Efficacy of heat-inactivated Mycobacterium bovis vaccine delivered to European badgers (Meles meles) through edible bait.

Badgers (Meles meles) are a major tuberculosis (TB) reservoir in Europe, with the potential to transmit infection to cattle. Here we assessed whether a recently described oral tuberculosis vaccine based on heat-inactivated Mycobacterium bovis (HIMB), delivered as edible baits, can protect badgers from infection. Eight badgers were given individually five baits, each one consisting of a ball of peanut butter, natural peanut and oat flakes including a dose of the vaccine containing 5 × 107 colony-forming units. In parallel, a control group of seven badgers did not receive the vaccine. One month and a half later a second dose of the vaccine was offered to the vaccinated group. Ninety-four days after the second dose, all badgers were challenged with M. bovis (103 colony-forming units per animal) delivered endobronchially to the right middle lung lobe. Clinical, immunological, pathological and bacteriological variables were measured throughout the whole study to assess the efficacy of the vaccine. Two vaccinated animals showed high bacterial load of M. bovis and worsening of pathological lesions of TB. Conversely, the other six vaccinated animals showed slight improvement in bacterial load and pathology with respect to the control group. These results suggest that delivering the TB vaccine via food bait can partially protect wild badger populations, although vaccination can lead to either protection or tolerization, likely depending on the animal's immune status and general condition at the time of vaccination. Further optimization of the vaccination trial/strategy is needed to reduce the rate of tolerization, such as altering vaccine dose, number of doses, type of bait, use of adjuvants or route of administration.

Beyond tuberculosis: Diversity and implications of non-tuberculous mycobacteria at the wildlife-livestock interface.

Non-tuberculous mycobacteria (NTM) circulate between the environment, animals and humans entailing a double concern: their ability to interfere with tuberculosis diagnosis and their potential to cause infections in their hosts. However, published records on NTM infections in animals are still scarce. The aims of the present study were to describe the diversity of NTM circulating among wild and domestic species from Spain and to analyze their implications as potential pathogenic microorganisms or as sources of interferences in the diagnosis of bovine tuberculosis. Overall, 293 NTM isolates of 277 animals were obtained from tissue samples collected between 2012 and 2019, and analyzed through a multigene approach for mycobacteria identification. Thirty-one species were identified, being Mycobacterium avium subsp. avium (Maa) and M. avium subsp. hominissuis (Mah), but also M. bouchedurhonense, M. nonchromogenicum and M. lentiflavum, the most abundant ones. Maa and M. lentiflavum were isolated in several animals showing tuberculosis-like lesions. Maa, Mah and M. nonchromogenicum were recovered from many cattle that had reacted to the tuberculin skin test. Other NTM were also associated to these phenomena. These four mycobacterial species were geographically associated between wild boar and other hosts. The findings of the present study suggest that a high diversity of NTM circulates among wildlife and livestock. Wild boar and M. avium seem to play a relevant role in this epidemiological scenario.

Ticks on wild boar in the metropolitan area of Barcelona (Spain) are infected with spotted fever group rickettsiae.

Tick-borne pathogens (TBPs) constitute an emerging public health concern favoured by multidimensional global changes. Amongst these, increase and spread of wild boar (Sus scrofa) populations are of special concern since this species can act as a reservoir of zoonotic pathogens and promote tick abundance. Thus, we aimed to make a first assessment of the risk by TBPs resulting from wild boar and ticks in the vicinity of a highly populated area. Between 2014 and 2016, we collected spleen samples and 2256 ticks from 261 wild boars (out of 438 inspected) in the metropolitan area of Barcelona (MAB; northeast Spain). We morphologically identified four tick species: Hyalomma lusitanicum (infestation prevalence: 33.6%), Dermacentor marginatus (26.9%), Rhipicephalus sanguineus sensu lato (18.9%) and R. bursa (0.2%). Ticks were pooled according to species and individual host. A total of 180 tick pools and 167 spleen samples were screened by real-time PCR and/or reverse line blot hybridization assay for Ehrlichia sp., Anaplasma sp., Babesia sp., Rickettsia sp., Borrelia burgdorferi sensu lato and Coxiella burnetii. Seventy-two out of the 180 tick pools were positive to Rickettsia spp. (minimum prevalence of 8.7%), including Rickettsia massiliae, R. slovaca and R. raoultii. We did not detect Rickettsia spp. in wild boar spleens nor other TBPs in ticks or wild boars. Since the ticks identified can bite humans, and the recorded spotted fever group (SFG) rickettsiae are zoonotic pathogens, there is a risk of SFG rickettsiae transmission for MAB inhabitants. Our results suggest a broader distribution of H. lusitanicum, competent vector for the Crimean-Congo haemorrhagic fever virus than previously known. Wild boar is not a Rickettsia spp. reservoir according to the spleen negative results. However, its abundance could favour tick life cycle and abundance, and its proximity to humans could promote the infection risk by Rickettsia spp.

