• Comparative genomics
• Klebsiella pneumoniae sensu stricto
• Mastitis
• Opportunistic pathogen

• genome
• huanglongbing
• plant pathogens
• prophage
• “Candidatus Liberibacter asiaticus”

• Brazil
• Phylogeny
• Genome, Bacterial
• Plant Diseases/microbiology
• Citrus/microbiology
• Whole Genome Sequencing
• Rhizobiaceae/genetics
• Rhizobiaceae/classification
• Rhizobiaceae/virology
• Prophages/genetics

• 88887.341888/2019-00.88887.817472/2013-00 Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)
• 2015/07011-3 Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)
• 2016/01993-1.2019/17735-0 Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)
• 04 3069 Fundo de Defesa da Citricultura (Fundecitrus)
• 313812/2021-7 Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

• Illumina sequencing
• Peribacillus frigoritolerans
• bactericidal activity
• biosynthetic gene clusters
• draft genome
• fungicidal properties
• mobile genetic elements

• 075-15-2021-1055 date 28 September 2021 Ministry of Science and Higher Education of the Russian Federation

• Bead-beating
• DNA extraction
• Enzymatic lysis
• GridION
• Long-read sequencing
• Pathogen

• Genome, Bacterial
• Plasmids/genetics
• DNA, Bacterial/genetics
• DNA, Bacterial/isolation & purification
• Nanopores
• Nanopore Sequencing/methods
• High-Throughput Nucleotide Sequencing/methods
• Bacteria/genetics
• Bacteria/isolation & purification
• Sequence Analysis, DNA/methods

• HSRI 65-118 Health Systems Research Institute of Thailand under the Genomics Thailand Initiative
• HSRI 65-118 Health Systems Research Institute of Thailand under the Genomics Thailand Initiative
• HSRI 65-118 Health Systems Research Institute of Thailand under the Genomics Thailand Initiative
• HSRI 65-118 Health Systems Research Institute of Thailand under the Genomics Thailand Initiative
• N42A660897 National Research Council of Thailand (NRCT)
• N42A660897 National Research Council of Thailand (NRCT)
• B13F660073 NSRF via the Program Management Unit for Human Resources & Institutional Development, Research Innovation
• B13F660073 NSRF via the Program Management Unit for Human Resources & Institutional Development, Research Innovation

• Dellaglioa algida
• Dellaglioa carnosa
• High-oxygen modified-atmosphere packaged meat (HiOx-MAP)
• Underestimated genus

• Animals
• Cattle
• Food Packaging/methods
• Oxygen/analysis
• Meat/analysis
• Red Meat/analysis
• Atmosphere/analysis

• Antimicrobial resistance
• Entrapment vector
• Insertion sequence
• Intracellular
• Mobile genetic elements
• Plasmids

• Chloramphenicol/pharmacology
• Escherichia coli
• Anti-Bacterial Agents/pharmacology
• Plasmids/genetics
• Interspersed Repetitive Sequences
• Drug Resistance, Bacterial/genetics
• Microbial Sensitivity Tests

• MR/W030578/1 Medical Research Council

• Chlamydia psittaci
• Genome analysis
• Host preference
• Inclusion proteins
• Phylogeny
• Plasmid
• Plasticity zone
• Polymorphic membrane proteins

• Animals
• Humans
• Horses
• Swine
• Chlamydophila psittaci/genetics
• Psittacosis/veterinary
• Phylogeny
• Chlamydia/genetics
• Birds
• Genomics

• FZT 118 - iDiv 202548816 Deutsche Forschungsgemeinschaft
• Friedrich-Schiller-Universität Jena (1010)

• Commensal Neisseria
• Neisseria spp
• T6SS
• Toxin antitoxin system
• Type 6 secretion system
• Type VI secretion system

Transposable elements (TEs) are major genomic components in most eukaryotic genomes and play an important role in genome evolution. However, despite their relevance the identification of TEs is not an easy task and a number of tools were developed to tackle this problem. To better understand how they perform, we tested several widely used tools for de novo TE detection and compared their performance on both simulated data and well curated genomic sequences. As expected, tools that build TE-models performed better than k-mer counting ones, with RepeatModeler beating competitors in most datasets. However, there is a tendency for most tools to identify TE-regions in a fragmented manner and it is also frequent that small TEs or fragmented TEs are not detected. Consequently, the identification of TEs is still a challenging endeavor and it requires a significant manual curation by an experienced expert. The results will be helpful for identifying common issues associated with TE-annotation and for evaluating how comparable are the results obtained with different tools.

