RIBAP: a comprehensive bacterial core genome annotation pipeline for pangenome calculation beyond the species level.

Microbial pangenome analysis identifies present or absent genes in prokaryotic genomes. However, current tools are limited when analyzing species with higher sequence diversity or higher taxonomic orders such as genera or families. The Roary ILP Bacterial core Annotation Pipeline (RIBAP) uses an integer linear programming approach to refine gene clusters predicted by Roary for identifying core genes. RIBAP successfully handles the complexity and diversity of Chlamydia, Klebsiella, Brucella, and Enterococcus genomes, outperforming other established and recent pangenome tools for identifying all-encompassing core genes at the genus level. RIBAP is a freely available Nextflow pipeline at github.com/hoelzer-lab/ribap and zenodo.org/doi/10.5281/zenodo.10890871.

Detection and characterization of Brucella species in rodents: A threat for the persistence of brucellosis in livestock farms.

Brucellosis, caused by various Brucella species, poses a significant threat to global public health and livestock industries. This study aims to fill the knowledge gap concerning the presence of Brucella spp. in rodents on livestock farms in Iran. Both bacteriological and molecular surveys were conducted to assess the prevalence of Brucella spp. in these rodent populations. A total of 16 rodents were captured in four seropositive dairy cattle farms (n = 7) and two seropositive sheep farms (n = 9) and were then examined for the presence of the Brucella-infection. Five cow milk samples and 53 bovine lymph node samples from these farms were also tested for Brucella spp. Lymph node samples from dairy cattle farms contained 32 B. abortus biovar 3 isolates and one B. melitensis Rev1 vaccine isolate. The bacterial culture of rodents identified 12.5% of them (Mus musculus and Rattus norvegicus) harboring Brucella strains in dairy cattle farms. The rodents had B. abortus biovar 3 and B. melitensis biovar 1, suggesting a reservoir for these bacteria. A two-step molecular assay, utilizing the Omp28 sequences in tissue samples of rodents, demonstrated that 68.75% (n = 11) of the tested rodents yielded positive results. Bruce-ladder PCR and wboA typing on isolated bacteria revealed a close relationship to field strain of Brucella species. The study reveals that rodents on seropositive livestock farms in Iran harbor Brucella spp., indicating a potential reservoir for these bacteria. This highlights the importance of monitoring rodent populations through the molecular and bacterial methods to manage and control brucellosis in livestock.

Molecular Characterization and Analysis of Risk Factors Associated with Brucellosis in Central Indian and Meghalaya Population.

Objective: In this study we evaluated the utility of Abortus Melitensis Ovis Suis Brucella PCR (AMOS PCR) for the molecular characterization of Brucella species and analyzed the associated risk factors for brucellosis in Central Indian and Meghalayan population. Methods: AMOS PCR was carried out in a total of 160 BSCP-31 PCR-positive DNA samples isolated previously from the blood of Central Indian (n = 90) and Meghalayan cohorts (n = 70). Clinical and associated risk factors recorded earlier were used to establish strain-specific disease outcomes in study cohorts. Results: Brucella melitensis was found to be the dominant strain in both Central Indian and Meghalayan cohorts (57.7% and 54.28%, respectively) followed by Brucella abortus (42.22% and 38.57%). Although rare, brucellosis cases in the Meghalayan population also showed the presence of Brucella suis (7.14%) and Brucella ovis (2.85%). Febrile illness was a major clinical risk factor in both study cohorts, while occupational risk factors like exposure to animals and raw milk consumption were major mediating factors for brucellosis in Central Indian cohorts. On the contrary, meat consumption was found to be significant predisposing factor for brucellosis in Meghalaya. Conclusion: Molecular characterization of Brucella species provides important public health data for mitigation, advocacy, and antimicrobial stewardship.

Equine brucellosis in Iran: serological, bacteriological and molecular analysis.

