Scientific Opinion on bluetongue monitoring and surveillance 1

Transcription

1 EFSA Journal 2011;9(6):2192 SCIENTIFIC OPINION Scientific Opinion on bluetongue monitoring and surveillance 1 EFSA Panel on Animal Health and Welfare 2,3 European Food Safety Authority (EFSA), Parma, Italy ABSTRACT Following a request from the Commission, the Panel on Animal Health and Welfare was asked to deliver a Scientific Opinion on: 1) the expected prevalence (design prevalence) under different circumstances, and, 2) an updated scientific assessment of the size of the relevant geographical area for the purpose of monitoring and surveillance programmes for bluetongue. A systematic literature review and a review of monitoring and surveillance data from European Union Member States was performed in order to estimate the prevalences observed in the Member States. The prevalences observed in areas that have been infected for several years were slightly lower than the design prevalence of 2 % currently used for monthly testing of sentinel animals, but much lower than the design prevalences of 20 % and 10 % for annual surveys in populations of unvaccinated and vaccinated ruminants, respectively. Currently there is no scientific evidence that suggests an optimal size of the relevant geographic unit for BTV monitoring and surveillance, since it depends on many factors, including the goal of the surveillance programmes. Early warning based on passive surveillance will take place irrespective of the size of the geographical unit but, when based on active surveillance, it is best targeted at regions considered at risk for introduction, using small geographical units, a high sampling frequency and sample size. For estimating the impact of interventions on the prevalence of infected animals, smaller areas result in more precise estimates of the prevalence and also take better account of local differences. For establishing freedom from infection, smaller areas result in lower design prevalence for a region as a whole and take better account of local differences in infection dynamics. European Food Safety Authority, 2011 KEY WORDS Bluetongue, epidemiology, monitoring and surveillance, expected prevalence, relevant geographical area 1 On request from the European Commission, Question No EFSA-Q , adopted on 16 May Panel members: Anette Bøtner, Donald Broom, Marcus G. Doherr, Mariano Domingo, Jörg Hartung, Linda Keeling, Frank Koenen, Simon More, David Morton, Pascal Oltenacu, Albert Osterhaus, Fulvio Salati, Mo Salman, Moez Sanaa, James M. Sharp, Jan A. Stegeman, Endre Szücs, Hans-H. Thulke, Philippe Vannier, John Webster and Martin Wierup. Correspondence: ahaw@efsa.europa.eu. 3 Acknowledgement: The Panel wishes to thank the members of the Working Group on bluetongue monitoring and surveillance: Arjan Stegeman (chair), Annette Bøtner, Giovanni Savini, Jordi Casal, Rudolf Meiswinkel, Yves van der Stede for the preparatory work on this Scientific Opinion and the hearing expert Jean-Philippe Amat and EFSA staff Andrea Gervelmeyer, Sofie Dhollander, Diane Lefebvre, Francesca Riolo and Jane Richardson for the support provided to this scientific opinion. Suggested citation: EFSA Panel on Animal Health and Welfare (AHAW; Scientific Opinion on Bluetongue monitoring and surveillance. EFSA Journal 2011;9(6):2192. [61 pp.] doi: /j.efsa Available online: European Food Safety Authority, 2011

2 SUMMARY Following a request from the Commission, the Panel on Animal Health and Welfare was asked to deliver a Scientific Opinion on: 1) the expected prevalence (design prevalence) under different circumstances, and, 2) an updated scientific assessment of the size of the relevant geographical area for the purpose of monitoring and surveillance programmes for bluetongue. Five epidemiological phases of a bluetongue virus (BTV) infection in a population were distinguished, each with a specific goal for monitoring and surveillance (Table 1). Phase 1 is a BTV free population without a history of infection (i.e. fully susceptible). Upon introduction of BTV, virus transmission will result in a rise in the prevalence of BTV positive animals (phase 2), followed by a rise in the prevalence of seropositive animals until it plateaus (phase 3). In phase 4, the prevalence drops again to an endemic equilibrium or to zero. In the latter case, the area is again free from BTV infection, but at this point there is a history of infection (phase 5). This pattern will also be reflected in the prevalence of virus positive animals and (to a lesser extent) seropositive animals. Moreover, because there are many BTV serotypes, an area can be in more than one phase at the same time, for example, in phase 4 for BTV-1 and in phase 1 for the other serotypes. The expected prevalence in a region infected for several years (phase 4) is a suitable target for the design prevalence to declare freedom from infection in phase 5, whereas the expected plateau prevalence (phase 3) is a useful target for the design prevalence to demonstrate a region is infection free once a large part of the population becomes susceptible to infection again. To provide scientific advice on the expected prevalence (design prevalence) under different circumstances To obtain estimates of the expected prevalences, a systematic literature review (SLR) and a review of monitoring and surveillance data from European Union (EU) Member States were performed in order to obtain the prevalences observed in the Member States. While prevalences related to BTV-8 were overrepresented in the SLR, the data from the Member States had a focus on other serotypes, thus this Opinion addresses the expected prevalence for BTV-8 as well as for other BTV serotypes. On average, the median of the observed prevalences was 0.02 in epidemic phase 2 (Table 1), which indicated that new BTV incursions were more likely to be detected by passive surveillance. In phase 3, the median of the observed prevalences was 0.3 (0.38 in the SLR and 0.24 in Member States data). Here, the prevalence of BTV-8 infected ruminants in North-western Europe was markedly higher than that of other serotypes in Southern Europe. In phase 4, the median of the observed prevalences was and in vaccinated and unvaccinated populations, respectively. Although Culicoides seems to have a higher preference for cattle than for sheep, this assessment showed no clear indications that the expected prevalence was substantially different in cattle to that in small ruminants. In addition, the relation between ruminant density and BTV prevalence has not been established, although indoor housing appears to be a prevalence reducing factor. Moreover, it is not possible to advise differences on expected prevalence for the different epidemiological phases based on the current knowledge of the distribution and abundance of different vector species in EU Member States. The reason is that a vector system exists across Europe within which various serotypes of BTV can be transmitted efficiently both in the presence and absence of C. imicola and, moreover, absolute vector abundance cannot be used as a reliable parameter. EFSA Journal 2011;9(6):2192 2

3 The prevalences observed in areas that have been infected for several years (phase 4) were slightly lower than the design prevalence of 2 % currently used for monthly testing of sentinel animals, but much lower than the design prevalences of 20 % and 10 % for annual surveys in populations of unvaccinated and vaccinated ruminants, respectively. Although repeated testing obviously increases the power of the current monitoring and surveillance programmes for demonstrating freedom from infection, it seems appropriate to reduce the currently used design prevalence for the annual surveys to demonstrate such freedom. To provide an updated scientific assessment of the size of the relevant geographical area for the purpose of monitoring and surveillance programmes for bluetongue. Bluetongue monitoring and surveillance programmes take the geographical area as the relevant epidemiological unit, rather than the farm. However, there is no scientific evidence that suggests an optimal size of the relevant geographic unit for BTV monitoring and surveillance. This is because it depends on many factors, including the goal and the design of surveillance programmes, as well as the dynamics of the infection in the area which are in turn influenced by the density of the vector and the host, and the interactions between them. Moreover, although the interaction of climatic, vegetation and topographical factors influence the population dynamics of BTV infections, their relation to the optimal size of the relevant geographical unit for the purpose of monitoring and surveillance has not yet been established. Table 1 summarizes the importance of the size of the geographical region in each of the epidemic phases. According to the rate of spatial spread of BTV, the size of the currently defined geographical unit for monitoring and surveillance (45 km x 45 km) is sufficient to encompass local spread of BTV infection for a few weeks. However, for early warning based on passive surveillance the size of the geographical unit is irrelevant. If active surveillance is considered for the purpose of early warning, it is best targeted at regions considered at risk from introduction, using small geographical units in addition to a high sampling frequency and sample size. For estimating the impact of interventions on the prevalence of infected animals, smaller areas result in more precise estimates of the prevalence than large regions and also take better account of local differences. For establishing freedom from infection, smaller areas result in lower design prevalence for a region as a whole and take better account of local differences in infection dynamics. In the case of effective vaccination, enlarging the geographical unit for monitoring and surveillance might be an option, however, this can only be considered if the infection is homogeneously distributed across the zone. Where this distribution is heterogeneous or unknown, smaller units provide better precision of the actual prevalence. Since there are many factors involved in a monitoring and surveillance programme, developing a mathematical model is recommended in order to evaluate the current monitoring and surveillance programme, taking into account the dynamics of infection, the design of the programme (including sampling frequency, sample size, repeated testing and test characteristics) and the geographical unit. EFSA Journal 2011;9(6):2192 3

4 Table 1: Phases of BTV infection in a population, their objectives and requirements for monitoring and surveillance related to their expected prevalence and the size of the relevant geographical unit EFSA Journal 2011;9(6):2192 4

5 TABLE OF CONTENTS Abstract... 1 Summary... 2 Table of contents... 5 Background as provided by the European Commission... 6 Terms of reference as provided by the European Commission... 7 Assessment Introduction Phases of BTV infection in a population and possibly related aspects of vector and ruminant host Course of BTV infection in a population Culicoides Northward movement of C. imicola Possible sub-episystems in Europe Vector abundance Ruminant species and density Diagnostics for detection of antibodies and virus Clinical signs Virus detection Antibody detection Expected prevalence Systematic literature review data Methodology Prevalence at the animal level Prevalence at the herd level Data from EU Member States Prevalence at the animal level Prevalence at the herd level Size of the relevant geographical area for the purpose of monitoring and surveillance Spatial spread of bluetongue virus Spatial spread Associated factors Geographical unit related to the phases of BTV infection in a population Relevant geographical unit in phase Relevant geographical unit in phase Relevant geographical unit in phase Relevant geographical unit in phase Relevant geographical unit in phase Conclusions and Recommendations References Appendices Appendix 1: Model used to combine the ruminant density and the number of days with an average temperature >15 C shown in Figure Appendix 2: Protocol for Systematic Literature Review on monitoring and surveillance of bluetongue Appendix 3: Diagrams showing the distribution of the prevalences and their 95 % CI (exact binomial) obtained by data extraction of the papers from the SLR Glossary and abbreviations EFSA Journal 2011;9(6):2192 5

