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We analyzed West Nile Virus (WNV) exposure from 1,222 blood donors during 2017–2018 from an area of south-central Spain. Results revealed WNV seroprevalence of 0.08% (95% CI 0.004%–0.4%) in this population. Our findings underscore the need for continued surveillance and research to manage WNV infection in this region.

To estimate the determinants of spatial variation in Crimean-Congo hemorrhagic fever virus (CCHFV) transmission and to create a risk map as a preventive public health tool, we designed a survey of small domestic ruminants in Andalusia, Spain. To assess CCHFV exposure spatial distribution, we analyzed serum from 2,440 sheep and goats by using a double-antigen ELISA and modeled exposure probability with environmental predictors by using generalized linear mixed models. CCHFV antibodies detected in 84 samples confirmed low CCHFV prevalence in small domestic ruminants in the region. The best-fitted statistical model indicated that the most significant predictors of virus exposure risk were cattle/horse density and the normalized difference vegetation index. Model validation showed 99.7% specificity and 10.2% sensitivity for identifying CCHFV circulation areas. To map CCHFV exposure risk, we projected the model at a 1 × 1-km spatial resolution. Our study provides insight into CCHFV ecology that is useful for preventing virus transmission.

Crimean-Congo haemorrhagic fever (CCHF) virus (CCHFV) is a tick-borne zoonotic pathogen that can cause a lethal haemorrhagic disease in humans. Although the virus appears to be endemically established in the Iberian Peninsula, CCHF is an emerging disease in Spain. Clinical signs of CCHFV infection are mainly manifested in humans, but the virus replicates in several animal species. Understanding the determinants of CCHFV exposure risk from animal models is essential to predicting high-risk exposure hotspots for public health action. With this objective in mind, we designed a cross-sectional study of Eurasian wild boar (Sus scrofa) in Spain and Portugal. The study analysed 5,291 sera collected between 2006 and 2022 from 90 wild boar populations with a specific double-antigen ELISA to estimate CCHFV serum prevalence and identify the main determinants of exposure probability. To do so, we statistically modelled exposure risk with host- and environment-related predictors and spatially projected it at a 10 × 10 km square resolution at the scale of the Iberian Peninsula to map foci of infection risk. Fifty-seven (63.3 %) of the 90 populations had at least one seropositive animal, with seroprevalence ranging from 0.0 to 88.2 %. Anti-CCHFV antibodies were found in 1,026 of 5,291 wild boar (19.4 %; 95 % confidence interval: 18.3–20.5 %), with highest exposure rates in southwestern Iberia. The most relevant predictors of virus exposure risk were wild boar abundance, local rainfall regime, shrub cover, winter air temperature and soil temperature variation. The spatial projection of the best-fit model identified high-risk foci as occurring in most of western and southwestern Iberia and identified recently confirmed risk foci in eastern Spain. The results of the study demonstrate that serological surveys of CCHFV vector hosts are a powerful, robust and highly informative tool for public health authorities to take action to prevent human cases of CCHF in enzootic and emergency settings.

A significant gap in exposure data for most livestock and zoonotic pathogens is common for several Latin America deer species. This study examined the seroprevalence against 13 pathogens in 164 wild and captive southern pudu from Chile between 2011 and 2023. Livestock and zoonotic pathogen antibodies were detected in 22 of 109 wild pudus (20.18%; 95% CI: 13.34–29.18) and 17 of 55 captive pudus (30.91%; 95% CI: 19.52–44.96), including five Leptospira interrogans serovars (15.38% and 10.71%), Toxoplasma gondii (8.57% and 37.50%), Chlamydia abortus (3.03% and 12.82%), Neospora caninum (0.00% and 9.52%), and Pestivirus (8.00% and 6.67%). Risk factors were detected for Leptospira spp., showing that fawn pudu have statistically significantly higher risk of positivity than adults. In the case of T. gondii, pudu living in “free-range” have a lower risk of being positive for this parasite. In under-human-care pudu, a Pestivirus outbreak is the most strongly suspected as the cause of abortions in a zoo in the past. This study presents the first evidence of Chlamydia abortus in wildlife in South America and exposure to T. gondii, L. interrogans, and N. caninum in wild ungulate species in Chile. High seroprevalence of livestock pathogens such as Pestivirus and Leptospira Hardjo in wild animals suggests a livestock transmission in Chilean template forest.