Equine brucellosis significantly impacts the health and functionality of horses, leading to complications such as bursitis infection, septic tenosynovitis, septic arthritis, and non-specific lameness resulting from joint infections. In the present study, we used the Rose Bengal plate agglutination test (RBPT), serum agglutination test (SAT), and the 2-mercaptoethanol (2-ME) assays to find equine brucellosis. From June 2018 to September 2022, 876 blood samples were randomly taken from apparently healthy racing horses in certain parts of Iran, such as Kerman, Isfahan, Tehran, Qom, and Kurdistan. DNA extraction was carried out directly on all 63 serum samples identified as seropositive through RBPT. An additional 30 seronegative serum samples were also randomly chosen for study. Bacterial culture was also done on milk, blood, and vaginal swabs taken from seropositive horses.The bacteria that were found in the samples were then put through Bruce-ladder PCR. Our results indicated that 63 (7.1%), 21 (2.3%), and 2 (0.2%) of horses were seropositive using RBPT, SAT, and 2-ME, respectively. Also, none of the 30 DNA-extracted serum samples from seronegative horses tested positive for Brucella DNA, while 44.5% (28/63) of the DNA samples from seropositive horses yielded positive results for Brucella DNA. Out of the seropositive samples, 26 had DNA from Brucella abortus and 2 had DNA from Brucella melitensis. Also, B. melitensis biovar 1 was found in two milk samples from mares in the provinces of Kerman and Isfahan. It was identified using classical biotyping, and molecular assays. It was seen that some of healthy racing horses in some parts of Iran had antibodies against Brucella. The bacteriology and PCR methodologies provide a more comprehensive and reliable means of identifying Brucella spp. infections in horse, especially when the RBPT test came back positive. This underscores the imperative for employing molecular, bacterial, and serological methods in the diagnosis and monitoring of this zoonotic infection. Additionally, this finding suggests that Brucella is being transmitted to equine hosts as a result of its presence in ruminants. The mechanism of transmission may involve interactions between infected ruminants and susceptible equines. This discovery is significant as it underscores the potential cross-species transmission of Brucella and highlights the importance of understanding and managing the spread of the pathogen in both ruminant and equine populations.

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.

Diagnostic utility of LAMP PCR targeting bcsp-31 gene for human brucellosis infection.

PURPOSE: Human brucellosis is a neglected zoonotic disease of significant public health concern. Molecular diagnosis of brucella remains challenging in low resource settings, due to the high infrastructure and cost involved. Loop-mediated isothermal amplification (LAMP) is a rapid point of care polymerase chain reaction (PCR) with the utility of on-field molecular diagnosis and offers a convenient alternative to conventional PCR. In the present study, we developed and evaluated the diagnostic utility of in house LAMP PCR targeting the Brucella genus-specific bcsp-31 gene in patients having febrile illness. MATERIALS AND METHODS: The analytical sensitivity and specificity of bcsp-31 LAMP PCR was first evaluated using brucella (n â€‹= â€‹8) and non-brucella cultures (n â€‹= â€‹5), along with spiked clinical samples. The overall diagnostic utility of developed LAMP PCR was then further evaluated in 393 human samples suspected of brucellosis. RESULTS: The developed LAMP PCR could detect as low as 8 â€‹fg of DNA by visual detection within 35min. We report sensitivity and specificity of the developed LAMP PCR as 90.91% and 99.37%.The accuracy of the developed test assay was found to be 98.60%. In clinical samples, LAMP gave positivity of 20% with the concordance of 89% with conventional PCR. CONCLUSION: To conclude, a rapid, efficacious, sensitive LAMP PCR targeting the bcsp 31 gene was developed. The existing LAMP PCR can be used as a point of care screening test in various low resource endemic setting in lieu of conventional PCR for estimation of prevalence data, diagnosis and treatment of brucellosis.

Characterization of Brucella spp. and other abortigenic pathogens from aborted tissues of cattle and goats in Rwanda.

BACKGROUND: Abortions cause tremendous economic losses in food-producing animals and may lead to food insecurity. OBJECTIVES: This study aimed to characterize Brucella spp. and other abortigenic pathogens from aborted tissues of cattle. METHODS: For cattle, aborted tissues (n = 19) were cultured, and Brucella spp. were detected using the genus-specific 16S-23S ribosomal DNA interspacer region (ITS) assay and speciated using Brucella abortus, Brucella melitensis, Brucella ovis, and Brucella suis (AMOS) and Bruce-ladder PCR assays. Brucella negative samples were screened using the eight abortigenic pathogens PCR panel. Samples from an abortion outbreak that occurred within a goat tribe were included in this investigation. Sera of females (n = 8) and males (n = 2) were analyzed using the Rose Bengal Test (RBT) and indirect enzyme-linked immunosorbent assay (i-ELISA), while vaginal swabs (n = 3) and aborted tissues (n = 1) were cultured and characterized. RESULTS: The ITS-PCR detected Brucella DNA in cultures from two aborted tissues of cattle (10.5%, [2/19]), which were identified as B. melitensis (n = 1), and B. abortus (n = 1) using AMOS and Bruce-ladder PCR assays. Campylobacter fetus (n = 7) and Leptospira spp. (n = 4) including co-infections (n = 2) of C. fetus and Leptospira spp. were identified from the Brucella negative samples of cattle. Goats (100.0%, 10/10) were brucellosis seropositive on RBT and i-ELISA. Mixed infections caused by B. melitensis and B. abortus were isolated from the vaginal swabs (n = 3) and aborted tissues (n = 1). DISCUSSION AND CONCLUSIONS: This is the first identification of abortion-associated pathogens in aborted cattle indicating the enormous financial losses and a threat to public health. It is therefore essential to include these identified pathogens in the surveillance scheme of veterinary and human services.