Interaction Patterns between Wildlife and Cattle Reveal Opportunities for Mycobacteria Transmission in Farms from North-Eastern Atlantic Iberian Peninsula.

Interactions taking place between sympatric wildlife and livestock may contribute to interspecies transmission of the Mycobacterium tuberculosis complex or non-tuberculous mycobacteria, leading to the spread of relevant mycobacterioses or to interferences with the diagnosis of tuberculosis. The aim of this study was to characterize the spatiotemporal patterns of interactions between wildlife and cattle in a low bovine tuberculosis prevalence Atlantic region. Camera traps were set during a one-year period in cattle farms with a history of tuberculosis and/or non-tuberculous mycobacterioses. The frequency and duration of wildlife visits, and the number of individuals per visit, were analysed through generalized linear mixed models. The seasons, type of place, type of point, and period of the day were the explanatory variables. A total of 1293 visits were recorded during 2741 days of camera observation. Only 23 visits showed direct contacts with cattle, suggesting that mycobacteria transmission at the wildlife-livestock interface would occur mainly through indirect interactions. Cattle pastures represented the most appropriate habitat for interspecies transmission of mycobacteria, and badgers' latrines appear to be a potential hotspot for mycobacteria circulation between badgers, wild boars, foxes, and cattle. According to both previous epidemiological information and the interaction patterns observed, wild boars, badgers, foxes, and small rodents are the species or group most often in contact with livestock, and thus may be the most involved in the epidemiology of mycobacterioses in the wildlife-livestock interface in this area.

Risk factors associated to a high Mycobacterium tuberculosis complex seroprevalence in wild boar (Sus scrofa) from a low bovine tuberculosis prevalence area.

Animal tuberculosis is a worldwide zoonotic disease caused principally by Mycobacterium bovis, a member of the Mycobacterium tuberculosis complex (MTC). In southern Iberian Peninsula, wild reservoirs such as the wild boar, among other factors, have prevented the eradication of bovine tuberculosis. However, most of the studies have been focused on south-central Spain, where the prevalence of tuberculosis is high among wild ungulates and cattle herds. In northern regions, where wild boar density and bovine tuberculosis prevalence are lower, fewer studies have been carried out and the role of this species is still under debate. The aim of this study was to describe the temporal and spatial distribution of antibodies against MTC in wild boar from the Basque Country, northern Spain. Sera from 1902 animals were collected between 2010 and 2016. The seroprevalence was determined with an in house enzyme-linked immunosorbent assay and the search of risk factors was assessed by Generalized Linear Models. Overall, 17% of wild boars (326/1902; 95%CI, [15.5%-18.9%]) showed antibodies against MTC. Risk factors associated with seropositivity were the year and location of sampling, the number of MTC positive cattle, the distance to positive farms and the percentage of shrub cover. Younger age classes were associated with increased antibody titres among seropositive individuals. The seroprevalence detected was higher than those previously reported in neighbouring regions. Hence, further studies are needed to better understand the role of wild boar in the epidemiology of tuberculosis in low tuberculosis prevalence areas and consequently, its relevance when developing control strategies.

Detection of Wood Mice (Apodemus sylvaticus) Carrying Non-Tuberculous Mycobacteria Able to Infect Cattle and Interfere with the Diagnosis of Bovine Tuberculosis.