• Nanopore
• SAP11
• effector
• genome sequencing
• phytoplasma
• potential mobile unit

• Enterobacterales
• Lactobacillales
• Rhodospirillales
• bacterial metabolism
• comparative genomics
• functional redundancy

• Animals
• Drosophila/genetics
• Drosophila melanogaster/genetics
• Drosophila melanogaster/microbiology
• Gastrointestinal Microbiome/genetics
• Metagenome
• Phylogeny

• R01 GM095372 NIGMS NIH HHS

Antimicrobial resistance (AMR) is a significant public health threat. With the rise of affordable whole genome sequencing, in silico approaches to assessing AMR gene content can be used to detect known resistance mechanisms and potentially identify novel mechanisms. To enable accurate assessment of AMR gene content, as part of a multi-agency collaboration, NCBI developed a comprehensive AMR gene database, the Bacterial Antimicrobial Resistance Reference Gene Database and the AMR gene detection tool AMRFinder. Here, we describe the expansion of the Reference Gene Database, now called the Reference Gene Catalog, to include putative acid, biocide, metal, stress resistance genes, in addition to virulence genes and species-specific point mutations. Genes and point mutations are classified by broad functions, as well as more detailed functions. As we have expanded both the functional repertoire of identified genes and functionality, NCBI released a new version of AMRFinder, known as AMRFinderPlus. This new tool allows users the option to utilize only the core set of AMR elements, or include stress response and virulence genes, too. AMRFinderPlus can detect acquired genes and point mutations in both protein and nucleotide sequence. In addition, the evidence used to identify the gene has been expanded to include whether nucleotide or protein sequence was used, its location in the contig, and presence of an internal stop codon. These database improvements and functional expansions will enable increased precision in identifying AMR genes, linking AMR genotypes and phenotypes, and determining possible relationships between AMR, virulence, and stress response.

• Chlamydia
• Chlamydia psittaci
• Chlamydia trachomatis
• annotation tool
• core genome
• genome analysis
• host preference
• plasticity zone
• variable genomic sites

• add-on fellowship for interdisciplinary life science Joachim Herz Foundation

• PblB
• Streptococcus pneumoniae
• lysogeny
• phage integration
• phage phylogeny
• temperate bacteriophages

• NA Swedish Research Council
• NA Swedish Foundation for Strategic research (SSF)
• NA Stockholm County Council
• NA Knut and Alice Wallenberg foundation

Ralstonia solanacearum, a soil-borne plant pathogen, encompasses a large number of strains known as R. solanacearum species complex (RSSC). Although it has been suggested that mobile genetic elements (MGEs) may play an important role in the RSSC genome, the evolutionary impact of these elements remains unknown. Here, we identified and analysed Integrative and Conjugative Elements (ICEs) and Genomic Islands (GIs) in the 121 genomes published for Ralstonia spp., including RSSC strains and three other non-plant pathogenic Ralstonia spp. Our results provided a dataset of 12 ICEs and 31 GIs distributed throughout Ralstonia spp. Four novel ICEs in RSSC were found. Some of these elements cover 5% of the host genome and carry accessory genes with a potential impact on the fitness and pathogenicity of RSSC. In addition, phylogenetic analysis revealed that these MGEs clustered to the same species, but there is evidence of strains from different countries that host the same element. Our results provide novel insight into the RSSC adaptation, opening new paths to a better understanding of how these elements affect this soil-borne plant pathogen.