6 BACKGROUND AS PROVIDED BY THE EUROPEAN COMMISSION Commission Regulation (EC) No 1266/2007 of 26 October 2007 on implementing rules for Council Directive 2000/75/EC as regards the control, monitoring, surveillance and restrictions on movements of certain animals of susceptible species in relation to bluetongue 4, provides for a detailed regulatory framework on bluetongue that entered into force on 2 November Aiming for sustainable, proportionate and science-based rules, the Regulation has been amended nine times following its first publication. Since 2007, EFSA has published one epidemiological analysis 5 and four Scientific Opinions on bluetongue 6,7,8,9 which support the Commission in the process of legislative decision making and risk management. In the past 10 years, bluetongue has evolved from an 'exotic disease' to being a more widespread disease situation with the potential of becoming endemic in certain areas. However, at the same time, in the last few years in many areas the situation has significantly improved, due to massive vaccination campaigns using recently developed inactivated vaccines. There are still some gaps in the scientific knowledge on bluetongue disease. Scientific advice is needed to assure that the rules on bluetongue are effective, and at the same time proportionate. International standards are taken into consideration to reduce as far as possible obstacles to trade that bluetongue may cause while maintaining the adequate level of guarantees. Taking this into account, the Commission requested recommendations on epidemiological parameters, such as expected prevalence under different circumstances and the size of a relevant geographical area for the purpose of monitoring and surveillance programmes for bluetongue. For the purpose of a review of Annex I (Minimum requirements for bluetongue monitoring and surveillance programmes) from Commission Regulation (EC) No 1266/2007, the Commission was in need of scientific advice on key epidemiological parameters, such as expected prevalence under different epidemiological circumstances and the appropriate size of a relevant geographical area for the purpose of monitoring and surveillance programmes for bluetongue. Geographic unit of reference Based on past experience with bluetongue surveillance, point 1 of Annex I describes a "geographic unit of reference for the purpose of bluetongue monitoring and surveillance as a grid of 45 x 45 km, (approximately 2,000 km 2 ) unless specific environmental conditions justify a different size". Member States may also use the "region" as defined in Article 2(p) of Directive 64/432/EEC as the geographical reference for monitoring and surveillance purposes. The EFSA was requested to review these criteria on geographical units. 4 OJ L 283, , p EFSA (European Food Safety Authority), 2007a Technical report on epidemiological analysis of the 2006 bluetongue virus serotype 8 epidemic in north-western Europe. EFSA Journal, 5(4), 42 pp 6 EFSA Panel on Animal Health and Welfare (AHAW), 2007b. Scientific Opinion on bluetongue vectors and vaccines. EFSA Journal, 479, EFSA Panel on Animal Health and Welfare (AHAW), 2007c. Scientific Opinion on the EFSA self-mandate on bluetongue origin and occurrence. EFSA Journal, 480, EFSA Panel on Animal Health and Welfare (AHAW), 2008a. Scientific Opinion on a request from the European Commission (DG SANCO) on bluetongue. EFSA Journal, 735, EFSA Panel on Animal Health and Welfare (AHAW), 2008b. Scientific Opinion on risk of bluetongue transmission in animal transit. EFSA Journal, 795, 1-56 EFSA Journal 2011;9(6):2192 6

7 Design prevalence for different situations In addition, an Opinion was needed on the expected prevalence under the different, specific epidemiological circumstances that may exist within the EU. Under the current rules, Annex I lays down a design prevalence (expected prevalence) for several situations, depending for example on: - the presence or absence of virus circulation; - the purpose of the programme: demonstrate absence of virus circulation or basic monitoring; - the vaccination status of the target population; that are critical to determine the surveillance strategy and the sample size. For targeted risk-based monitoring, however, the sampling strategy may be adjusted to the risks and the defined target population. Currently, the epidemiological situations as regards bluetongue may differ greatly in different areas of the EU due to different levels of virus circulation and different vaccination strategies of the Member States. The Commission therefore asked for a scientific review of the appropriate design prevalence for different situations, taking into account not only the situations as described above, but also other factors that could be important, such as: - recent virus circulation (the last season) versus not so recent virus circulation; - circulating serotype(s) and other serotypes; - Culicoides species and local distribution. TERMS OF REFERENCE AS PROVIDED BY THE EUROPEAN COMMISSION - To provide scientific advice on the expected prevalence (design prevalence) under different circumstances. - To provide an updated scientific assessment of the size of the relevant geographical area for the purpose of monitoring and surveillance programmes for bluetongue. EFSA Journal 2011;9(6):2192 7

8 ASSESSMENT 1. Introduction Bluetongue (BT) is an infectious, non-contagious disease of ruminants caused by an Orbivirus (Parsonon, 1990; Mertens et al., 2004) transmitted by Culicoides, small biting midges. At the moment, 24 different bluetongue virus (BTV) serotypes have been identified (Bonneau et al., 1999; Pritchard et al., 2004; Hofmann et al., 2008). Before 2006, it was generally believed that the global distribution of bluetongue occurred between latitudes of approximately 40-50ºN and 35ºS (Zhang et al., 1999; Mellor et al., 2008). In Africa and the Mediterranean Basin, C. imicola is considered the principal vector of BTV, however, other Culicoides species have also been associated with BTV transmission in Southern Europe (Caracappa et al., 2003; Savini et al., 2003). Between 1998 and 2005, BTV spread further northwards and occurred in many countries of the Mediterranean Basin (Calistri et al., 2004a; Giovannini et al., 2004; Mellor et al., 2008), and five different serotypes (1, 2, 4, 9 and 16) were detected. In August 2006, BT outbreaks occurred in North-western Europe and BTV-8 was identified as the causal serotype. The introduction of this serotype came as a surprise, because it was not circulating in the Mediterranean basin at that time. In North-western Europe, C. imicola is not present, and C. scoticus, C. obsoletus, C. chiopterus and C. dewulfi were probably the most important vectors for BTV-8 (Meiswinkel et al., 2007; Meiswinkel et al., 2008). Annex I of Regulation (EC) No 1266/2007 lays down the minimal harmonized requirements for monitoring and surveillance programmes for BTV. The programmes should contain at least passive (clinical) surveillance and an active laboratory-based programme. The active programme may be based on a longitudinal serological or virological (or both) survey of sentinel animals, on an annual survey of virological or serological testing of a random sample of the population in the period of the year when infection or seroconversion is more likely to be detected, or on targeted risk-based surveillance. If the aim of the programmes is to demonstrate freedom from infection, the sample size should be calculated according to a design prevalence (minimal detectable prevalence) and desired confidence level. The value of the design prevalence can be an estimate of the prevalence likely to be present in an epidemiological unit if the infection had been present, which can be derived from field observations in endemically infected zones, or from disease modelling (Cannon, 2002). In Annex I of Regulation 1266/2007, several design prevalences for BTV monitoring and surveillance programmes are given. A design prevalence of 2 % is given for use in a monthly cross-sectional survey of known seronegative sentinel animals during the period when the vector is most active, whereas a design prevalence of 20 % is given for annual serological or virological surveys in geographical units with unvaccinated ruminants. However, if there is evidence that the annual rate of seroconversion in the epidemiologically relevant geographical unit is lower than 20 % the sample size has to be calculated to detect the lower estimated prevalence. Member States can also establish a design prevalence according to a target population of susceptible animals at a relatively high risk. For monitoring and surveillance programmes, a relevant geographical unit is defined in the Regulation and comprises a grid of 45 x 45 km or ± 2,000 km 2, unless specific environmental conditions justify a different size. Member States may also use the administrative region as defined in Article 2(p) of Directive 64/432/EEC (for tuberculosis, brucellosis and leukosis) as the geographical unit of reference for monitoring and surveillance of BTV. EFSA Journal 2011;9(6):2192 8

9 In this Opinion, scientific advice on the expected prevalence under different circumstances is provided. Moreover, a scientific assessment of the size of the relevant geographical area for the purpose of monitoring and surveillance is given. Advice on the design of a monitoring and surveillance programme is outside this mandate, but it is obvious that, besides an appropriate design prevalence and relevant geographical unit, a valid design of the monitoring and surveillance programme, as well as good diagnostic quality of the tests used, are pivotal. The starting point for this Opinion was that prevalences that have been observed in the past are the best estimates for the expected prevalences in the future. To obtain observed prevalences, a systematic literature review (SLR) and a review of data collected during BTV monitoring and surveillance programmes in EU Member States were performed. In regard to the different circumstances mentioned in the mandate, vaccination, BTV serotype, animal species (cattle or small ruminants) and their density, Culicoides species, and local distribution were taken into account. Although this Opinion aimed to address all BTV serotypes, BTV-8 was specifically compared with other serotypes. The reasons for this are: 1) that the Scientific Opinion on BTV-8 (EFSA, 2011) identified that this serotype differed from others in its ability for transplacental transmission and contamination of semen, and, 2) the rapid and extensive spread of this serotype in North-western Europe (EFSA, 2007), both of which may have an effect on the expected prevalence. Wild ruminants were not addressed in this opinion. Before specifically addressing the ToRs in Sections 4 and 5, the epidemiological concept that was used to address the ToRs (Section 2.1), background information on Culicoides species (Section 2.2), domestic ruminant species and their density (Section 2.3), and diagnostic methods used in EU Member States monitoring and surveillance programmes (Section 3) are described first. 2. Phases of BTV infection in a population and possibly related aspects of vector and ruminant host For a detailed description of the epidemiology of bluetongue, a previous EFSA Opinion (EFSA, 2007) and more recent publications (Hartemink et al., 2009; Wilson and Mellor, 2009) should be consulted. In view of the current mandate, this section serves to introduce the epidemiological concept that was applied to answer the ToRs. Furthermore, based on the literature, the section addresses the possible influence of Culicoides and ruminant species and density Course of BTV infection in a population In the course of a BTV infection in a region, three fundamental steps can be distinguished (Randolph and Rogers, 2010): introduction, establishment and spread in a geographical sense. During these steps, the prevalence of infected animals in a region changes, since, upon introduction into a BTVfree region, the prevalence in a geographical unit rises from zero to a maximum (plateau prevalence) and subsequently drops again either to zero, in case the infection fades out, or to a level determined by endemic infection in the region (Giovannini et al., 2004; Schaik et al., 2008; Santman-Berends et al., 2010). The value of the plateau prevalence depends on the density of susceptible ruminants, the density of active and competent vectors (Hartemink et al., 2009) and the contact pattern between them. Whether the virus is able to establish an endemic infection in the area depends on the ability of the virus to overwinter and the rate at which susceptible ruminants are introduced into the population (note that vaccination reduces this rate). Based on the pattern described above, with respect to the expected prevalence, five phases are distinguished in this Opinion (Table 2). Phase 1 is a BTV-free population without a history of infection (i.e. fully susceptible). Upon introduction of BTV, phase 2 starts and subsequent virus transmission will result in a rise in the prevalence of BTV positive animals, followed by a rise in the prevalence of seropositive animals until this has reached a plateau (phase 3). In phase 4, the prevalence drops again to an endemic equilibrium or to zero, and in the latter case, phase 5 starts, at which point, the area is again free from BTV infection, but now there is a history of infection. EFSA Journal 2011;9(6):2192 9

