Brucella spp. in Wildlife of the Lombardy Region, Northern Italy.

Surveillance data collected in the period 2017-20 for Brucella spp. in wildlife of the Lombardy Region in northern Italy were used to describe the exposure of the wildlife species to Brucella spp. in wild boar (Sus scrofa), European brown hare (Lepus europaeus), fallow deer (Dama dama), red deer (Cervus elaphus), and roe deer (Capreolus capreolus). Among the tested species, wild boar (n=6,440) showed the highest percentage of seropositive samples (5.9%). Notably, wild boars of perifluvial area of the Po River showed higher percentages of positivity than those of the pre-Alpine district. In addition, during the hunting season in 2018, 95 organs (uterus or testes, spleen, and submandibular lymph nodes) from wild boar of the perifluvial area of the Po River were collected for bacteriological examination. Brucella suis was isolated in culture from 18.9% of tested lymph nodes. These serological and microbiological results highlight the presence of B. suis in wild boar and suggest the importance of wild boar as a reservoir for B. suis. Comparison of the spatial distribution of Brucella-seropositive wild boars with the location of backyard swine farms revealed a higher chance of contact between the two populations only in the areas where the lower percentage of seropositive samples was observed. Conversely, the high percentage of seropositive samples observed in the Po River area coupled with positive microbiological cultures suggest a greater risk of infection for the humans directly or indirectly involved in wild boar hunting activity. These results may serve as a basis to establish sound wildlife management and to adopt education campaigns aimed at reducing the risk of human infection in people involved in wild boar hunting related activities.

The Use of Flocked Swabs with a Protective Medium Increases the Recovery of Live Brucella spp. and DNA Detection.

Brucellosis is a worldwide zoonosis caused by bacteria from the genus Brucella. Once established, it is very hard to eradicate this disease, since it contaminates animals, the environment, and humans, causing problems for veterinary and public health as well as wildlife protection programs. Swabs are used for sampling in bacteriological and/or molecular diagnostics, from seropositive animals with disease symptoms, from genitalia or tissue lesions, as well as from contaminated environments. The aim of this study was to compare main of the commercially used swab types for sampling and diagnostics of Brucella spp. and determine the optimal storage conditions and time frame for testing. To achieve this, we tested bacterial and molecular methods for detection of Brucella abortus, Brucella melitensis, and Brucella suis using nine swab types, all with different tip materials, treated immediately after spiking, after 72 h at +4°C, and after 72 h at -20°C. Flocked swabs showed the highest capacity to preserve bacterial viability and DNA quality, regardless the storage conditions. Flocked swabs immersed in a protective medium provided the best conditions for Brucella survival in all three storage conditions. At the same time, the efficacy of quantitative PCR (qPCR) detection for all swabs, including the positive control, was above 50%, irrespective of the storage conditions, while bacterial survival was significantly lowered when swabs were kept at +4°C or -20°C for 72 h (48.2% and 27.5%, respectively). Compared to the positive control and other types, the flocked swabs maintained higher reproducibility regarding their capacity to preserve live bacteria in all three storage conditions. IMPORTANCE In order to protect public and veterinary health from highly zoonotic bacteria such as members of the genus Brucella and prevent their dissemination into the environment, direct diagnostics are of utmost importance. However, in addition to the highly specific diagnostic tests, the sampling methods, time necessary for specimens to reach the laboratories, and transport conditions are important factors to consider in order to increase the sensitivity of performed tests, especially bacterial culturing and qPCR. This paper shows how different swab types and storage conditions influence classical bacteriological diagnostics of the most prevalent Brucella species - B. melitensis, B. abortus, and B. suis - but have little impact on molecular methods. The presented results highlight (i) the choice of swab regarding the storage and transport conditions, (ii) the importance of immediate swab treatment upon sampling, and (iii) that molecular methods do not depend on storage conditions, unlike classical bacteriological isolation.

First isolation and characterization of Brucella suis from yak.

Brucella spp., facultative intracellular pathogens that can persistently colonize animal host cells and cause zoonosis, affect public health and safety. A Brucella strain was isolated from yak in Qinghai Province. To detect whether this isolate could cause an outbreak of brucellosis and to reveal its genetic characteristics, several typing and whole-genome sequencing methods were applied to identify its species and genetic characteristics. Phylogenetic analysis based on MLVA and whole-genome sequencing revealed the genetic characteristics of the isolated strain. The results showed that the isolated strain is a B. suis biovar 1 smooth strain, and this isolate was named B. suis QH05. The results of comparative genomics and MLVA showed that B. suis QH05 is not a vaccine strain. Comparison with other B. suis strains isolated from humans and animals indicated that B. suis QH05 may be linked to specific animal and human sources. In conclusion, B. suis QH05 does not belong to the Brucella epidemic species in China, and as the first isolation of B. suis from yak, this strain expands the host range of B. suis.

