Draft Genome Sequence of Environmental Isolate Acinetobacter nosocomialis U20-HoPe-S34-3 from Germany

The draft genome sequence of Acinetobacter nosocomialis U20-HoPe-S34-3, isolated from soil sampled from the banks of the river Holtemme in Germany, is provided. The strain has an average nucleotide identity of 98.3% to the type strain of the species.Peer Reviewe

Flagellin genes of Yersinia enterocolitica biotype 1A: playground of evolution towards novel flagellin functions

Yersinia enterocolitica strain 8081, representing the high-pathogenic biotype 1B, harbours three flagellin genes arranged in tandem in the order fliC3, fliC, fliC2. The genes are organized monocistronic but coordinately expressed under the control of the flagellar sigma factor. No sequence data is available on flagellins of low-pathogenic Y. enterocolitica biotypes 2-5 and of biotype 1A strains, appearing non-pathogenic in the mouse infection model. We sequenced the flagellin genes of ten biotype 1A and biotype 4 isolates, respectively. While we could not identify any sequence polymorphism among flagellin genes of biotype 4 isolates, we found that biotype 1A strains harbour three variable flagellin genes. Moreover, three biotype 1A isolates exhibited a rearranged flagellin gene order and at least one rearranged flagellin gene was apparently acquired by horizontal gene transfer. The variability of flagellin genes seems to mirror evolution towards novel flagellin functions. By contrast, strictly conserved flagellins of biotype 4 isolates point at a strong selection pressure such as expected to be imposed by an important function in the context of infection

Interrelationship between type three secretion system and metabolism in pathogenic bacteria

Before the advent of molecular biology methods, studies of pathogens were dominated by analyses of their metabolism. Development of molecular biology techniques then enabled the identification and functional characterisation of the fascinating toolbox of virulence factors. Increasing, genomic and proteomic approaches form the basis for a more systemic view on pathogens’ functions in the context of infection. Re-emerging interest in the metabolism of pathogens and hosts further expands our view of infections. There is increasing evidence that virulence functions and metabolism of pathogens are extremely intertwined. Type three secretion systems (T3SSs) are major virulence determinants of many Gram-negative pathogens and it is the objective of this review to illustrate the intertwined relationship between T3SSs and the metabolism of the pathogens deploying them

Evaluation of CHROMagar Acinetobacter and MacConkey media for the recovery of Acinetobacter baumannii from soil samples

The opportunistic pathogen Acinetobacter baumannii occupies niches in human and veterinary clinics and other environments. This comparative study was designed to assess the recovery rate of Ac. baumannii from 20 soil samples collected in Germany (from which Ac. baumannii had been previously isolated) on selective CHROMagar Acinetobacter (CMA) and MacConkey Agar (MCA). Suspected Ac. baumannii were confirmed by detection and sequencing of blaOXA-51-like genes. Overall, the recovery rate of Ac. baumannii from both media was similar. Out of the 20 soil samples enriched in mineral salt medium (MM) plus 0.2% acetate for 5 h, Ac. baumannii was recovered from 12 (60%) on CMA and 11 (55%) on MCA, and from 14 (70%) with both selective media after enrichment for 24 h. Typical and distinct colonies of Ac. baumannii were more often observed on CMA with soil enriched in MM for 5 h, while on MCA distinct colonies were more obvious after 24 h. In five soil samples (25%), strains harbouring different OXA-51-like variants were recovered on MCA. Late lactose fermentation (48 h) was observed on MCA. The study suggests that no single growth medium would efficiently recover Ac. baumannii from all soil samples.Peer Reviewe