10 In the case of BTV-8, Sweden and Denmark have gone from phase 1 to 5 in a few years, but in these countries the plateau prevalence was low (Rasmussen et al., 2010; Lewerin et al., 2010) in comparison to, for example, the Netherlands (Santman-Berends et al., 2010). The reason for the low plateau prevalence may have been the rapid start of the vaccination campaign in these Scandinavian countries, but aspects related to the vector, such as abundance of competent vectors or average daily temperature, may also have played a role. The seasonal pattern of BTV infections will also be reflected in the prevalence of virus positive animals and (to a lesser extent) seropositive animals. Moreover, because there are many BTV serotypes, an area can be in more than one phase at the same time, for example in phase 4 for BTV-1 and in phase 1 for the other serotypes. From the perspective of the risk manager, the goals for a monitoring and surveillance programme depend on the epidemic situation (Thulke et al., 2009). In phase 1, the aim is early warning in case BTV (or a new BTV serotype) enters the region. Since the ruminant population has no history of BTV infection, it is fully susceptible and introduction of BTV will most likely result in clinical disease, and passive surveillance (notification of clinical suspicions) is the surveillance programme that will detect clinical disease. The contribution of active surveillance to early warning of new BTV introductions into free regions of the EU is unknown, and it was not possible to estimate it from data of primary BTV outbreaks in the European Animal Disease Notification System (ADNS). Firstly, it was unclear which of the notified primary outbreaks were new introductions of BTV into a free region and, secondly, for the vast majority of the primary outbreaks the reason for testing (passive, active, other) was not documented in the ADNS. However, active surveillance using laboratory tests for the purpose of early warning is hampered by the fact that it is unlikely that such a survey will happen exactly at the initial time and place of the outbreak(s) that start an epidemic. Active surveillance might, however, be worthwhile in areas at risk for BTV incursion (for example, because they are adjacent to a zone considered to be BTV-infected, or located in areas at risk for introduction of exotic BTV), especially when a strain predominantly causes subclinical infections. However, in such a situation, sample collection would be most appropriate during the risk period and the frequency of sample collection should be sufficiently high. In phase 2, the goal of the surveillance programme will be to establish the extent of the area infected by BTV, and the observed prevalence may also provide relevant information for a decision regarding intervention measures. This also applies to phase 3, since, in addition, the prevalence of infected animals provides information that serves as the starting point for detecting a drop of prevalence in the subsequent year(s). Detecting a year to year drop in prevalence is the goal of the surveys in phase 4 and for that purpose the sample size should allow for detection of a meaningful change in the prevalence of infected animals. In phase 5, demonstrating freedom from BTV infection is the objective. After a population has been demonstrated to be free from infection, the focus of the risk manager will move again to early warning of new incursions of BTV. Obviously, the expected prevalence in phases 1 and 5 is zero. However, initially, it is unclear whether or when an area has moved from phase 4 to phase 5. For the purpose of demonstrating freedom from infection, the risk manager will need to define a design prevalence as the target to be detected by surveillance at the desired level of confidence. The value of this design prevalence can be based on the expected prevalence in an epidemiological unit if BTV had been present (Cannon, 2002). For that purpose, prevalences observed in phases 2, 3 and 4 from the literature and from the data of Member States BTV monitoring and surveillance programmes were assessed and considered as the prevalences to be expected in each of these three phases. The expected prevalence in phase 4 of the epidemic can be considered a target for the design prevalence in phase 5 (if the infection had been EFSA Journal 2011;9(6):

11 present, the prevalence would have been according to phase 4). However, if virus circulation has been absent for several years and the majority of the population is susceptible again, because of natural removal of immune animals, the expected prevalence in phase 3 is more appropriate as a design prevalence for demonstrating freedom from infection. The epidemiological concept of the five phases described above will be used to answer the ToRs on the expected prevalence (Section 4) and the relevant geographical unit (Section 5), because from the above it follows that the expected prevalence, as well as the size of the relevant geographical unit, are likely to be associated with the phase undergoing the epidemic. Table 2: Five phases of a BTV epidemic and their relation with the objective and type of monitoring and surveillance 2.2. Culicoides Culicoides biting midges were reviewed in a previous Scientific Opinion on bluetongue (EFSA, 2007). Therefore, this paragraph is mainly focused on research conducted on the vector since then, including the incursion of BTV-8 into North-western Europe and findings made during ongoing studies in the Mediterranean region. Three issues are briefly reviewed to assess whether vector distribution and abundance can be used as factors in establishing the expected prevalence and relevant geographical unit: (i) possible movement of C. imicola northwards into Europe, (ii) possible bluetongue sub-episystems in Europe, and, iii) vector abundance Northward movement of C. imicola Numerous and intensive vector surveillance programmes have mapped in detail the presence of C. imicola in Morocco, Portugal, Spain, southern France, Italy, Bulgaria and Greece. These programmes show its distribution to be mosaical, and largely restricted to lower-lying, planar areas. The northernmost records of C. imicola are from just below 45 N, and published data on the distribution of C. imicola are most likely reliable due to its easy taxonomic identification. In the 1980s, C. imicola was identified in countries located to the north of the Mediterranean Sea. This finding is interpreted in some publications (Purse et al., 2005; Purse et al., 2008) as evidence for its recent invasion of Europe, northwards from Africa, and that it will continue to advance northwards under the influence of climate change. However, the absence of C. imicola in equatorial Africa indicates that a warm climate alone is not sufficient for the presence of C. imicola, as is the finding that C. imicola occurs abundantly throughout the island of Sardinia, but is virtually absent from neighbouring Sicily. Based on these discrepancies some workers (Calistri et al., 2003; Conte et al., 2009) remain circumspect about the recent invasion of C. imicola and its march northwards. They EFSA Journal 2011;9(6):

12 propose instead that the discovery of C. imicola may have been delayed due to its narrow seasonality (August-November) and fragmented geographic range, and the past use of non-systematic vector collecting using inferior tools. The pattern of expansion has been explored further in more sophisticated studies conducted in Spain (Calvo et al., 2009) and in Italy (Conte et al., 2009), however, they have only provoked further debate by presenting conflicting results showing moderate, or rapid, expansion of C. imicola in Spain, as opposed to stasis in Italy Possible sub-episystems in Europe There are now six proven or potential Culicoides vectors recognized in Western Europe: C. imicola and five potential vectors of the Obsoletus and Pulicaris species complexes. At a distance, Europe would seem to consist of two BT sub-episystems: southern, with C. imicola, and northern, without C. imicola (Wilson and Mellor, 2009). However, the rate of spread of BTV-1 on both sides of the Pyrenees, prior to vaccination, was uniformly rapid, suggesting the southern and northern subepisystems comprise equally competent Culicoides vector populations. The spread of BTV-9 as far north as Kosovo where C. imicola is absent, is yet another example of a BTV strain circulating efficiently in Europe both in the presence and absence of C. imicola (Wilson and Mellor, 2009). Consequently, these examples nullify the north-south dual sub-episystems concept. In addition, since 2006, it has become clear that the vector populations of North-western Europe provide a system through which viruses may spread efficiently. We can only speculate why BTV-8 did not move further south or east. Prompt vaccination, depressed vector abundances, and possible refractoriness in C. imicola, may have hindered BTV-8 in its spread within parts of the Mediterranean region and, likewise, the apparent rarity of C. chiopterus and C. dewulfi in Eastern Europe (particularly in Bulgaria and Romania), and in the Mediterranean region, may explain why BTV-8 has not penetrated further east and south. Overall, too little is known about the precise in-field vector competencies of individual species (Carpenter et al., 2008) and their geographic ranges. Consequently, upon incursion of BTV in an area it is impossible to circumscribe zones of lower or higher risk for subsequent spread. The existence of zones with a higher risk of virus introduction is, however, recognised. Three principal routes of virus introduction into Europe have been identified, all of them situated in areas where multiple vector species occur (Calvete et al., 2008; Pili et al., 2010). These routes are: i) Morocco Spain, ii) North-Africa Western and Central Mediterranean islands and Italy, iii) Turkey Greece (Wilson and Mellor, 2008). Nevertheless, the incursion of BTV-8 into North-western Europe has demonstrated that virus introduction outside these three routes is also possible. As for BTV serotypes circulating within Europe, free zones adjacent to zones considered infected can be regarded to be at risk for BTV introduction Vector abundance The vector Culicoides occurs within every farmyard surveyed, breeds in a great variety of natural and man-made biotopes (Zimmer et al., 2008, 2010; Foxi and Delrio 2010) and bites a wide range of vertebrate hosts (Lassen et al., 2011; Ninio et al., 2011). Vector densities and species diversity, however, fluctuate seasonally and geographically. This is due to differing vector ecologies, uneven livestock densities, and variations in climate (Conte et al., 2007; Purse et al., 2008). Although undoubtedly important, the precise role played by each of these circumstances in the establishment and maintenance of an incursive virus has not been identified yet. Generally, the size of an outbreak is considered proportionate to the vector pressure that is being exerted. This holds true for C. imicola in Africa, where its super-abundance after heavy rains coincides with large epidemics (Nevill, 1971; Meiswinkel, 1998; Baylis et al., 1999a). However, it has been shown that vector abundances throughout the season, as determined by light trap catches, in North-western Europe can be considerably lower than those encountered in Southern Europe, suggesting a high competence of these northern vectors (Meiswinkel et al., 2008). The northern populations were able to ignite and sustain an epizootic that was exceptionally large. Four factors may EFSA Journal 2011;9(6):