Phylogeography and epidemiology of Brucella suis biovar 2 in wildlife and domestic swine.

Swine brucellosis due to Brucella suis biovar 2 (bv2) is enzootic in wild boar and hare in continental Europe and may cause major economic losses to the pig industry, mainly in free-ranged pig farms. The high nucleotide identity found among the B. suis biovar 2 isolates has long hindered the full understanding of the epidemiology and the phylogeography of the disease. Here, we used multilocus variable-number tandem-repeat (VNTR) analysis (MLVA) and whole-genome analysis to identify single-nucleotide polymorphisms (SNPs) in order to gain insights from the largest B. suis bv2 dataset analyzed so far composed of domestic pigs and wildlife isolates collected throughout Europe since the 1970s. We found four major clades with a specific phylogeographic pattern. The Iberian clade contains isolates exclusively from the Iberian Peninsula. The Central European clade includes most isolates from France, Northern Italy, Switzerland and an important proportion of those of Northern Spain. The Eastern European clade clustered isolates from Croatia and Hungary mainly but also from areas of France, Germany, Italy and Poland. Finally, a separated Sardinian clade grouped three isolates from this island. At fine scale, MLVA demonstrated an endemic status of the infection in Europe and it allowed tracking a large outbreak formed by different farms from Spain linked to the same infection source. The whole genome SNP analysis showed that the strains form genetically distinct clades, shared between wild boar and pigs, in agreement with the MLVA clades. Interestingly, all hare isolates clustered together within two groups composed exclusively of wildlife isolates. Our results support the hypothesis that maintenance and spread of B. suis bv2 in Europe is a dynamic process linked to the natural expansion of wild boar as the main wild reservoir of the infection, while spread over long distances is found largely dependent on anthropogenic activities.

A headache with surprising outcome: first case of brucellosis caused by Brucella suis biovar 1 in Germany.

In July 2018, brucellosis was diagnosed in a German patient without a travel history to regions endemic for Brucella. Microbiological analysis, including whole-genome sequencing, revealed Brucella suis biovar 1 as the etiologic agent. Core-genome-based multilocus sequence-typing analysis placed the isolate in close proximity to strains originating from Argentina. Notably, despite a strong IgM response, the patient did not develop Brucella-specific IgG antibodies during infection. Here, we describe the clinical course of infection, the extensive epidemiological investigations, and discuss possible routes of transmission.

Fluorescence polarization assay: Diagnostic evaluation for porcine brucellosis.

Brucellosis in pigs, caused by the bacterium Brucella suis, is an important zoonotic infection. In the present study, fluorescence polarization assay (FPA) was standardized and compared with indirect enzyme linked immunosorbent assay (iELISA) and competitive ELISA (cELISA) for diagnosis of porcine brucellosis. Test performances were evaluated using representative panel (n = 100), samples from swine brucellosis outbreak (n = 300), samples from brucellosis suspected animals (n = 291) and sera samples from apparently healthy animals (n = 1121). With panel samples, the FPA cut-off ≥11ΔmP was arrived with sensitivity (Se) and specificity (Sp) of 95.00 and 98.75%, respectively. Testing of samples from swine brucellosis outbreak, the diagnostic Se and Sp of 100 and 95.14% by iELISA, 73.91 and 100% by cELISA and 86.96 and 100% by FPA, respectively were recorded. Similarly, in case of swine brucellosis suspected samples, relative performance of FPA with cELISA had revealed higher kappa value of 0.864 with an accuracy of 93.47. Indirect ELISA was found to be highly sensitive but showed cross reactivity mainly for Yersinia enterocolitica O9 antibodies than cELISA and FPA. The high specificity of FPA test recorded in various types of samples in the study indicated that, FPA could serve as confirmatory test for individual animal diagnosis, outbreak confirmation, surveillance and quarantine of swine brucellosis cases.

A case of Brucella aortitis associated with development of thoracic aortic aneurysm and aortobronchial fistula.

