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.

Proteomics-based screening and antibiotic resistance assessment of clinical and sub-clinical Brucella species: An evolution of brucellosis infection control.

Brucellae are intracellular sneaky bacteria and they can elude the host's defensive mechanisms, resulting in therapeutic failure. Therefore, the goal of this investigation was to rapid identification of Brucella species collected from animals and humans in Saudi Arabia, as well as to evaluate their resistance to antibiotics. On selective media, 364 animal samples as well as 70 human blood samples were cultured. Serological and biochemical approaches were initially used to identify a total of 25 probable cultured isolates. The proteomics of Brucella species were identified using the MALDI Biotyper (MBT) system, which was subsequently verified using real-time polymerase chain reaction (real-time PCR) and microfluidic electrophoresis assays. Both Brucella melitensis (B. melitensis) and Brucella abortus (B. abortus) were tested for antimicrobial susceptibility using Kirby Bauer method and the E-test. In total, 25 samples were positive for Brucella and included 11 B. melitensis and 14 B. abortus isolates. Twenty-two out of 25 (88%) and 24/25 (96%) of Brucella strains were recognized through the Vitek 2 Compact system. While MBT was magnificently identified 100% of the strains at the species level with a score value more than or equal to 2.00. Trimethoprim-sulfamethoxazole, rifampin, ampicillin-sulbactam, and ampicillin resistance in B. melitensis was 36.36%, 31.82%, 27.27%, and 22.70%, respectively. Rifampin, trimethoprim-sulfamethoxazole, ampicillin, and ampicillin-sulbactam resistance was found in 35.71%, 32.14%, 32.14%, and 28.57% of B. abortus isolates, correspondingly. MBT confirmed by microfluidic electrophoresis is a successful approach for identifying Brucella species at the species level. The resistance of B. melitensis and B. abortus to various antibiotics should be investigated in future studies.

The VirB System Plays a Crucial Role in Brucella Intracellular Infection.

Brucellosis is a highly prevalent zoonotic disease caused by Brucella. Brucella spp. are gram-negative facultative intracellular parasitic bacteria. Its intracellular survival and replication depend on a functional virB system, an operon encoded by VirB1-VirB12. Type IV secretion system (T4SS) encoded by the virB operon is an important virulence factor of Brucella. It can subvert cellular pathway and induce host immune response by secreting effectors, which promotes Brucella replication in host cells and induce persistent infection. Therefore, this paper summarizes the function and significance of the VirB system, focusing on the structure of the VirB system where VirB T4SS mediates biogenesis of the endoplasmic reticulum (ER)-derived replicative Brucella-containing vacuole (rBCV), the effectors of T4SS and the cellular pathways it subverts, which will help better understand the pathogenic mechanism of Brucella and provide new ideas for clinical vaccine research and development.

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.

Importance of brucellosis control programs of livestock on the improvement of one health.

Brucellosis not only represents an important health restraint on livestock but also causes high economic losses in many developing countries worldwide. Despite considerable efforts made for the control of brucellosis, the disease is still spreading in many regions (such as the Middle East) where it represents one of the most important health hazards impacting both animals and humans. The present review aims to investigate the efficacy of veterinary control programs regarding brucellosis, with a special focus on current prevention, control, and eradication approaches. The reasons for unsuccessful control programs such as the absence of highly effective vaccines and non-certified bulls are also debated, to understand why the prevalence of brucellosis in livestock is not decreasing in many areas despite considerable efforts taken to date. The importance of governmental and regional investment in brucellosis control remains one of the main limiting factors owing to the limited budget allocated to tackle this disease. In this context, one health concept has generated novel comprehensive approaches with multiple economic implications across the livestock industry and public health. However, the implementation of such global preventive strategies appears to be a key issue for many endemic and low-income countries. According to the collected data, epidemiological contexts including management and trade systems along with well-defined agro-ecological zones should be evaluated in brucellosis endemic countries to improve milk production and to enhance the sustainability of the livestock sector at both national and regional levels.