13 neutralise the negative effect that low abundances might exert on vector capacity: (i) high animal density (and susceptibility) across a fairly homogeneous geographic and climatic zone; (ii) increased vector longevity due to milder ambient temperatures; (iii) involvement of multiple vector species, and, (iv) unrestricted short- and long-range movement of BTV-infected livestock (initiating fresh foci of disease). Clearly then, absolute vector abundance cannot be used alone as a reliable parameter when assessing future risk. What is apparent, however, is that autochthonous vector distribution patterns are far more stable and remain so over the longer term and, therefore, should prove to be important for risk management decisions, if properly established. The unresolved taxonomy of the five potential vectors of the Obsoletus and Pulicaris species complexes means that detailed mapping of their respective geographic ranges has hardly begun. Recent molecular studies (Cêtre-Sossah et al., 2004; Gomulski et al., 2005; Pàges et al., 2009) show the taxonomy of the more than 100 species of Culicoides found in Western Europe to be incompletely resolved. This unfinished business (Meiswinkel et al., 2004) has various consequences: (i) merging multiple species under a single label compromises our understanding of the role (or non-role) of each individual species in the transmission of BTV; (ii) an imperfect nomenclature encourages the misinterpretation of field data, which translates into a loss at many levels (and here also financial loss must be included because field data are collected at enormous cost); (iii) the ability to map a vector s range is undermined, and, (iv) the inability to compare the vector s range against that of the disease, the first step required for unravelling the spatial relationship that exists between a virus and its putative vector(s) Ruminant species and density Ruminant species and their husbandry systems may have an influence on BTV prevalence. Culicoides have been shown to have a higher preference for cattle than for sheep (Nevill, 1979). Moreover, Ninio et al. (2011), while examining blood fed midges of the Obsoletus complex, found that 54 % of midges had fed on cattle, while only 4 % had fed on sheep. Therefore, higher prevalences might be expected in cattle than in small ruminants. Moreover, taking into account that a high proportion of Culicoides are exophilic, it is reasonable to assume that animals reared indoors would be less exposed to infection. Santman-Berends et al. (2010) observed, in the Netherlands in 2007, that the monthly increase in seroprevalence in cattle pastured a few hours per day or throughout the day was higher (5.6 % to 11.4 %; p<0.5) relative to that for cattle kept indoors. For cattle that grazed outdoors throughout the day and the night, the monthly increase in seroprevalence was 13.6 %. Moreover, in the same study, cattle herds kept indoors year round stayed significantly (p<0.05) more often BTVfree (48 %) than herds kept outdoors (14 %). In addition, C. imicola has been considered as an outdoor species. Nevertheless, a recent paper from Calvete et al. (2009) describes a higher number of captured C. imicola (two to six fold) in traps located indoors than in traps located outdoors. Although animal density probably plays a role in the spread of BTV, to our knowledge, no articles have been published until now indicating a ruminant density threshold that allows efficient spread of BTV. Actually, it is unlikely that there is a single threshold of ruminant density because, according to the biological mechanism, it is interrelated with vector density. However, given the same density of competent vectors, zones with higher ruminant densities would allow for a more efficient spread of BTV, resulting in a higher prevalence than in zones with a lower density of ruminants. Figures 1 and 2 show the number of cattle and small ruminants per hectare of agricultural land. It has to be noted that some countries (e.g. Norway) have a low agricultural area compared to their total size, which results in a high density of animals per hectare of agricultural land. The high cattle densities of Belgium and the Netherlands and surrounding areas could have contributed to a more extensive spread observed in these BTV-8 infected countries compared to that observed in zones infected by other serotypes. However, this difference could also be related to other factors, such as serotype specific features or differences in vectors. According to Figure 2, the density of small ruminants is EFSA Journal 2011;9(6):

14 higher in Mediterranean countries, the UK and Ireland than in other areas of Europe. Ruminant densities are generally low in Eastern European countries, which could have reduced the probability of the serotypes that affect Balkan countries spreading into Western and Central Europe. Greece has a high density of small ruminants, which provides a good opportunity for the maintenance of BTV infections. Figure 1: Cattle density per hectare of agricultural area in Europe in 2007 Figure 2: Density of small ruminants per hectare of agricultural land in Europe in 2007 Both De Koeijer et al. (2011) and Gubbins et al. (2008) observed that transmission of BTV-8 between farms was efficient (Rh>1) only on days where the average temperature exceeded 15 C, suggesting a relation of temperature with activity of competent vectors. Although these studies are limited with regard to the time and geographic area studied and, hence, caution must be applied when generalising EFSA Journal 2011;9(6):

15 their findings, Welby et al. (1996) demonstrated that at temperatures below around 12 C, virus replication ceases altogether. Figure 3 shows the distribution of the number of days where the average daily temperature exceeded 15 C in According to this map, the number of days with an average temperature above 15 C is lower in Norway, large parts of Sweden, Ireland and the northern part of the UK, as compared to the Netherlands, Belgium and Germany, suggesting that conditions for spread of BTV-8 are favourable only for a short period in the former countries. More research is needed to determine if and how this parameter can be used to identify risk regions. Figure 4 is the result of ranking a qualitative combination of the overall ruminant density data in 2007 with a map of the distribution of the number of days where the average daily temperature exceeded 15 C (the combination rule is shown in Appendix 1). Although Figure 4 shows risk spots in the Netherlands and Belgium, consistent with efficient spread of BTV-8, and shows that, for example, Scandinavia and large parts of Eastern Europe are low risk areas, whereas Greece is a high risk area, in several countries the colours shown on the map are not consistent with the actual risk of BTV infection in place. For example, it does not explain why BTV-8 spread efficiently in large parts of France, but not in Hungary or the Czech Republic. Moreover, the high risk predicted for the north of Italy compared to the south on this map is not a reflection of the existing risk of BTV infection in these areas. Figure 3: Distribution of the number of days with an average daily temperature >15 C. EFSA Journal 2011;9(6):

16 Figure 4: Ranking of administrative units in Europe by their risks of BTV infection, based on a qualitative combination of ruminant density and number of days with an average daily temperature >15 C (higher ranking scores represent are higher risks) 3. Diagnostics for detection of antibodies and virus Monitoring and surveillance programmes within the EU include clinical investigations and laboratorybased surveys. As a result of the Regulation supporting this approach, the National Reference Laboratories throughout Europe have upgraded their testing procedures in order to be able to offer a timely and effective service, not only because of the necessity to control the disease but also to allow farmers to move animals safely from restricted zones into bluetongue-free areas. To be efficient, monitoring and surveillance programmes require a valid and accurate diagnostic system. In recent years, the diagnostics of BT have improved significantly. The diagnosis, which means the capability of recognising an animal that is or has previously been infected with BTV, usually involves the detection and the identification of a BTV-specific antigen, antibodies or RNA in diagnostic samples collected from potentially infected animals, using virus isolation and serological or molecular assays. Presence of typical clinical signs might also indeed facilitate the diagnostic process. The selection of an assay in fact depends on many factors. Amongst them, the intended purpose and the different setting in which the test has to be used, are the most important to be considered when selecting an assay. For disease eradication or surveillance programmes, a highly sensitive test capable of testing high numbers of samples is generally required, whereas to confirm a clinical suspicion, high throughput and high sensitivity are less essential. The performance of an assay is also closely linked to, among other factors, the test itself (direct or indirect), dynamics of the infection (acute, persistent, chronic, etc.) and the status of the target population (unvaccinated, vaccinated) Clinical signs Recognition of clinical signs of BT could provide an early indication of infection, particularly in regions where BTV appears for the first time. The disease, which occurs mainly in sheep and some species of wild ruminants, is characterised by various clinical forms, ranging from acute to chronic. However, BTV serotype 8, which was introduced to North-western Europe in 2006, has also been EFSA Journal 2011;9(6):

17 shown to cause serious disease in cattle. Acute clinical signs include fever, serous to bloody nasal and ocular discharge, ulceration of the mucosa, facial oedema and limp, while weakness, emaciation, muscle stiffness, fragile fleece with consequent alopecia might characterise the chronic forms (Verwoerd and Erasmus, 2004). However, none of these signs are pathognomonic of BT and, consequently, the clinical diagnosis requires confirmation by laboratory testing. However, in the first stage of the epidemic (phase 2 according to the Table 2 presented in 2.1), countries have used clinical signs to determine the prevalence of BTV infection in sheep. Although not as sensitive as a serological assay, recognition of clinical signs in sheep followed by direct laboratory confirmation could represent a valid method to determine the BT prevalence at the flock level in the early stage of an epidemic Virus detection Various techniques have been used to detect the presence of the virus, antigen or viral RNA. The most commonly used were: real time RT-PCR, conventional RT-PCR on agarose gel, capture ELISA, viral isolation from embryonated chicken eggs and viral isolation from mammal or insect cells. RT-PCR and, more recently, real time RT-PCR assays provide a versatile system able to give information on virus serogroup and serotype within a few hours. They are also highly sensitive and capable of detecting very low concentrations of viral RNA. In addition, the real time assay is able to quantify the viral genome. Due to all these advantages, in most laboratories, these new techniques are preferred to classical viral detection techniques, which require three to four weeks to be completed. However, RT- PCRs are actually not able to distinguish whether the RNA detected in the animal is part of an infectious virus or just RNA of degraded virus no longer able to infect the vector. The classical virus isolation technique (inoculation of embryonated chicken eggs/mammal cells or insect cells/mammal cells) is still the only method able to reveal the presence of infectious virus in an animal. Several studies have confirmed that, at least for some BTV serotypes, RNA could be detected in the blood longer than infectious virus (MacLachlan et al., 1994; Singer et al., 2001; Bonneau et al., 2002; Di Gialleonardo et al., 2011). The sensitivity of the RT-PCR in determining BTV circulation is dependent on the duration of viral RNA persistence in the blood of the host. Based on the detection of BTV RNA in cattle for months following infection (Bonneau et al., 2002; Di Gialleonardo et al., 2011), real time RT-PCR could be considered a valid and sensitive method for determining BTV circulation in cattle. Quantitative assessment of the diagnostic sensitivity and specificity of tests used in the EU monitoring and surveillance programmes is important to allow the establishment of the predictive values of positive and negative results for a given prevalence. Vandenbussche et al. (2008) reported a diagnostic sensitivity and specificity for the real time RT-PCR of 99.5 % (95 % CI: ) and 98.5 % ( ), respectively. Based on these figures, this test is good for using during monitoring and surveillance in phases 2-4, however, when testing large numbers of negative animals (phase 1 or 5) some false positive test results may be observed. Specificity can be increased if positive results are subsequently tested by a serotype specific PCR. Moreover, the epidemiological situation can be taken into account in case of unforeseen positive results. Consequently, PCR is a good test for the purpose of monitoring and surveillance. However, standardisation of this assay between different countries and laboratories is required Antibody detection Several techniques can be used to detect the presence of BTV-specific humoral antibodies in animals which have either been infected with BTV or vaccinated against the virus. The methods enabling the detection of serogroup-specific and serotype-specific antibodies in the serum and milk of infected animals most commonly used were: competitive ELISA (c-elisa), milk ELISA and micromethod plate seroneutralisation (SN). The OIE has prescribed the competitive ELISA (c-elisa) serological test for bluetongue diagnosis for international trade (OIE, 2010). Over time, the accuracy of this assay has been progressively EFSA Journal 2011;9(6):

