Genes and genome editing tools for breeding desirable phenotypes in ornamentals.

KEY MESSAGE: We review the main genes underlying commercial traits in cut flower species and critically discuss the possibility to apply genome editing approaches to produce novel variation and phenotypes. Promoting flowering and flower longevity as well as creating novelty in flower structure, colour range and fragrances are major objectives of ornamental plant breeding. The novel genome editing techniques add new possibilities to study gene function and breed new varieties. The implementation of such techniques, however, relies on detailed information about structure and function of genomes and genes. Moreover, improved protocols for efficient delivery of editing reagents are required. Recent results of the application of genome editing techniques to elite ornamental crops are discussed in this review. Enabling technologies and genomic resources are reviewed in relation to the implementation of such approaches. Availability of the main gene sequences, underlying commercial traits and in vitro transformation protocols are provided for the world's best-selling cut flowers, namely rose, lily, chrysanthemum, lisianthus, tulip, gerbera, freesia, alstroemeria, carnation and hydrangea. Results obtained so far are described and their implications for the improvement of flowering, flower architecture, colour, scent and shelf-life are discussed.

Development of food safety risk assessment tools based on molecular typing and WGS of Campylobacter jejuni genome.

The 'learning-by-doing' EU-FORA fellowship programme in the development of risk assessment tools based on molecular typing and WGS of Campylobacter jejuni genome was structured into two main activities: the primary one focused on training on risk assessment methodology and the secondary one in starting and enhancing the cooperation between the hosting and home organisations, or other joint activities. The primary activities had three subsequent work packages (WPs): WP1 data organisation, WP2 cluster and association analyses, and WP3 development of risk assessment models. The secondary activities have branched into one workshop and the initiation of a cooperation programme between the hosting and home organisations. In the last quarter, the fellow had contributed to the characterisation of some pathogens in possible response to a changing climate, part of the CLEFSA project. The fellow attended various forms of training: online and on-site courses, and also participated at several conferences and meetings for improving his knowledge and skills, contributing to performing the Campylobacter risk assessment and source attribution.

Development of a forecasting model for brucellosis spreading in the Italian cattle trade network aimed to prioritise the field interventions.

Brucellosis caused by Brucella abortus is an important zoonosis that constitutes a serious hazard to public health. Prevention of human brucellosis depends on the control of the disease in animals. Livestock movement data represent a valuable source of information to understand the pattern of contacts between holdings, which may determine the inter-herds and intra-herd spread of the disease. The manuscript addresses the use of computational epidemic models rooted in the knowledge of cattle trade network to assess the probabilities of brucellosis spread and to design control strategies. Three different spread network-based models were proposed: the DFC (Disease Flow Centrality) model based only on temporal cattle network structure and unrelated to the epidemiological disease parameters; a deterministic SIR (Susceptible-Infectious-Recovered) model; a stochastic SEIR (Susceptible-Exposed-Infectious-Recovered) model in which epidemiological and demographic within-farm aspects were also modelled. Containment strategies based on farms centrality in the cattle network were tested and discussed. All three models started from the identification of the entire sub-network originated from an infected farm, up to the fifth order of contacts. Their performances were based on data collected in Sicily in the framework of the national eradication plan of brucellosis in 2009. Results show that the proposed methods improves the efficacy and efficiency of the tracing activities in comparison to the procedure currently adopted by the veterinary services in the brucellosis control, in Italy. An overall assessment shows that the SIR model is the most suitable for the practical needs of the veterinary services, being the one with the highest sensitivity and the shortest computation time.

Prioritization of Companion Animal Transmissible Diseases for Policy Intervention in Europe.