• Antimicrobial resistance
• Biomarker
• Enterococci
• Genome size
• Genomics
• Mobilome
• Pangenome
• Vancomycin resistant Enterococcus (VRE)
• Wastewater

• Bacterial Proteins/genetics
• Drug Resistance, Multiple, Bacterial
• Enterococcus faecium/genetics
• Genome Size
• Genomics/methods
• Interspersed Repetitive Sequences
• Multilocus Sequence Typing
• Phylogeny
• Vancomycin Resistance
• Virulence Factors/genetics
• Wastewater/microbiology
• Whole Genome Sequencing

• 001 Genomics Research and Development Initiative
• Beef Cattle Research Council

• Antimicrobial resistance (AMR)
• MinION
• Oxford nanopore
• S. suis
• Sequence typing
• Sequencing

• Bacteroides fragilis
• Oxford Nanopore
• antimicrobial resistance
• genome sequencing
• hybrid assembly
• insertion sequences
• plasmid

• Bacteroides fragilis/genetics
• Drug Resistance, Multiple, Bacterial/genetics
• Genome, Bacterial/genetics
• Genomics/methods
• High-Throughput Nucleotide Sequencing/methods
• Humans
• Sequence Analysis, DNA/methods
• Whole Genome Sequencing/methods

• microbiology techniques
• next-generation sequencing
• infectious-disease diagnostics

• Anti-Bacterial Agents/pharmacology
• Bacteria/genetics
• Computational Biology/methods
• Drug Resistance, Bacterial/genetics
• Drug Resistance, Microbial/genetics
• Genetic Engineering
• Genome, Bacterial/drug effects
• Genomics/methods
• High-Throughput Nucleotide Sequencing/methods
• Nanopore Sequencing/methods
• Sequence Analysis, DNA/methods
• Virulence/drug effects
• Whole Genome Sequencing/methods

Generating a complete, de novo genome assembly for prokaryotes is often considered a solved problem. However, we here show that Pseudomonas koreensis P19E3 harbors multiple, near identical repeat pairs up to 70 kilobase pairs in length, which contained several genes that may confer fitness advantages to the strain. Its complex genome, which also included a variable shufflon region, could not be de novo assembled with long reads produced by Pacific Biosciences’ technology, but required very long reads from Oxford Nanopore Technologies. Importantly, a repeat analysis, whose results we release for over 9600 prokaryotes, indicated that very complex bacterial genomes represent a general phenomenon beyond Pseudomonas. Roughly 10% of 9331 complete bacterial and a handful of 293 complete archaeal genomes represented this ‘dark matter’ for de novo genome assembly of prokaryotes. Several of these ‘dark matter’ genome assemblies contained repeats far beyond the resolution of the sequencing technology employed and likely contain errors, other genomes were closed employing labor-intense steps like cosmid libraries, primer walking or optical mapping. Using very long sequencing reads in combination with assembly algorithms capable of resolving long, near identical repeats will bring most prokaryotic genomes within reach of fast and complete de novo genome assembly.

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.

• Brucella abortus
• comparative genomics
• genome typing
• phage resistance
• phylogenetic analysis

Bacteria can acquire an accessory genome through the horizontal transfer of genetic elements from non-parental lineages. This leads to rapid genetic evolution allowing traits such as antibiotic resistance and virulence to spread through bacterial communities. The study of complete genomes of bacterial strains helps to understand the genomic traits associated with virulence and antibiotic resistance. We aimed to investigate the complete accessory genome of an ocular isolate of Pseudomonas aeruginosa strain PA34. We obtained the complete genome of PA34 utilising genome sequence reads from Illumina and Oxford Nanopore Technology followed by PCR to close any identified gaps. In-depth genomic analysis was performed using various bioinformatics tools. The susceptibility to heavy metals and cytotoxicity was determined to confirm expression of certain traits. The complete genome of PA34 includes a chromosome of 6.8 Mbp and two plasmids of 95.4 Kbp (pMKPA34-1) and 26.8 Kbp (pMKPA34-2). PA34 had a large accessory genome of 1,213 genes and had 543 unique genes not present in other strains. These exclusive genes encoded features related to metal and antibiotic resistance, phage integrase and transposons. At least 24 genomic islands (GIs) were predicated in the complete chromosome, of which two were integrated into novel sites. Eleven GIs carried virulence factors or replaced pathogenic genes. A bacteriophage carried the aminoglycoside resistance gene (AAC(3)-IId). The two plasmids carried other six antibiotic resistance genes. The large accessory genome of this ocular isolate plays a large role in shaping its virulence and antibiotic resistance.