The incidence of West Nile fever (WNF) is highly variable in emerging areas, making it difficult to identify risk periods. Using clinical case records has important biases in understanding the transmission dynamics of West Nile virus (WNV) because asymptomatic infections are frequent. However, estimating virus exposure in sentinel species could help achieve this goal at varying spatiotemporal scales. To identify the determinants of inter-annual variation in WNV transmission rates, we designed a 15-year longitudinal seroepidemiological study (2005–2020) in five environmentally diverse areas of southwestern Spain. We modeled individual annual area-dependent exposure risk based on potential environmental and host predictors using generalized linear mixed models. Further, we analyzed the weight of predictors on exposure probability by variance partitioning of the model components. The analysis of 2418 wild ungulate sera (1168 red deer - Cervus elaphus - and 1250 Eurasian wild boar - Sus scrofa) with a highly sensitive commercial blocking ELISA identified an average seroprevalence of 24.9% (95% confidence interval (CI): 23.2–26.7%). Antibody prevalence was slightly higher in wild boar (27.5%; CI: 25.1–30.1%) than in deer (22.2%; CI: 19.8–24.7%). We observed a spatial trend in exposure, with higher frequency in the southernmost areas and a slight, although area-dependent, increasing temporal trend. Host-related predictors were important drivers of exposure risk. The environmental predictor with the highest weight was annual cumulative precipitation, while temperature variations were also relevant but with less weight. We observed a coincidence of spatiotemporal changes in exposure with the notification of WNF outbreaks in horses and humans. That indicates the usefulness of wild ungulates as sentinels for WNV transmission and as models to understand its spatiotemporal dynamics. These results will allow the development of more accurate predictive models of spatiotemporal variations in transmission risk that can inform health authorities to take appropriate action.

Aedes albopictus is an invasive mosquito species that can maintain and transmit several arboviruses causing disease in humans. Understanding the determinants of its ecology and population dynamics to predict its abundance was the main objective of this study.Adult mosquitoes were captured weekly between 2009 and 2016 with BG sentinel traps baited with BG-Lure outdoors at a collection site within the urban area of Palermo (southern Italy). In parallel, between 2012 and 2016, we monitored the uninterrupted weekly abundance of Ae. albopictus at four additional sites nearby over an area of about two hectares. Catches were collected three times per week and mosquitoes were identified morphologically.To identify the determinants of mosquito abundance, seasonal autoregressive integrated moving-average and Poisson regression models were fitted to the weekly abundance of Ae. albopictus with a series of weather predictors that potentially modulate its activity and population dynamics. The time lag of the influence of predictors was analysed to identify the intergenerational environmental determinants of Ae. albopictus population dynamics. A cross-validation of the predictive accuracy of the different models was carried out to select the best predictive model.Over 7 years we captured 12,152 Ae. albopictus in the first trap and another 58,710 in four years of trapping in four additional traps. Aedes albopictus abundance was highly seasonal, with activity between mid-March and late December, highest abundances between July and September, and peak abundances in autumn. The predictive potential of the best model was further externally validated with four years of data from the other four traps, showing a high predictive capacity and a very good fit of seasonality and abundance peaks. Relative humidity, vapour saturation deficit and wind speed were identified as the main determinants of the weekly abundance of Ae. albopictus. The results obtained will allow accurate prediction of the abundance of this invasive mosquito in coastal Mediterranean areas and the design of ad-hoc measures for efficient and environmentally sustainable control.

Abstract Background The ‘snow-melt mosquito’ aedes (rusticoidus) refiki is a rare species with a wide distribution in Europe that is usually defined as an aggressive mosquito for mammals, including humans. During a mosquito survey in a peri-urban area in south-central mainland Spain, adult Ae. refiki females were captured and identified by morphological traits. The presence of this species of mosquito has never been molecularly confirmed under continental dry Mediterranean climatic influence with scarce number of days with snow on soil. The aim of this study was to confirm by amplification and sequencing of mitochondrial cytochrome c oxidase subunit I (COI) and internal transcribed spacer 2 (ITS2) region. Results We also successfully amplified and typed the species molecularly by COI and ITS2 regions. The peri-urban area where Ae. refiki was found contrasts with the reported cold, humid and snowy environments required by the species to breed. Conclusions This finding suggests that the species is already adapted to continental dry Mediterranean environments, questioning whether it is a truly stenotopic species of cold snowy environments.

