If You're Not Confused, You're Not Paying Attention: Ochrobactrum Is Not Brucella.

Bacteria of the genus Brucella are facultative intracellular parasites that cause brucellosis, a severe animal and human disease. Recently, a group of taxonomists merged the brucellae with the primarily free-living, phylogenetically related Ochrobactrum spp. in the genus Brucella. This change, founded only on global genomic analysis and the fortuitous isolation of some opportunistic Ochrobactrum spp. from medically compromised patients, has been automatically included in culture collections and databases. We argue that clinical and environmental microbiologists should not accept this nomenclature, and we advise against its use because (i) it was presented without in-depth phylogenetic analyses and did not consider alternative taxonomic solutions; (ii) it was launched without the input of experts in brucellosis or Ochrobactrum; (iii) it applies a non-consensus genus concept that disregards taxonomically relevant differences in structure, physiology, population structure, core-pangenome assemblies, genome structure, genomic traits, clinical features, treatment, prevention, diagnosis, genus description rules, and, above all, pathogenicity; and (iv) placing these two bacterial groups in the same genus creates risks for veterinarians, medical doctors, clinical laboratories, health authorities, and legislators who deal with brucellosis, a disease that is particularly relevant in low- and middle-income countries. Based on all this information, we urge microbiologists, bacterial collections, genomic databases, journals, and public health boards to keep the Brucella and Ochrobactrum genera separate to avoid further bewilderment and harm.

Core Genome Multilocus Sequence Typing and Single Nucleotide Polymorphism Analysis in the Epidemiology of Brucella melitensis Infections.

The use of whole-genome sequencing (WGS) using next-generation sequencing (NGS) technology has become a widely accepted method for microbiology laboratories in the application of molecular typing for outbreak tracing and genomic epidemiology. Several studies demonstrated the usefulness of WGS data analysis through single-nucleotide polymorphism (SNP) calling from a reference sequence analysis for Brucella melitensis, whereas gene-by-gene comparison through core-genome multilocus sequence typing (cgMLST) has not been explored so far. The current study developed an allele-based cgMLST method and compared its performance to that of the genome-wide SNP approach and the traditional multilocus variable-number tandem repeat analysis (MLVA) on a defined sample collection. The data set was comprised of 37 epidemiologically linked animal cases of brucellosis as well as 71 isolates with unknown epidemiological status, composed of human and animal samples collected in Italy. The cgMLST scheme generated in this study contained 2,704 targets of the B. melitensis 16M reference genome. We established the potential criteria necessary for inclusion of an isolate into a brucellosis outbreak cluster to be ≤6 loci in the cgMLST and ≤7 in WGS SNP analysis. Higher phylogenetic distance resolution was achieved with cgMLST and SNP analysis than with MLVA, particularly for strains belonging to the same lineage, thereby allowing diverse and unrelated genotypes to be identified with greater confidence. The application of a cgMLST scheme to the characterization of B. melitensis strains provided insights into the epidemiology of this pathogen, and it is a candidate to be a benchmark tool for outbreak investigations in human and animal brucellosis.

Origins and global context of Brucella abortus in Italy.

BACKGROUND: Brucellosis is a common and chronic disease of cattle and other bovids that often causes reproductive disorders. Natural infection in cattle is caused by Brucella abortus and transmission typically occurs during abortions, calving, or nursing. Brucellosis is also a major zoonotic disease due to contamination of dairy products or contact with the tissues of infected animals. Brucellosis has been eradicated from most of the developed world in the last 40 years but persists in many regions-the disease remains prevalent in portions of Africa, the Middle East, Asia, and Central and South America, as well as in the Mediterranean basin. In Italy, B. abortus has persisted in southern regions in both cattle and water buffalo. Previous attempts at analyzing the phylogenetics of B. abortus in Italy have been challenging due to limited genetic variability and unresolved global population genetic structure of this pathogen. RESULTS: We conducted genome-wide phylogenetic analyses on 11 representative strains of B. abortus from Italy, and compared these sequences to a worldwide collection of publically available genomes. Italian isolates belong to three clades that are basal to the main and global B. abortus lineage. Using six SNP-based assays designed to identify substructure within the Italian clades, we surveyed a collection of 261 isolates and found that one clade predominates throughout endemic districts in the country, while the other two clades are more geographically restricted to portions of southern Italy. CONCLUSIONS: Although related strains exist worldwide, B. abortus isolates from Italy are substantially different than those found in much of the rest of Europe and North America, and are more closely related to strains from the Middle East and Asia. Our assays targeting genetic substructure within Italy allowed us to identify the major lineages quickly and inexpensively, without having to generate whole genome sequences for a large isolate collection. These findings highlight the importance of genetic studies to assess the status and the history of pathogens.