• Adult
• Corneal Diseases/genetics
• Corneal Diseases/microbiology
• Drug Resistance, Multiple/genetics
• Genome, Bacterial
• Genomic Islands
• Humans
• Keratitis/genetics
• Keratitis/microbiology
• Male
• Pseudomonas Infections/genetics
• Pseudomonas Infections/microbiology
• Pseudomonas aeruginosa/genetics
• Pseudomonas aeruginosa/isolation & purification
• Young Adult

• Nanopore
• RNA-Seq
• adaptive laboratory evolution
• multireplicon
• platinum resistance

• Cupriavidus/drug effects
• Cupriavidus/genetics
• Drug Resistance, Bacterial
• Evolution, Molecular
• Genome, Bacterial
• Platinum/toxicity
• Transcriptome

The large and complex genome of Pseudomonas aeruginosa, which consists of significant portions (up to 20%) of transferable genetic elements contributes to the rapid development of antibiotic resistance. The whole genome sequences of 22 strains isolated from eye and cystic fibrosis patients in Australia and India between 1992 and 2007 were used to compare genomic divergence and phylogenetic relationships as well as genes for antibiotic resistance and virulence factors. Analysis of the pangenome indicated a large variation in the size of accessory genome amongst 22 stains and the size of the accessory genome correlated with number of genomic islands, insertion sequences and prophages. The strains were diverse in terms of sequence type and dissimilar to that of global epidemic P. aeruginosa clones. Of the eye isolates, 62% clustered together within a single lineage. Indian eye isolates possessed genes associated with resistance to aminoglycoside, beta-lactams, sulphonamide, quaternary ammonium compounds, tetracycline, trimethoprims and chloramphenicols. These genes were, however, absent in Australian isolates regardless of source. Overall, our results provide valuable information for understanding the genomic diversity of P. aeruginosa isolated from two different infection types and countries.

• Cupriavidus
• metal
• mobile genetic elements
• phenotype microarray
• resistance

Lactobacillus casei, Lactobacillus paracasei, and Lactobacillus rhamnosus are phenotypically and genotypically closely related, and together comprise the L. casei group. Although the strains of this group are commercially valuable as probiotics, the taxonomic status and nomenclature of the L. casei group have long been contentious because of the difficulties in identifying these three species by using the most frequently used genotypic methodology of 16S rRNA gene sequencing. Long used as the gold standard for species classification, DNA–DNA hybridization is laborious, requires expert skills, and is difficult to use routinely in laboratories. Currently, genome-based comparisons, including average nucleotide identity (ANI) and digital DNA–DNA hybridization (dDDH), are commonly applied to bacterial taxonomy as alternatives to the gold standard method for the demarcating phylogenetic relationships. To establish quick and accurate methods for identifying strains in the L. casei group at the species and subspecies levels, we developed species- and subspecies-specific identification methods based on housekeeping gene sequences and whole-cell matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) spectral pattern analysis. By phylogenetic analysis based on concatenated housekeeping gene sequences (dnaJ, dnaK, mutL, pheS, and yycH), 53 strains were separated into four clusters corresponding to the four species: L. casei, L. paracasei and L. rhamnosus, and Lactobacillus chiayiensis sp. nov. A multiplex minisequencing assay using single nucleotide polymorphism (SNP)-specific primers based on the dnaK gene sequences and species-specific primers based on the mutL gene sequences provided high resolution that enabled the strains at the species level to be identified as L. casei, L. paracasei, and L. rhamnosus. By MALDI-TOF MS analysis coupled with an internal database and ClinProTools software, species- and subspecies-level L. casei group strains were identified based on reliable scores and species- and subspecies-specific MS peaks. The L. paracasei strains were distinguished clearly at the subspecies level based on subspecies-specific MS peaks. This article describes the rapid and accurate methods used for identification and classification of strains in the L. casei group based on housekeeping gene sequences and MALDI-TOF MS analysis as well as the novel speciation of this group including L. chiayiensis sp. nov. and ‘Lactobacillus zeae’ by genome-based methods.