A number of papers have been published on the prioritization of transmissible diseases in farm animals and wildlife, based either on semiquantitative or truly quantitative methods, but there is no published literature on the prioritization of transmissible diseases in companion animals. In this study, available epidemiological data for diseases transmissible from companion animals to man were analysed with the aim of developing a procedure suitable for their prioritization within a European framework. A new method and its associated questionnaire and scoring system were designed based on methods described by the World Organisation for Animal Health (OIE). Modifications were applied to allow for the paucity of specific information on companion animal transmissible diseases. The OIE method was also adapted to the subject and to the regional scope of the interprofessional network addressing zoonotic diseases transmitted via companion animals in Europe: the Companion Animals multisectoriaL interprofessionaL Interdisciplinary Strategic Think tank On zoonoses (CALLISTO). Adaptations were made based on information collected from expert groups on viral, bacterial and parasitic diseases using a structured questionnaire, in which all questions were closed-ended. The expert groups were asked to select the most appropriate answer for each question taking into account the relevance and reliability of the data available in the scientific literature. Subsequently, the scoring of the answers obtained for each disease covered by the questionnaire was analysed to obtain two final overall scores, one for human health impact and one for agricultural economic impact. The adapted method was then applied to select the 15 most important pathogens (five for each pathogen group: viral, bacterial and parasitic) on the basis of their overall impact on public health and agriculture. The result of the prioritization exercise was a joint priority list (available at www.callistoproject.eu) of relevant pathogens according to these two criteria. As the scope of CALLISTO was comprehensive in terms of geographical area, animal species involved and impact of the diseases, the list of prioritized diseases had to accommodate the realities in different European countries and the differences in biology and animal-human relationships in a wide range of species including cats and dogs, pet pigs and sheep as well as captive reptiles. The methodology presented in this paper can be used to generate accurate priority lists according to narrower and more specific objectives.

Disease Risk Assessments Involving Companion Animals: an Overview for 15 Selected Pathogens Taking a European Perspective.

Prioritization of companion animal transmissible diseases was performed by the Companion Animals multisectoriaL interprofessionaL Interdisciplinary Strategic Think tank On zoonoses (CALLISTO) project. The project considered diseases occurring in domesticated species commonly kept as pets, such as dogs and cats, but also included diseases occurring in captive wild animals and production animal species. The prioritization process led to the selection of 15 diseases of prime public health relevance, agricultural economic importance, or both. An analysis was made of the current knowledge on the risk of occurrence and transmission of these diseases among companion animals, and from companion animals to man (zoonoses) or to livestock. The literature was scanned for risk assessments for these diseases. Studies were classified as import risk assessments (IRAs) or risk factor analyses (RFAs) in endemic areas. For those pathogens that are absent from Europe, only IRAs were considered; for pathogens present throughout Europe, only RFAs were considered. IRAs were identified for seven of the eight diseases totally or partially absent from Europe. IRAs for classical rabies and alveolar echinococcosis found an increased risk for introduction of the pathogen into officially disease-free areas as a consequence of abandoning national rules and adopting the harmonized EU rules for pet travel. IRAs for leishmaniosis focused on risk associated with the presence of persistently infected dogs in new geographical areas, taking into consideration the risk of disease establishment should a competent vector arise. IRAs for Crimean-Congo haemorrhagic fever and West Nile fever indicated that the likelihood of introduction via companion animals was low. IRAs for bluetongue paid no attention to the risk of introduction via companion animals, which was also the case for IRAs for foot-and-mouth disease, the only disease considered to be absent from Europe. RFAs dealing with the risk factors for companion animals to become infected were identified for eight of the 14 diseases found in Europe or parts of it. RFAs for leptospirosis were most numerous (four studies). The host related risk factor 'age' was identified as significant for dogs in at least two RFAs for cystic echinococcosis and giardiasis. Among husbandry and healthcare related factors, 'eating (uncooked) offal', 'being free roaming' and 'poor deworming practice' were associated with risk for dogs in at least two RFAs for cystic echinococcosis, while 'having received recent veterinary treatment' was identified as a risk factor in at least two studies on infection with extended spectrum beta lactamase-producing bacteria, one in horses and the other in dogs and cats. Finally, although the environmental factors 'season' and 'hydrological density' were identified as significant risk factors for dogs in at least two RFAs for leptospirosis, the inconsistent case definitions used in those studies made comparison of study results problematic. RFAs considering the risk of people becoming infected from companion animals were identified for eight of the 14 diseases found in Europe or parts of it. RFAs for human campylobacteriosis were the most numerous (n = 6). Most studies made an assessment as to whether keeping a pet per se, or keeping a pet with supposed or known risk factors, was a risk factor for people relative to other risks. This allowed some studies to report the population attributable risk or population attributable fraction of the incidence of human disease due to companion animals (for campylobacteriosis, salmonellosis and toxoplasmosis), which is a measure that is easy to perceive for laymen and policy makers. No RFAs were found that dealt with the risk to food animals from companion animals for any of the 15 pathogens investigated. Few risk method-based studies were identified that provided information on risk factors for companion animals and on their role as a source of these 15 selected diseases, indicating a clear knowledge gap. There were not enough assessments for any of the 15 diseases to allow meta-analyses, whether these assessments dealt with companion animal disease risk or companion animal-associated human disease risk. Important method and technology gaps were the lack of harmonization in the case definitions used for a given disease and the lack of good diagnostics allowing pathogen identification to taxonomic levels that are meaningful for risk analysis. Molecular epidemiology studies on zoonotic pathogens, which included companion animals among the potential human risk factors, were not found, although such studies would provide good preliminary insights without requiring any tracing of people or any interviews. In addition to performing further risk studies that take into account these issues, there is a need for responsible pet ownership and continued education of professionals in companion animal zoonoses. Additional risk assessment studies should allow more targeted actions to reduce the risk of zoonotic diseases transferred via companion animals and provide information that will promote risk-awareness in healthy human-animal relationships.

