GSC/BRC Metadata Standards ProjectSetup: Example CSV file for setting up Project registration and update event for GSC/BRC metadata standards. User as load the setup file using CLI interface or user can setup project using metadata setup GUI. (CSV 14 kb)
Enterobacter aerogenes was recently renamed Klebsiella aerogenes . This study aimed to identify differences in clinical characteristics, outcomes, and bacterial genetics among patients with K. aerogenes versus Enterobacter species bloodstream infections (BSI). We prospectively enrolled patients with K. aerogenes or Enterobacter cloacae complex ( Ecc ) BSI from 2002 to 2015.
By targeting penicillin binding protein-3, the AmpC β-lactamase, and MurA, another enzyme involved in cell wall synthesis, with the ceftazidime–avibactam–fosfomycin combination, we previously overcame multidrug resistance (MDR) in vitro in an archived collection of Pseudomonas aeruginosa clinical isolates. Here, we further validate the ceftazidime–avibactam–fosfomycin combination using the MDR P. aeruginosa clinical isolate, CL232. Whole genome and transcriptome sequencing, checkerboard analysis, and determination of mutation frequency as well as mutation prevention concentration were conducted. In addition, the ceftazidime–avibactam–fosfomycin combination was tested in a neutropenic thigh murine infection model with a high bacterial burden (2 × 107 colony forming units (CFUs)) of MDR P. aeruginosa clinical isolate CL232. Checkerboard analysis revealed slight synergy with fractional inhibitory concentration index of 0.53 for 25–6.25 μg/mL of ceftazidime–avibactam combined with 12.5 μg/mL of fosfomycin. Accordingly, the resistance elements in P. aeruginosa CL232 were analyzed via whole-genome sequencing (WGS) and transcriptome sequencing (RNAseq). WGS of CL232 revealed mutations in genes (e.g., oprD, ampR) that contribute to β-lactam resistance. Moreover, expression of the AmpC β-lactamase and the MexAB-OprM efflux pump were upregulated (~2–6-fold). The potential for the development of ceftazidime–avibactam-fosfomycin resistance was assessed in vitro. Fosfomycin alone was found to have a high mutation frequency 1.9 × 10−5; however, the addition of ceftazidime–avibactam reduced this frequency by 3-logs. In addition, the ceftazidime–avibactam–fosfomycin combination possessed the lowest mutation prevention concentration at 64 mg/L–4 mg/L–64 mg/L. In a neutropenic thigh murine infection model, the ceftazidime–avibactam–fosfomycin combination was found to reduce CFUs by 5–6 logs compared with vehicle-treated mice, while ceftazidime–avibactam and fosfomycin dosed separately decreased CFUs by ~1 log and 2–3 logs, respectively. The combination of ceftazidime–avibactam–fosfomycin is highly likely to offer patients who suffer from infections with a high bacteria burdens (i.e., pneumonia) a therapeutic hope against MDR P. aeruginosa. K. M. Papp-Wallace, F. Hoffmann-La Roche Ltd: Grant Investigator, Research grant. E. J. Ellis-Grosse, Zavante Therapeutics, Inc.,: Employee and Shareholder, Salary.
The understanding of marine microbial ecology and metabolism has been hampered by the paucity of sequenced reference genomes. To this end, we report the sequencing of 137 diverse marine isolates collected from around the world. We analysed these sequences, along with previously published marine prokaryotic genomes, in the context of marine metagenomic data, to gain insights into the ecology of the surface ocean prokaryotic picoplankton (0.1-3.0 μm size range). The results suggest that the sequenced genomes define two microbial groups: one composed of only a few taxa that are nearly always abundant in picoplanktonic communities, and the other consisting of many microbial taxa that are rarely abundant. The genomic content of the second group suggests that these microbes are capable of slow growth and survival in energy-limited environments, and rapid growth in energy-rich environments. By contrast, the abundant and cosmopolitan picoplanktonic prokaryotes for which there is genomic representation have smaller genomes, are probably capable of only slow growth and seem to be relatively unable to sense or rapidly acclimate to energy-rich conditions. Their genomic features also lead us to propose that one method used to avoid predation by viruses and/or bacterivores is by means of slow growth and the maintenance of low biomass.
Developing and evaluating "connectionist models" in artificial intelligence, cognitive science, and neurophysiological model ing is currently a difficult and time-consuming task. To address this issue, as part of a larger research effort, we implemented a software system called MIRRORS/II for developing connec tionist models. MIRRORS/II is distinguished by its support of a very high-level non-procedural language along with user and system libraries for the accumulation of system resources. We describe these features in detail, give a small example of the use of MIRRORS/II, and outline details of the implementation. Fi nally, we discuss current work using MIRRORS/II, future plans for the system and other related connectionist modeling systems.