Genomics of Ochrobactrum pseudogrignonense (newly named Brucella pseudogrignonensis) reveals a new blaOXA subgroup.

Ochrobactrum pseudogrignonense (newly named Brucella pseudogrignonensis) is an emerging pathogen in immunodeficient and immunocompetent patients. Most documented cases associated with Ochrobactrum are frequently catheter-related and exhibit wide-spectrum ß-lactam resistance. Misidentification of this pathogen using commercial bacterial identification kits is common. We identified a case of O. pseudogrignonense infection associated with cholelithiasis. The O. pseudogrignonense genome was sequenced and reconstructed using a Nanopore and Illumina hybrid strategy. A novel blaOXA-919 divergent from existing OXA members was identified and subsequent analysis revealed its existence in all available O. pseudogrignonense genomes, which forms a new phylogenetic subgroup distinct from other OXA clusters. Further analysis demonstrated the presence of the novel blaOXA-919 in the chromosome of several other Ochrobactrum species. Our study indicated that Ochrobactrum chromosomes may be a reservoir of blaOXA-919 ß-lactamases.

Herd-level contamination of Neospora caninum, Toxoplasma gondii and Brucella in milk of Iranian dairy farms.

The bulk milk examination is a reliable screening tool for monitoring the quality of milk in the farms. The infection to Neospora caninum, Toxoplasma gondii and Brucella sp. Was evaluated in bulk milk samples of dairy farms in Hamedan province, West part of Iran. All the dairy farms (n = 149) were examined for N. caninum, T. gondii and Brucella infections using milk ring test (MRT), microbiology, serology (Enzyme-linked Immunosorbent Assay), and molecular techniques. Based on molecular methods, Brucella-infection was negative in all farms; while, 55 %, 5.4 % and 2.7 % of samples were positive for N. caninum, T. gondii and mix infection, respectively. The highest Neospora-infection was detected in the farms with history of abortion in fall and winter. There was significant association between Neospora-infection and the presence of dogs and rodents in the farms, herd size, and age of the animals. Also, a significant association was seen between Toxoplasma-infection and the presence of cats and rodents in the farms, as well as age of the animals. Average total bacterial count (TBC) was calculated 1.14 × 106±1.1 × 106. The highest TBC was in the farms from Central locations of studied area (5.7 × 106±2.24 × 106), farms with more than 120 animals (7.9 × 106±2.8 × 106), and farms with ≥50-months age (1.74 × 106±6.3 × 105) in spring and summer (6.9 × 106±3.7 × 106). The number of somatic cells was estimated between 1 × 104 and 2 × 106 (Average = 4.2 × 105±3.39 × 105). The current study was a comprehensive evaluation of Neospora, Toxoplasma and Brucella infections in milk samples of Iranian dairy farms for the first time. Neospora-infection is responsible for economic losses in the region. Health education and milk pasteurization are so helpful for inhibiting the milk borne diseases. To reduce the risk factors, predict and design the appropriate schemes like redundant of heterogeneous animals are recommended.

Multicenter evaluation of the VITEK MS matrix-assisted laser desorption/ionization-time of flight mass spectrometry system for identification of bacteria, including Brucella, and yeasts.

The use of matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) mass spectrometry has proven to be rapid and accurate for the majority of clinical isolates. Some gaps remain concerning rare, emerging, or highly pathogenic species, showing the need to continuously expand the databases. In this multicenter study, we evaluated the accuracy of the VITEK MS v3.2 database in identifying 1172 unique isolates compared to identification by DNA sequence analysis. A total of 93.6% of the isolates were identified to species or group/complex level. A remaining 5.2% of the isolates were identified to the genus level. Forty tests gave a result of no identification (0.9%) and 12 tests (0.3%) gave a discordant identification compared to the reference identification. VITEK MS is also the first MALDI-TOF MS system that is able to delineate the four members of the Acinetobacter baumannii complex at species level without any specific protocol or special analysis method. These findings demonstrate that the VITEK MS v3.2 database is highly accurate for the identification of bacteria and fungi encountered in the clinical laboratory as well as emerging species like Candida auris and the highly pathogenic Brucella species.