Uncovering the Hidden Credentials of Brucella Virulence.

Bacteria in the genus Brucella are important human and veterinary pathogens. The abortion and infertility they cause in food animals produce economic hardships in areas where the disease has not been controlled, and human brucellosis is one of the world's most common zoonoses. Brucella strains have also been isolated from wildlife, but we know much less about the pathobiology and epidemiology of these infections than we do about brucellosis in domestic animals. The brucellae maintain predominantly an intracellular lifestyle in their mammalian hosts, and their ability to subvert the host immune response and survive and replicate in macrophages and placental trophoblasts underlies their success as pathogens. We are just beginning to understand how these bacteria evolved from a progenitor alphaproteobacterium with an environmental niche and diverged to become highly host-adapted and host-specific pathogens. Two important virulence determinants played critical roles in this evolution: (i) a type IV secretion system that secretes effector molecules into the host cell cytoplasm that direct the intracellular trafficking of the brucellae and modulate host immune responses and (ii) a lipopolysaccharide moiety which poorly stimulates host inflammatory responses. This review highlights what we presently know about how these and other virulence determinants contribute to Brucella pathogenesis. Gaining a better understanding of how the brucellae produce disease will provide us with information that can be used to design better strategies for preventing brucellosis in animals and for preventing and treating this disease in humans.

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.

Sero-prevalence and associated risk factors of bovine brucellosis in Sendafa, Oromia Special Zone surrounding Addis Ababa, Ethiopia.

Bovine brucellosis is an infectious bacterial disease caused by members of genus Brucella, affecting both animals and humans, and resulting in a serious economic loss in animal production sector and deterioration of public health. A cross-sectional study was conducted from November 2014 to April 2015 to determine the seroprevalence and associated risk factors of bovine brucellosis in Sendafa, Oromia Special Zone, Ethiopia. A total of 503 blood samples were collected using a simple random sampling technique from dairy cattle of above 6 months of age with no history of previous vaccination against brucellosis. All sera samples were subjected to both Rose Bengal Plate Test for screening and Complement Fixation Test for confirmation. Accordingly, the overall seroprevalence of bovine brucellosis in the study area was 0.40%. The result showed that the seroprevalence of bovine brucellosis in the study area was not statistically significant with all proposed risk factors. No reactors were observed in male animals. The seroprevalence was observed in animals without previous history of abortion. Moreover, information was gathered on individual animal and farm-level risk factors and other farm characteristics using a questionnaire. Awareness among society was poor, so the positive animals can be a potential hazard to animals and humans in the study area. Therefore, public education should be done to improve the awareness of the community on bovine brucellosis and its public health impact with due consideration on the safe consumption of food of animal origin.

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.

Enhancing the Detection of Brucella-Specific CD4+ T Cell Responses in Cattle via in vitro Antigenic Expansion and Restimulation.

Bovine brucellosis, cause by infection with Brucella abortus, causes reproductive failure in cattle, has a major economic impact to producers, and as a zoonoses, it is a disease of public health concern. Characterization of the protective immune response against Brucella infection is important to our understanding of disease pathogenesis and for the development of diagnostic assays and vaccines. Most of the knowledge regarding protection against Brucella comes from studies in the murine model, but less is known about the immune responses in cattle. Assessment of antigen-specific T cell frequency and functional phenotype are critical to understand the immune status of the host, characterize mechanisms of protective immunity and immunopathology, and to predict immune protection. The frequency of circulating T cells specific for a particular pathogen is often very low, making analysis of such responses difficult. Our goal was to develop a flow-cytometry based approach to better track Brucella-specific T cell responses. Using peripheral blood mononuclear cells (PMBC) from Brucella abortus strain RB51-vaccinated cattle, we optimized an in vitro stimulation protocol based on a combination of antigen and pan-T cell stimulation. We then assessed RB51-specific T cell responses by concurrently measuring proliferation and cytokine production using flow-cytometry. This methodology enhances the detection of peripheral, Brucella-specific responses in cattle following RB51 vaccination. This protocol is versatile in that it can be modified to fit other in vitro stimulation systems and additional functional or phenotypic parameters can be added for flow cytometric detection and characterization of antigen-specific T cells.