The optimisation of Salmonella surveillance programmes for pullet and layer farms using local farm density as a risk factor.

Human salmonellosis cases are often caused by Salmonella serovars Enteritidis and Typhimurium and associated with the consumption of eggs and egg products. Many countries therefore implemented general surveillance programmes on pullet and layer farms. The identification of risk factors for Salmonella infection may be used to improve the performance of these surveillance programmes. The aims of this study were therefore to determine 1) whether local farm density is a risk factor for the infection of pullet and layer farms by Salmonella Enteritidis and Typhimurium and 2) whether the sampling effort of surveillance programmes can be reduced by accounting for this risk factor, while still providing sufficient control of these serovars. We assessed the importance of local farm density as a risk factor by fitting transmission kernels to Israeli surveillance data during the period from June 2017 to April 2019. The analysis shows that the risk of infection by serovars Enteritidis and Typhimurium significantly increased if infected farms were present within a radius of approximately 4 km and 0.3 km, respectively. We subsequently optimized a surveillance programme that subdivided layer farms into low and high risk groups based on the local farm density with and allowed the sampling frequency to vary between these groups. In this design, the pullet farms were always sampled one week prior to pullet distribution. Our analysis shows that the risk-based surveillance programme is able to keep the between-farm R0 of serovars Enteritidis and Typhimurium below 1 for all pullet and layer farms, while reducing the sampling effort by 32% compared to the currently implemented surveillance programme in Israel. The results of our study therefore indicate that local farm density is an important risk factor for infection of pullet and layer farms by Salmonella Enteritidis and Typhimurium and can be used to improve the performance of surveillance programmes.

Survey of Mycobacterium spp. in Eurasian Badgers (Meles meles) in Central Italy.

A survey to determine the presence of Mycobacterium spp. in the Abruzzo and Molise regions was conducted by testing samples from 124 badgers found dead or road-killed during the 2013-2021 period. Head lymph nodes were collected from all carcasses, as well as mediastinal lymph nodes from 20 of them, for bacteriological and molecular tests; tissues were inoculated onto a set of solid egg-based Lowenstein-Jensen media and in a liquid culture system (BACTEC) and were analyzed by polymerase chain reactions (PCRs). Organs and lymph nodes from 31 carcasses were collected for histological tests. During post-mortem examinations, macroscopic lesions consistent with a Mycobacterium tuberculosis complex (MTBC) and with nontuberculous mycobacteria (NTM) infections were not detected. Mycobacteria were isolated from four animals (3.22%). M. avium subsp. avium was isolated by head lymph nodes from two badgers (1.61%), M. avium subsp. paratuberculosis (0.80%) from one, and Mycobacterium spp. from another (0.80%). The significance of nontuberculous mycobacteria (NTM) in wildlife hosts in the absence of clinical signs and gross pathology has yet to be assessed. The most critical aspect came from isolates belonging to the Mycobacterium avium complex infection in wildlife due to the possible interference with tuberculin skin tests in cattle.

A Retrospective Study (2015-2020) on the Risk Factors Associated with the Persistence and Spread of Brucellosis in Buffalo Farms in Caserta Province, Italy.

