeting Figure 5 displays the percentage of birds sampled by MS each quarter. Raw numbers of birds sampled by quarter and MS are shown below. Overall, across all participating MS similar numbers of birds were tested in each of the first three quarters with a significant increase in the final quarter. Temporal targeting of sampling varied among MS. While some MS increased the number of birds tested throughout the year other MS decreased the numbers. Belgium focused most of their survey effort in the first quarter whereas Estonia and Denmark s sampling efforts were much increased in the final quarter. 100% 90% 80% 70% 60% 50% 40% 30% 20% 10% 0% AT BE BG CY CZ DE DK EE ES FI FR GR HU IE IT LT LU LV NL PL PT RO SE SI SK UK EU Jan - Mar Apr - Jun Jul - Sep Oct - Dec Figure 5: Proportion of all birds sampled in 2008 by quarter and MS Figure 6 displays the percentage of birds sampled by quarter in MS. Very similar numbers of birds were tested in each of the first three quarters with the highest numbers being in the final quarter. Ja n - Mar 24.1% Oct - Dec 36.1% Apr - Jun 18.7% Ju l - Se p 21.2% Figure 6: Proportion of birds sampled by quarter for the 26 MS during

16 Figure 7 displays the overall number of birds sampled in MS throughout 2008 by surveillance type. Active surveillance was increased in the early autumn to winter months in Passive surveillance however remained at a relatively constant level throughout the year with only a slight increase during the autumn and winter. Outbreaks in poultry that occurred between January and February did not trigger an overall increase in passive surveillance of wild birds. Tables displaying the number of birds sampled according to surveillance type and MS are displayed in Annex II Number of birds sam JAN FEB MAR APR MAY JUN JUL AUG SEP OCT NOV DEC bird positive for H5N1 HPAIV Passive Active Figure 7: Total number of birds sampled by surveillance type during 2008 showing the incidents of H5N1 HPAI. (Each positive bird is indicated with a single red arrow). Targeting of bird species Table 3: Bird Orders most frequently sampled in 2008 Order Number sampled Species Table 4: Bird Species most frequently sampled in 2008 Number sampled Anseriformes Anas platyrhynchos Charadriiformes 4832 Anser anser 3211 Passeriformes 2194 Cygnus olor 2766 Gruiformes 1809 Larus ridibundus 2038 Falconiformes 1362 Anser albifrons 1653 Ciconiiformes 1081 Anas crecca 1616 Galliformes 765 Anas sp Columbiformes 694 Fulica atra 1179 Pelecaniformes 533 Anas penelope 1130 Strigiformes 430 Cygnus sp Cygnus cygnus 984 Branta canadensis 748 Anser cygnoides 675 Anser spp. 605 Anser fabalis

17 In total 45,923 birds of 23 orders and at least 366 species were sampled in Table 3 displays the ten most frequently sampled orders. As in 2006 and 2007, the three Orders in which most birds were sampled were: Anseriformes (Ducks, Geese and Swans), Charadriiformes (Gulls and waders) and Passeriformes (perching/ songbirds). Table 4 displays the top 15 species sampled in 2008 throughout all 26 MS. Mallards (Anas platyrhynchos) were the most frequently sampled species in 2008 (n=14340) as in 2006 and Greylag Goose (Anser anser) (n=3051) and Mute Swans (Cygnus olor) (n=2362) were also sampled in high numbers. The ten most frequently sampled species were all HRS. Figure 9 also indicates that the top 15 species account for over 80% of all birds tested in The large majority of non high risk species were sampled in very low numbers. H5N1 HPAI Positives A differentiation is made between H5N1 HPAI infections and LPAI. Unless otherwise stated, all references made to H5N1 refer to highly pathogenic H5N1. In total 11 cases of H5N1 HPAI infections were observed in 2008 in one MS. Overall 0.03% of the sampled birds tested positive for H5N1 HPAI. In the MS experiencing H5N1 infections, the proportion of H5N1 HPAI positive birds was 0.23% (UK) (Table 5). Table 5: Number and proportion of wild birds positive for H5N1 HPAI in 2008 Member State No. of birds sampled birds positive Percentage of sampled birds testing positive H5N1 HPAI UK % Table 6: Details of H5N1 HPAI incidents in 2008 Member State Incident Duration First species detected UK 07/01/08-25/02/08 Cygnus olor Number and species of birds H5N1 HPAI Positive (bird status ) 10 Cygnus olor (found dead) 1 Branta canadensis (found dead) Descriptive overview of H5N1 HPAI incidents in wild birds A single incident occurred in the UK in January - February of 2008, no further incidents in wild birds were observed. A single outbreak of H5N1 HPAI was reported in poultry in the EU in 2008, this occurred during October in Germany. Figure 8 displays the location of the H5N1 HPAI incidents in wild birds. Cases of wild birds were grouped into an incident if they were located within 10 km of each other. A single incident in wild birds occurred. More detailed information regarding how they were detected is displayed in Tables 5 and 6. The map also shows the location of H5/H7 LPAI findings in wild dealt with in the section on LPAI