Pathogenesis and immune response in Brucella infection acquired by the respiratory route.

Brucella infection is frequently acquired through the respiratory route. The pathogen disseminates systemically from the lungs to infect peripheral organs. In this review we summarize the existing data on the pathogenesis of inhalational Brucella infection, the pulmonary immune response to the pathogen, and potential strategies for inducing protective lung immunity.

Brucellar reproductive system injury: A retrospective study of 22 cases and review of the literature.

OBJECTIVE: We aimed to describe the clinical characteristics and prognosis of 22 patients with Brucella-induced reproductive system injury. METHODS: We assessed 22 patients with reproductive system injury between 2010 and 2018 at The First Affiliated Hospital of Xinjiang Medical University. RESULTS: The disease is predominant in men. Male patients had orchitis, erectile dysfunction, prostatitis, and urethral stricture, while female patients had vaginitis and cervicitis. Some patients had laboratory abnormalities and liver injury. Patients received combination therapy of rifampicin and doxycycline. Doxycycline combined with levofloxacin or moxifloxacin was administered to patients with rifampicin intolerance. All patients had received antibiotic therapy for at least 6 weeks. One patient was lost to follow-up, one patient relapsed because of osteoarthropathy, and one patient had dysuria resulting from chronic prostatitis. The clinical symptoms resolved in the other patients, and the overall patient prognosis was good. CONCLUSION: Clinicians should pay attention to brucellosis-induced reproductive system damage. The two-drug regimen of rifampicin+doxycycline is recommended for these patients. Doxycycline combined with levofloxacin or moxifloxacin should be used in patients with brucellosis-induced reproductive system damage who have rifampicin intolerance. The treatment course should be at least 6 weeks.

Brucellar spondylodiscitis: A case series with focus on histopathological features.

Brucellar spondylodiscitis (BS) is the most common form of musculoskeletal brucellosis. The isolation of Brucella spp from blood, other body fluids or tissue cultures is the gold standard for definitive diagnosis of BS. BS shows a large histopathological spectrum of lesions with non-specific and granulomatous forms, and its histopathological features are not widely reported. This case series study reported the histopathological features of a series of 21 BS. Ten cases revealed nonspecific forms of BS. Lymphocytes were the predominant inflammatory cells in this group. In one case neutrophils were predominant. The eleven remaining were granulomatous. The 11 exhibited histiocytic type granulomas. Ten were non-necrotizing. One case, taken from abscess wall, contained minimal deposition of caseous like necrotic material. This case and 2 others corresponding to abscess wall biopsies, showed histiocytic granulomas mixed with small aggregates of epithelioid cells without well-formed epithelioid granuloma nor giant cell. The histopathological diagnosis of BS is challenging. Nonspecific forms of BS, mimiking pyogenic spondylodiscitis, are observed in about half of the cases. Although nonspecific forms of BS are characterized by the predominance of lymphocytes and plasmocytes, BS forms with predominant neutrophil cell infiltration do exist. Histiocytic granuloma is highly suggestive of BS. Purely epithelioid forms of BS were not observed in the present series. Caseous like necrosis suggestive of tuberculosis is a possible feature of BS.