OBJECTIVES: The purpose of this case report is to document the occurrence of granulomatous aortitis complicated by formation of a saccular aneurysm and aortobronchial fistula due to Brucella infection. METHODS: A 65-year-old man with a history of feral swine hunting presented with hemoptysis and was found to have a saccular thoracic aortic aneurysm and associated aortobronchial fistula. The aneurysm underwent operative repair with closure of the aortobronchial fistula. RESULTS: Histopathological examination of the aneurysm wall revealed evidence of granulomatous aortitis. Cultures of the blood and aortic wall tissue were positive for Brucella suis. CONCLUSIONS: Although rare, Brucella infection should be considered in the differential diagnosis of aortic aneurysm with granulomatous aortitis.

Detection of Brucella suis, Campylobacter jejuni, and Escherichia coli Strains in Feral Pig (Sus scrofa) Communities of Georgia.

Feral pigs (Sus scrofa) are an environmentally destructive invasive species that act as a reservoir for zoonotic pathogens. The aim of this study was to determine the presence of Brucella suis, Campylobacter jejuni, and of Escherichia coli in feces of feral pigs from Georgia. Fecal samples were collected from 87 feral pigs from forested and agricultural regions of Georgia. DNA was extracted from the fecal samples and quantitative PCR (qPCR) was used to screen for each of the four pathogens. The qPCR assays indicated that B. suis and eaeA-containing strains of E. coli was present in about 22% and 28% of the samples, respectively. C. jejuni was undetected in any of the feral pig fecal samples. The incidence of B. suis was higher in the pigs from forested region, whereas E. coli strains possessing eaeA gene incidence was higher in the pigs from agricultural regions. In Georgia, feral pigs harbor infectious agents and are a growing threat to the transmission of pathogens to native wildlife, humans, and food crops.

Seroprevalence of Brucella suis in eastern Latvian wild boars (Sus scrofa).

Brucellosis due to Brucella suis biovar 2 is one of the most important endemic diseases in wild boar (Sus scrofa) populations in Europe. The aim of the present study was to determine the seroprevalence of brucellosis in wild boars in the eastern part of Latvia. Wild boars killed by hunters in the period from January to April 2015 (n = 877) and from March to April in 2016 (n = 167) were examined for antibodies against B. suis by the Rose Bengal test (RBT), a complement fixation test (CFT), and by enzyme-linked immunosorbent assays. In 2015, 199 samples (22.7%) were positive by RBT and/or CFT while 36 samples (21.6%) were seropositive in 2016. Of the Brucella seropositive samples from 2015 and 2016 (n = 235), 162 (68.9%) were also seropositive to Yersinia enterocolitica. Considering cross-reactivity of serological tests, the seroprevalence of B. suis biovar 2 exposure in wild boars in the eastern part of Latvia was calculated to 14.0% in 2015 and 9.6% in 2016. From selected seropositive samples (42 in 2015 and 36 in 2016) total DNA was extracted and analyzed with an IS711-based nested polymerase chain reaction (PCR) assay. Species and biovar identification was conducted for bacteria isolated in monoculture from PCR positive samples by species specific primers and Bruce-ladder multiplex PCR. Brucella suis biovar 2 was isolated from 12/20 samples in 2015 and 9/9 samples in 2016. The average seroprevalence was relatively low compared to that found in certain other European countries. Males and females had an equal level of seropositivity, but a positive age-trend was observed for both males and females.

Brucella suis biovar 2 infection in humans in France: emerging infection or better recognition?

Brucellosis is usually acquired by humans through contact with infected animals or the consumption of raw milk from infected ruminants. Brucella suis biovar 2 (BSB2) is mainly encountered in hares and wild boars (Sus scrofa), and is known to have very low pathogenicity to humans with only two case reports published in the literature. Human cases of brucellosis caused by BSB2 were identified through the national mandatory notification of brucellosis. The identification of the bacterium species and biovar were confirmed by the national reference laboratory. Epidemiological data were obtained during medical follow-up visits. Seven human cases were identified between 2004 and 2016, all confirmed by the isolation of BSB2 in clinical specimens. All patients had direct contact with wild boars while hunting or preparing wild boar meat for consumption. Five patients had chronic medical conditions possibly responsible for an increased risk of infection. Our findings suggest that BSB2 might be an emerging pathogen in hunters with massive exposure through the dressing of wild boar carcasses. Hunters, especially those with chronic medical conditions, should be informed about the risk of BSB2 infection and should receive information on protective measures.