An experimental infection approach was used to estimate the competence of the common mosquito, Culex pipiens, for hepatitis E virus replication and transmission, using an isolate of hepatitis E virus genotype 3 of human origin in varying infectious doses. The experimental approach was carried out in biosafety level 2 conditions on three batches of 120 Cx. pipiens females, each using an artificial feeding system containing the virus in aliquots of fresh avian blood. Mosquitoes from each batch were collected 1, 7, 14, and 21 days post-infection (dpi) and dissected. The proboscis was subjected to forced excretion of saliva to estimate potential virus transmission. HEV RNA presence in abdomen, thorax, and saliva samples was analyzed by PCR at the selected post-infection times. HEV RNA was detected in the abdomens of Cx. pipiens females collected 1 dpi in the two experimentally-infected batches, but not in the saliva or thorax. None of the samples collected 7–21 dpi were positive. Our results show that Cx. pipiens is not a competent vector for HEV, at least for zoonotic genotype 3.

Tick-borne encephalitis (TBE) is a viral disease endemic in Eurasia. The virus is mainly transmitted to humans via ticks and occasionally via the consumption of unpasteurized milk products. The European Centre for Disease Prevention and Control reported an increase in TBE incidence over the past years in Europe as well as the emergence of the disease in new areas. To better understand this phenomenon, we investigated the drivers of TBE emergence and increase in incidence in humans through an expert knowledge elicitation. We listed 59 possible drivers grouped in eight domains and elicited forty European experts to: (i) allocate a score per driver, (ii) weight this score within each domain, and (iii) weight the different domains and attribute an uncertainty level per domain. An overall weighted score per driver was calculated, and drivers with comparable scores were grouped into three terminal nodes using a regression tree analysis. The drivers with the highest scores were: (i) changes in human behavior/activities; (ii) changes in eating habits or consumer demand; (iii) changes in the landscape; (iv) influence of humidity on the survival and transmission of the pathogen; (v) difficulty to control reservoir(s) and/or vector(s); (vi) influence of temperature on virus survival and transmission; (vii) number of wildlife compartments/groups acting as reservoirs or amplifying hosts; (viii) increase of autochthonous wild mammals; and (ix) number of tick species vectors and their distribution. Our results support researchers in prioritizing studies targeting the most relevant drivers of emergence and increasing TBE incidence.

West Nile virus (WNV) transmission rate is shaped by the interaction between virus reservoirs and vectors, which may be maximized in farm environments. Based on this hypothesis, we screened for WNV in wild birds in three scenarios with decreasing gradient of interaction with horses: (i) the farm (A1); (ii) the neighborhood (A2); and (iii) a wild area (A3). We captured wild birds and analyzed their sera for WNV antibodies by blocking ELISA and micro-virus neutralization test. Flavivirus infections were tested with generic and specific PCR protocols. We parameterized linear mixed models with predictors (bird abundance and diversity, vector abundance, vector host abundance, and weather quantities) to identify Flavivirus spp. and WNV exposure risk factors. We detected a low rate of Flavivirus infections by PCR (0.8%) and 6.9% of the birds were seropositive by ELISA. Exposure to Flavivirus spp. was higher in A1 (9%) than in A2 and A3 (5.6% and 5.8%, respectively). Bird diversity was the most relevant predictor of exposure risk and passerines dominated the on-farm bird community. Our results suggest that measures deterring the use of the farm by passerines should be implemented because the environmental favorability of continental Mediterranean environments for WNV is increasing and more outbreaks are expected.

High impact, mosquito-borne flaviviruses such as West Nile virus (WNV), Usutu virus (USUV), Japanese encephalitis virus (JEV), Tembusu virus (TMUV), and Bagaza/Israel turkey meningoencephalomyelitis virus (BAGV/ITV) are emerging in different areas of the world. These viruses belong to the Japanese encephalitis (JE) serocomplex (JEV, WNV, and USUV) and the Ntaya serocomplex (TMUV and BAGV/ITV). Notably, they share transmission route (mosquito bite) and reservoir host type (wild birds), and some of them co-circulate in the same areas, infecting overlapping mosquito and avian population. This may simplify epidemiological surveillance, since it allows the detection of different infections targeting the same population, but also represents a challenge, as the diagnostic tools applied need to detect the whole range of flaviviruses surveyed, and correctly differentiate between these closely related pathogens. To this aim, a duplex real-time RT-PCR (dRRT-PCR) method has been developed for the simultaneous and differential detection of JE and Ntaya flavivirus serocomplexes. The method has been standardized and evaluated by analyzing a panel of 49 flaviviral and non-flaviviral isolates, and clinical samples of different bird species obtained from experimental infections or from the field, proving its value for virus detection in apparently healthy or suspicious animals. This new dRRT-PCR technique is a reliable, specific and highly sensitive tool for rapid detection and differentiation of JE and Ntaya flavivirus groups in either domestic or wild animals. This novel method can be implemented in animal virology diagnostic laboratories as screening tool in routine surveillance and in the event of bird encephalitis emergence.