Bovine and bubaline brucellosis is still present in some regions of Italy. Although control and eradication measures have been implemented for several years, the brucellosis situation remains problematic in the Campania region. The infection is present in the provinces of Salerno and Caserta, with the latter experiencing a drastic increase in the prevalence and incidence of infection in buffalo species (Bubalus bubalis) in recent years. The brucellosis eradication plan in Italy is subject to the European co-financing system, and failure to achieve the objectives of the plan has resulted in economic cuts for the Campania Region for years. This study aimed to evaluate the possible risk factors associated with the spread and persistence of brucellosis infection on buffalo farms in the Province of Caserta. The results of official controls carried out from 2015 to 2020 on the buffalo farms of the Province were analyzed. Statistical analysis was performed by means of the R software (version 4.1.0) on a final dataset consisting of 4583 observations. The possible association between covariates and outcome (presence/absence of infection) was evaluated (T-Fisher and Wilcoxon). A logistic regression model with mixed effects was carried out. The study shows that the risk of infection is statistically associated with the density of farms per square km and previous notifications of abortions on the same farms. Furthermore, animal movements constitute a risk factor for the permanence of infection over time (OR > 1), and herds already infected prior to 2015 were seen to have an almost three-fold higher risk of developing the disease (OR = 3.35).

Genetic Diversity of Brucella melitensis Isolated from Domestic Ruminants in Iraq.

The control and eradication of brucellosis represents a critical objective for Veterinary and Health Authorities across several countries globally. Efficient surveillance programs play a pivotal role in detecting and managing outbreaks. Epidemiological investigations significantly benefit from standardized and efficient molecular typing techniques and analytical tools, enabling public health laboratories to identify the origin of outbreaks. This study aimed to sequence Brucella spp. strains isolated in Iraq from different ruminant species to verify their molecular epidemiological correlations and, above all, to shed a light on how these Iraqi isolates are positioned in the phylogenetic context of Brucella spp. The 35 isolates under study were from abortion, milk, placenta, and the fetal membranes of sheep, cattle, and buffalo. Genotyping involved various techniques: MLVA-16, Whole Genome Sequencing, MLST, and cgMLST. All the Iraqi isolates from our study clustered in MLVA-16 within the East Mediterranean clade, and all but one grouped together in the same branch of the MST tree. MST analysis showed the minimum distance of one allele between the studied isolates, except for one strain from buffalo, which was positioned farther away from the rest of the isolates. In cgMLST, the majority of strains grouped within a large cluster predominantly comprising genotypes from the Middle East. The application of different control measures in different territories based on molecular epidemiological studies would increase the chances of maximizing public health benefits and minimizing the spread of infection to disease-free or lower prevalence areas.

Brucellosis and One Health: Inherited and Future Challenges.

One Health is the collaborative efforts of multiple disciplines to attain optimal health for people, animals and the environment, a concept that historically owes much to the study of brucellosis, including recent political and ethical considerations. Brucellosis One Health actors include Public Health and Veterinary Services, microbiologists, medical and veterinary practitioners and breeders. Brucellosis awareness, and the correct use of diagnostic, epidemiological and prophylactic tools is essential. In brucellosis, One Health implementation faces inherited and new challenges, some aggravated by global warming and the intensification of breeding to meet growing food demands. In endemic scenarios, disease awareness, stakeholder sensitization/engagement and the need to build breeder trust are unresolved issues, all made difficult by the protean characteristics of this zoonosis. Extended infrastructural weaknesses, often accentuated by geography and climate, are critically important. Capacity-building faces misconceptions derived from an uncritical adoption of control/eradication strategies applied in countries with suitable means, and requires additional reference laboratories in endemic areas. Challenges for One Health implementation include the lack of research in species other than cattle and small ruminants, the need for a safer small ruminant vaccine, the need to fill in the infrastructure gap, the need for realistic capacity-building, the creation of reference laboratories in critical areas, and the stepwise implementation of measures not directly transposed from the so-called developed countries.

Insights for brucellosis eradication in Italy through a model-based spread evaluation in grazing livestock - Sicily case study.