Aflibercept as primary treatment for myopic choroidal neovascularisation: a retrospective study.

AIM The aim of this study is to evaluate long-term efficacy of intravitreal injections of aflibercept as primary treatment for subfoveal/juxtafoveal myopic choroidal neovascularisation (CNV).METHODS Thirty-eight treatment-naive eyes of thirty-eight patients with subfoveal/juxtafoveal myopic CNV received initial intravitreal aflibercept injections and were followed for at least 18 months. Aflibercept was applied again for persistent or recurrent CNV, as required. Statistical analysis was carried out using SPSS.RESULTS Mean patient age was 45.8 years, and mean eye refractive error was -7.79 D. For the total patient group (n=38 eyes), mean logMAR best-corrected visual acuity (BCVA) significantly improved from 0.69 at baseline to 0.15 at 18 months (P<0.01). Over half of the treated eyes obtained resolution with one aflibercept injection. Patients were also grouped according to age, as <50 years (n=20 eyes) and ≥50 years (n=18 eyes). Mean BCVA improvement was significantly greater in eyes of the younger myopic CNV group, compared with those of ≥50 years (0.21 vs 0.35; P<0.05). The mean number of aflibercept injections was 1.8 for the <50 years myopic CNV group, and 3.6 for the ≥50 years myopic CNV group (P<0.001). Correlation between spherical equivalent refraction and final visual acuity reached statistical significance only for the <50 years myopic CNV group (P<0.001; Levene's correlation).CONCLUSIONS Intravitreal aflibercept provides long-term visual acuity improvement in myopic CNV. The <50 years old myopic CNV group had significantly fewer injections, with greater visual acuity improvement. Intravitreal aflibercept in myopic CNV does not require the three-injection loading phase used for aflibercept treatment of neovascular age-related macular degeneration.

Psoriasis beyond the skin surface: a pilot study on the ocular involvement.