Mycobacterial infections caused by the Mycobacterium tuberculosis complex (MTC) and non-tuberculous mycobacteria (NTM) are of great medical and veterinary relevance. The aim of this research was to study whether small mammals play a role in the epidemiology of mycobacterioses. Four samplings of 100 traps were performed in each of three cattle farms with previous history of tuberculosis or NTM between 2017 and 2018. A total of 108 animals belonging to seven species were trapped, classified, and necropsied, and tissues were submitted to microbiological and molecular methods for mycobacteria identification. The wood mouse (Apodemus sylvaticus) was the most abundant species (87%). No MTC was detected but six different NTM were identified (M. intracellulare, M. avium subsp. paratuberculosis, M. gordonae, M. celatum, M. fortuitum, and a not determined Mycobacterium sp.), showing a prevalence of 6.5%. No significant association was found between mycobacteria prevalence and the analyzed factors. Although a role in the epidemiology of MTC could not be attributed to small mammals, A. sylvaticus carries NTM that could be pathogenic or interfere with the diagnosis of tuberculosis. According to our results, there is a risk of NTM transmission at the wildlife-livestock interface through potential indirect contacts between small mammals and cattle.

A review on the eco-epidemiology and clinical management of human granulocytic anaplasmosis and its agent in Europe.

Anaplasma phagocytophilum is the agent of tick-borne fever, equine, canine and human granulocytic anaplasmosis. The common route of A. phagocytophilum transmission is through a tick bite, the main vector in Europe being Ixodes ricinus. Despite the apparently ubiquitous presence of the pathogen A. phagocytophilum in ticks and various wild and domestic animals from Europe, up to date published clinical cases of human granulocytic anaplasmosis (HGA) remain rare compared to the worldwide status. It is unclear if this reflects the epidemiological dynamics of the human infection in Europe or if the disease is underdiagnosed or underreported. Epidemiologic studies in Europe have suggested an increased occupational risk of infection for forestry workers, hunters, veterinarians, and farmers with a tick-bite history and living in endemic areas. Although the overall genetic diversity of A. phagocytophilum in Europe is higher than in the USA, the strains responsible for the human infections are related on both continents. However, the study of the genetic variability and assessment of the difference of pathogenicity and infectivity between strains to various hosts has been insufficiently explored to date. Most of the European HGA cases presented as a mild infection, common clinical signs being pyrexia, headache, myalgia and arthralgia. The diagnosis of HGA in the USA was recommended to be based on clinical signs and the patient's history and later confirmed using specialized laboratory tests. However, in Europe since the majority of cases are presenting as mild infection, laboratory tests may be performed before the treatment in order to avoid antibiotic overuse. The drug of choice for HGA is doxycycline and because of potential for serious complication the treatment should be instituted on clinical suspicion alone.

On the possible role of ticks in the eco-epidemiology of Coxiella burnetii in a Mediterranean ecosystem.

Ruminant livestock is the main reservoir of Coxiella burnetii (Cb), but little is known about the role of wildlife and ticks in its epidemiology. The Iberian ibex (Capra pyrenaica, Schinz 1838) population of "Ports de Tortosa i Beseit" (NE Spain) suffers intense tick infestations and low reproduction rates. This study aims to (1) assess the relationship between infection in ibexes (detection of serum antibodies and/or of Cb DNA in tissues) and Cb DNA presence in ticks hosted by the same ibexes; and (2) identify Cb associated risk factors. Between 2011 and 2015, serum (n = 130), spleen (n = 72), lymph node (n = 89) and tick (n = 669) samples from 134 hunter-harvested ibexes were collected. Antibody detection was performed by ELISA and Cb DNA presence was assessed by PCR. Potential risk factors were assessed with regression tree models. Although 30% of the ibexes (39/130; 95%CI, [10%-29.8%]) had antibodies, Cb DNA was detected in only 9.8% of the ibexes (11/112; 95%CI [7.6%-27.25%]). The prevalence of Cb-carrier ticks averaged 10% and exceeded 20% for the genus Haemaphysalis. However, lacking correlation between infection in ibexes and their ticks does not support tick-to-ibex transmission or vice versa. Tree modelling points to host, population and environmental factors as drivers of Cb infection in ticks and suggests connections with the domestic cycle. The percentage of Cb-carrier ticks detected is noteworthy. Along with heavy tick infestations, it suggests vector potential for these tick species, especially for the genera Rhipicephalus and Haemaphysalis. Since vector competence has not been assessed in these tick species, a classic vector role cannot be proposed nor discarded, but promoter factors of vector capacity occur. In addition, the risk of tick-borne infection through tick excreta should not be neglected. While the airborne route is the preeminent route for Cb infection, ticks' contribution to Cb epidemiology deserves further attention.