18 improved through the use of antigens obtained from recombinant structural proteins of BTV. The competitive ELISA, using direct antibodies against the VP7 protein common to all BTV serotypes, detects the presence of antibodies to all 24 BTV serotypes. The technique is highly sensitive and specific, with its specificity being due to the use of the monoclonal anti-vp7, the protein that distinguishes the bluetongue serogroup from other Orbivirus serogroups. It is able to determine the presence of antibodies no matter which serotype the BTV strain belongs to and represents the preferred method to determine and monitor BTV circulation, since it is cheap, sensitive and specific. Vandenbussche et al. (2008) reported a diagnostic sensitivity and specificity for the c-elisa of 87.8 % ( ) and 98.2 % ( ), respectively. However, in cases where the population is vaccinated, it cannot be used to identify infections, unless seronegative sentinels are tested as well. Although some kits have been developed (see, for example, Barros et al., 2009), no DIVA tests for discriminating BTV-infected from vaccinated animals are at the moment commercially available. Milk ELISAs have also been developed that successfully assess the antibody status of animals with respect to BTV infection. The advantage of these tests is that they can be used on bulk milk samples, which has the obvious benefit of testing groups of animals in one step, avoiding the need for blood sampling (Kramps et al., 2007). Its specificity was recently improved by precipitating the milk protein prior to carrying out the ELISA (Chaignat et al., 2009). Mars et al. (2010) reported a herd sensitivity of 88 % (95 % CI 80-94) at a within-herd prevalence of 1 %, accompanied by a specificity of 100 % (95 % CI ), and a herd sensitivity of 100 % (95 % CI ) at a within-herd prevalence of 10 %, accompanied by a specificity of 93 % (86-98). As ELISAs are not able to distinguish between BTV serotypes, in areas where more than one serotype has been circulating, the serum neutralization assay is needed to determine the prevalence of each BTV serotype. Serum neutralisation (SN) is the only serological method so far able to determine the viral serotype responsible for the infection (serotype-specific test). The above indicates that the c-elisa is a good test to be used for monitoring and surveillance in phases 1-3 and 5. In case an unforeseen positive result is observed in phases 1 or 5, the test result can be confirmed by a test based on a different diagnostic principle. In phase 4, the c-elisa can only be used if applied on sentinel animals that are seronegative at the start of the vector season. 4. Expected prevalence To retrieve expected BTV prevalences, a systematic literature review (SLR) on prevalences observed within the EU was performed (Section 4.1). Anticipating that studies into the BTV-8 prevalence would be overrepresented in the published literature, which is most likely due to the attention paid to this serotype since 2006, data collected in monitoring and surveillance programmes of Member States was also assessed (Section 4.2). In the assessment in this Opinion, BTV 8 was distinguished from the other serotypes, which, because, of it quickly becoming widespread in North-western Europe, and its difference in certain aspects of transmission (such as transplacental transmission and contamination of semen), might be considered a worst case scenario for the prevalence compared to the other serotypes. Conclusions from both Section 4.1 and Section 4.2 are combined at the end of section Systematic literature review data Methodology An SLR was carried out in the CAB abstracts database, the Web of Science and in PubMed for published literature on the prevalence of bluetongue virus in domesticated ruminants in EU Member States in the period from 2000 to The goal of the SLR was to retrieve all relevant publications in a structured and reproducible way, and extract information uniformly from the papers. The detailed EFSA Journal 2011;9(6):

19 protocol can be found in Appendix 2. According to the infection mechanism, the rate of infection likely depends on both the density of ruminants in a region and the density of active and competent vectors (Hartemink et al., 2009). As a consequence, extrapolating prevalences observed on other continents to EU Member States is difficult because of differences in animal husbandry systems, differences in wildlife populations across the continents, differences in climate and differences in competent vector species (MacLachlan et al., 2006). Therefore, no reference is made to data from outside the EU. Thus, the review question asked was: Review Question: What was the prevalence of bluetongue virus in domesticated ruminants in European Union Member States in the period from 2000 to 2010? Relevance criteria: published scientific articles with a title and abstract indicating that they were primary research papers, Ph.D. theses or conference proceedings with an article body in English, French, German, Dutch, Italian, Danish or Spanish, and testing a hypothesis regarding or describing the prevalence of bluetongue in an EU Member State. Eligibility criteria: published scientific articles in which the ruminants studied (restricted to domestic animals), the number of animals sampled (in case of animal level prevalence), the number of herds sampled (in case of herd level prevalence), the applied sampling procedure, the diagnostic method used, the year of the study, the country/region of the study and the studied BTV-serotype were mentioned. Data extraction: extraction parameters included the animal species, the age of animals, the husbandry system, the vaccination status of the sampled animals, the country of study, and if the entire country was not covered by the study, the region in which the study was done, the year of study, the BTV serotype present in the country, whether the BTV serotype was new in the region/country or present for a longer time (>2 years) (epidemic or endemic situation), the Culicoides species present in the country, the diagnostic test and diagnostic test method/assay used in the study, the number of animals in the study area, the number of animals tested, the number of positive animals, the number of herds in the study area, the number of herds tested, the number of positive herds, the prevalence at the animal level within the region or country (including variation parameters, for example, the p-value, confidence interval, mean, median, minimum and maximum values), the prevalence at the herd level, and the incidence. The search was limited to publications with a title and abstract in English. A total of 761 references were retrieved and checked for duplicates. After removing duplicates, of the remaining 486 references, 80 were considered relevant, and the full text of the relevant publications was further assessed for eligibility. In all, 35 peer-reviewed publications were considered eligible. These papers provided data that enabled the prevalence to be calculated in 105 regions, originating from nine European countries (Belgium, Bulgaria, Denmark, France, Germany, Italy, Netherlands, Sweden and Switzerland). The data were either based on clinical diagnosis (3), virus detection (15, mostly RT- PCR) or antibody detection (87, mostly c-elisa). Seventy-nine observations concerned BTV-8 infections, 10 BTV-2, 8 BTV-9, and at least one for BTV-4, BTV-6, BTV-11, BTV-16 and Toggenburg virus. Thus, studies into BTV-8 prevalence were overrepresented, which is most likely due to the attention paid to this serotype since However, in the data from the Member States presented in Section 4.2, the proportion of observations on BTV-8 was limited compared to the other serotypes. Thus, section 4.1 and section 4.2 give a balanced view of the expected prevalence. Since the region is considered the relevant epidemiological unit (and not the herd) for BT, the animal and the herd level prevalence in the region are addressed in this Opinion. The animal level prevalence is the number of test positive animals of the total number of animals tested in a region, whereas the herd prevalence is the number of test positive herds of the total number of tested herds in that region. In all, 93 observations of the animal level prevalence and 74 observations of the herd prevalence were retrieved. Additionally, prevalences were distinguished between those obtained by antibody test and EFSA Journal 2011;9(6):

20 those obtained by virus test. The importance of the phase of the epidemic, vaccination, BTV serotype, vector species and density, and ruminant species (cattle versus small ruminants) was also taken into account. The WG experts indicated for each of the observed prevalences whether they were originating from phase 2, 3 or 4, according to the scheme presented in Table 2. Since the number of observations was low for each of the non-btv-8 serotypes individually, they were grouped together in this Opinion to enable a meaningful comparison between these combined other serotypes and BTV- 8. However, the number of papers giving useful information on Culicoides species and abundance was too small to enable an assessment. In Appendix 3 diagrams presenting the distribution of the observed animal and herd seroprevalences according to the phase, BTV serotype and animal species, each with their individual 95 % confidence interval, are shown. Phases with few observations and prevalences based on virus tests are not shown, because the low numbers did not allow any conclusions to be drawn. The median values of the prevalences (prevalences are not normally distributed) and their accompanying ranges are shown in Table 3 Cases where no single positive samples were observed in a region (zero prevalence) were not included in the table. The reason for this was that no distinction could be made between a region with a low prevalence (below the detection limit of the sample size) or an infection-free region. However, in Appendix 3 those observations showing a prevalence equal to 0 are shown with their 95 % CI. As concluded from Section 3, there is still limited quantitative data on the diagnostic sensitivity and specificity of the tests used. Therefore, due to the lack of this data, this assessment was based on the apparent prevalences as derived from the test results Prevalence at the animal level Prevalence at the animal level using antibody tests Phase 2: The SLR revealed 48 observed animal level prevalences based on antibody tests in phase 2, although 12 of these 48 observations did not include a single positive animal, which gave an overall count of 36 (Table 3). However, the seroprevalence was generally low, as indicated by the median seroprevalence of A total of 32 of the 48 observations were BTV-8 seroprevalences. In the case of BTV-8, the observed seroprevalence (median 0.020) was lower than compared to the other BTV serotypes (median 0.20). However, the latter group comprised only four observations with a prevalence >0. A total of nine of the 48 observed prevalences were from small ruminants, and six of them exceeded 0. In cattle, lower seroprevalences (median 0.008) were observed compared to small ruminants (median 0.20). However, it should be noted that the highest observed prevalence in small ruminants was associated with BTV-8. Phase 3: All 19 seroprevalences of phase 3 exceeded 0, with a median of 0.38 and a range of However, 18 of the 19 observations dealt with BTV-8, which did not enable any inferences to be made about the other serotypes. The BTV-8 seroprevalence in phase 3 was clearly higher than in phase 2 of the epidemic. Moreover, the observations that showed a low prevalence originated from sentinel animals in the first months after the start of the vector season. By September the mean seroprevalence within those animals exceeded 50 %. Phase 4: Only two observed prevalences were found in phase 4, both for BTV-2 infection and both in a vaccinated population. The prevalences in these two groups of sentinel animals were 0.13 and Prevalence at the animal level using virus detection Phase 2: Eleven observations of the prevalence at the animal level by virus detection were retrieved in phase 2. Seven observations concerned BTV-8 (median prevalence 0.05) and two other serotypes (0.024 and 0.64, respectively). In the case of BTV-8, the prevalence of virus positive animals seemed EFSA Journal 2011;9(6):