Emerging diversity and ongoing expansion of the genus Brucella.

Remarkable genetic diversity and breadth of host species has been uncovered in the Brucella genus over the past decade, fundamentally changing our concept of what it means to be a Brucella. From ocean fishes and marine mammals, to pond dwelling amphibians, forest foxes, desert rodents, and cave-dwelling bats, Brucella have revealed a variety of previously unknown niches. Classical microbiological techniques have been able to help us classify many of these new strains but at times have limited our ability to see the true relationships among or within species. The closest relatives of Brucella are soil bacteria and the adaptations of Brucella spp. to live intracellularly suggest that the genus has evolved to live in vertebrate hosts. Several recently discovered species appear to have phenotypes that are intermediate between soil bacteria and core Brucella, suggesting that they may represent ancestral traits that were subsequently lost in the traditional species. Remarkably, the broad relationships among Brucella species using a variety of sequence and fragment-based approaches have been upheld when using comparative genomics with whole genomes. Nonetheless, genomes are required for fine-scale resolution of many of the relationships and for understanding the evolutionary history of the genus. We expect that the coming decades will reveal many more hosts and previously unknown diversity in a wide range of environments.

New Brucella variant isolated from Croatian cattle.

BACKGROUND: A novel Brucella strain closely related to Brucella (B.) melitensis biovar (bv) 3 was found in Croatian cattle during testing within a brucellosis eradication programme. CASE PRESENTATION: Standardised serological, brucellin skin test, bacteriological and molecular diagnostic screening for Brucella infection led to positive detection in one dairy cattle herd. Three isolates from that herd were identified to species level using the Bruce ladder method. Initially, two strains were typed as B. melitensis and one as B. abortus, but multiplex PCR based on IS711 and the Suis ladder showed that all of them to belong to B. melitensis, and the combination of whole-genome and multi-locus sequencing as well as Multi-Locus Variable numbers of tandem repeats Analysis (MLVA) highlighted a strong proximity within the phylogenetic branch of B. melitensis strains previously isolated from Croatia, Albania, Kosovo and Bosnia and Herzegovina. Two isolates were determined to be B. melitensis bv. 3, while the third showed a unique phylogenetic profile, growth profile on dyes and bacteriophage typing results. This isolate contained the 609-bp omp31 sequence, but not the 723-bp omp31 sequence present in the two isolates of B. melitensis bv. 3. CONCLUSIONS: Identification of a novel Brucella variant in this geographic region is predictable given the historic endemicity of brucellosis. The emergence of a new variant may reflect a combination of high prevalence among domestic ruminants and humans as well as weak eradication strategies. The zoonotic potential, reservoirs and transmission pathways of this and other Brucella variants should be explored.

Comparative study of IS711 and bcsp31-based polymerase chain reaction (PCR) for the diagnosis of human brucellosis in whole blood and serum samples.

BACKGROUND: Clinical diagnosis of human brucellosis (HB) is often difficult due to non-specific symptoms. Immunological tests have been the most common method used in HB diagnosis, but molecular methods based on quantitative polymerase chain reaction (qPCR) have largely replaced these diagnostic methods. The aim of this study was to validate a HB diagnostic qPCR method; assessing different target Brucella genes, and the influence of biological matrices (serum vs. whole blood) on analytical parameters. MATERIAL AND METHODS: Two target genes, IS711 and bcsp31, for HB molecular diagnosis were evaluated, together with biological matrix type (whole blood and serum) using samples spiked with Brucella abortus. In addition, diagnostic parameters of this qPCR method were evaluated in paired whole blood and serum samples from patients with suspected HB. RESULTS: Both genes could be potential diagnostic targets, but IS711 showed a lower limit of detection. In spiked matrix experiments, whole blood showed a lower limit of detection than serum after probit regression (224 vs. 3681 CFU/mL) and ANOVA analysis showed a significant (p < 0.001) difference between the Cq of whole blood at all dilutions and that of serum. In 12 paired clinical samples, no serum samples and only one whole blood sample tested positive for Brucella using this qPCR detection method. CONCLUSIONS: This standardized qPCR-based Brucella detection method could improve diagnosis of HB, serving as a rapid, highly sensitive, and specific test. Whole blood is better suited to qPCR-based HB diagnosis due to the presence of higher target DNA loads in this matrix, compared to serum.