Marcadores moleculares para la taxonomía e identificación del género Brucella (Alphaproteobacteria) / Molecular markers for the taxonomy and identification of the genus Brucella (Alphaproteobacteria)

Introducción: El género Brucella está incluido en la familia Brucellaceae que pertenece al orden Rhizobiales y es reconocido por su alto grado de patogenicidad. Las bacterias de este género son responsables de la brucelosis, enfermedad que ha sido reportada como una de las zoonosis más importantes a nivel mundial por su incidencia en el ganado y el hombre. Los estudios previos para la clasificación taxonómica del género, se han basado fundamentalmente en el análisis del gen 16S ARNr. Sin embargo, pocas investigaciones se han dirigido a la identificación de marcadores moleculares que distingan a sus miembros de otros grupos de bacterias. Objetivo: Identificar inserciones en secuencias de proteínas conservadas, que pudieran ser utilizados como marcadores moleculares para la taxonomía y diagnóstico de especies del género Brucella. Métodos: Las secuencias homólogas de las proteínas analizadas fueron obtenidas de bases de datos internacionales y, posteriormente, alineadas con el programa ClustalX2, para ello fueron considerados los parámetros sugeridos en la literatura. Resultados: Se identificaron inserciones en las proteínas oxoglutarato deshidrogenasa (componente E1) y ADN ligasa A específicas del género Brucella. Conclusiones: Las inserciones halladas pueden ser empleadas como complemento a los métodos tradicionales de clasificación taxonómica y para el diagnóstico molecular de bacterias incluidas en el género Brucella(AU)

Introduction: Brucella is a genus from the Brucellaceae family, Rhizobiales order. This genus is recognized for its high pathogenicity. Brucella bacteria cause brucellosis, a disease reported as one of the most important zoonoses worldwide due to its incidence in cattle and people. Previous studies on taxonomic classification of the genus have been mainly based on the analysis of gene 16S rDNA. However, few studies have been aimed at identification of molecular markers distinguishing its members from other groups of bacteria. Objective: Identify insertions in preserved protein sequences which could be used as molecular markers for the taxonomy and diagnosis of species from the Brucella genus. Methods: The homologous sequences for the proteins analyzed were obtained from international databases and aligned with the software ClustalX2, considering the parameters suggested in the literature. Results: Insertions were identified in the proteins oxoglutarate dehydrogenase (component E1) and DNA ligase A, specific of the genus Brucella. Conclusions: The insertions found may be used as complements to the traditional methods for taxonomic classification and for the molecular diagnosis of bacteria from the genus Brucella(AU)

Dendritic cells and Brucella spp. interaction: the sentinel host and the stealthy pathogen.

As dendritic cells (DCs) are among the first cells to encounter antigens, these cells trigger both innate and T cell responses, and are the most potent antigen-presenting cells. Brucella spp., which is an intracellular facultative and stealthy pathogen, is able to evade the bactericidal activities of professional phagocytes. Several studies have demonstrated that Brucella can survive and replicate intracellularly, thereby provoking impaired maturation of DCs. Therefore, the interaction between DCs and Brucella becomes an interesting model to study the immune response. In this review, we first will describe the most common techniques for DCs differentiation in vitro as well as general features of brucellosis. Then, the interaction of DCs and Brucella, including pathogen recognition, molecular mechanisms of bacterial pathogenesis, and intracellular trafficking of Brucella to subvert innate response, will be reviewed. Finally, we will debate diversity in immunological DC response and the controversial role of DC activation against Brucella infection.

Lethality of Brucella microti in a murine model of infection depends on the wbkE gene involved in O-polysaccharide synthesis.

Brucella microti was isolated a decade ago from wildlife and soil in Europe. Compared to the classical Brucella species, it exhibits atypical virulence properties such as increased growth in human and murine macrophages and lethality in experimentally infected mice. A spontaneous rough (R) mutant strain, derived from the smooth reference strain CCM4915T, showed increased macrophage colonization and was non-lethal in murine infections. Whole-genome sequencing and construction of an isogenic mutant of B. microti and Brucella suis 1330 revealed that the R-phenotype was due to a deletion in a single gene, namely wbkE (BMI_I539), encoding a putative glycosyltransferase involved in lipopolysaccharide (LPS) O-polysaccharide biosynthesis. Complementation of the R-strains with the wbkE gene restored the smooth phenotype and the ability of B. microti to kill infected mice. LPS with an intact O-polysaccharide is therefore essential for lethal B. microti infections in the murine model, demonstrating its importance in pathogenesis.