Abstract Bulk tank milk (BTM) samples were collected from 81 sheep flocks in the Basque Country, Spain, in 2015 and were analysed for antibodies against Coxiella burnetii by ELISA and for C. burnetii DNA by real-time PCR. Thirty-two percent of the flocks had BTM antibodies against C. burnetii. Presence of C. burnetii DNA in BTM was detected in 23% of the flocks, suggesting recent C. burnetii infections. Retrospective data of BTM samples obtained from 154 sheep flocks investigated in 2005 in the same geographic area were compiled to assess temporal changes in C. burnetii infection. The overall percentage of infected sheep flocks did not significantly change after the 10-year period. Among the 46 flocks sampled in both periods, 11 flocks that were negative in 2005 were positive in 2015, 18 maintained their initial status (positive or negative), and 17 positive flocks were negative in 2015. These findings indicate that C. burnetii infection is a dynamic process in dairy sheep in northern Spain. Single nucleotide polymorphism (SNP) genotyping of positive samples identified three genotypes, SNP1 being the most prevalent in 2015 and SNP8 in 2005; SNP4 was only detected once in 2005. These results suggest possible changes in the pattern of genotype infection over time.

We extend the classical compartmental frameworks for susceptible-infected-susceptible (SIS) and susceptible-infected-recovered (SIR) systems to include an exposed/latent class or a chronic class of infection. Using a suite of stochastic continuous-time Markov chain models we examine the impact of latent and chronic infection on the mean time to extinction of the infection. Our findings indicate that the mean time to pathogen extinction is increased for infectious diseases which cause exposed/latent infection prior to full infection and that the extinction time is increased further if these exposed individuals are also capable of transmitting the infection. A chronic infection stage can decrease or increase the mean time to pathogen extinction and in particular this depends on whether chronically infected individuals incur disease-induced mortality and whether they are able to transmit the infection. We relate our findings to specific infectious diseases that exhibit latent and chronic infectious stages and argue that infectious diseases with these characteristics may be more difficult to manage and control.

Coxiella burnetii, the causal agent of human Q fever and animal Coxiellosis, is a zoonotic infectious bacterium with a complex ecology that results from its ability to replicate in multiple (in)vertebrate host species. Spain notifies the highest number of Q fever cases to the ECDC annually and wildlife plays a relevant role in C. burnetii ecology in the country. However, the whole picture of C. burnetii hosts is incomplete, so this study seeks to better understand the role of micromammals in C. burnetii ecology in the country. Spleen samples from 816 micromammals of 10 species and 130 vaginal swabs from Microtus arvalis were analysed by qPCR to detect C. burnetii infection and shedding, respectively. The 9.7% of the spleen samples were qPCR positive. The highest infection prevalence (10.8%) was found in Microtus arvalis, in which C. burnetii DNA was also detected in 1 of the 130 vaginal swabs (0.8%) analysed. Positive samples were also found in Apodemus sylvaticus (8.7%), Crocidura russula (7.7%) and Rattus rattus (6.4%). Positive samples were genotyped by coupling PCR with reverse line blotting and a genotype II+ strain was identified for the first time in one of the positive samples from M. arvalis, whereas only partial results could be obtained for the rest of the samples. Acute Q fever was diagnosed in one of the researchers that participated in the study, and it was presumably linked to M. arvalis handling. The results of the study are consistent with previous findings suggesting that micromammals can be infected by C. burnetii. Our findings additionally suggest that micromammals may be potential sources to trace back the origin of human Q fever and animal Coxiellosis cases in Europe.

We studied the role of European rabbits (Oryctolagus cuniculus) as a reservoir for Coxiella burnetii in the Iberian region. High individual and population seroprevalences observed in wild and farmed rabbits, evidence of systemic infections, and vaginal shedding support the reservoir role of the European rabbit for C. burnetii.