Detección serológica de Brucella suis, virus de influenza y virus de la enfermedad de Aujeszky en criaderos porcinos familiares de menos de 100 madres en Argentina / Serological detection of Brucella suis, influenza virus and Aujeszky's disease virus in backyard and small swine holders in Argentina

Los criaderos porcinos de menos de 100 madres representan más del 99% de los de todo el país; sin embargo, existen escasos reportes sobre su situación sanitaria y productiva. Se recabó información productiva y se tomaron muestras para detectar anticuerpos contra Brucella suis (Bs), virus de la enfermedad de Aujeszky (VA) y virus de influenza (VI) en 68 establecimientos de menos de 100 madres ubicados en la región norte, centro y sur del país. El 80% de los establecimientos fueron positivos al VI H1 pandémico 2009, el 11% al H3 clúster 2, mientras que el 11,7% presentó anticuerpos contra el VA y el 6% contra Bs. Ninguno de los productores conocía los factores de riesgo para la transmisión de enfermedades del cerdo al humano. El 47% compra sus reproductores a pares o en ferias. En lo que respecta a normas de bioseguridad, solo el 16% de los establecimientos tenía cerco perimetral y el 37% de las granjas contaba con asesoramiento veterinario. Los resultados de este estudio demuestran que la caracterización productiva y el relevamiento sanitario son de suma importancia para mejorar la productividad y reducir el riesgo de transmisión de enfermedades. El conocimiento de la situación sanitaria y de los factores de riesgo es necesario para conseguir un mejor control y la erradicación de enfermedades en sistemas de baja tecnificación. Se deberían llevar a cabo estudios más representativos a nivel país para detectar los agentes circulantes y, sobre la base de esta información, implementar medidas de prevención y control.

Farmers raising less than 100 sows represent more than 99% of swine producers in Argentina, although little is known about their sanitary status and productive characteristics in the country. Sanitary and productive information was obtained. Furthermore, samples for serological studies were taken to detect antibodies against Brucella suis (Bs), Aujeszky's disease virus (AV) and influenza virus (IV) in 68 backyard and small producers with less than 100 sows located in the north, central and south regions of Argentina. Antibodies against H1 pandemic were detected in 80% of the farms while 11%, 11.7% and 6.0% of the producers were positive to influenza H3 cluster 2, AV and Bs, respectively. None of the producers was aware of the risk factors concerning the transmission of diseases from pigs to humans. A percentage of 47% of them buy pigs for breeding from other farmers and markets. With regard to biosecurity measures, only 16% of the farms had perimeter fences. The results of this study demonstrate that productive characterization and disease surveys are important to improve productivity and to reduce the risk of disease transmission among animals and humans. The study of sanitary status and risk factors is necessary for better control and eradication of diseases in backyard or small producers. More representative studies at country level should be carried out to detect the pathogensthat circulate and, with this knowledge, to implement prevention and control measures.

[Serological detection of Brucella suis, influenza virus and Aujeszky's disease virus in backyard and small swine holders in Argentina]. / Detección serológica de Brucella suis, virus de influenza y virus de la enfermedad de Aujeszky en criaderos porcinos familiares de menos de 100 madres en Argentina.

Farmers raising less than 100 sows represent more than 99% of swine producers in Argentina, although little is known about their sanitary status and productive characteristics in the country. Sanitary and productive information was obtained. Furthermore, samples for serological studies were taken to detect antibodies against Brucella suis (Bs), Aujeszky's disease virus (AV) and influenza virus (IV) in 68 backyard and small producers with less than 100 sows located in the north, central and south regions of Argentina. Antibodies against H1 pandemic were detected in 80% of the farms while 11%, 11.7% and 6.0% of the producers were positive to influenza H3 cluster 2, AV and Bs, respectively. None of the producers was aware of the risk factors concerning the transmission of diseases from pigs to humans. A percentage of 47% of them buy pigs for breeding from other farmers and markets. With regard to biosecurity measures, only 16% of the farms had perimeter fences. The results of this study demonstrate that productive characterization and disease surveys are important to improve productivity and to reduce the risk of disease transmission among animals and humans. The study of sanitary status and risk factors is necessary for better control and eradication of diseases in backyard or small producers. More representative studies at country level should be carried out to detect the pathogensthat circulate and, with this knowledge, to implement prevention and control measures.

Clinical management of Brucella suis infection in dogs and implications for public health.