18 Figure 8: Sampling intensity and distribution of H5N1 HPAI incidents and LPAI detections (H5 and H7) in wild birds in EU MS in 2008 (Q1 data randomized at NUTS 0 level)

19 Temporal pattern of H5N1 wild bird incidents The timing of the H5N1 HPAI incidents in wild birds presented in Figure 9 shows the number of wild bird H5N1 HPAI cases as well as the number of birds tested in the EU in 2008 by week. Number of positive samples Jan Feb Mar Apr May Jun Jul Aug Sep Oct Nov Dec 2008 CONFIRMED H5N1 HP Birds sampled Number of birds sampled Figure 9: Number of H5N1 HPAI incidents in wild birds and number of wild birds sampled in the EU by week in

20 Origin of the HPAI H5N1 infected birds All incidents in 2008 were detected through passive surveillance and all positive birds were found dead or would have died if not euthanized on welfare grounds. In 2006 and 2007 all incidents were detected through passive surveillance and the majority of birds were found dead. Order and species of birds affected by HPAI H5N1 HPAI infections Table 7 shows the Order of birds in which H5N1 HPAI cases were found in 2008 and the apparent prevalence. Anseriformes were the only order in which H5N1 HPAI was detected. Anseriformes also had positives in 2006 and Table 7: Number tested and percentage positive for H5N1 HPAI in 2008 by Order Order Total number tested Number positive for H5N1 HPAI (percentage of birds testing positive) Anseriformes (0.032%) Table 8 below displays the apparent overall prevalence of H5N1 HPAI in the two species in which H5N1 HPAI was detected. Detailed information regarding the number of birds tested and positive birds by MS and species that were either of the HRS or tested positive for H5N1 HPAI or LPAI H5/H7 is displayed in Annex IV. Table 8: Number of birds tested and number positive for H5N1 HPAI in 2008 by species Species Total number tested Number positive for H5N1 HPAI (percentage of birds testing positive) Cygnus olor (mute swan) (0.36%) Branta canadensis (Canada goose) (0.13%) Phylogenetic analysis of HPAI H5N1 viruses relevant to this surveillance Phylogenetic analysis is an invaluable tool for studying precise relationships and addressing the question of long term persistence or new introduction of viruses that may then spread to the European Union. The analysis presented here supports a new introduction of virus rather than virus persisting through continuous circulation at undetected levels in wild bird populations. The analysis is supported by preliminary information using molecular clock methods, whereby more precise inferences can be made on the relationship between viruses based on exact dates of sampling in the field. This analysis underpins the observation that the H5N1 viruses present in wild birds in the UK in early 2008 shared a common progenitor with viruses present in poultry populations in Turkey and Ukraine in the same period. Whilst these viruses are closely related to strains from elsewhere in the EU, identified in 2007, which themselves represented a new introduction of virus, they can still be clearly differentiated. These data indicate that the virus present in the single EU wild bird incident can be clearly differentiated from clade 2.2 viruses present in Africa and the Far East but more closely related to contemporary viruses from Central Asia/Eastern Europe. The virus detected in the UK incident was less similar to virus detected in poultry in Germany in 2008 which showed a higher degree of similarity to that which was detected in poultry and wild birds in These results raise the possibility of a complex dynamic whereby the virus may not be maintained in wild bird populations but may spread to wild birds through frequent and persistent exposure to poultry populations that may be endemically or frequently infected, especially in closer geographical proximity to the EU