The ocular involvement in psoriasis is not a completely well-known problem. The ophthalmologic involvement occurs in about 10 % of patients, particularly in case of arthropathic or pustular psoriasis. Ocular lesions are more common in males, and they often occur during psoriasis exacerbations. Our study aimed to assess the prevalence and type of ocular involvement in psoriasis, by a comparison between psoriasis and healthy subjects, and if/how a 12-week long systemic immunosuppressive therapy is able to modify them. This study involved thirty-two psoriatic patients and thirty-two healthy subjects. Dermatological evaluation was done using Psoriasis Area and Severity Index, Physician Global Assessment, and Dermatology Life Quality Index (PASI, PGA, and DLQI score). Ophthalmological evaluation included ocular surface involvement (Schirmer, Jones, break-up time--BUT, DR-1 camera), retinal pathologies, and ocular surface disease index. Laboratory investigations including the C-reactive protein (CRP) of all the patients were performed. At baseline, the values of Schirmer, Jones, and BUT tests in the patient group were significantly lower compared to controls; moreover, conjunctival hyperemia was more frequent in psoriatic patients than in healthy subjects. Ocular involvement was more prominent in the subset of psoriatic patients with sebo-psoriasis than in general psoriatic population. A statistically significant correlation was found in sebo-psoriasis between PASI and Schirmer, between PASI and Jones, and between PASI and BUT. On the other hand, the results obtained from DR1 camera showed statistically significant difference between psoriatic and sebo-psoriatic patients at the end of the follow-up. After 12 weeks of treatment, the mean values of PASI, PGA, DLQI, CRP, and BUT showed significant changes in psoriatic patients. Our findings suggest a high rate of ocular involvement in psoriatic patients, emphasizing the need of performing periodic ophthalmological examinations in order to avoid underestimating eye diseases and to allow early diagnosis and treatment of patients.

Development of an active risk-based surveillance strategy for avian influenza in Cuba.

The authors designed a risk-based approach to the selection of poultry flocks to be sampled in order to further improve the sensitivity of avian influenza (AI) active surveillance programme in Cuba. The study focused on the western region of Cuba, which harbours nearly 70% of national poultry holdings and comprise several wetlands where migratory waterfowl settle (migratory waterfowl settlements - MWS). The model took into account the potential risk of commercial poultry farms in western Cuba contracting from migratory waterfowl of the orders Anseriformes and Charadriiformes through dispersion for pasturing of migratory birds around the MWS. We computed spatial risk index by geographical analysis with Python scripts in ESRI(®) ArcGIS 10 on data projected in the reference system NAD 1927-UTM17. Farms located closer to MWS had the highest values for the risk indicator pj and in total 31 farms were chosen for targeted surveillance during the risk period. The authors proposed to start active surveillance in the study area 3 weeks after the onset of Anseriformes migration, with additional sampling repeated twice in the same selected poultry farms at 15 days interval (Comin et al., 2012; EFSA, 2008) to cover the whole migration season. In this way, the antibody detectability would be favoured in case of either a posterior AI introduction or enhancement of a previous seroprevalence under the sensitivity level. The model identified the areas with higher risk for AIV introduction from MW, aiming at selecting poultry premises for the application of risk-based surveillance. Given the infrequency of HPAI introduction into domestic poultry populations and the relative paucity of occurrences of LPAI epidemics, the evaluation of the effectiveness of this approach would require its application for several migration seasons to allow the collection of sufficient reliable data.

An integrated web system to support veterinary activities in Italy for the management of information in epidemic emergencies.

The management of public health emergencies is improved by quick, exhaustive and standardized flow of data on disease outbreaks, by using specific tools for data collection, registration and analysis. In this context, the National Information System for the Notification of Outbreaks of Animal Diseases (SIMAN) has been developed in Italy to collect and share data on the notifications of outbreaks of animal diseases. SIMAN is connected through web services to the national database of animals and holdings (BDN) and has been integrated with tools for the management of epidemic emergencies. The website has been updated with a section dedicated to the contingency planning in case of epidemic emergency. EpiTrace is one such useful tool also integrated in the BDN and based on the Social Network Analysis (SNA) and on network epidemiological models. This tool gives the possibility of assessing the risk associated to holdings and animals on the basis of their trade, in order to support the veterinary services in tracing back and forward the animals in case of outbreaks of infectious diseases.

The components of 'One World - One Health' approach.

The interaction between living beings, including men, animals and pathogens, sharing the same environment, should be considered as a unique dynamic system, in which the health of each component is inextricably interconnected and dependent with the others. Nowadays, a new integrated One Health approach is reflecting this interdependence with a holistic view to the ecological system. The One Health approach can be defined as a collaborative and a multidisciplinary effort at local, national and global level to guarantee an optimal healthy status for humans, animals and environment. Strictly related to the One Health concept is to be considered the control of infectious diseases, which have influenced the course of human history. Four different components might be identified as key elements within the 'One World - One Health' (OWOH) approach: the geographical component, the ecological one, the human activities and the food-agricultural ones.