Immune Response to Mucosal Brucella Infection.

Brucellosis is one of the most prevalent bacterial zoonosis of worldwide distribution. The disease is caused by Brucella spp., facultative intracellular pathogens. Brucellosis in animals results in abortion of fetuses, while in humans, it frequently manifests flu-like symptoms and a typical undulant fever, being osteoarthritis a common complication of the chronic infection. The two most common ways to acquire the infection in humans are through the ingestion of contaminated dairy products or by inhalation of contaminated aerosols. Brucella spp. enter the body mainly through the gastrointestinal and respiratory mucosa; however, most studies of immune response to Brucella spp. are performed analyzing models of systemic immunity. It is necessary to better understand the mucosal immune response induced by Brucella infection since this is the main entry site for the bacterium. In this review, some virulence factors and the mechanisms needed for pathogen invasion and persistence are discussed. Furthermore, some aspects of local immune responses induced during Brucella infection will be reviewed. With this knowledge, better vaccines can be designed focused on inducing protective mucosal immune response.

Prediction of Structure and Molecular Interaction with DNA of BvrR, a Virulence-Associated Regulatory Protein of Brucella.

Brucellosis, also known as "undulant fever" is a zoonotic disease caused by Brucella, which is a facultative intracellular bacterium. Despite efforts to eradicate this disease, infection in uncontrolled domestic animals persists in several countries and therefore transmission to humans is common. Brucella evasion of the innate immune system depends on its ability to evade the mechanisms of intracellular death in phagocytic cells. The BvrR-BvrS two-component system allows the bacterium to detect adverse conditions in the environment. The BvrS protein has been associated with genes of virulence factors, metabolism, and membrane transport. In this study, we predicted the DNA sequence recognized by BvrR with Gibbs Recursive Sampling and identified the three-dimensional structure of BvrR using I-TASSER suite, and the interaction mechanism between BvrR and DNA with Protein-DNA docking and molecular dynamics (MD) simulation. Based on the Gibbs recursive Sampling analysis, we found the motif AAHTGC (H represents A, C, and T nucleotides) as a possible sequence recognized by BvrR. The docking and EMD simulation results showed that C-terminal effector domain of BvrR protein is likely to interact with AAHTGC sequence. In conclusion, we predicted the structure, recognition motif, and interaction of BvrR with DNA.

Multi-pathogen serological survey of migratory caribou herds: A snapshot in time.

Pathogens can impact host survival, fecundity, and population dynamics even when no obvious disease is observed. Few baseline data on pathogen prevalence and diversity of caribou are available, which hampers our ability to track changes over time and evaluate impacts on caribou health. Archived blood samples collected from ten migratory caribou herds in Canada and two in Greenland were used to test for exposure to pathogens that have the potential to effect population productivity, are zoonotic or are emerging. Relationships between seroprevalence and individual, population, and other health parameters were also examined. For adult caribou, the highest overall seroprevalence was for alphaherpesvirus (49%, n = 722), pestivirus (49%, n = 572) and Neospora caninum (27%, n = 452). Lower seroprevalence was found for parainfluenza virus type 3 (9%, n = 708), Brucella suis (2%, n = 758), and Toxoplasma gondii (2%, n = 706). No animal tested positive for antibodies against West Nile virus (n = 418) or bovine respiratory syncytial virus (n = 417). This extensive multi-pathogen survey of migratory caribou herds provides evidence that caribou are exposed to pathogens that may have impacts on herd health and revealed potential interactions between pathogens as well as geographical differences in pathogen exposure that could be linked to the bio-geographical history of caribou. Caribou are a keystone species and the socio-economic cornerstone of many indigenous cultures across the North. The results from this study highlight the urgent need for a better understanding of pathogen diversity and the impact of pathogens on caribou health.