21 A/swan/Austria/AV917(P4227)-7/2006 A/swan/Czech/AV1324/06 A/chicken/Ukraine/AV6766-1/2006 A/goose/Greece/AV /2006 A/pochard/France/AV (2)/2006 A/Swan/Slovenia/AV136/2006 A/chicken/Nigeria/SO493/2006 Wild Birds 2008 Domestic Wild Domestic Wild Domestic Birds birds Birds Birds birds EU incidents 100 A/chicken/Egypt/1081-NAMRU3/ A/duck/Egypt/1301-NAMRU3/2007 A/chicken/Israel/1055/2008 A/pigeon/Turkey/AV73-14/ A/chicken/Turkey/AV73-16/06 A/goose/Hungary/AV232-2/ A/turkey/ England/250/2007 A/turkey/England/ /2007 A/chicken/Burkina Faso/AV /2006 A/chicken/Sudan/ / A/turkey/IvoryCoast/4372-4/ A/chicken/Burkina Faso/AV1583-1/2007 A/Swan/Scotland/1430/06 95 A/goose/Hungary/AV /2006 A/chicken/Hungary/AV6740/2006 A/common coot/switzerland/v544/ A/swan/Bavaria/21/2006 A/turkey/France/AV1142-1/2006 A/duck/Germany/R839/2008* A/black swan/germany/r1970-1/2006 A/tufted duck/germany/r1240/2006 A/chicken/Romania/AV170-2/06 A/peacock/Denmark/AV6014/2006A A/buzzard/Denmark/AV1075/2006 A/chicken/Albania/AV1079-1/2006 A/Turkey/Germany/Av1077/06 A/swan/Serbia/AV (1)/ A/chicken/Serbia/AV (2)/ A/swan/Azerbaijan/AV107-k2-2/2006 A/chicken/Moscow-2/ A/chicken/India/Jalgaon/8824/2006 A/chicken/Afghanistan/ /2006 A/common goldeneye/mongolia/12/2006 A/goose/Turkey/AV972-10/ A/ck/Bangladesh/FD(J)-2550BL/411/ A/chicken/Assam/140187/2008 A/ck/Bangladesh/AV543-10/ A/mute Swan/France/07 72 A/cygnus olor/germany/r1359/ /2007 A/chicken/Czech/AV1787-1/2007 A/turkey/England/AV /2007 A Chicken/Romania/10705/ A/chicken/Ukraine/TR 67 / A/chicken/Turkey/TR 67 Merkez-2/2008 A/swan/England/26-70/2008 A/goose/England/AV /2008 A/chicken/Romania/AV /2006 A/chicken/Romania/AV5780-1/ A/goose/Romania/AV (3)/2006 A/Crow/Myanmar/1661/07 A/ck/Anglong/NIAH101204/ A/Chicken/ Uthai Thani/NIAH115067/ Figure 10: Neighbor-Joining phylogenetic tree of a 959 base pair HA1 fragment of H5 gene