Surveillance strategies for foot and mouth disease to prove absence of disease and absence of viral circulation.

Free trade of animals and their products is based on the international or bilateral recognition of the health status of the animal populations being traded. This recognition is based on documentation of their health status by the exporting country, based on the results of continuing surveillance. According to the Terrestrial Animal Health Code of the World Organisation for Animal Health (OIE), this may be based on various methods of surveillance, such as: documenting non-specific surveillance (clinical surveillance, passive notification of suspect cases, etc.); documenting activities that increase the sensitivity of non-specific surveillance (training activities, rewards/sanctions for notification/failure to notify, etc.); documenting all specific surveillance and its results (random surveys, targeted and risk-based surveillance, convenience-testing activities, etc.). Usually, the infection is the subject of the declaration of freedom. While clinical and passive surveillance can provide a high level of confidence that foot and mouth disease (FMD) infection is absent, this is not the case in vaccinated populations. In these populations, specific surveillance becomes much more important than non-specific clinical surveillance. Specific surveillance is severely restricted by the performance of the test(s) employed. The imperfect specificity of any serological test is further complicated when techniques to differentiate infected from vaccinated animals (DIVA) are used, because imperfect purification of the antigen used for vaccination may foster the production of undesired antibodies in the vaccinated animals. The authors discuss various approaches to overcome this problem; their merits and flaws in documenting the absence of infection or virus circulation for animal diseases in general, and for FMD in particular. Particular attention is paid to finding methods that can be applied in a variety of epidemiological conditions and organisational structures, since these vary greatly among OIE Members.

CD1A and CD1E gene polymorphisms are associated with susceptibility to multiple sclerosis.

Multiple sclerosis (MS) is thought to be an autoimmune T-cell-mediated disease directed at myelin antigens of the central nervous system. Besides myelin proteins, lipid components of CNS are supposed to play a role as antigens for T cells in MS. CD1 is a family of MHC-like glycoproteins specialized in capturing and presenting a variety of microbial and self lipids and glycolipids to antigen-specific T cells. CD1-restricted T cells specific for gangliosides and sulfatide have been isolated from subjects with MS and in mice with experimental allergic encephalopathy. We genotyped exon 2 of CD1A and CD1E in 205 MS patients and 223 unrelated healthy controls and determined their association with the presence of anti-ganglioside and anti-sulfatide antibodies. CD1E 01-01 is associated with a reduced risk of MS (OR 0.54, p=0.001); CD1A 02-02 (OR 1.99, p=0.012) or CD1E 02-02 (OR 2.45, p=0.000) with an increased risk. The combination of the genotypes CD1A 02-02 and CD1E 02-02 is present in 90.7% of patients but in only 9.4% controls (OR 94.16, p= 0.000). CD1A and CD1E polymorphisms contribute to the polygenic susceptibility to MS. The functional effects of CD1 polymorphisms are unknown, however changes in CD1 alleles may affect numerous immunological functions.

Antibody response in cattle vaccinated against bluetongue serotype 8 in Italy.

To assess the immunogenicity of Zulvac 8 Bovis (a commercial inactivated vaccine against bluetongue virus serotype 8 - BTV8) under field conditions, 71 cattle vaccinated according to manufacturer schedule in Verona province (Italy) were tested for the presence of BTV8 neutralizing antibodies at 21, 29, 36, 43, 49, 102 and 201 days post-vaccination (dpv). Another group of 528 BTV8 vaccinated cattle in Mantova province (Italy) was also tested once between 113 and 174 dpv. The vaccine was able to elicit an immune response in 69 (97.2%) and 346 (65.5%) animals of the Verona and Mantova groups, respectively.

West Nile virus transmission in 2008 in north-eastern Italy.