The red deer (Cervus elaphus) is a relevant reservoir for Coxiella burnetii in Iberia. C. burnetii genotypes that infect red deer also infect humans and domestic animals. Integrated control approaches that target both domestic and wild ruminants are, therefore, required to reduce C. burnetii infection risks in Iberia, especially in wildlife–livestock–human interaction scenarios. The aim of this field experiment was to test the efficacy of an inactivated phase I vaccine [Inactivated phase I vaccine (IPIV); Coxevac®] when used to control C. burnetii shedding prevalence and burden in red deer as a tool to prevent transmission to livestock and humans. A semi-extensively bred red deer population in which C. burnetii is endemic was used as a model of the Iberian context. Around 75% of the reproductive hinds (>1 year old; N = 441) in the population were first vaccinated early in 2012 and were then revaccinated 3 weeks later; they were subsequently revaccinated biannually until January 2014. 75% of the yearling females left as replacement in 2012 and 2013 were vaccinated in June and revaccinated thereafter following the same protocol. 25% of the population, including the replacement females, was kept as a control group throughout the study. Changes in the humoral immune response after vaccination were estimated by analyzing sera collected at 10 different times between January 2011 and January 2015. The vaccinated and control hinds were surveyed at 2.5, 3.5, and 4.5 months after calving in 2012, 2013, and 2014 to collect vaginal swabs, milk, and feces. The presence and burden of C. burnetii DNA in swabs, milk, and feces was evaluated by means of real-time PCR. Vaccination induced high antibody prevalence and levels. The proportion of animals shedding C. burnetii in vaginal secretions and milk did not change over time in the vaccination group with respect to the control group. In contrast, there was a significant reduction in the proportion of deer shedding C. burnetii in feces in both the vaccinated and control groups. The decrease in the proportion of fecal shedders coincided with a significant reduction in the incidence of infection of non-vaccinated yearling females in the population. This finding suggests that long-term vaccination with IPIV could reduce environmental contamination with C. burnetii and control transmission, perhaps making this a promising tool with which to control C. burnetii in red deer in the future.

Crimean-Congo hemorrhagic fever (CCHF) is an emerging tick-borne zoonotic disease caused by the CCHF virus (CCHFV). In this study, an experimental approach combining RT-PCR and proteomics was used for the identification and characterization of CCHFV in 106 ticks from 7 species that were collected from small ruminants in Greece. The methodological approach included an initial screening for CCHFV by RT-PCR followed by proteomics analysis of positive and control negative tick samples. This novel approach allowed the identification of CCHFV-positive ticks and provided additional information to corroborate the RT-PCR findings using a different approach. Two ticks, Dermacentor marginatus and Haemaphysalis parva collected from a goat and a sheep, respectively were positive for CCHFV. The sequences for CCHFV RNA segments S and L were characterized by RT-PCR and proteomics analysis of tick samples, respectively. These results showed the possibility of combining analyses at the RNA and protein levels using RT-PCR and proteomics for the characterization of CCHFV in ticks. The results supported that the CCHFV identified in ticks are genetic variants of the AP92 strain. Although the AP92-like strains probably do not represent a high risk of CCHF to the population, the circulation of genetically diverse CCHFV strains could potentially result in the appearance of novel viral genotypes with increased pathogenicity and fitness.

The genus Culicoides Latreille 1809 is a well-known vector for protozoa, filarial worms and, above all, numerous viruses. The Bluetongue virus (BTV) and the recently emerged Schmallenberg virus (SBV) are responsible for important infectious, non-contagious, insect-borne viral diseases found in domestic ruminants and transmitted by Culicoides spp. Both of these diseases have been detected in wild ruminants, but their role as reservoirs during the vector-free season still remains relatively unknown. In fact, we tend to ignore the possibility of wild ruminants acting as a source of disease (BTV, SBV) and permitting its reintroduction to domestic ruminants during the following vector season. In this context, a knowledge of the composition of the Culicoides species communities that inhabit areas where there are wild ruminants is of major importance as the presence of a vector species is a prerequisite for disease transmission. In this study, samplings were conducted in areas inhabited by different wild ruminant species; samples were taken in both 2009 and 2010, on a monthly basis, during the peak season for midge activity (in summer and autumn). A total of 102,693 specimens of 40 different species of the genus Culicoides were trapped; these included major BTV and SBV vector species. The most abundant vector species were C. imicola and species of the Obsoletus group, which represented 15% and 11% of total numbers of specimens, respectively. At the local scale, the presence of major BTV and SBV vector species in areas with wild ruminants coincided with that of the nearest sentinel farms included in the Spanish Bluetongue Entomological Surveillance Programme, although their relative abundance varied. The data suggest that such species do not exhibit strong host specificity towards either domestic or wild ruminants and that they could consequently play a prominent role as bridge vectors for different pathogens between both types of ruminants. This finding would support the hypothesis that wild ruminants could act as reservoirs for such pathogens, and subsequently be involved in the reintroduction of disease to livestock on neighbouring farms.