22 LPAI This section focuses on the analysis of H5/H7 LPAI unless specifically mentioned. Since some positive birds were reported with pathotype unknown, pending, missing etc, birds that tested positive on PCR or virus isolation are reported in four groups in this section: 1) LPAI H5 are birds positive for LPAI H5 2) LPAI H7 are birds positive for LPAI H7 3) LPAI other are birds reported as LPAI of other H subtypes 4) Other positives are birds positive for influenza A by PCR or Virus isolation but that were not clearly reported as either LPAI or HPAI. Overview of LPAI In total 1441 birds tested positive for subtypes other than H5N1 HPAI. LPAI H5 was detected in 71 birds from 8 MS: CZ (5) DE (34), DK (6), FR (7), IT(2), NL (4), PT (9) and UK (4). LPAI H7 was identified in 27 birds from 8 MS: AT (3) DK (14), FR (1), IT (3), LT (2), NL (2), PT (1) and SE (1). LPAI of other subtypes was detected in 614 birds. Other positives were detected in 729 birds. Table 9 indicates the total number and proportion of wild birds testing positive for LPAI H5, LPAI H7, other LPAI cases and other positives by Member State. Figure 15 maps the geographical distribution of LPAI H5 and H7 positives. Overall a very low apparent prevalence of LPAI H5 of 0.14% was detected, which reflects, in spite of minor variations at MS level, the findings of 2006 and 2007 when an overall LPAI H5 apparent prevalence of 0.11% and 0.13% were observed respectively (Annex 2 b). The highest apparent prevalence of LPAI H5 was found in PT. Cases of LPAI H7 found in 2008 in the EU increased slightly to 0.05% from 0.02% (2006) and 0.01% (2007). Table 9 shows the total number and proportion of birds that tested positive for LPAI H5, H7 and other subtypes in those MS that detected LPAI cases during

23 Table 9: Total number and proportion of birds testing positive for LPAI H5, H7 and other subtypes for MS in 2008 Member State Total number of birds sampled Number of H5 LPAI cases (proportion of total sampled) Number of H7 LPAI cases (proportion of total sampled) Other LPAI cases (proportion of total sampled) "Other positives" with pathotype unspecified (proportion of total sampled) AT (0.15%) - 79 (3.92%) BE (8.11%) BG (0.65%) CZ (0.61 %) - 52 (6.39%) - DE (0.19%) - 46 (0.26%) 124 (0.70%) DK (0.42%) 14 (0.99%) 57 (4.03%) - ES (0.3%) FI (5.36%) 5 (2.98%) FR (0.51%) 1 (0.07%) (14.84%) IE (0.51%) 2 (0.51%) IT (0.05%) 3 (0.07%) 27 (0.67%) 86 (2.14%) LT (0.28%) 85 (11.84%) 1 (0.14%) LV (0.21%) NL (0.06%) 2 (0.03%) 336 (4.96%) 72 (1.06%) PL (0.39%) PT (1.54%) 1 (0.17%) - - SE (0.1%) - 67 (6.39%) SI (2.72%) UK (0.08%) (0.44%) EU TOTAL (0.14%) 27 (0.05%) 614 (1.24%) 729 (1.47%)

24 Timing of LPAI H5/H7 detections Figure 11 displays the calendar week of LPAI H5 and H7 detections by MS. Figure 12 displays the number of LPAI H5/H7 detections and the number of birds sampled by week. LPAI H5 was mainly found in the autumn, while LPAI H7 infections occurred throughout the year but were mainly found earlier in the year. JAN FEB MAR APR MAY JUN JUL AUG SEP OCT NOV DEC MS AT 2 1 CZ 5 DE DK FR IT LT 1 1 NL PT SE 1 UK Figure 11: Number and week of detection of LPAI H5/ H7 positive birds by MS Number of positive samples Jan Feb Mar Apr Ma Jun Jul Aug Sep Oct Nov Dec Number of birds sampled H5 H7 Birds sampled 2008 Figure 12: Number of LPAI H5 and H7 detections in wild birds and number of wild birds sampled in the EU by week in