After 10 years, West Nile virus (WNV) re-emerged in Italy in August 2008. As on 31 December 2008, the infection affected eight Provinces in three Regions (Emilia Romagna, Veneto, Lombardy), where a total of 794 cases of WNV infection in 251 equine stables were detected on the basis of the clinical signs and as a result of a serological screening in horses living in the area. Only 4.0% (32/794) of the serologically positive animals showed clinical signs, and the 32 clinical cases were reported in 18 different farms. The observed case-fatality rate was 15.6% (5/32). The confirmed clinical cases were detected from end August to mid October. Significant levels of positivity by RT-PCR were also observed in magpies (Pica pica) (9.1%, 95% confidence levels: 6.1-13.4%), carrion crows (Corvus corone) (7.4%, 95% confidence levels: 3.6-14.4%) and rock pigeons (Columba livia) (12.9%, 95% confidence levels: 7.6-21.2%).

Bluetongue control strategy, including recourse to vaccine: a critical review.

The bluetongue (BT) epidemic that has prevailed in Europe since 2000 is the first example of continental spread of the BT virus (BTV) in large naive populations of susceptible animals. Based on the results of intensive surveillance and research in countries of the southern Mediterranean that were affected by the infection early on in the epidemic, a new strategy for prevention and control of the disease was developed to limit direct losses and to reduce the consequences due to movement restrictions. The basic innovations that were introduced were the use of mass vaccination of all domestic ruminant species to limit the spread of BTV and the use of intensive active surveillance to limit, as far as possible, the zone where movement restrictions must be applied. The novel strategy that was adopted dramatically reduced the number of clinical outbreaks in southern Europe and the Mediterranean Basin and ensured safer animal trade. In 2006, the first BTV-8 epidemic occurred, this time in north-western Europe. During this epidemic, affected countries adopted a 'wait and see' approach. No vaccination was implemented until 2008 and, in many instances, the movement of animals was authorised within restricted areas, thereby facilitating the spread of infection. The delay in administering vaccination was due to the decision to avoid the use of modified live virus vaccines, although this type of vaccine performed satisfactorily in the previous BT epidemics in southern Europe. Bluetongue has demonstrated that the infectious agents present in southern Africa can easily spread to the Mediterranean Basin, which should be considered a single entity as far as the epidemiology of animal diseases is concerned. Therefore, any effective strategy for the prevention and control of animal disease in Europe must take into account this reality and recognise the need for regional surveillance networks that include all the countries that border the Mediterranean.

Long-term control of choroidal neovascularisation secondary to angioid streaks treated with intravitreal bevacizumab (Avastin).

AIM: To evaluate the efficacy of intravitreal bevacizumab (IB) in the long-term control of subfoveal choroidal neovascularisation (CNV) associated with angioid streaks (AS). METHODS: Patients with unilateral active CNV associated with AS were enrolled. Exclusion criteria were previous treatment for CNV and comorbidity. Postoperative visual acuity was defined as a gain or loss of two or more lines of best-corrected visual acuity (BCVA). Post-treatment CNV size was dichotomised into "increased," if the CNV area had grown by > or =200 microm(2), and "stable/reduced" if it had decreased by > or =200 microm(2) or had not changed by more than 200 microm(2). Patients were retreated if no further improvement or worsening was noted. RESULTS: Patients were five males and six females aged 33 to 58 years (mean 46.8 (SD 9.2)), who received a mean number of 3.5 (1.3) IB treatments (min: 2; max: 6). The mean retreatment interval was 3 (1.5) months (min: 1; max: 6). The mean follow-up duration was 23.8 (2.9) months. At 20 months all patients had stable/reduced CNV size and stable/improved BCVA. The mean BCVA rose significantly from 0.28 (0.2) at baseline to 0.56 (0.29) at 20 months (p<0.0001). CONCLUSION: IB is a promising tool for the long-term control of CNV in AS. Further studies are required to validate these findings.

Intravitreal micronized triamcinolone versus triamcinolone acetonide: a clinical and morphological comparative study.

