Project description:This is a report of whole-genome sequences of a Brucella abortus strain and two Brucella suis strains isolated from bovine in Zimbabwe. These strains were selected based on their origin and data obtained when using multiplex PCR assays, then sequenced using next-generation sequencing technologies.
Project description:Brucellosis is a zoonotic disease caused mainly by the bacteria belonging to the genus Brucella, most common of them is Brucella abortus. Genome sequencing of Brucella was completed in 2005. While majority of the proteins were assigned function, a large number of the peptides remained un-annotated and were referred as 'hypothetical'. These hypothetical proteins may contain crucial information about the biology and pathogenesis of the B. abortus. Therefore, it is of interest to annotate one such hypothetical protein as a multiple antibiotic resistance regulator protein, MarR. The physiological parameters, localization and the structural features were predicted for this protein which corroborated as the winged-helix type DNA-binding domain superfamily of transcription factors.
Project description:As the causative agent of cattle brucellosis, Brucella abortus commonly exhibits smooth phenotype (by virtue of colony morphology) that is characteristically sensitive to specific Brucella phages, playing until recently a major role in taxonomical classification of the Brucella species by the phage typing approach. We previously reported the discrepancy between traditional phenotypic typing and MLVA results of a smooth phage-resistant (SPR) strain Bab8416 isolated from a 45-year-old custodial worker with brucellosis in a cattle farm. Here, we performed whole genome sequencing and further obtained a complete genome sequence of strain Bab8416 by a combination of multiple NGS technologies and routine PCR sequencing. The detailed genetic differences between B. abortus SPR Bab8416 and large smooth phage-sensitive (SPS) strains were investigated in a comprehensively comparative genomic study. The large indels between B. abortus SPS strains and Bab8416 showed possible divergence between two evolutionary branches at a far phylogenetic node. Compared to B. abortus SPS strain 9-941 (Bab9-941), the specific re-arrangement event in Bab8416 displaying a closer linear relationship with B. melitensis 16M than other B. abortus strains resulted in the truncation of c-di-GMP synthesis, and 3 c-di-GMP-metabolizing genes, were present in Bab8416 and B. melitensis 16M, but absent in Bab9-941 and other B. abortus strains, indicating potential SPR-associated key determinants and novel molecular mechanisms. Moreover, despite almost completely intact smooth LPS related genes, only one mutated OmpA family protein of Bab8416, functionally related to flagellar and efflux pump, was newly identified. Several point mutations were identified to be Bab8416 specific while a majority of them were verified to be B. abortus ST2 characteristic. In conclusion, our study therefore identifies new SPR-associated factors that could play a role in refining and updating Brucella taxonomic schemes and provides resources for further detailed analysis of mechanism for Brucella phage resistance.
Project description:Brucellosis is a worldwide disease of humans and livestock that is caused by a number of very closely related classical Brucella species in the alpha-2 subdivision of the Proteobacteria. We report the complete genome sequence of Brucella abortus field isolate 9-941 and compare it to those of Brucella suis 1330 and Brucella melitensis 16 M. The genomes of these Brucella species are strikingly similar, with nearly identical genetic content and gene organization. However, a number of insertion-deletion events and several polymorphic regions encoding putative outer membrane proteins were identified among the genomes. Several fragments previously identified as unique to either B. suis or B. melitensis were present in the B. abortus genome. Even though several fragments were shared between only B. abortus and B. suis, B. abortus shared more fragments and had fewer nucleotide polymorphisms with B. melitensis than B. suis. The complete genomic sequence of B. abortus provides an important resource for further investigations into determinants of the pathogenicity and virulence phenotypes of these bacteria.
Project description:Brucella abortus is the etiological agent of brucellosis, a disease that affects bovines and human. We generated DNA random sequences from the genome of B. abortus strain 2308 in order to characterize molecular targets that might be useful for developing immunological or chemotherapeutic strategies against this pathogen. The partial sequencing of 1,899 clones allowed the identification of 1,199 genomic sequence surveys (GSSs) with high homology (BLAST expect value < 10(-5)) to sequences deposited in the GenBank databases. Among them, 925 represent putative novel genes for the Brucella genus. Out of 925 nonredundant GSSs, 470 were classified in 15 categories based on cellular function. Seven hundred GSSs showed no significant database matches and remain available for further studies in order to identify their function. A high number of GSSs with homology to Agrobacterium tumefaciens and Rhizobium meliloti proteins were observed, thus confirming their close phylogenetic relationship. Among them, several GSSs showed high similarity with genes related to nodule nitrogen fixation, synthesis of nod factors, nodulation protein symbiotic plasmid, and nodule bacteroid differentiation. We have also identified several B. abortus homologs of virulence and pathogenesis genes from other pathogens, including a homolog to both the Shda gene from Salmonella enterica serovar Typhimurium and the AidA-1 gene from Escherichia coli. Other GSSs displayed significant homologies to genes encoding components of the type III and type IV secretion machineries, suggesting that Brucella might also have an active type III secretion machinery.