25 Origin of LPAI H5/H7 positive birds Table 10 displays LPAI H5/H7 cases detected by MS and shows the percentage of these cases detected by surveillance type. Very few LPAI H5/H7 positive birds were identified through passive surveillance. Active surveillance accounted for the vast majority of LPAI cases during Member State Table 10: Number and percentage of Birds positive for LPAI H5 or H7 by surveillance type in 2008 Total number of birds sampled Total number of H5 / H7 LPAIV Positive birds Percentage of birds positive for LPAIV H5 or H7 Proportion of LPAIV H5 and H7 detected by Active surveillance (n= total number of birds actively sampled) Proportion of LPAIV H5 and H7 detected by Passive surveillance (n= total number of birds passively sampled) AT % 0.30% (n=1008) 0% (n=1005) CZ % 0.90% (n=555) 0% (n=259) DE % 0.23% (n=14103) 0.03% (n=3707) DK % 1.49% (n=1342) 0% (n=74) FR % 0.83% (n=968) 0% (n=393) IT % 0.14% (n=3622) 0% (n-356) LT % 0.32% (n=630) 0% (n=88) NL % 0.10% (n=6262) 0% (n= 507) PT % 2.31% (n=433) 0% (n=152) SE % 0.11 (n=941) 0% (n=107) UK % 0.12% (n=3344) 0% (n=1419) EU TOTAL % 0.26% (n=39785) 0.01% (n=9812) Order and species of birds positive for LPAI H5 H7 Order LPAI H5 was found in Anseriformes and Gruiformes, while LPAI H7 was exclusively observed in Anseriformes. Other positives were observed in a Ciconiiformes, Charadriformes and Passeriformes (Table 11). Order Table 11: Apparent prevalence of LPAI by order in EU MS in 2008 Total sampled Birds Positive for LPAI H5 Birds Positive for LPAI H7 Other LPAI Positive Birds "Other Positive" birds pathotype undetermined Anseriformes (0.20%) 27 (0.08%) 605 (1.76%) 559 (1.61%) Charadriiformes (0.14%) 92 (1.90%) Ciconiiformes (6.29%) Passeriformes (0.05%) Gruiformes (0.11%) 9 (0.50%)

26 Species Further details and tables regarding sampling and results for high-risk and other species by MS can be found in Annex IV. All observations of LPAI H5 in 2008 were made in dabbling ducks and swans (Table 12). With the exception of two species (Anser albifrons and Aythya fuligula) all observations of LPAI H5 in 2006 and 2007 were also made in dabbling ducks and swans. Most observations of LPAI H5 were made in Mallards (Anas platyrhynchos). In all species in 2008 the percentage detected was below one percent, which is in line with the detection rate in both years 2006 and All detections of LPAI H7 were also made only in dabbling ducks and swans in 2008 (Table 12). Results from 2007 were also limited to dabbling ducks and swans. In total 33 species tested positive for AI subtypes other than HPAI. Of these species, 19 were Anseriformes. Nine species were in the order Charadriformes with two species coming from both Ciconiiformes and Gruiformes and just one species in the order Passeriformes. Species Table 12: Apparent prevalence of LPAI H5 by species in EU MS in 2008 Total birds sampled Total Birds LPAIV H5 Positive Total Birds LPAIV H7 Positive Total positive for AIV (excluding HPAI) Anas crecca (0.43%) 1 (0.06%) 86 (5.32%) Anas platyrhynchos (0.39%) 24 (0.16%) 949 (6.13%) Anas penelope (0.09%) 49 (4.34%) Cygnus olor (0.11%) 1 (0.04%) 14 (0.51%)