News from the Inner Tube of Life A major initiative by the U.S. National Institutes of Health to sequence 900 genomes of microorganisms that live on the surfaces and orifices of the human body has established standardized protocols and methods for such large-scale reference sequencing. By combining previously accumulated data with new data, Nelson et al. (p. 994 ) present an initial analysis of 178 bacterial genomes. The sampling so far barely scratches the surface of the microbial diversity found on humans, but the work provides an important baseline for future analyses.
The influence of bacterial sequence type (ST) in Escherichia coli bloodstream infections (BSI) is incompletely understood. Adult inpatients with E. coli BSI at Duke University were prospectively enrolled from 2002–2015. Bacterial genotypes were determined by whole genome sequencing and multilocus sequence typing. Clinical outcomes associated with STs were evaluated using multivariate regression models adjusted for patient demographics, comorbidities, antibiotic treatment, BSI source, and BSI route (e.g., hospital-acquired). PanOCT was used to define flexible genomic islands (fGI) associated with STs. 193 patients with E. coli BSI were enrolled. The most common STs were ST131 (68/193 [35%]), ST95 (19/193 [10%]), and ST69 (14/193 [7%]) (Figure 1). E. coli BSI with ST393, relative to non-ST393, was associated with increased in-hospital mortality (5/10 [50%] vs.. 35/183 [19%]; P = 0.03). ST393 was not associated with an outbreak, and patient characteristics were similar in those with ST393 vs. non-ST393 BSI. Adjusted analysis similarly revealed an association between ST393 BSI and in-hospital mortality (Odds ratio 4.99; 95% Confidence interval 1.00–24.84; P = 0.05). Genomic analysis revealed 14 fGI associated with ST393 vs. non-ST393 genomes, ranging from 5–35 putative genes. The largest fGI (present in 10/10 [100%] ST393 vs. 43/183 [23%] non-ST393) included 15 genes (15/35 [43%]) with homology to a type III secretion system (T3SS). Additional T3SS components, including genes homologous to a chaperone and two effector genes, were present on a second fGI (10/10 [100%] ST393 vs. 35/183 [19%] non-ST393). Another fGI (10/10 [100%] ST393 vs. 43/183 [23%] non-ST393) contained 8 genes encoding a type 1 CRISPR-Cas system. E. coli ST393 BSI was uncommon but associated with increased mortality. There were substantial genetic differences in ST393 that could account for increased mortality. V. Fowler Jr., Karius, Inc: Grant Investigator, Grant recipient; Pfizer, Novartis, Galderma, Novadigm, Durata, Debiopharm, Genentech, Achaogen, Affinium, Medicines Co., Cerexa, Tetraphase, Trius, MedImmune, Bayer, Theravance, Cubist, Basilea, Affinergy, Janssen, xBiotech, Contrafect: Consultant, Consulting fee; NIH, MedImmune, Cerexa/Forest/Actavis/Allergan, Pfizer, Advanced Liquid Logics, Theravance, Novartis, Cubist/Merck; Medical Biosurfaces; Locus; Affinergy; Basilea; Contrafect; Karius: Grant Investigator, Grant recipient; Green Cross, Cubist, Cerexa, Durata, Theravance; Debiopharm: Consultant, Consulting fee; UpToDate: Royalties, Royalties
Since its introduction a decade ago, whole-genome shotgun sequencing (WGS) has been the main approach for producing cost-effective and high-quality genome sequence data. Until now, the Sanger sequencing technology that has served as a platform for WGS has not been truly challenged by emerging technologies. The recent introduction of the pyrosequencing-based 454 sequencing platform (454 Life Sciences, Branford, CT) offers a very promising sequencing technology alternative for incorporation in WGS. In this study, we evaluated the utility and cost-effectiveness of a hybrid sequencing approach using 3730xl Sanger data and 454 data to generate higher-quality lower-cost assemblies of microbial genomes compared to current Sanger sequencing strategies alone.
Background: Synthetic engineering of bacteria to produce industrial products is a burgeoning field of research and application. In order to optimize genome design, designers need to understand which genes are essential, which are optimal for growth, and locations in the genome that will be tolerated by the organism when inserting engineered cassettes.Methods: We present a pan-genome based method for the identification of core regions in a genome that are strongly conserved at the species level.Results: We show that the core regions determined by our method contain all or almost all essential genes. This demonstrates the accuracy of our method as essential genes should be core genes. We show that we outperform previous methods by this measure. We also explain why there are exceptions to this rule for our method.Conclusions: We assert that synthetic engineers should avoid deleting or inserting into these core regions unless they understand and are manipulating the function of the genes in that region. Similarly, if the designer wishes to streamline the genome, non-core regions and in particular low penetrance genes would be good targets for deletion. Care should be taken to remove entire cassettes with similar penetrance of the genes within cassettes as they may harbor toxin/antitoxin genes which need to be removed in tandem. The bioinformatic approach introduced here saves considerable time and effort relative to knockout studies on single isolates of a given species and captures a broad understanding of the conservation of genes that are core to a species.