21 slightly higher than that of BTV-8 antibody positive animals in the same phase, which was not unexpected given that antibody formation follows replication of the virus. The difference between both prevalences was, however, only small. Phase 3: Only four observations based on virus detection in phase 3 were found, with prevalences ranging from Two observations were associated with BTV-8 (0.1 and 0.02) and two of another serotype (0.002 and 0.01). Phase 4: No data were available that demonstrated a prevalence based on a virological assay in phase Prevalence at the herd level Prevalence at the herd level using antibody tests Phase 2: Of the 26 observations at the herd level in phase 2, six were without a single positive herd. The median herd prevalence of the other 20 observations was 0.15 (range ). Seventeen observations concerned BTV-8 infection, 16 of which exceeded zero with a median prevalence of 0.03 (range ). Observations on the other serotypes included four that exceeded 0, with a median prevalence of 0.61 (range ). The median herd prevalence was 0.04 (range ) in cattle and 0.5 in small ruminants. Phase 3: Nine observations, ranging from , with a median prevalence of 0.8 were retrieved. All but one (BTV-2, prevalence 0.49) of these observations concerned BTV-8. Three observations described the herd prevalence in small ruminants (range ) and the other six the herd prevalence in cattle (range ). Obviously, these numbers of observations did not allow a comparison between species, or between serotypes. Phase 4: Only a single observation was retrieved from the literature describing a (sentinel) herd prevalence of 0.53 in a vaccinated population Prevalence at the herd level using virus detection Phase 2: At the herd level, the SLR retrieved only one observation that could be used to estimate a herd level prevalence based on virus detection. The estimated herd prevalence of concerned cattle herds in a BTV-8 infected region. Phase 3: No data were available demonstrating a herd prevalence based on a virological assay in phase 3. Phase 4: No data were available demonstrating a herd prevalence based on a virological assay in phase 4. EFSA Journal 2011;9(6):

22 herd level prevalence virus test herd level prevalence antibodies test animal level prevalence virus test animal level prevalence antibodies test Bluetongue monitoring and surveillance Table 3: Observed prevalence extracted from the systematic literature review (only those data with a prevalence >0 are included) Phase 1 Infection free propulation without history of BTV infection Observed prevalence derived from the Systematic Literature Review (only observations >0 included) Phase 2 Phase 3 Infected population with prevalence having reached a plateau Infected population with rising prevalence Phase 4 a Infected population with reduced prevalence (endemically infected or fade out of infection), no vaccination against BTV Phase 4 b Phase 5 Infected population with reduced prevalence (endemically infected or fade out of infection), vaccination against BTV Infection free propulation with history of BTV infection Count Median Minimum Maximum Count Median Minimum Maximum Count Median Minimum Maximum Count Median Minimum Maximum Count Median Minimum Maximum Count Median Minimum Maximum Overall Cattle Small ruminants n.a n.d. n.a. BTV BTV other than BTV ST unknown Overall Cattle Small ruminants n.a n.d. n.d. n.a. BTV BTV other than BTV ST unknown Overall Cattle Small ruminants n.a n.d n.a. BTV BTV other than BTV ST unknown Overall Cattle Small ruminants n.a n.d. n.d. n.d. n.a. BTV BTV other than BTV ST unknown n.a. = not applicable, n.d. no data 4.2. Data from EU Member States BTV infected EU Member States have to perform a monitoring (within restriction zones) and surveillance (outside restriction zones) programme according to Annex I of Regulation (EC) No 1266/2007. Results of these programmes are then presented to the EC. Unfortunately, the data as they are submitted to the EC are unsuitable for epidemiological analysis without proper knowledge of the national situation. As a consequence, this Opinion focused on those Member States of which at least one of the WG members had enough knowledge of the national programme to make the data useful for the analysis. In addition, the help of the French agency for food, environmental and occupational health and safety (ANSES) for elucidating data collected in the French monitoring and surveillance system is acknowledged (ANSES, 2011). Member States' data from Belgium ( ), Denmark (2009), France ( ), Italy ( ), Spain (2009), Sweden (2009) and the Netherlands (2007) were kindly made available to EFSA. However, data collected in the Swedish and Danish programme only contained negative results and, since both countries were declared free from BTV-8 recently, they were excluded from this assessment because they could not provide information on the expected prevalence. In all, 1281 observations of a prevalence in a geographical unit were included. In contrast to the data derived from the literature, this data were not predominantly on BTV-8, since 115 were related to BTV-8, 510 to other serotypes and in 656 observations the BTV serotype was unknown. However, given the geographical origin of the latter, only a small proportion of the associated prevalences was likely to reflect BTV-8 infection. Approximately 2/3 of the results originated from cattle and 1/3 from small ruminants. In phase 4, serological monitoring and surveillance was carried out using a sentinel system, and only the last sampling of the season was used in order to obtain the prevalence reached during that season. Due to the high numbers of observations, analyses using stack diagrams are not shown in the appendix. However, this information is available upon request. Furthermore, the approach followed here is the same as in Section Prevalence at the animal level Prevalence at the animal level using antibody tests Phase 2: The data from the EU Member States comprised 78 observations of seroprevalences at the animal level in phase 2, 44 of which exceeded 0 (Table 3). These 44 observations ranged from , with a median of The median prevalence in cattle was slightly higher than in EFSA Journal 2011;9(6):

23 small ruminants. This is in contrast to the findings of the SLR. Also, the median prevalence in BTV-8 infected populations was slightly higher than in populations infected with other or unknown types. Phase 3: Ninety-three out of 100 observations exceeded 0, ranging from with a median of The seroprevalence in this phase was clearly higher than in the initial phase of the epidemic, however, the number of observations with a low prevalence was remarkable. Although the data for this phase showed the results of sentinels at the end of the season, the low prevalence could still be explained by the use of sentinel animals. Since sentinel animals or herds are (in part) surrounded by immune animals and herds with immune animals, the actual transmission in such a population may be lower than in a naïve population, resulting in a lower prevalence (de Jong, 1995). In contrast, however, the sentinel data of BTV-8 derived from Belgium and the Netherlands in 2007 resulted in a median prevalence of 0.48, which is close to the overall median value of BTV-8 seroprevalence in phase 3. Apparently, the seroprevalences associated with non-btv-8 serotypes, as observed for example in Italy and Spain, were lower than those associated with BTV-8 in the Netherlands and Belgium. Whether this difference is the result of the serotype, different geographical location or other factors remains to be elucidated. The difference in the median prevalence in phase 3 between SLR and data from Member States is most likely due to the fact that BTV-8 was overrepresented in the SLR. Although median prevalence in cattle was lower than in small ruminants, this might also be explained by an unequal distribution of the data for these species across the serotypes. Phase 4: Phase 4 data of BT in an unvaccinated population (4a) were sparse in the EU due to the fact that in most European countries, the population is vaccinated in phase 4. The results of phase 4a presented here originated from Italy, and the median prevalence was 0.014, ranging from The median did not differ between cattle and small ruminants. Little information was available on the serotype in this phase, although it was most likely not BTV-8. The dataset included 651 observations of the prevalence at the animal level in (unvaccinated) sentinel animals in vaccinated populations (phase 4b) that exceeded zero. The seroprevalence ranged from with a median of 0.016, which was a similar value to that shown above in phase 4a. The data showed little difference between cattle and small ruminants. The seroprevalence was a bit lower in the case of BTV-8 compared to the other serotypes (although it was even lower in the case of unknown serotypes) Prevalence at the animal level using virus tests Phase 2: The dataset included only two virological observations showing an animal level prevalence in phase 2, one without any positive finding (small ruminants) and one with a prevalence of (cattle). Phase 3: No data were available demonstrating a prevalence based on a virological assay in phase 3. Phase 4: The dataset contained 149 virological observations in phase 4, all of them originating from populations vaccinated with inactivated vaccines. Forty three were without any positive test result, and in the other 106, prevalences ranged from , with a median value of (phase 4b). Small ruminants were only sparsely present in this dataset (four observations, median of the positive values ). Twenty observations concerned BTV-8, and all of them had at least one positive sample, with a median of ( ). Fifteen observations concerned a non-btv- 8 serotype, and eight had at least one positive sample with a median prevalence of ( ). In all the other observations, the serotype of BTV was not determined. No data were available on the prevalence in unvaccinated populations (phase 4a). EFSA Journal 2011;9(6):