The one hundred year journey of the genus Brucella (Meyer and Shaw 1920).

The genus Brucella, described by Meyer and Shaw in 1920, comprises bacterial pathogens of veterinary and public health relevance. For 36 years, the genus came to include three species that caused brucellosis in livestock and humans. In the second half of the 20th century, bacteriologists discovered five new species and several 'atypical' strains in domestic animals and wildlife. In 1990, the Brucella species were recognized as part of the Class Alphaproteobacteria, clustering with pathogens and endosymbionts of animals and plants such as Bartonella, Agrobacterium and Ochrobactrum; all bacteria that live in close association with eukaryotic cells. Comparisons with Alphaproteobacteria contributed to identify virulence factors and to establish evolutionary relationships. Brucella members have two circular chromosomes, are devoid of plasmids, and display close genetic relatedness. A proposal, asserting that all brucellae belong to a single species with several subspecies debated for over 70 years, was ultimately rejected in 2006 by the subcommittee of taxonomy, based on scientific, practical, and biosafety considerations. Following this, the nomenclature of having multiples Brucella species prevailed and defined according to their molecular characteristics, host preference, and virulence. The 100-year history of the genus corresponds to the chronicle of scientific efforts and the struggle for understanding brucellosis.

Brucella Genomics: Macro and Micro Evolution.

Brucella organisms are responsible for one of the most widespread bacterial zoonoses, named brucellosis. The disease affects several species of animals, including humans. One of the most intriguing aspects of the brucellae is that the various species show a ~97% similarity at the genome level. Still, the distinct Brucella species display different host preferences, zoonotic risk, and virulence. After 133 years of research, there are many aspects of the Brucella biology that remain poorly understood, such as host adaptation and virulence mechanisms. A strategy to understand these characteristics focuses on the relationship between the genomic diversity and host preference of the various Brucella species. Pseudogenization, genome reduction, single nucleotide polymorphism variation, number of tandem repeats, and mobile genetic elements are unveiled markers for host adaptation and virulence. Understanding the mechanisms of genome variability in the Brucella genus is relevant to comprehend the emergence of pathogens.

Detection of Brucella spp. in raw milk from various livestock species raised under pastoral production systems in Isiolo and Marsabit Counties, northern Kenya.

INTRODUCTION: Brucellosis is an important zoonotic disease in Kenya, and identifying the bacteria in milk is important in assessing the risk of exposure in people. METHODS: A cross-sectional study that involved 175 households was implemented in the pastoral counties of Marsabit and Isiolo in Kenya. Pooled milk samples (n = 164) were collected at the household level, and another 372 were collected from domesticated lactating animals (312 goats, 7 sheep, 50 cattle and 3 camels). Real-time polymerase chain reaction (qPCR) testing of the milk samples was performed to identify Brucella species. Brucella anti-LPS IgG antibodies were also detected in bovine milk samples using an indirect enzyme-linked immunosorbent assay (ELISA). RESULTS: Based on the qPCR, the prevalence of the pathogen at the animal level (considering samples from individual animals) was 2.4% (95% confidence interval (CI) 1.1-4.5) and 3.0% (CI: 1.0-7.0) in pooled samples. All 14 samples found positive by qPCR were from goats, with 10 contaminated with B. abortus and 4 with B. melitensis. The Brucella spp. antibody prevalence in bovine milk using the milk ELISA was 26.0% (95% CI: 14.6-40.3) in individual animal samples and 46.3% (95% CI: 30.7-62.6) in pooled samples. CONCLUSION: The study is the first in Kenya to test for Brucella spp. directly from milk using qPCR without culturing for the bacteria. It also detected B. abortus in goats, suggesting transmission of brucellosis between cattle and goats. The high prevalence of Brucella spp. is a significant public health risk, and there is a need for intervention strategies necessary in the study area.

Expanding the host range: infection of a reptilian host (Furcifer pardalis) by an atypical Brucella strain.

Atypical brucellae show deviant phenotypes and/or genotypes. Besides Brucella inopinata, B. microti and B. vulpis, atypical strains have been described infecting humans, rodents, amphibians and fish. They represent potential zoonotic agents. Here, we provide evidence that reptiles as the remaining poikilothermic vertebrate class also represent susceptible hosts for atypical Brucella.

Molecular characterization of Brucella species detected in humans and domestic ruminants of pastoral areas in Kagera ecosystem, Tanzania.