Wild and domestic ruminants are susceptible to Bluetongue virus (BTV) infection. Three BTV serotypes (BTV-4, BTV-1 and BTV-8) have been detected in Spain in the last decade. Even though control strategies have been applied to livestock, BTV circulation has been frequently detected in wild ruminant populations in Spain. The aim of the present study is to assess the role for wild ruminants in maintaining BTV after the vaccination programs in livestock in mainland Spain. A total of 931 out 1,914 (48.6%) serum samples, collected from eight different wild ruminant species between 2006 and 2011, were BTV positive by ELISA. In order to detect specific antibodies against BTV-1, BTV-4 and BTV-8, positive sera were also tested by serumneutralisation test (SNT). From the ELISA positive samples that could be tested by SNT (687 out of 931), 292 (42.5%) showed neutralising antibodies against one or two BTV serotypes. For each BTV serotype, the number of outbreaks in livestock (11,857 outbreaks in total) was modelled with pure autoregressive models and the resulting smoothed values, representing the predicted number of BTV outbreaks in livestock at municipality level, were positively correlated with BTV persistence in wild species. The strength of this relationship significantly decreased as red deer (Cervus elaphus) population abundance increased. In addition, BTV RNA was detected by real time RT-PCR in 32 out of 311 (10.3%) spleen samples from seropositive animals. Although BT outbreaks in livestock have decreased substantially after vaccination campaigns, our results indicated that wild ruminants have been exposed to BTV in territories where outbreaks in domestic animals occurred. The detection of BTV RNA and spatial association between BT outbreaks in livestock and BTV rates in red deer are consistent with the hypothesis of virus circulation and BTV maintenance within Iberian wild ruminant populations.

We studied the potential of red deer as bluetongue maintenance hosts and sentinels. Deer maintained detectable bluetongue virus (BTV) serotype 4 RNA for 1 year after the virus was cleared from livestock. However, the virus was not transmitted to yearlings. BTV serotype 1 RNA was detected in red deer immediately after its first detection in cattle.

We confirmed the emergence of bluetongue virus (BTV) in 5 wild ruminant species in Spain. BTV seroprevalence was high and dispersed with time, with a south-to-north gradient. Our results suggest a complex epidemiology of BTV and underline the need for additional research on wildlife in Europe.

Tuberculosis (TB) caused by Mycobacterium bovis and closely related members of the Mycobacterium tuberculosis complex continues to affect humans and animals worldwide and its control requires vaccination of wildlife reservoir species such as Eurasian wild boar (Sus scrofa). Vaccination efforts for TB control in wildlife have been based primarily on oral live BCG formulations. However, this is the first report of the use of oral inactivated vaccines for controlling TB in wildlife. In this study, four groups of 5 wild boar each were vaccinated with inactivated M. bovis by the oral and intramuscular routes, vaccinated with oral BCG or left unvaccinated as controls. All groups were later challenged with a field strain of M. bovis. The results of the IFN-gamma response, serum antibody levels, M. bovis culture, TB lesion scores, and the expression of C3 and MUT genes were compared between these four groups. The results suggested that vaccination with heat-inactivated M. bovis or BCG protect wild boar from TB. These results also encouraged testing combinations of BCG and inactivated M. bovis to vaccinate wild boar against TB. Vaccine formulations using heat-inactivated M. bovis for TB control in wildlife would have the advantage of being environmentally safe and more stable under field conditions when compared to live BCG vaccines. The antibody response and MUT expression levels can help differentiating between vaccinated and infected wild boar and as correlates of protective response in vaccinated animals. These results suggest that vaccine studies in free-living wild boar are now possible to reveal the full potential of protecting against TB using oral M. bovis inactivated and BCG vaccines.