24 Table 4: Observed prevalence derived from the EU Member States monitoring and surveillance data (only those data with a prevalence >0 are included) Prevalence at the herd level Prevalence at the herd level using antibody tests Phase 2: The dataset from the EU Member States contained no observations of the seroprevalence at the herd level in phase 2 (Table 4). Phase 3: Thirty observations were available from phase 3 with a median prevalence of 1 (range ) but all these observations were related to BTV-8, with ten originating from small ruminants (0.86-1) and the other 20 from cattle (0.47-1). Phase 4: Observations on the herd prevalence in phase 4 were not available Prevalence at the herd level using virus tests Phase 2: Similar to the animal level prevalence, only two observations on the herd prevalence based on a virological test were available for phase 2. The observed herd prevalences were 0 (small ruminants) and 0.2 (cattle), and neither were related to BTV-8. Phase 3: Observations on the herd prevalence in phase 3 were not available. Phase 4: Thirty four observations on the herd prevalence in vaccinated populations (phase 4b) were available, with 28 including at least one herd that tested positive that had a median of 0.19 and a range of Twenty observations concerned BTV-8 (median 0.20, range ) and 8 observations concerned another serotype (median 0.13, range ). Twenty four observations were of cattle populations with generally higher herd prevalence than in small ruminants. The assessment of both SLR and data from Member States is summarised in Table 4. Conclusions from Section 4.1 and Section A design prevalence for early warning of new BTV incursions in a region cannot be based on the expected prevalence in phase 2 alone, because it is also necessary to take into account the dynamics of the infection in the area (influenced by vector and host density and the interactions between them), the EFSA Journal 2011;9(6):

25 frequency of sampling and the objective of surveillance (for example, detecting an epidemic before a certain number of herds have become infected). - The plateau prevalence (phase 3) of BTV-8 infected ruminants in North-western Europe was markedly higher than that of other serotypes in Southern Europe. - The literature, as well as Member States data, do not demonstrate a clear indication that the expected prevalence is substantially different in cattle than in small ruminants. - The currently used design prevalence of 2 % for monthly testing of sentinel animals is slightly higher than the median expected prevalence in an endemically infected area (phase 4). - The currently used design prevalences of 20 % and 10 % for an annual survey in populations of unvaccinated and vaccinated ruminants, respectively, are high in comparison with the expected (sero)prevalences in phase Size of the relevant geographical area for the purpose of monitoring and surveillance Bluetongue monitoring and surveillance programmes take the geographical area as the relevant epidemiological unit, rather than the farm, as is common in the case of directly transmitted animal diseases. The reason is that besides ruminants, vectors are an integral part of the infection chain and they are not confined to a specific farm. In Europe, an area-based BTV monitoring and surveillance system was first developed in Italy to enable movement of animals between BTV-free geographical units (Giovannini et al., 2004). Based on the circle around an infected farm foreseen by legislation (control zone, 20 km radius = ± 1,256 km 2 ), the size of the unit (grid) was established at 40 x 40 km (=1,600 km 2 ). A thorough scientific evaluation of that monitoring and surveillance system has, however, not been published in the peer reviewed literature. In Regulation (EC) 1266/2007, the epidemiological unit for monitoring and surveillance of BTV is defined as a 45 x 45 km area (approximately 2,000 km 2 ), which is somewhat larger than the size of the Italian grid. However, 2,000 km 2 is the smallest possible size of the area of the administrative unit considered in Council Directive 64/432/EEC for monitoring campaigns on tuberculosis, brucellosis and leukosis. Member States may also use this administrative region for monitoring and surveillance of BTV. The optimal size of the relevant geographical unit for the purpose of monitoring and surveillance depends on the goal and design of the monitoring and surveillance programme and the spatial spread of the infection. In this Opinion, the first information on the spatial spread of BTV and associated factors is reviewed. Subsequently, the implications this might have for the size of the relevant geographical unit in each of the five phases of BTV infection in an area are discussed Spatial spread of bluetongue virus Spatial spread Gerbier et al. (2008) estimated, from the epidemic in the northern part of Western Europe, the rate of BTV-8 spread after introduction into a naïve population at 15 km/week. However, Calistri et al. (2004b) estimated the rate for BTV-2 in Sardinia at 30 km/week. Spread of BTV infected vectors by air across open water may even be beyond 30 km, as shown by the incursion of BTV into the UK (Gloster et al., 2010). Moreover, Ducheyne et al. (2007) reported that windborne-spread of BTV infected vectors over sea may be possible for a distance of up to 750 km. However, spread over such a long distance across land has never been published. Hendrickx et al. (2008) estimated that, in 2006, 50 % of the spread from an infected farm in Belgium took place within a 5 km radius and 95 % within EFSA Journal 2011;9(6):

26 a 30 km radius. Using a different model, De Koeijer et al. (2011) estimated that 85 % of the secondary infections of BTV-8 took place within a 15 km radius of an infected farm. However, Szmaragd et al. (2010) predicted even more localised spread than the De Koeijer et al. model. Both studies used the same outbreak dataset, including comparable assumptions on data, but assuming different spatial transmission kernels (probability of infection as a function of the distance to an infectious source) during model fitting. This likely explains the detailed differences between derived spreading distances. Nevertheless, the results of both studies suggest a local to intermediate spatial transmission. Taking the information above into account (and assuming that infection starts in the centre of the grid) means that the currently defined geographical unit for monitoring and surveillance (45 km x 45 km) covers spatial spread of BTV infection of a few weeks. According to Reynolds et al. (2006), local spread of BTV is mostly caused by active movements of the vectors themselves in all directions and is rather symmetric, while the medium and long distance spread is caused by windborne movement of vectors and animal movements and may follow a rather asymmetric pattern. Although increasing the size of the zone considered to be infected might give more protection to surrounding free zones, results from De Koeijer et al. (2011) suggest that doing this increases the amount of long distance spread of BTV within the zone considered to be infected Associated factors Information about important risk factors for the spread of BTV may help to define the optimal size of the geographical unit for the purpose of monitoring and surveillance. It is assumed that vaccination will reduce the rate of spread of BTV if a sufficiently high coverage (>80 %) is achieved (Szmaragd et al., 2010). Ducheyne et al. (2011) simulated BTV-1 outbreaks in France using wind spread models, taking into consideration the vaccination coverage achieved in each department of France in Their model predicted that the level of vaccination was sufficient to prevent spread of BTV. However, in the data from the Member States presented in Section 4.2, no striking difference was observed in the prevalences of BTV infected animals in vaccinated populations, as compared to non-vaccinated populations. One reason might be that in many of the vaccinated populations included in that dataset, vaccination coverage of 80 % was not reached. In case a coverage >80 % is reached using an efficacious vaccine, enlarging the geographical unit for monitoring and surveillance might be an option, however, this can only be considered if the infection is homogeneously distributed across the zone. In the case where the distribution is heterogeneous, or not known, smaller units provide a better precision of the actual prevalence. The same argument could be applied to cold regions (only a few days with an average temperature >15 C; see Section 2.3). Climatic and topographical factors play a role in the spread of BTV and such factors may be very different from one Member State to another. For example, many differences exist between the various Mediterranean regions in Europe (Walker 1977; Baylis et al., 1998, 1999a, 1999b; Purse et al. 2005). Moreover, in the north of Western Europe, the presence of BTV-8 was favoured in locations that were warmer on average, where temperatures varied less throughout the year and where temperatures rose quickly in spring reaching a peak earlier in the year (EFSA, 2007). Landscape elements, such as land-cover and topography, influence patterns in Culicoides-borne diseases, probably via their effects on habitat availability, both for Culicoides and their ruminant hosts, and also through disturbed wind dispersal over rough terrain. Even in small countries such landscape elements may show a huge variety. As an example, in Belgium it was shown that clusters of outbreaks occurred in woody areas (the Ardennes in the south-eastern part of Belgium), while clusters of outbreaks also occurred in areas with little forest (Ghent cluster in the north-western part of Belgium and the initially infected area in the vicinity of Maastricht). This supports findings that efficient spread may take place in various landscape patterns and that different vectors are capable of efficient transmission, because the abundance of specific vectors may vary from one area to another (Schroder and Schmidt, 2008). It also highlights that, even over relatively short distances, the EFSA Journal 2011;9(6):

27 differences in landscape patterns may be considerable. Due to our limited knowledge of the interplay between vector and landscape patterns it is not yet possible to indicate their effect on the optimal size of the relevant geographical unit Geographical unit related to the phases of BTV infection in a population The EU is subdivided into BTV-free zones and restricted zones, and the latter may differ from one BTV serotype to another (Figure 5). The relevant geographical unit should be a clearly defined part of a territory, containing an animal population with a single distinct health status. Consequently, a geographical unit cannot exceed the size of the restricted (or free) zone. Moreover, in a restricted zone such a unit can be considered to be infected (phases 2, 3, 4) by a certain BTV serotype (for example BTV-8), but be free (phases 1, 5) from others at the same time. As a result, different goals for monitoring and surveillance apply within a geographical unit at the same time. Figure 5: Bluetongue restricted zones in the European Union (5 April 2011) Relevant geographical unit in phase 1 Table 5 shows qualitatively how the size of the relevant geographical unit is related to the five phases of BTV infection in a region, and which additional factors are to be considered. In a BTV-free region without any history of BTV, the primary goal is to detect a BTV introduction quickly. Hereto, an early warning system must be in place for all relevant species, as well as measures to prevent introduction of the virus. In the case of a BTV strain with a clear clinically manifested infection, passive surveillance (notification of clinical suspicions) is the surveillance system of choice and, because it can be applied in the whole region, the size of the geographical unit is irrelevant. In general, active surveillance is not expected to outperform passive surveillance, because it is unlikely that sample collection will take place in space and time exactly at the very beginning of an epidemic. However, if an area is considered at risk for BTV introduction, where the infection does not lead to EFSA Journal 2011;9(6):