BACKGROUND: Brucellosis caused by Brucella suis is a notifiable disease that has recently emerged in dogs in New South Wales (NSW). Given the potential for zoonotic transmission, euthanasia of affected dogs is recommended, but this action is not mandatory. We report the clinical management of three dogs that underwent treatment at their owners' request. CASE REPORTS: A 14-month-old spayed female crossbreed originally obtained from an urban animal shelter underwent extensive investigations in 2011-12 for lameness and back pain, culminating in decompressive laminectomy. Diagnosis of multifocal discospondylitis and spinal empyema was made, with B. suis cultured from surgical biopsy specimens. The dog responded to long-term treatment using rifampicin and doxycycline. A second case of B. suis infection was diagnosed in January 2016 in a 3-year-old crossbreed pig-hunting dog with unilateral testicular enlargement. Following serological diagnosis the dog was given preliminary therapy using rifampicin and doxycycline, the affected testis was resected and the patient given a further month of combination therapy. In March 2016 a 7-year-old crossbreed pig-hunting dog with brucellosis was handled similarly, although both testes were removed. CONCLUSION: Brucellosis should be considered in the differential diagnosis of back pain, discospondylitis, lameness, abortion, prostatic abscessation and testicular/epididymal enlargement in dogs, especially if there is exposure to feral pigs or consumption of uncooked feral pig meat. Euthanasia is the only guarantee of reducing the public health risk to zero. However, where treatment is desired by the owner, combination therapy using rifampicin and doxycycline appears to be effective, when combined with surgical resection of infected tissues. Further monitoring of dogs during and after treatment is required to document cure.

Human brucellosis in France in the 21st century: Results from national surveillance 2004-2013.

Brucellosis is a bacterial zoonotic disease mainly transmitted to humans by ruminants. In France, brucellosis has disappeared from ruminants herds. Human brucellosis surveillance is performed through mandatory notification and the national reference center. METHODS: We report the results of human brucellosis surveillance from 2004 to 2013 with regards to epidemiological, clinical and microbiological data. RESULTS: A total of 250 cases were notified, making an annual incidence of 0.3 cases per million inhabitants. Brucella melitensis biovar 3 was the most frequently identified bacterium (79% of isolated strains). In total, 213 (85%) cases had been contaminated abroad in endemic countries. In 2012, an episode of re-emergence of brucellosis in cattle occurred in Haute-Savoie, in the French Alps, and was responsible for 2 human cases. CONCLUSION: Brucellosis has become a disease of travelers in France. However, maintaining a stringent epidemiological surveillance is necessary to be able to early detect any local re-emergence in humans or animals. The multidisciplinary surveillance was implemented in France years ago and is a successful example of the One Health Concept.

Emergence of Brucella suis in dogs in New South Wales, Australia: clinical findings and implications for zoonotic transmission.

BACKGROUND: Animal reservoirs of brucellosis constitute an ongoing threat to human health globally, with foodborne, occupational and recreational exposures creating opportunities for transmission. In Australia and the United States, hunting of feral pigs has been identified as the principal risk factor for human brucellosis due to Brucella suis. Following increased reports of canine B. suis infection, we undertook a review of case notification data and veterinary records to address knowledge gaps about transmission, clinical presentation, and zoonotic risks arising from infected dogs. RESULTS: Between 2011 and 2015, there was a 17-fold increase in the number of cases identified (74 in total) in New South Wales, Australia. Spatial distribution of cases largely overlapped with high feral pig densities in the north of the state. Ninety per cent of dogs had participated directly in pig hunting; feeding of raw feral pig meat and cohabitation with cases in the same household were other putative modes of transmission. Dogs with confirmed brucellosis presented with reproductive tract signs (33 %), back pain (13 %) or lameness (10 %); sub-clinical infection was also common (40 %). Opportunities for dog-to-human transmission in household and occupational environments were identified, highlighting potential public health risks associated with canine B. suis infection. CONCLUSIONS: Brucellosis due to B. suis is an emerging disease of dogs in Australia. Veterinarians should consider this diagnosis in any dog that presents with reproductive tract signs, back pain or lameness, particularly if the dog has a history of feral pig exposure. Moreover, all people in close contact with these dogs such as hunters, household contacts and veterinary personnel should take precautions to prevent zoonotic transmission.

Serological and molecular investigation for brucellosis in swine in selected districts of Uganda.