Nowadays many authors suggest the use of intravitreal triamcinolone acetonide (TA) for the treatment of vitreoretinal diseases, although it can be associated with a high risk of local toxicity. In order to develop a safer injection for clinical use, the purpose of our study is to evaluate the in situ safety of two different triamcinolone preparations, a commercially available TA and a micronized triamcinolone. The experiments were performed on 18 adult male age-matched New Zealand rabbits. The clinical examination included funduscopy with an indirect ophthalmoscope and intraocular pressure (IOP) measurement. At the end of the clinical observations, the animals were sacrificed and the eyes enucleated and processed for the morphological evaluation. In our study the main side effect observed was the IOP elevation in the group injected with triamcinolone acetonide. In addition, in the TA-injected group, one eye was enucleated following an endophthalmitis. Our study highlights that doses as low as 4 mg of triamcinolone acetonide injected into the rabbit vitreous may have a local toxic effect in terms of IOP elevation, endophthalmitis occurrence and changes in the retinal morphology. In contrast, the micronized triamcinolone injection shows a less toxic effect in situ, thus suggesting the alternative use of this more reliable preparation which seems to be safer for a clinical use.

An indirect ELISA for the detection of antibody in milk from sheep experimentally infected with Brucella melitensis biovar 3.

An indirect ELISA was evaluated for the detection of Brucella antibodies in milk (m-ELISA) from sheep experimentally infected with B. melitensis biovar 3. At the end of the second reproductive cycle (13 months post infection), the milk of 22 lactating sheep was tested using the m-ELISA. Sera from the same sheep were also tested for Brucella antibodies using the Rose Bengal test (RBT) and the complement fixation test (CFT). The first serum sampling after parturition showed 100% sensitivity in both the RBT and the CFT (confidence interval [CI] 94-100%), but in subsequent samplings the sensitivity of the RBT decreased to 73% (CI 55-85%). Similarly, the sensitivity of the CFT decreased two months after the first sampling, when respective sensitivities of 95% (CI 81-98%) and 81% (CI 61-93%) were recorded for the final two samplings. The sensitivity of the m-ELISA decreased initially (68% on the third sampling, CI 50-81%), but then increased to 95% (CI 81-98%) for the final sampling. When disease prevalence in a flock is below 5%, the estimated probability of not detecting an infected flock through m-ELISA bulk milk testing is over 25%. Under field conditions in Italy (average sheep flock size of 70), the probability that the infection will not be detected is over 25% when four (or less) infected milking sheep are present in the flock. The results show that the m-ELISA is not a reliable screening test for bulk milk samples when the prevalence of brucellosis in a sheep flock is low.

National monitoring and surveillance.

The author describes the characteristics of surveillance systems and the use of surveillance in the three following scenarios: absence of infection, appearance of an exotic/emerging infection and endemic infections. In a population free from an infection, surveillance is used mainly to protect, by means of early detection systems, the population itself from the introduction of the infectious agent from other populations, and to document the health status of the population for international trade purposes. When an exotic infection enters a country, the information required to foresee its possible spread and to plan control and eradication activities is very often missing. As an example of the use of surveillance to collect the information needed to plan control and eradication activities, the author describes the response to the incursion into Europe of bluetongue in the early 2000s. The European brucellosis programme (from 1964 to the present) is taken as an example of the use of surveillance to monitor the control/eradication activities and to steer the control/eradication programme. Finally, the principal challenges currently faced by animal health surveillance professionals are discussed, namely: the methods for gathering information from the wild animal populations, and the methods used to evaluate the equivalence between different surveillance systems based on structured non-random activities and random surveys.

The concept of compartmentalisation.

The rationale for establishing trade 'regions' and 'zones' is based on principles of epidemiological science and risk analysis that assess and manage animal disease risks so that the safety of trade can be ensured. However, the boundaries of geographical regions and zones may readily be breached through numerous epidemiological pathways. The concept of a 'compartment' extends the application of a 'risk boundary' beyond that of a geographical interface and considers all epidemiological factors that can contribute to the creation of an effective boundary. The fundamental requirement for application of either concept (regions/zones or compartments) is that the population considered for trade is maintained within management or geographical boundaries which allow clear epidemiological differentiation to be made between those animals and surrounding populations of higher risk. Seven factors are presented that an exporting country might use to guide the identification and documentation of a compartment. Additionally, the steps that would be undertaken to implement trade based on the compartmentalisation concept are discussed.