Bluetongue (BT) is still present in Europe and the introduction of new serotypes from endemic areas in the African continent is a possible threat. Culicoides imicola remains one of the most relevant BT vectors in Spain and research on the environmental determinants driving its life cycle is key to preventing and controlling BT. Our aim was to improve our understanding of the biotic and abiotic determinants of C. imicola by modelling its present abundance, studying the spatial pattern of predicted abundance in relation to BT outbreaks, and investigating how the predicted current distribution and abundance patterns might change under future (2011-2040) scenarios of climate change according to the Intergovernmental Panel on Climate Change. C. imicola abundance data from the bluetongue national surveillance programme were modelled with spatial, topoclimatic, host and soil factors. The influence of these factors was further assessed by variation partitioning procedures. The predicted abundance of C. imicola was also projected to a future period. Variation partitioning demonstrated that the pure effect of host and topoclimate factors explained a high percentage (>80%) of the variation. The pure effect of soil followed in importance in explaining the abundance of C. imicola. A close link was confirmed between C. imicola abundance and BT outbreaks. To the best of our knowledge, this study is the first to consider wild and domestic hosts in predictive modelling for an arthropod vector. The main findings regarding the near future show that there is no evidence to suggest that there will be an important increase in the distribution range of C. imicola; this contrasts with an expected increase in abundance in the areas where it is already present in mainland Spain. What may be expected regarding the future scenario for orbiviruses in mainland Spain, is that higher predicted C. imicola abundance may significantly change the rate of transmission of orbiviruses.

Foci of tick-borne pathogens occur at fine spatial scales, and depend upon a complex arrangement of factors involving climate, host abundance and landscape composition. It has been proposed that the presence of hosts that support tick feeding but not pathogen multiplication may dilute the transmission of the pathogen. However, models need to consider the spatial component to adequately explain how hosts, ticks and pathogens are distributed into the landscape. In this study, a novel, lattice-derived, behavior-based, spatially-explicit model was developed to test how changes in the assumed perception of different landscape elements affect the outcome of the connectivity between patches and therefore the dilution effect. The objective of this study was to explain changes in the exposure rate (ER) of red deer to Anaplasma spp. under different configurations of suitable habitat and landscape fragmentation in the presence of variable densities of the potentially diluting host, wild boar. The model showed that the increase in habitat fragmentation had a deep impact on Habitat Sharing Ratio (HSR), a parameter describing the amount of habitat shared by red deer and wild boar, weighted by the probability of the animals to remain together in the same patch (according to movement rules), the density of ticks and the density of animals at a given vegetation patch, and decreased the dilution effect of wild boar on deer Anaplasma ER. The model was validated with data collected on deer, wild boar and tick densities, climate, landscape composition, host vegetation preferences and deer seropositivity to Anaplasma spp. (as a measure of ER) in 10 study sites in Spain. However, although conditions were appropriate for a dilution effect, empirical results did not show a decrease in deer ER in sites with high wild boar densities. The model showed that the HSR was the most effective parameter to explain the absence of the dilution effect. These results suggest that host habitat usage may weaken the predicted dilution effect for tick-borne pathogens and emphasize the importance of the perceptual capabilities of different hosts in different landscapes and habitat fragmentation conditions for predictions of dilution effects.

Abstract Animal tuberculosis (TB) remains a major problem in some countries despite the existence of control programmes focused mainly on cattle. In this species, aerogenous transmission is accepted as the most frequent infection route, affecting mainly the respiratory system. Under the hypothesis that the oral route could be playing a more relevant role in transmission, diagnosis and disease persistence than previously thought, this study was performed to assess the course of TB infection in cattle and its effects on diagnosis depending on the route of entry of Mycobacterium bovis. Two groups of five calves each were either endotracheally (EC) or orally (OC) challenged. Necropsies were carried out 12 weeks after challenge except for three OC calves slaughtered 8 weeks later. All animals reacted to the tuberculin skin test and the entire EC group was positive to the interferon-gamma release assay (IGRA) 2 weeks after challenge and thereafter. The first positive IGRA results for OC calves (3/5) were recorded 4 weeks after challenge. Group comparison revealed significant differences in lesion and positive culture location and scoring. TB-compatible gross lesions and positive cultures were more frequently found in the thorax (p