Project description:Gene expression analysis of wild-type and STING knock-out mouse bone marrow-derived macrophages (mBMDM) infected with Brucella abortus or transfected with Brucella abortus DNA. Genes whose expression are affected by Brucella abortus in a STING-dependent manner will be identified and signaling pathways regulated by STING will be elucidated. Overall design: Mouse BMDMs were infected with Brucella abortus or transfected with Brucella abortus DNA for 17 hours. Total RNA was extracted for array analysis.
Project description:Brucellosis is a highly contagious zoonosis that occurs worldwide. Whole-genome sequencing (WGS) has become a widely accepted molecular typing method for outbreak tracing and genomic epidemiology of brucellosis. Twenty-nine <i>Brucella</i> spp. (eight <i>B. abortus</i> biovar 1 and 21 <i>B. melitensis</i> biovar 3) were isolated from lymph nodes, milk, and fetal abomasal contents of infected cattle, buffaloes, sheep, and goats originating from nine districts in Egypt. The isolates were identified by microbiological methods and matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS). Differentiation and genotyping were confirmed using multiplex PCR. Illumina MiSeq<sup>®</sup> was used to sequence the 29 <i>Brucella</i> isolates. Using MLST typing, ST11 and ST1 were identified among <i>B. melitensis</i> and <i>B. abortus</i>, respectively. <i>Brucella abortus</i> and <i>B. melitensis</i> isolates were divided into two main clusters (clusters 1 and 2) containing two and nine distinct genotypes by core-genome SNP analysis, respectively. The genotypes were irregularly distributed over time and space in the study area. Both Egyptian <i>B. abortus</i> and <i>B. melitensis</i> isolates proved to be genomically unique upon comparison with publicly available sequencing from strains of neighboring Mediterranean, African, and Asian countries. The antimicrobial resistance mechanism caused by mutations in rpoB, gyrA, and gyrB genes associated with rifampicin and ciprofloxacin resistance were identified. To the best of our knowledge, this is the first study investigating the epidemiology of <i>Brucella</i> isolates from livestock belonging to different localities in Egypt based on whole genome analysis.
Project description:Brucellosis is a zoonotic disease that affects both humans and animals. Its distribution is global, concentrated in the Mediterranean area, India, Central Asia, and Latin America. Here, we present a complete genome assembly of 10 Brucella abortus strains isolated from water buffaloes farmed in the Campania region of Italy.
Project description:Brucella abortus is a class III zoonotic bacterial pathogen able to survive and replicate inside host cells, including macrophages. Here we report a multidimensional transposon sequencing analysis to identify genes essential for Brucella abortus growth in rich medium and replication in RAW 264.7 macrophages. The construction of a dense transposon mutant library and mapping of 929,769 unique mini-Tn5 insertion sites in the genome allowed identification of 491 essential coding sequences and essential segments in the B. abortus genome. Chromosome II carries a lower proportion (5%) of essential genes than chromosome I (19%), supporting the hypothesis of a recent acquisition of a megaplasmid as the origin of chromosome II. Temporally resolved transposon sequencing analysis as a function of macrophage infection stages identified 79 genes with a specific attenuation phenotype in macrophages, at either 2, 5, or 24 h postinfection, and 86 genes for which the attenuated mutant phenotype correlated with a growth defect on plates. We identified 48 genes required for intracellular growth, including the virB operon, encoding the type IV secretion system, which supports the validity of the screen. The remaining genes encode amino acid and pyrimidine biosynthesis, electron transfer systems, transcriptional regulators, and transporters. In particular, we report the need of an intact pyrimidine nucleotide biosynthesis pathway in order for B. abortus to proliferate inside RAW 264.7 macrophages.