27 DISCUSSION Avian Influenza (AI) is a highly contagious viral infection, which can affect all species of birds. Highly Pathogenic Avian Influenza (HPAI) can spread rapidly causing serious disease with high mortality in many poultry species and has so far comprised only H5 or H7 subtypes of influenza A. The current H5N1 HPAI epizootic has affected over 60 countries across Asia, Africa and Europe, resulting in the loss of hundreds of millions of birds. Historically HPAI infection has been rarely observed in wild birds and then only in connection with poultry outbreaks. However, since the current H5N1 HPAI epizootic, wild birds have been implicated in the long distance spread of this virus. However the exact species involved and the role of wild birds in maintaining the virus is not clear. This virus is currently circulating in poultry in parts of Eurasia and Africa and was declared to be endemic in Egypt in mid Egypt is on major water-bird migratory flyways and is less than 400km from the EU. Low Pathogenicity Avian Influenza (LPAI) can comprise subtypes H1 to H16 and usually causes a mild disease in poultry. LPAI strains of the H5 and H7 subtypes have the potential to mutate into HPAI following introduction to poultry populations. Birds of the Orders Anseriformes and Charadriiformes are thought to be the major reservoirs for LPAI viruses. The first objective of AI surveillance in wild birds in the EU is to ensure early detection of H5N1 HPAI through the investigation of increased mortalities. The detection of incidents in wild birds without outbreaks in poultry illustrates the value and role of wild bird surveillance in the early detection of the presence of H5N1 HPAI in a country. Detections of such infections require the implementation of control measures (EC 2006a) and are important to maintain and raise vigilance amongst the poultry sector, especially keepers of free-range poultry. The second objective of EU wild bird surveillance relates to the investigation of asymptomatic carriers and bridge species in the event of H5N1 HPAI detection. Evaluation of the importance of carrier and bridge species is perhaps best fully investigated through a multidisciplinary approach with ornithologists, ecologists, virologists and epidemiologists; since such an approach can evaluate factors such as size of local populations, local and migratory bird movements and functional links with the affected site, all of which are relevant to the principle of bridge species. Information collection and dissemination on the epidemiology of wild bird incidents and influential factors such as details of resident wild bird populations and environmental sampling could play an important role in increasing the knowledge on the epidemiology of H5N1 HPAI in wild birds. The final objective of the EU surveillance, the baseline surveillance of LPAI H5 and H7 in wild birds appears to be best addressed through active surveillance of live birds, as consistent with previous years, the large majority of LPAI infections in 2008 were identified through active surveillance of High Risk Species, especially dabbling ducks and swans. A total of 49,597 wild birds, from 26 Member States of the European Union were tested during the 2008 survey. In contrast to 2007, where cases in wild birds were reported from four MS, in 2008, H5N1 HPAI was reported from only one MS. The incident was detected through testing of samples from Mute swans (Cygnus olor) and a Canada goose (Branta canadensis) that were found dead or would have died if not euthanized on welfare grounds. The incident was limited in size and duration with H5N1 HPAI virus detected in a total of 11 birds over approximately six weeks in January and February. H5N1 HPAI infection was also identified in mute swans and Canada geese in 2006 and However, the 2008 data suggest that the occurrence of the H5N1 HPAI incident in 2008 was the result of a new introduction rather than a continuous circulation of the virus. While the number of samples is too small to show a clear epidemic curve, the short temporal duration of the incident is consistent with a single introduction event, followed by infection which died out rapidly. In the 2008 incident, apparently healthy mute swans (n = 71), mallards (n = 68) and coots (n = 48) were tested with negative results for H5N1 HPAI. Well defined data were available on the local population size and movement, migratory behaviour and habitat use which together with epidemiological data suggested very localised spread of infection within the wild bird populations (VLA unpublished data, manuscript in preparation). If H5N1 HPAI had remained in the EU since 2007, it seems likely that other detections in wild birds would have occurred during the surveillance programme. Also phylogenetic analysis showed that the HPAI virus found in 2008, although closely related, could be clearly differentiated from those strains associated with previous incidents in poultry and wild birds in the EU. It appears possible that the 2008 HPAI virus arose from independent spill-over potentially from infected poultry populations in Turkey, Ukraine or the