Brucellosis is one of the world's major zoonotic pathogens and is responsible for enormous economic losses as well as considerable human morbidity in endemic areas. Definitive control of human brucellosis requires control of brucellosis in livestock through practical solutions that can be easily applied to the field. In Italy, brucellosis remains endemic in several southern provinces, particularly in Sicily Region. The purpose of this paper is to describe the developed brucellosis model and its applications, trying to reproduce as faithfully as possible the complex transmission process of brucellosis accounting for the mixing of grazing animals. The model focuses on the contaminated environment rather than on the infected animal, uses real data from the main grazing areas of the Sicily Region, and aims to identify the best control options for minimizing the spread (and the prevalence) and to reach the eradication within the concerned areas. Simulation results confirmed the efficacy of an earlier application of the controls, showed the control should take place 30 days after going to pasture, and the culling time being negligible. Moreover, results highlighted the importance of the timing of both births and grazing pastures (and their interaction) more than other factors. As these factors are region­specific, the study encourages the adoption of different and new eradication tools, tuned on the grazing and commercial behavior of each region. This study will be further extended to improve the model's adaptability to the real world, with the purpose of making the model an operational tool able to help decision makers in accelerating brucellosis eradication in Italy.

Proteomics and bioinformatics investigations to improve serological diagnosis of canine brucellosis.

PURPOSE: Brucella canis is pathogenic for dogs and humans. Serological diagnosis is a cost-effective approach for disease surveillance, but a major drawback of current serological tests is the cross-reactivity with other bacteria that results in false positive reactions. Development of indirect tests with improved sensitivity and specificity that use selected B. canis proteins instead of the whole antigen remain a priority. EXPERIMENTAL DESIGN: A western blotting assay was developed to define the serum antibody patterns associated to infection using a panel of positive and negative dog sera. B. canis positive sera recognized immunogenic bands ranging from 7 to 30 kDa that were then submitted to ESI-LC-MS/MS and analyzed by bioinformatics tools. RESULTS: A total of 398 B. canis proteins were identified. Bioinformatics tools identified 16 non cytoplasmic immunogenic proteins predicted as non-homologous with the most important Brucella cross-reactive bacteria and nine B. canis proteins non-homologous to B. ovis; among the latter, one resulted non-homologous to B. melitensis. Data are available via ProteomeXchange with identifier PXD042682. CONCLUSIONS AND CLINICAL RELEVANCE: The western blotting test developed was able to distinguish between infected and non-infected animals and may serve as a confirmatory test for the serological diagnosis of B. canis. The mass spectrometry and in silico results lead to the identification of specific candidate antigens that pave the way for the development of more accurate indirect diagnostic tests.

Risk Factors for the Spread of Brucellosis in Sheep and Goats in the Campania Region in the Years 2015-2020.

Brucella is a Gram-negative facultative intracellular pathogen that causes infection in sheep and goats (B. melitensis.); B. melitensis can also infect other animals. Sheep and goat brucellosis is still present in some regions of Italy, including Campania, and causes considerable economic losses and health threats. The aim of this study was to evaluate the possible risk factors influencing the spread of brucellosis among sheep and goat farms in the Campania region in order to provide the local veterinary services with practical support in evaluating and planning diagnostic, preventive and control interventions. The results of official controls for brucellosis carried out from 2015 to 2020 in the sheep and goat farms of the Campania Region were analyzed. Data were extracted from the National Veterinary Information Systems and the Laboratory Management System of the Experimental Zooprophylactic Institute of Southern Italy. Statistical analysis was carried out through the software R version 4.1.0; the dataset consisted of 37,442 observations, and 9 qualitative and quantitative variables were evaluated on 8487 farms, 248 of which were positive. The association between covariates and the outcome (presence/absence of the disease) was evaluated (Fisher and Wilcoxon tests). A logistic regression model with mixed effects was carried out. This study confirmed that brucellosis in sheep and goats in the Campania region mostly occurs through contact with infected animals imported from other farms (OR = 3.41-IC 95% [1.82-6.41]). Farms with a greater number of animals were seen to be at the greatest risk of infection (OR = 1.04-IC 95% [1.03-1.05]); previous suspension of healthy status also proved to be a risk factor (OR = 55.8-IC 95% [26.7-117]).

Evaluation of Three Serological Tests for Diagnosis of Canine Brucellosis.