Brucellosis is a notifiable zoonotic disease affecting livestock, humans, and wildlife in Uganda. Pigs can be infected with human pathogenic Brucella suis biovars 1 and 3 and can be a significant source of brucellosis for humans. Uganda has a rapidly growing pig population, and the pork consumption per capita is the highest in East Africa. The objective of this work was to determine the seroprevalence of brucellosis in Ugandan pigs. A cross-sectional serosurvey of pigs was conducted in three of the major pig-keeping districts in Uganda (Masaka (n = 381 samples), Mukono (n = 398), and Kamuli (n = 414)). In addition, pigs originating from these districts were sampled in the major pig abattoir in Kampala (n = 472). In total, 1665 serum samples were investigated by serological and molecular tests. Only three putative brucellosis-positive samples were detected serologically using indirect ELISA. These sera were found negative for Brucella antibodies by CFT; however, two had antibodies against Yersinia enterocolitica as determined by SAT. Presence of antibodies against Yersiniae was confirmed by Y. enterocolitica antibody-specific ELISA. The two Yersiniae ELISA-positive samples were brucellosis negative using real-time PCR. We tested additional 142 sera from the 1665 samples with real-time PCR. All tested negative. Under this type of production system, we expect a maximum B. suis prevalence of less than 1 % at 95 % confidence level, and therefore, the risk of acquiring brucellosis from the pigs or their products is negligible. However, pigs may harbor the zoonotic Y. enterocolitica. This is the first study to investigate the occurrence of brucellosis in pigs in Uganda and the first study to report Y. enterocolitica antibodies in swine in Uganda.

Brucella suis bacteremia misidentified as Ochrobactrum anthropi by the VITEK 2 system.

Ochrobactrum and Brucella are genetically related genera of the family Brucellaceae, sharing 98.8% rRNA similarity. Because of their phenotypic similarity, Ochrobactrum can be miscoded as Brucella by automated identification systems. The misidentification on blood cultures (BCs) of B. suis as O. anthropi by the VITEK 2 system is herein described. A 67-year-old male with a prosthetic mitral valve and fever was admitted with bacteremia due to a Gram-negative coccobacillus identified as O. anthropi by VITEK 2. The patient's fever persisted along with positive blood cultures despite specific antimicrobial treatment. Due to this adverse outcome, the patient was interrogated again and admitted having domestic swine. Serological tests were positive for acute brucellosis. Polymerase chain reaction restriction fragment length polymorphism (PCR-RFLP) of BC strains identified B. suis biovar 1. Timely identification of Brucella is essential for providing proper treatment to the patient and for advising safe handling of laboratory cultures in biological safety cabinets to prevent laboratory-acquired infection. Countries where brucellosis is endemic must be aware of this possibility.

The first report of Brucella suis biovar 1 isolation in human in Turkey.

BACKGROUND: Brucella melitensis and B. abortus are the species generally isolated from human samples in Turkey. Several studies have also demonstrated the presence of antibodies against B. canis. CASE REPORT AND STUDY: Brucella spp. was isolated from blood culture from a 35-year-old male with clinical signs and symptoms of acute meningitis, including fever lasting for 1 week. Multiplex PCR demonstrated B. suis, and biochemical features indicated biovar 1. CONCLUSIONS: This report is the first emphasizing that B. suis should be considered among the causes of brucellosis in Turkey.

First report of Brucella suis biovar 2 in a semi free-range pig farm, Italy.

This communication describes the isolation of Brucella suis (B. suis) biovar 2 in semi­free­range pigs located in the province of Rome, Italy. Sera of 28 pigs from a herd with reproductive problems were tested for brucellosis. Twenty-five sera (89%) were found positive to Rose Bengal Test (RBT), while 22 (79%) were positive to Complement Fixation Test (CFT). Two positive pigs were slaughtered, organs were collected and tested for the presence of bacteria. Brucella spp. was isolated from the spleens and the abdominal lymph nodes of the 2 subjects. The isolates were identified as B. suis biovar 2 by biochemical and Polymerase Chain Reaction (PCR) tests. The frequent infringement in the fences of the premises and the birth of striped piglets provided evidence that sows mated with wild boar, the major reservoir of B. suis biovar 2. Conversely, the isolation of B. suis biovar 2 from spleens and lymphnodes of seropositive slaughtered animals only, as well as the constant negative results from all vaginal swabs and the abortion materials tested, raise doubts on the implication of B. suis biovar 2 in the infertility of the holding.