28 clinical manifestations, both active surveillance, in addition to passive surveillance, may be worthwhile. Areas at risk for BTV introduction are zones in the vicinity of the three routes of incursion into Europe (Greece, the southern parts of Italy and Spain, the Balearic Islands, Sardinia and Corsica), as well as zones adjacent to zones considered to be infected within the EU. The size of such an area will depend on the surveillance programme in place. For example, in case of monthly testing, assuming a rate of spatial spread of 15 km/week (De Koeijer et al., 2011) and taking into account the time to reach the design prevalence in a herd, the time to seroconversion and some time for testing, an area of two currently used geographical units (a zone 90 km wide) would allow detection of the virus before it has left the risk area again. However, it has to be taken into account that the airborne spread of the vectors across water can be much further than across land. In the risk areas, in addition to a high sampling frequency (short interval between samplings to enable detection shortly after virus introduction) during the period at risk (for BTV between July and October) and an adequate sample size, relatively small geographical units within the area are to be preferred. The larger the area/population, the longer the average time between introduction and detection, given the same sample size and sampling frequency. Furthermore, to enhance the probability of detection in such a situation, surveillance should be focused on animal dense areas. Table 5: Phases of BTV infection in a population, their objectives and requirements for monitoring and surveillance, and their expected prevalence Relevant geographical unit in phase 2 Upon introduction of BTV into a region, the size of the relevant geographical unit is important if an attempt to control the infection is being considered and the demarcation of the infected area needs to be established. The latter can be undertaken by intensive investigations on farms surrounding the infected premises. For the actual size of the zone considered to be infected, the time between virus introduction and detection of the infection, as determined by an epidemiological assessment, is relevant, because it determines the spatial spread that has already taken place at the moment of EFSA Journal 2011;9(6):

29 detection. The size of the relevant geographical unit, however, should be determined by the desired precision. If control of infection is considered, animal movements are important, because movements of infected animals can result in new episodes of local spread elsewhere. If no actions are taken to control infection, the size of the geographical unit is not relevant in phase Relevant geographical unit in phase 3 Once the rise in the prevalence of infected animals has come to an end, monitoring and surveillance programmes can show the demarcation of the BTV-infected area (as in phase 2) and the prevalence that has been reached. Both may provide relevant information for the decision regarding intervention strategies. The size of the geographical unit for monitoring and surveillance is therefore related to the precision of the estimated prevalence across the region. Dividing a country into smaller areas will result in a more precise estimate of the prevalence in the country as a whole and, moreover, it may show differences in prevalences across the country Relevant geographical unit in phase 4 The precision issue discussed in is even more relevant for phase 4, where the objective is to identify meaningful year to year changes in infection prevalence in the presence or absence of intervention measures (such as vaccination) Relevant geographical unit in phase 5 Freedom from infection implies the absence of circulating BTV (serotype) in the country or zone. Monitoring and surveillance programmes cannot provide the absolute certainty of the absence of infection. Moreover, demonstrating freedom from infection occurs in retrospect, since it is only valuable for trading partners if it is associated with the presence of an early warning programme to quickly detect new incursions of the virus and measures to reduce the risks of new introductions. As a consequence, during this phase, no animals must be introduced from BTV infected regions, unless they have been demonstrated free from infection, or have been vaccinated before movement. In practice, the aim of demonstrating freedom from infection is to demonstrate that a sample size able to detect the design prevalence (with a certain level of confidence, for example, 95 %) does not show any positive result (in the case of BTV this needs to be undertaken in two consecutive years). However, the larger the size of the geographical unit chosen to conduct the surveillance, the lower the sample size will be per km 2. As an example, Spain and Belgium have, respectively, a surface of 506,000 km 2 (~ 250 grids of 45 x 45 km) and 33,500 km 2 (~ 15 grids of 45 x 45 km). To prove freedom from infection in Spain as a whole (with a 99 % confidence and a design prevalence of 0.1 %, and assuming a perfect test), a sample size of 4,600 is needed if it is considered as one geographical unit (calculations performed in Survey Toolbox version 1.0 Beta). This sample size is only slightly higher than the 4,470 samples needed in Belgium for the same purpose. However, on the level of the individual grid, the detectable prevalence (at a 95 % confidence) in this example is approximately 15 % in Spain and 1 % in Belgium. It is obvious that in this case prevalence common to phase 3 and phase 4 (Section 4) could be easily missed in Spain. Moreover, while designing a sampling strategy, the expected prevalences are usually assumed to be homogeneous across the geographical unit, both at the herd as well as at the animal level. However, this is often not true, and, although the effect of this heterogeneity is unclear, one would prefer to be on the safe side in regard to the sampling strategy. It can be argued that the confidence in freedom from BTV infection in a geographical unit is not only based on the test results and history of infection of that region, but also on the BTV status of surrounding geographical units. As a consequence, in the vicinity of areas where BTV is still EFSA Journal 2011;9(6):

30 circulating, a greater effort should be made to detect new outbreaks than if such areas are far away. This implies that the geographical units for monitoring and surveillance should be chosen smaller, and a more intensive sampling design (frequency and sample size) should be applied when located close to infected areas. Table 6 presents a summary of the assessment of the relevant geographical unit. Table 6: Phases of BTV infection in a population, their objectives and requirements for monitoring and surveillance related to the size of the relevant geographical unit CONCLUSIONS AND RECOMMENDATIONS CONCLUSIONS Regarding expected prevalence - Different phases can be distinguished in the course of a BTV epidemic, and monitoring and surveillance have a different purpose during each phase. - The expected prevalence in a region infected for several years (phase 4) can be considered a useful target for the design prevalence to demonstrate freedom from infection. - If virus circulation has been absent in a region for several years and the majority of the population is susceptible to infection, the plateau prevalence (phase 3) can be considered as a useful target for the design prevalence in order to demonstrate freedom from infection. - A design prevalence for early warning of new BTV incursions into a region cannot be based on the expected prevalence in phase 2 alone, because it is necessary to also take into account the dynamics of the infection in the area (influenced by vector and host density and the interactions between them), the frequency of sampling and the objective of surveillance (for example, detecting an epidemic before a certain number of herds have become infected). EFSA Journal 2011;9(6):

31 - The plateau prevalence (phase 3) of BTV-8 infected ruminants in North-western Europe was markedly higher than that of other serotypes in Southern Europe. - The literature, as well as data from EU Member States, does not demonstrate a clear indication that the expected prevalence is substantially different in cattle than in small ruminants. - The currently used design prevalence of 2 % for monthly testing of sentinel animals is slightly higher than the median expected prevalence in an endemically infected area (phase 4). - The currently used design prevalences of 20 % and 10 % for an annual survey in populations of unvaccinated and vaccinated ruminants, respectively, are high in comparison with the expected (sero)prevalences in phase 4. Regarding expected prevalence and vectors - The northward movement of C. imicola in Europe is not unequivocal. - BTV-1, BTV-8 and BTV-9 have been transmitted efficiently in the absence of C. imicola. - The rate of spread of BTV-1 on both sides of the Pyrenees, prior to vaccination, was uniformly rapid, suggesting the southern and northern sub-episystems comprise equally competent Culicoides vector populations. - All of the six potential vector species within Southern and Northern Europe share an equal infield transmission potential. - Absolute vector abundance cannot be used alone as a reliable parameter when assessing future risk. - The quantitative relation between the distribution and the abundance of different vector species in the EU Member States, and the expected prevalence and optimal size of the relevant geographical unit is still unknown. Thus, it is not possible to provide advice on differences in expected prevalence for the different epidemiological phases based on the current knowledge of the distribution and abundance of different vector species in Member States. - Indoor housing appears to be a prevalence reducing factor for BTV-8 in cattle in the Netherlands. - The relation between ruminant density and BTV prevalence has not been established yet. Given the same density of competent vectors, zones with higher ruminant densities would allow for a more efficient spread of BTV, resulting in a higher prevalence than in zones with a lower density of ruminants. Regarding expected prevalence and areas with high risk of introduction - Considering the three routes of introduction of exotic BTV serotypes into Europe, Greece, the southern parts of Italy and Spain, the Balearic Islands, Sardinia and Corsica are at the highest risk for introduction of exotic BTV and, consequently, the most appropriate regions for targeted (active) surveillance for early detection of incursions of new BTV serotypes, in addition to the usually implemented and necessary passive surveillance. - Considering BTV serotypes circulating within Europe, free zones adjacent to infected zones are the most appropriate regions for targeted (active) surveillance for early detection of incursions of new BTV serotypes, in addition to the usually implemented and necessary passive surveillance. EFSA Journal 2011;9(6):

32 - After incursion of BTV in an area, there is no scientific basis to circumscribe zones of lower or higher risk for further spread of the virus. Regarding expected prevalence and diagnosis - PCR is a good test for use during monitoring and surveillance in phases The c-elisa is a good test to be used for monitoring and surveillance in phases 1-3 and 5. In case an unforeseen positive result is observed in phases 1 or 5, the test result can be confirmed by a test based on a different diagnostic principle. - In phase 4, the c-elisa can only be used if applied on sentinel animals that are seronegative at the start of the vector season. Regarding the relevant geographical unit - Local spread of BTV is mostly caused by active movements of vectors. - Increasing the size of the zone considered to be infected increases the amount of long distance spread of BTV, because of uncontrolled movement of animals within that zone. - The size of the currently defined geographical unit for monitoring and surveillance (45 km x 45 km) encompasses local spread of BTV infection for a few weeks. - Climatic, vegetation and topographical factors and their interaction undoubtedly influence the population dynamics of BTV infections, but it has not yet been established how they quantitatively affect the optimal size of the relevant geographical unit for the purpose of monitoring and surveillance. - The size of the relevant geographical unit will depend on the goal of surveillance: - For the purpose of early warning, passive surveillance using disease notifications takes place independent of the size of the epidemiological unit. - For the purpose of monitoring the impact of interventions on the prevalence of infected animals, smaller units result in a more precise estimate for the region or country as a whole and take better account of local differences. - For the purpose of establishing freedom from infection, smaller areas result in a lower design prevalence for the area as a whole and take better account of local differences in infection dynamics. - There is no scientific evidence suggesting an optimal size of the relevant geographical unit for BTV monitoring and surveillance, because it depends on many factors including the goal of surveillance, the dynamics of the infection in the area and the design of the surveillance programme. RECOMMENDATIONS Regarding expected prevalence - A lower design prevalence should be used for the purpose of demonstrating freedom from infection than those currently used. EFSA Journal 2011;9(6):

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39 APPENDICES APPENDIX 1: MODEL USED TO COMBINE THE RUMINANT DENSITY AND THE NUMBER OF DAYS WITH AN AVERAGE TEMPERATURE >15 C SHOWN IN FIGURE 4. EFSA Journal 2011;9(6):