• Lactobacillus casei group
• MALDI-TOF MS
• MLSA
• species-specific PCR
• whole genome sequence

• PacBio
• Pseudomonas aeruginosa
• arrivée d’eau d’un appareil de rinçage dentaire
• dental unit waterlines
• genomic islands
• insertion sequences
• introns
• mobilome
• prophages
• séquences d’insertion
• îlots génomiques

Although complete genome sequences hold particular value for an accurate description of core genomes, the identification of strain-specific genes, and as the optimal basis for functional genomics studies, they are still largely underrepresented in public repositories. Based on an assessment of the genome assembly complexity for all lactobacilli, we used Pacific Biosciences' long read technology to sequence and de novo assemble the genomes of three Lactobacillus helveticus starter strains, raising the number of completely sequenced strains to 12. The first comparative genomics study for L. helveticus—to our knowledge—identified a core genome of 988 genes and sets of unique, strain-specific genes ranging from about 30 to more than 200 genes. Importantly, the comparison of MiSeq- and PacBio-based assemblies uncovered that not only accessory but also core genes can be missed in incomplete genome assemblies based on short reads. Analysis of the three genomes revealed that a large number of pseudogenes were enriched for functional Gene Ontology categories such as amino acid transmembrane transport and carbohydrate metabolism, which is in line with a reductive genome evolution in the rich natural habitat of L. helveticus. Notably, the functional Clusters of Orthologous Groups of proteins categories “cell wall/membrane biogenesis” and “defense mechanisms” were found to be enriched among the strain-specific genes. A genome mining effort uncovered examples where an experimentally observed phenotype could be linked to the underlying genotype, such as for cell envelope proteinase PrtH3 of strain FAM8627. Another possible link identified for peptidoglycan hydrolases will require further experiments. Of note, strain FAM22155 did not harbor a CRISPR/Cas system; its loss was also observed in other L. helveticus strains and lactobacillus species, thus questioning the value of the CRISPR/Cas system for diagnostic purposes. Importantly, the complete genome sequences proved to be very useful for the analysis of natural whey starter cultures with metagenomics, as a larger percentage of the sequenced reads of these complex mixtures could be unambiguously assigned down to the strain level.

• CRISPR/Cas
• PacBio
• comparative genomics
• dairy industry
• metagenomics
• natural whey starter cultures
• strain-specific genes
• whole genome sequencing

• Rhizobium leguminosarum
• genome
• nitrogen fixation
• plasmid
• symbiosis

• R01 GM080227 NIGMS NIH HHS

• microbial genetics
• molecular microbiology
• genomics
• microbiology
• draft genomes

Bacteriophages are the most abundant biological entities on the planet, playing crucial roles in the shaping of bacterial populations. Phages have smaller genomes than their bacterial hosts, yet there are currently fewer fully sequenced phage than bacterial genomes. We assessed the suitability of Illumina technology for high-throughput sequencing and subsequent assembly of phage genomes. In silico datasets reveal that 30× coverage is sufficient to correctly assemble the complete genome of ~98.5% of known phages, with experimental data confirming that the majority of phage genomes can be assembled at 30× coverage. Furthermore, in silico data demonstrate it is possible to co-sequence multiple phages from different hosts, without introducing assembly errors.

• Assembly
• Bacteriophage
• Genome
• Illumina
• Sequencing

Many new Drosophila genomes have been sequenced in recent years using new-generation sequencing platforms and assembly methods. Transposable elements (TEs), being repetitive sequences, are often misassembled, especially in the genomes sequenced with short reads. Consequently, the mobile fraction of many of the new genomes has not been analyzed in detail or compared with that of other genomes sequenced with different methods, which could shed light into the understanding of genome and TE evolution. Here we compare the TE content of three genomes: D. buzzatii st-1, j-19, and D. mojavensis.