Canine brucellosis caused by Brucella canis, is an infectious disease affecting dogs and wild Canidae. Clinical diagnosis is challenging, and laboratory testing is crucial for a definitive diagnosis. Various serological methods have been described, but their accuracy is uncertain due to limited validation studies. The present study aimed to evaluate the performances of three serological tests for the diagnosis of B. canis in comparison with bacterial isolation (gold standard), in order to establish a protocol for the serological diagnosis of canine brucellosis. A panel of sera from naturally infected dogs (n = 61), from which B. canis was isolated, and uninfected dogs (n = 143), negative for B. canis isolation, were tested using microplate serum agglutination (mSAT), complement fixation performed using the Brucella ovis antigen (B. ovis-CFT), and a commercial immunofluorescence assay (IFAT). The sensitivity and specificity of the three serological methods were, respectively, the following: 96.7% (95% CI 88.8-98.7%) and 92.3 (95% CI 86.7-95.1%) for mSAT; 96.7% (95% CI 88.8-98.7%) and 96.5 (95% CI 92.1-98.2%) for B. ovis-CFT; 98.4% (95% CI 91.3-99.4%) and 99.3 (95% CI 96.2-99.8%) for IFAT. The use in of the three methods in parallel, combined with bacterial isolation and molecular methods, could improve the diagnosis of the infection in dogs.

Brucella abortus Strain RB51 Administered to Prepubescent Water Buffaloes, from Vaccination to Lactation: Kinetics of Antibody Response and Vaccine Safety.

Brucella RB51 is a live modified vaccine. Its use in water buffalo has been proposed using a vaccination protocol different to that used for cattle, but knowledge of the long-term effects of RB51 vaccination in this species remains incomplete. The aim of the study was to evaluate the safety and kinetics of antibody responses in water buffaloes vaccinated according to the protocol described for the bovine species in the WOAH Manual, modified with the use of a triple dose. Water buffaloes were vaccinated with the vaccine RB51. A booster vaccination was administered at 12 months of age. When turning 23-25 months old, female animals were induced to pregnancy. RB51-specific antibodies were detected and quantified using a CFT based on the RB51 antigen. Vaccinated animals showed a positive serological reaction following each vaccine injection, but titers and the duration of the antibody differed among animals. For 36 weeks after booster vaccination, the comparison of CFT values between vaccinated and control groups remained constantly significant. Afterwards, antibody titers decreased. No relevant changes in antibody response were recorded during pregnancy or lactation. In conclusion, results indicated that the vaccination schedule applied is safe and allows for vaccinated and unvaccinated controls to be discriminated between for up to 8 months after booster vaccination.

The emergence of Brucella canis as a public health threat in Europe: what we know and what we need to learn.

The zoonotic bacteria, Brucella canis, is becoming the leading cause of canine brucellosis in Europe. In dogs, it causes reproductive problems as well as non-specific lameness or discospondilitis. In humans, B. canis can be origin of chronic debilitating conditions characteristic to its genus such as undulant fever, splenomegaly, and lymphadenopathy. Although B. canis shows some pathogenic characteristics similar to B. abortus and B. melitensis, it lacks surface O-polysaccharide, like nonzoonotic B. ovis. This review shows that host-B. canis interactions are still poorly understood, with many knowledge and capability gaps, causing relatively poor sensitivity and specificity of existing diagnostic tools. Currently, there is no vaccine for this rough Brucella species. Besides, antimicrobial therapy does not guarantee bacterial elimination, and infection relapses are frequently reported, increasing the risks of antibiotic resistance development. B. canis has been detected in dogs in almost all European countries which increased human exposure, but currently there is no systematic surveillance. Moreover, B. canis caused brucellosis is not included in Animal Health Law, and therefore there is no legal framework to tackle this emerging infectious disease. To map out the diagnostic strategies, identify risks for human infections and propose management scheme for infected pet and kennel dogs, we present current understanding of canine B. canis caused brucellosis, outline major knowledge gaps and propose future steps. To address and highlight challenges veterinary and public health services encounter in Europe, we developed two B. canis infection scenarios: of a single household pet and of a kennel dog in larger group.

Brucella ceti Infection in Striped Dolphins from Italian Seas: Associated Lesions and Epidemiological Data.