Novel Genes Required for Surface-Associated Motility in Acinetobacter baumannii

Acinetobacter baumannii is an opportunistic and increasingly multi-drug resistant human pathogen rated as a critical priority one pathogen for the development of new antibiotics by the WHO in 2017. Despite the lack of flagella, A. baumannii can move along wet surfaces in two different ways: via twitching motility and surface-associated motility. While twitching motility is known to depend on type IV pili, the mechanism of surface-associated motility is poorly understood. In this study, we established a library of 30 A. baumannii ATCC® 17978™ mutants that displayed deficiency in surface-associated motility. By making use of natural competence, we also introduced these mutations into strain 29D2 to differentiate strain-specific versus species-specific effects of mutations. Mutated genes were associated with purine/pyrimidine/folate biosynthesis (e.g. purH, purF, purM, purE), alarmone/stress metabolism (e.g. Ap4A hydrolase), RNA modification/regulation (e.g. methionyl-tRNA synthetase), outer membrane proteins (e.g. ompA), and genes involved in natural competence (comEC). All tested mutants originally identified as motility-deficient in strain ATCC® 17978™ also displayed a motility-deficient phenotype in 29D2. By contrast, further comparative characterization of the mutant sets of both strains regarding pellicle biofilm formation, antibiotic resistance, and virulence in the Galleria mellonella infection model revealed numerous strain-specific mutant phenotypes. Our studies highlight the need for comparative analyses to characterize gene functions in A. baumannii and for further studies on the mechanisms underlying surface-associated motility.Peer Reviewe

Yersinia enterocolitica type III secretion: Evidence for the ability to transport proteins that are folded prior to secretion

BACKGROUND: Pathogenic Yersinia species (Y. enterocolitica, Y. pestis, Y. pseudotuberculosis) share a type three secretion system (TTSS) which allows translocation of effector proteins (called Yops) into host cells. It is believed that proteins are delivered through a hollow needle with an inner diameter of 2–3 nm. Thus transport seems to require substrates which are essentially unfolded. Recent work from different groups suggests that the Yersinia TTSS cannot accommodate substrates which are folded prior to secretion. It was suggested that folding is prevented either by co-translational secretion or by the assistance of specific Yop chaperones (called Sycs). RESULTS: In this study we have fused YopE secretion signals of various length to the mouse dihydrofolate reductase (DHFR) in order to analyse the DHFR folding state prior to secretion. We could demonstrate that secretion-deficient as well as secretion-competent YopE-DHFR fusions complexed to SycE can be efficiently purified from Yersinia cytosol by affinity chromatography using methotrexate-agarose. This implies the folding of the DHFR fusion moiety despite SycE binding and contradicts the previously presented model of folding inhibition by chaperone binding. Secretion-deficient YopE-DHFR fusions caused severe jamming of the TTSS. This observation contradicts the co-translational secretion model. CONCLUSIONS: We present evidence that the Yersinia TTSS is familiar with the processing of transport substrates which are folded prior to secretion. We therefore predict that an unfoldase is involved in type III secretion

Impact of Acinetobacter baumannii Superoxide Dismutase on Motility, Virulence, Oxidative Stress Resistance and Susceptibility to Antibiotics

Acinetobacter baumannii is a Gram-negative bacterium appearing as an opportunistic pathogen in hospital settings. Superoxide dismutase (SOD) contributes to virulence in several pathogenic bacteria by detoxifying reactive oxygen species released in the course of host defense reactions. However, the biological role of SODs in A. baumannii has not yet been elucidated. Here, we inactivated in A. baumannii ATCC 17978 gene A1S_2343, encoding a putative SOD of the Fe-Mn type by transposon insertion, resulting in mutant ATCC 17978 sod2343::Km. The mutation was also introduced in two naturally competent A. baumannii isolates by transformation with chromosomal DNA derived from mutant ATCC 17978 sod2343::Km. We demonstrate that inactivation of sod2343 leads to significant motility defects in all three A. baumannii strains. The mutant strains were more susceptible to oxidative stress compared to their parental strains. Susceptibility to colistin and tetracycline was increased in all mutant strains while susceptibility of the mutants to gentamicin, levofloxacin and imipenem was strain-dependent. In the Galleria mellonella infection model the mutant strains were significantly attenuated. In conclusion, sod2343 plays an important role in motility, resistance to oxidative stress, susceptibility to antibiotics and virulence in A. baumannii

The Yersinia enterocolitica type three secretion chaperone SycO is integrated into the Yop regulatory network and binds to the Yop secretion protein YscM1