We have sequenced a new D. buzzatii genome (j-19) that complements the D. buzzatii reference genome (st-1) already published, and compared their TE contents with that of D. mojavensis. We found an underestimation of TE sequences in Drosophila genus NGS-genomes when compared to Sanger-genomes. To be able to compare genomes sequenced with different technologies, we developed a coverage-based method and applied it to the D. buzzatii st-1 and j-19 genome. Between 10.85 and 11.16 % of the D. buzzatii st-1 genome is made up of TEs, between 7 and 7,5 % of D. buzzatii j-19 genome, while TEs represent 15.35 % of the D. mojavensis genome. Helitrons are the most abundant order in the three genomes.

TEs in D. buzzatii are less abundant than in D. mojavensis, as expected according to the genome size and TE content positive correlation. However, TEs alone do not explain the genome size difference. TEs accumulate in the dot chromosomes and proximal regions of D. buzzatii and D. mojavensis chromosomes. We also report a significantly higher TE density in D. buzzatii and D. mojavensis X chromosomes, which is not expected under the current models. Our easy-to-use correction method allowed us to identify recently active families in D. buzzatii st-1 belonging to the LTR-retrotransposon superfamily Gypsy.

The online version of this article (doi:10.1186/s12864-016-2648-8) contains supplementary material, which is available to authorized users.

• Buzzatii
• Drosophila
• Genome
• Transposable elements

• Burkholderia
• Chlorocatechol degradation genes
• Genome assembly
• Mobile elements
• Next generation sequencing
• Plasmid

• Burkholderia/genetics
• Catechols/metabolism
• Genome, Bacterial
• High-Throughput Nucleotide Sequencing
• Operon
• Repetitive Sequences, Nucleic Acid
• Replicon
• Sequence Analysis, DNA
• Species Specificity

Plasmids play critical roles in bacterial fitness and evolution of Pseudomonas aeruginosa. Here two plasmids found in a drug-resistant P. aeruginosa clinical isolate HS87 were completely sequenced. The pHS87b plasmid (11.2 kb) carries phage-related genes and function-unknown genes. Notably, pHS87b encodes an integrase and has an adjacent tRNA^Thr-associated attachment site. A corresponding integrated form of pHS87b at the tRNA^Thr locus was identified on the chromosome of P. aeruginosa, showing that pHS87b is able to site-specifically integrate into the 3’-end of the tRNA^Thr gene. The pHS87a plasmid (26.8 kb) displays a plastic structure containing a putative replication module, stability factors and a variable region. The RepA of pHS87a shows significant similarity to the replication proteins of pPT23A-family plasmids. pHS87a carries a transposon Tn6049, a truncated insertion sequence ΔIS1071 and a Tn402-like class 1 integron which contains an aacA4 cassette that may confer aminoglycoside resistance. Thus, pHS87b is a site-specific integrative plasmid whereas pHS87a is a plastic antibiotic resistance plasmid. The two native plasmids may promote the fitness and evolution of P. aeruginosa.

• Aminoglycosides
• Chromosomes, Bacterial/genetics
• DNA Transposable Elements/genetics
• Drug Resistance, Bacterial/genetics
• Evolution, Molecular
• Plasmids/genetics
• Pseudomonas aeruginosa/genetics
• RNA, Transfer, Thr/genetics
• Replication Origin/genetics

While the extent and impact of horizontal transfers in prokaryotes are widely acknowledged, their importance to the eukaryotic kingdom is unclear and thought by many to be anecdotal. Here we report multiple recent transfers of a huge genomic island between Penicillium spp. found in the food environment. Sequencing of the two leading filamentous fungi used in cheese making, P. roqueforti and P. camemberti, and comparison with the penicillin producer P. rubens reveals a 575 kb long genomic island in P. roqueforti—called Wallaby—present as identical fragments at non-homologous loci in P. camemberti and P. rubens. Wallaby is detected in Penicillium collections exclusively in strains from food environments. Wallaby encompasses about 250 predicted genes, some of which are probably involved in competition with microorganisms. The occurrence of multiple recent eukaryotic transfers in the food environment provides strong evidence for the importance of this understudied and probably underestimated phenomenon in eukaryotes.

Horizontal gene transfers are known to play an important role in prokaryote evolution but their impact and prevalence in eukaryotes is less clear. Here, the authors sequence the genomes of cheese making fungi P. roqueforti and P. camemberti, and provide evidence for recent horizontal transfers of a large genomic region.