Brucella ceti infections have been increasingly reported in cetaceans. In this study, we analyzed all cases of B. ceti infection detected in striped dolphins stranded along the Italian coastline between 2012 and 2021 (N = 24). We focused on the pathogenic role of B. ceti through detailed pathological studies, and ad hoc microbiological, biomolecular, and serological investigations, coupled with a comparative genomic analysis of the strains. Neurobrucellosis was observed in 20 animals. The primary histopathologic features included non-suppurative meningoencephalitis (N = 9), meningitis (N = 6), and meningoencephalomyelitis (N = 5), which was also associated with typical lesions in other tissues (N = 8). Co-infections were detected in more than half of the cases, mostly involving Cetacean Morbillivirus (CeMV). The 24 B. ceti isolates were assigned primarily to sequence type 26 (ST26) (N = 21) and, in a few cases, ST49 (N = 3). The multilocus sequence typing (cgMLST) based on whole genome sequencing (WGS) data showed that strains from Italy clustered into four genetically distinct clades. Plotting these clades onto a geographic map suggests a link between their phylogeny and the topographical distribution. These results support the role of B. ceti as a primary neurotropic pathogen for striped dolphins and highlight the utility of WGS data in understanding the evolution of this emerging pathogen.

Canine brucellosis due to Brucella canis: description of the disease and control measures.

Brucellosis is a contagious disease caused by bacteria of the genus Brucella, which can affect different animal species. Dogs may occasionally be infected with B. abortus, B. melitensis or B. suis, or by the endemic form of the disease, caused by B. canis. Among the brucellosis­affecting domestic animals, that of the dog is certainly the least frequent, but also the least studied. Canine brucellosis due to B. canis represents the dog­specific brucellosis, both because it is the main susceptible animal species, and because it constitutes its fundamental reservoir of infection. The disease can also affect humans, although its course does not assume the characteristics of severity typical of the infection determined by the 'classical' species of the genus Brucella. In Italy, there are frequent imports of dogs from countries where the disease is present, often with non­controlled movements and without sanitary controls. Considering that the zoonotic potential of the disease can be favored by the close cohabitation between man and dog, which occurs especially in urban environments, canine brucellosis has to be regarded as a public health problem susceptible to introduction and spread in the Italian territory.

Detection of Brucella abortus Vaccine Strain RB51 in Water Buffalo (Bubalus bubalis) Milk.

The isolation of B. abortus RB51 vaccine strain from a milk sample in a water buffalo farm in southern Italy emphasizes the risk to public health of consuming contaminated milk or milk products following illegal vaccination.

Campylobacter and risk factors associated with dog ownership: a retrospective study in household and in shelter dogs.

Campylobacteriosis has been the most frequently reported zoonotic disease in humans in Europe. The scientific literature has reported that the role of dogs may be relevant. The objectives of this work are to improve the knowledge about Campylobacter spp. carriage, infection and antimicrobial resistance in household and shelter dogs in Italy, and to assess risk factors at the dog/human interface. During the 2015­2016 period, rectal swabs were collected from 431 household vet­visiting dogs and 173 dogs housed in shelters. A total of 3 veterinary clinics, located in three Italian regions (Abruzzo, Molise and Tuscany) and 10 shelters, five in Abruzzo and five in Molise, were included in the study. Relevant risk factors for the transmission of Campylobacter spp. from dogs to humans were assessed by means of a questionnaire administered to owners of household dogs. For Campylobacter spp. isolation, selective cultivation methods were used, followed by confirmation and species identification with the PCR method. Phenotypic antibiotic resistance profiles assayed using antimicrobial susceptibility testing were combined. Campylobacter spp. were isolated from 9 household dogs (2.1% CI 1.1% ­ 3.9%) and from 13 shelter dogs (7.5 % CI 4.5% ­ 12.4%). In household dogs C. jejuni was the most represented species (0.9%). In shelter dogs, the most common species was C. jejuni (5.2%). Campylobacter spp. isolates were resistant to ciprofloxacin (22.73%), nalidixic acid (22.73%), tetracyclines (27.27%), streptomycin (9.09%) and erythromycin (4.55%). The main C. jejuni Clonal Complex identified in dogs were CC21, CC45, CC206, CC403, CC42 and CC658. The risk of contracting Campylobacteriosis from dogs remains a concrete reality. This risk is increased in the presence of common habits, as shown by the data from the questionnaire. Prevalence control of Campylobacter spp. in household and shelter dogs would be important in order to reduce the transmission to humans.