<p>Abstract</p> <p>Background</p> <p>Pathogenic yersiniae (<it>Y. pestis</it>, <it>Y. pseudotuberculosis</it>, <it>Y. enterocolitica</it>) share a virulence plasmid encoding a type three secretion system (T3SS). This T3SS comprises more than 40 constituents. Among these are the transport substrates called Yops (<it>Yersinia </it>outer proteins), the specific Yop chaperones (Sycs), and the Ysc (Yop secretion) proteins which form the transport machinery. The effectors YopO and YopP are encoded on an operon together with SycO, the chaperone of YopO. The characterization of SycO is the focus of this study.</p> <p>Results</p> <p>We have established the large-scale production of recombinant SycO in its outright form. We confirm that <it>Y. enterocolitica </it>SycO forms homodimers which is typical for Syc chaperones. SycO overproduction in <it>Y. enterocolitica </it>decreases secretion of Yops into the culture supernatant suggesting a regulatory role of SycO in type III secretion. We demonstrate that <it>in vitro </it>SycO interacts with YscM1, a negative regulator of Yop expression in <it>Y. enterocolitica</it>. However, the SycO overproduction phenotype was not mediated by YscM1, YscM2, YopO or YopP as revealed by analysis of isogenic deletion mutants.</p> <p>Conclusion</p> <p>We present evidence that SycO is integrated into the regulatory network of the <it>Yersinia </it>T3SS. Our picture of the <it>Yersinia </it>T3SS interactome is supplemented by identification of the SycO/YscM1 interaction. Further, our results suggest that at least one additional interaction partner of SycO has to be identified.</p

Adding to Yersinia enterocolitica Gene Pool Diversity: Two Cryptic Plasmids from a Biotype 1A Isolate

We report the nucleotide sequence of two novel cryptic plasmids (4357 and 14 662 base pairs) carried by a Yersinia enterocolitica biotype 1A strain isolated from pork. As distinguished from most biotype 1A strains, this isolate, designated 07-04449, exhibited adherence to eukaryotic cells. The smaller plasmid pYe4449-1 carries five attributable open reading frames (ORFs) encoding the first CcdA/CcdB-like antitoxin/toxin system described for a Yersinia plasmid, a RepA-like replication initiation protein, and mobilizing factors MobA and MobC. The deduced amino acid sequences showed highest similarity to proteins described in Salmonella (CcdA/B), Klebsiella (RepA), and Plesiomonas (MobA/C) indicating genomic fluidity among members of the Enterobacteriaceae. One additional ORF with unknown function, termed ORF5, was identified with an ancestry distinct from the rest of the plasmid. While the C+G content of ORF5 is 38.3%, the rest of pYe4449-1 shows a C+G content of 55.7%. The C+G content of the larger plasmid pYe4449-2 (54.9%) was similar to that of pYe4449-1 (53.7%) and differed from that of the Y. enterocolitica genome (47.3%). Of the 14 ORFs identified on pYe4449-2, only six ORFs showed significant similarity to database entries. For three of these ORFs likely functions could be ascribed: a TnpR-like resolvase and a phage replication protein, localized each on a low C+G island, and DNA primase TraC. Two ORFs of pYe4449-2, ORF3 and ORF7, seem to encode secretable proteins. Epitope-tagging of ORF3 revealed protein expression at 4°C but not at or above 27°C suggesting adaptation to a habitat outside swine. The hypothetical protein encoded by ORF7 is the member of a novel repeat protein family sharing the DxxGN(x)nDxxGN motif. Our findings illustrate the exceptional gene pool diversity within the species Y. enterocolitica driven by horizontal gene transfer events

Complete Genome Sequencing of Acinetobacter sp. Strain LoGeW2-3, Isolated from the Pellet of a White Stork, Reveals a Novel Class D Beta-Lactamase Gene

Whole-genome sequencing of Acinetobacter sp. strain LoGeW2-3, isolated from the pellet of a white stork (Ciconia ciconia), reveals the presence of a plasmid of 179,399 bp encoding a CRISPR-Cas (clustered regularly interspaced short palindromic repeats and associated genes) system of the I-F type, and the chromosomally encoded novel class D beta-lactamase OXA-568