Advances in sequencing technology have drastically increased the depth and feasibility of bacterial genome sequencing. However, little information is available that details the specific techniques and procedures employed during genome sequencing despite the large numbers of published genomes. Shotgun approaches employed by second-generation sequencing platforms has necessitated the development of robust bioinformatics tools for in silico assembly, and complete assembly is limited by the presence of repetitive DNA sequences and multi-copy operons. Typically, re-sequencing with multiple platforms and laborious, targeted Sanger sequencing are employed to finish a draft bacterial genome. Here we describe a novel strategy based on the identification and targeted sequencing of repetitive rDNA operons to expedite bacterial genome assembly and finishing. Our strategy was validated by finishing the genome of Paenibacillus polymyxa strain CR1, a bacterium with potential in sustainable agriculture and bio-based processes. An analysis of the 38 contigs contained in the P. polymyxa strain CR1 draft genome revealed 12 repetitive rDNA operons with varied intragenic and flanking regions of variable length, unanimously located at contig boundaries and within contig gaps. These highly similar but not identical rDNA operons were experimentally verified and sequenced simultaneously with multiple, specially designed primer sets. This approach also identified and corrected significant sequence rearrangement generated during the initial in silico assembly of sequencing reads. Our approach reduces the required effort associated with blind primer walking for contig assembly, increasing both the speed and feasibility of genome finishing. Our study further reinforces the notion that repetitive DNA elements are major limiting factors for genome finishing. Moreover, we provided a step-by-step workflow for genome finishing, which may guide future bacterial genome finishing projects.

• bacterial genomics
• contigs assembly
• genome finishing
• ribosomal DNA
• second-generation sequencing (SGS)

• n. nucifera
• chloroplast genome sequencing
• basal eudicots
• systematic position
• divergence time
• pacbio rs ii

• Base Sequence
• Chloroplasts/genetics
• Codon/genetics
• Evolution, Molecular
• Genome, Chloroplast/genetics
• Molecular Sequence Annotation
• Molecular Sequence Data
• Nelumbo/genetics
• Phylogeny
• Plastids/genetics
• RNA, Ribosomal/genetics
• RNA, Transfer/genetics
• Sequence Analysis, DNA/instrumentation
• Sequence Analysis, DNA/methods

Escherichia coli ST131 is now recognised as a leading contributor to urinary tract and bloodstream infections in both community and clinical settings. Here we present the complete, annotated genome of E. coli EC958, which was isolated from the urine of a patient presenting with a urinary tract infection in the Northwest region of England and represents the most well characterised ST131 strain. Sequencing was carried out using the Pacific Biosciences platform, which provided sufficient depth and read-length to produce a complete genome without the need for other technologies. The discovery of spurious contigs within the assembly that correspond to site-specific inversions in the tail fibre regions of prophages demonstrates the potential for this technology to reveal dynamic evolutionary mechanisms. E. coli EC958 belongs to the major subgroup of ST131 strains that produce the CTX-M-15 extended spectrum β-lactamase, are fluoroquinolone resistant and encode the fimH30 type 1 fimbrial adhesin. This subgroup includes the Indian strain NA114 and the North American strain JJ1886. A comparison of the genomes of EC958, JJ1886 and NA114 revealed that differences in the arrangement of genomic islands, prophages and other repetitive elements in the NA114 genome are not biologically relevant and are due to misassembly. The availability of a high quality uropathogenic E. coli ST131 genome provides a reference for understanding this multidrug resistant pathogen and will facilitate novel functional, comparative and clinical studies of the E. coli ST131 clonal lineage.

• Anti-Bacterial Agents/pharmacology
• Chromosomes, Bacterial
• Computational Biology
• Drug Resistance, Multiple, Bacterial/genetics
• England
• Escherichia coli/classification
• Escherichia coli/drug effects
• Escherichia coli/genetics
• Escherichia coli Infections/microbiology
• Gene Order
• Genome, Bacterial
• Genomics
• Humans
• Molecular Sequence Annotation
• Phylogeny
• Sequence Analysis, DNA
• Urinary Tract Infections/microbiology