28 wider region into wild birds (see Figure 12). Phylogenetic analysis of recent H5N1 HPAI viruses also indicates that an outbreak in poultry that occurred in Germany in 2008 was genetically very similar to the H5N1 virus detected in a Tufted duck (Aythya fuligula) in Germany in 2006 and raises interesting questions and perspectives for the apparent maintenance of such viruses essentially unchanged over two years (see Figure 12). LPAI of subtypes H5 or H7 was detected in 98 birds from 11 Member States: Austria (3), Czech Republic (5), Germany (34), Denmark (20), France (8), Italy (5), Lithuania (2), Netherlands (6), Portugal (10), Sweden (1) and the United Kingdom (4). Consistent with previous years, the large majority of these infections were identified through active surveillance of High Risk Species, especially dabbling ducks (Anas spp.) and swans (Cygnus spp.). There are several aspects of the data that should be considered when interpreting the results: cluster effects occur and were not accounted for in the analysis. In addition when small sample numbers are collected for a MS or for a species, the uncertainty around a proportion positive increases. Surveillance programmes were quite variable between MS in respect to a number of parameters including sample size, weighting between active and passive surveillance and targeting of species and areas. Therefore, as with most surveillance data collected through various sources with heterogeneous design, limited direct comparisons can be made regarding the prevalence between MS and the apparent prevalence observed in a species that cannot be assumed to be the true underlying prevalence in the population. In 2008, as for 2006 and 2007, almost all of the detections of H5N1 HPAI were made through the testing of dead birds via passive surveillance. Passive surveillance appears more sensitive for the detection of HPAI, which is intuitive as birds may be more likely to die from infection with a virus of higher pathogenicity as has been demonstrated for a variety of bird species. However, it should be recognised that the dynamic is complex and influenced by a number of factors, such that the direct consequence of infection in some species will not necessarily result in mortality. Active surveillance appears more sensitive than passive surveillance for the detection of LPAI, which could be interpreted as suggesting that infection with an LPAI virus is somehow protective against mortality. A more plausible explanation however, is that localised infection with LPAI virus is in contrast to the systemic infection that occurs with HPAI infection resulting in a significantly lower detectability of LPAI virus in birds found dead. Experimental infections have shown that swans that were serologically positive to LPAI of a subtype other than H5 did not show clinical signs when consequently infected with H5N1 HPAI i.e. heterosubtypic immunity can occur. Although many factors such as bird density and conditions for virus survival are likely to influence the scale of observed mortalities, previous infection with an unrelated LPAI could be another important influential factor. This may mean that in areas where a high exposure to LPAI is thought to occur, mortality events due to HPAI are less likely. However, more knowledge is required on the level of clinical cross-protection provided by different LPAI strains for infection with H5N1 HPAI. The proportion of all wild bird samples that yielded any AI virus is small (around 2-3%) and a minority are HPAI or LPAI H5 or H7; the targets of this surveillance. Therefore, the majority of the resource dedicated to wild bird surveillance detects an absence of infection in individual birds. While this negative data is of value, the extremely low proportion of wild bird populations that are sampled allows only modest confidence in determining a population to be free of infection at any point in time. Surveillance efficacy for H5N1 HPAI could be improved by the targeting of passive surveillance through combining existing wild bird ecological and migratory data with AI detection data to determine areas, times and species for surveillance. This approach, utilising information on the host, environment and agent, can help resolve the otherwise apparently sporadic distribution of AI. A further action to improve targeting of passive surveillance could be to use existing migratory information in conjunction with outbreak data to identify priority areas for surveillance in real time. If, for example, an outbreak occurs in an area neighbouring the EU or within the EU, migratory data such as available through the BTO migration mapping tool (Atkinson et al., 2007) could be used to identify areas of increased vigilance for passive surveillance, based upon the areas and times migratory species are likely to arrive from affected regions. Also, following the detection of an HPAI outbreak within or outside the EU, increased surveillance could be conducted based on the locations and times that migratory species are likely to arrive from the affected area and the proximity of these species to poultry holdings. However, as some migratory species introducing the virus to the EU may not die or display clinical signs when infected, a balance of passive and active surveillance may continue to prove valuable in the future. A current EU research project New Flubird ( is implementing targeted surveillance for early detection incorporating data on bird migration patterns and movement dynamics and will provide further insights into this type of approach