Brucellosis is a zoonotic disease of importance to both public health and the livestock industry. The disease is likely to be endemic in Tanzania and little is reported on molecular characterization of Brucella species in pastoral settings. This study aimed at characterizing Brucella species (targeting genus Brucella) infecting humans, cattle and goat in Kagera region (Ngara and Karagwe districts) using real-time PCR, PCR amplification of 16S rRNA genes and Sanger sequencing. Brucella spp. were detected in 47 samples (19 sera and 28 milk) out of 125 samples (77 sera, 35 milk and 13 aborted materials) using real-time PCR. All aborted materials (13 samples) were negative to real-time PCR. Out of the 47 real-time PCR positive samples (28 milk and 19 sera), 20 samples (10 milk and 10 sera) showed an expected 16S rRNA gene PCR product. Sequence analysis and blasting confirmed the presence of Brucella spp. in pastoral areas of Kagera region. The Brucella spp. from Kagera were phylogenetically grouped in two clades and three branches all closer to B. melitensis, B. abortus and B. suis from USA, Sudan and Iran. However, they were distinct from other species isolated also in USA, New Zealand, Germany and Egypt. This was expected based on the distance between the geographical regions from which the data (nucleotides sequences from 16S gene sequencing) for the phylogeny reconstruction were obtained. This is the first study to report Brucella species identified using 16S rRNA gene sequencing in East and Central Africa. A livestock vaccination program re-inforced with a high index of Brucella diagnosis is needed to eradicate brucellosis in animals and minimize suffering from Brucella infections in humans in Tanzania.

Brucellosis in the Middle East: Current situation and a pathway forward.

Brucellosis is a bacterial endemic zoonotic disease of global significance with detrimental impacts on public health and food animal production. It is caused by Brucella spp., an expanding group of pathogens able to infect various host species. Bovines and small ruminants, which excrete the bacteria in milk and in reproductive discharges, are major sources of infection for humans and other animals. Contact with contaminated animals and consumption of unpasteurized dairy products are the main routes for human infection. In spite of the considerable progress of knowledge gained and success achieved in brucellosis control in the developed world, this disease continues to be an important burden in the Middle East (ME). Common risk factors implicated in the difficulty and complexity of brucellosis control within the region include (1) social and political instabilities; (2) insufficient resources and infrastructure for appropriate diagnosis, reporting, and implementation of control measures; (3) variation of livestock husbandry systems and their commingling with other livestock and wildlife; and (4) traditional cultural practices, including consumption of unpasteurized dairy products. Development of core interdisciplinary competencies is required for a true One Health-based endeavor against the disease. National awareness and educational programs addressing all population sectors from consumers to decision-makers seem to be the next logical, sustainable, and economically viable approach toward improving disease status in this region. In the present review, we describe the current situation of brucellosis in the ME, focusing on the major limitations and shortcomings regarding disease control. We propose a regional approach toward public awareness of brucellosis as the first step in mitigating the disease and discuss the potential benefits, and components of such a strategy, which can further be used as a model for other endemic zoonotic diseases.

Survival of Brucella abortus S19 and other Brucella spp. in the presence of oxidative stress and within macrophages.

The evolutionary "success" of the genus Brucella depends on the ability to persist both in the environment as well as inside of even activated macrophages of the animal host. For that, the Brucellae produce catalase and superoxide dismutase to defend against oxidative stress. Since the deletion of the mglA gene in the B. abortus S19 vaccine strain resulted not only in an increased tolerance to H2O2 but also in the induction of cytokines in macrophages, we here investigated the effect of oxidative stress (Fe2+ and H2O2) on the survival of B. abortus S19 and the isogenic B. abortus S 19 ∆mglA 3.14 deletion mutant in comparison with B. neotomae 5K33, Brucella strain 83/13, and B. microti CCM4915. These Brucellae belong to different phylogenetic clades and show characteristic differences in the mgl-operon. From the various Brucellae tested, B. abortus S19 showed the highest susceptibility to oxidative stress and the lowest ability to survive inside of murine macrophages. B. abortus S19 ∆mglA 3.14 as well as B. neotomae, which also belongs to the classical core clade of Brucella and lacks the regulators of the mgl-operon, presented the highest degree of tolerance to H2O2 but not in the survival in macrophages. The latter was most pronounced in case of an infection with B. 83/13 and B. microti CCM4915. The various Brucellae investigated here demonstrate significant differences in tolerance against oxidative stress and different survival in murine macrophages, which, however, do not correlate directly.