Distribution of Salmonella spp. Serotypes Isolated from Poultry in Abruzzo and Molise Regions (Italy) during a 6-Year Period.

Human salmonellosis incidence is increasing in the European Union (EU). Salmonellaenterica subsp. enterica serovar Enteriditis, Salmonellaenterica subsp. enterica serovar Typhimurium (including its monophasic variant) and Salmonellaenterica subsp. enterica serovar Infantis represent targets in control programs due to their frequent association with human cases. This study aimed to detect the most prevalent serotypes circulating in Abruzzo and Molise Regions between 2015 and 2020 in the framework of the Italian National Control Program for Salmonellosis in Poultry (PNCS)]. A total of 332 flocks of Abruzzo and Molise Regions were sampled by veterinary services in the period considered, and 2791 samples were taken. Samples were represented by faeces and dust from different categories of poultry flocks: laying hens (n = 284), broilers (n = 998), breeding chickens (n = 1353) and breeding or fattening turkeys (n = 156). Breeding and fattening turkeys had the highest rate of samples positive for Salmonella spp. (52.6%; C.I. 44.8%-60.3%). Faeces recovered through boot socks represented the greatest number of positive samples (18.2%). Salmonellaenterica subsp. enterica serovar Infantis was the prevalent serotype in breeding and fattening turkeys (32.7%; C.I. 25.8%-40.4%) and in broiler flocks (16.5%; C.I. 14.4%-19.0%). Salmonellaenterica subsp. enterica serovar Typhimurium was detected at low levels in laying hens (0.7%; C.I. 0.2%-2.5%) followed by breeding and fattening turkeys (0.6%; C.I. 0.2%-2.5%). Salmonellaenterica subsp. enterica serovar Enteriditis was also detected at low levels in laying hens (2.5%; C.I. 1.2%-5.0%). These findings highlight the role of broilers and breeding/fattening turkeys as reservoirs of Salmonella spp. and, as a consequence, in the diffusion of dangerous serotypes as Salmonellaenterica subsp. enterica serovar Infantis. This information could help veterinary services to analyze local trends and to take decisions not only based on indications from national control programs, but also based on real situations at farms in their own competence areas.

First Isolation of Brucella canis from a breeding kennel in Italy.

Brucella canis has been isolated for the first time in Italy in a commercial breeding kennel. It was diagnosed after a deep investigation related to the onset of reproductive disorders. Animals were tested with direct and indirect techniques. The agent was first detected in two Chihuahua aborted foetuses by direct culture. Further, it was also isolated from blood samples of dogs hosted in the kennel, which also showed reaction to conventional serological tests (microplate serum agglutination test). The isolates were identified as B. canis by standard microbiological methods and a Bruce­ladder multiplex PCR. To investigate the genomic diversity, whole genome sequencing was used, applying the core genome Multilocus Sequence Typing (cgMLST ). In a first round of serological testing performed on 598 animals, 269 (46.1%) tested positive. In the second round of laboratory testing carried out 4­5 weeks apart, the number of serologically positive dogs was 241 out of 683 tested (35.3%), while the number of dogs positive to isolation was 68 out of 683 tested (10.0%). The PCR showed a lack of sensitivity when compared to direct isolation. The epidemiological investigation did not identify the source of the infection, given the time elapsed from the onset of abortions to the definitive diagnosis of B. canis infection in the kennel. The genomic analyses featured the strains as ST21 and, according to the cgMLST, revealed the presence of a tight cluster with a maximum diversity of four allelic differences. The observed limited genomic variation, largely within the known outbreak cut­offs, suggests that the outbreak herein described was likely caused by a single introduction. Moreover, in a broader scale comparison using the public available genomes, we found that the closest genome, isolated in China, differed by more than 50 alleles making not possible to find out the likely origin of the outbreak. The lack of updated data on B. canis genome sequences in the public databases, together with the limited information retrieved from the epidemiological investigations on the outbreak, hampered identification of the source of B. canis infection.