scholarly journals Pseudomonas aeruginosa core metabolism exerts a widespread growth-independent control on virulence

2019 ◽  
Author(s):  
Stavria Panayidou ◽  
Kaliopi Georgiades ◽  
Theodoulakis Christofi ◽  
Stella Tamana ◽  
Vasilis Promponas ◽  
...  
Keyword(s):  
Mouse Lung ◽  
Infection Model ◽  
Beta Oxidation ◽  
Metabolic Gene ◽  
The Core ◽  
Gene Level ◽  
Acid Succinate ◽  
Metabolism Analysis ◽  
Core Metabolism ◽  
Pathway Level

AbstractBacterial virulence may rely on secondary metabolism, but core metabolism genes are assumed to be necessary primarily for bacterial growth. To assess this assumption, we correlated the genome, the transcriptome and the pathogenicity of 30 fully sequenced Pseudomonas strains using two Drosophila and one mouse infection assay. In accordance with previous studies gene presence-absence does not explain differences in virulence among P. aeruginosa strains, but merely between P. aeruginosa and other Pseudomonas species. Similarly, classical gene expression analysis of highly vs. lowly pathogenic P. aeruginosa strains identifies many virulence factors, and only a few metabolism genes related to virulence. Nevertheless, assessing the virulence of 553 core metabolic and 95 random non-metabolic gene mutants of P. aeruginosa PA14, we found 16.5% of the core metabolic and 8.5% of the non-metabolic genes to be necessary for full virulence. Strikingly, 11.8% of the core metabolism genes exhibit defects in virulence that cannot be attributed to auxotrophy. The compromised in virulence metabolic gene mutants were mapped in multiple pathways and exhibited further defects in acute virulence phenotypes and in a mouse lung infection model. Functional transcriptomics re-analysis of core metabolism at the pathway level, reveals amino-acid, succinate, citramalate, and chorismate biosynthesis and beta-oxidation as important for full virulence and expression of these pathways indicative of virulence in various strains. Thus, P. aeruginosa virulence variation, which to this point remains unpredictably combinatorial at the gene level, can be dissected at the pathway level via combinatorial trancriptome and functional core metabolism analysis.

scholarly journals Core Metabolism Shifts during Growth on Methanol versus Methane in the Methanotroph Methylomicrobium buryatense 5GB1

mBio ◽  
2019 ◽  
Vol 10 (2) ◽  
Author(s):  
Yanfen Fu ◽  
Lian He ◽  
Jennifer Reeve ◽  
David A. C. Beck ◽  
Mary E. Lidstrom
Keyword(s):  
Flux Balance Analysis ◽  
Metabolic Pathways ◽  
Scale Model ◽  
Obligate Methylotroph ◽  
Content Type ◽  
Advantages And Disadvantages ◽  
The Core ◽  
Flux Balance ◽  
Balance Analysis ◽  
Core Metabolism

ABSTRACT Methylomicrobium buryatense 5GB1 is an obligate methylotroph which grows on methane or methanol with similar growth rates. It has long been assumed that the core metabolic pathways must be similar on the two substrates, but recent studies of methane metabolism in this bacterium suggest that growth on methanol might have significant differences from growth on methane. In this study, both a targeted metabolomics approach and a 13C tracer approach were taken to understand core carbon metabolism in M. buryatense 5GB1 during growth on methanol and to determine whether such differences occur. Our results suggest a systematic shift of active core metabolism in which increased flux occurred through both the Entner-Doudoroff (ED) pathway and the partial serine cycle, while the tricarboxylic acid (TCA) cycle was incomplete, in contrast to growth on methane. Using the experimental results as constraints, we applied flux balance analysis to determine the metabolic flux phenotype of M. buryatense 5GB1 growing on methanol, and the results are consistent with predictions based on ATP and NADH changes. Transcriptomics analysis suggested that the changes in fluxes and metabolite levels represented results of posttranscriptional regulation. The combination of flux balance analysis of the genome-scale model and the flux ratio from 13C data changed the solution space for a better prediction of cell behavior and demonstrated the significant differences in physiology between growth on methane and growth on methanol. IMPORTANCE One-carbon compounds such as methane and methanol are of increasing interest as sustainable substrates for biological production of fuels and industrial chemicals. The bacteria that carry out these conversions have been studied for many decades, but gaps exist in our knowledge of their metabolic pathways. One such gap is the difference between growth on methane and growth on methanol. Understanding such metabolism is important, since each has advantages and disadvantages as a feedstock for production of chemicals and fuels. The significance of our research is in the demonstration that the metabolic network is substantially altered in each case and in the delineation of these changes. The resulting new insights into the core metabolism of this bacterium now provide an improved basis for future strain design.

scholarly journals In Vivo Efficacies and Pharmacokinetics of DX-619, a Novel Des-Fluoro(6) Quinolone, against Streptococcus pneumoniae in a Mouse Lung Infection Model

2006 ◽  
Vol 50 (3) ◽  
pp. 1122-1122
Author(s):  
Yuichi Fukuda ◽  
Katsunori Yanagihara ◽  
Hideaki Ohno ◽  
Yasuhito Higashiyama ◽  
Yoshitsugu Miyazaki ◽  
...  
Keyword(s):  
Streptococcus Pneumoniae ◽  
Lung Infection ◽  
Mouse Lung ◽  
Infection Model

scholarly journals Pyrrolamide DNA Gyrase Inhibitors: Fragment-Based Nuclear Magnetic Resonance Screening To Identify Antibacterial Agents

2011 ◽  
Vol 56 (3) ◽  
pp. 1240-1246 ◽  
Author(s):  
Ann E. Eakin ◽  
Oluyinka Green ◽  
Neil Hales ◽  
Grant K. Walkup ◽  
Shanta Bist ◽  
...  
Keyword(s):  
Nuclear Magnetic Resonance ◽  
Magnetic Resonance ◽  
Antibacterial Agents ◽  
Dna Gyrase ◽  
Mouse Lung ◽  
Infection Model ◽  
Lead Compound ◽  
Content Type ◽  
Gyrase Inhibitors ◽  
Nuclear Magnetic

ABSTRACTDNA gyrase is an essential enzyme in bacteria, and its inhibition results in the disruption of DNA synthesis and, subsequently, cell death. The pyrrolamides are a novel class of antibacterial agents targeting DNA gyrase. These compounds were identified by a fragment-based lead generation (FBLG) approach using nuclear magnetic resonance (NMR) screening to identify low-molecular-weight compounds that bind to the ATP pocket of DNA gyrase. A pyrrole hit with a binding constant of 1 mM formed the basis of the design and synthesis of a focused library of compounds that resulted in the rapid identification of a lead compound that inhibited DNA gyrase with a 50% inhibitory concentration (IC50) of 3 μM. The potency of the lead compound was further optimized by utilizing iterative X-ray crystallography to yield DNA gyrase inhibitors that also displayed antibacterial activity. Spontaneous mutants were isolated inStaphylococcus aureusby plating on agar plates containing pyrrolamide 4 at the MIC. The resistant variants displayed 4- to 8-fold-increased MIC values relative to the parent strain. DNA sequencing revealed two independent point mutations in the pyrrolamide binding region of thegyrBgenes from these variants, supporting the hypothesis that the mode of action of these compounds was inhibition of DNA gyrase. Efficacy of a representative pyrrolamide was demonstrated againstStreptococcus pneumoniaein a mouse lung infection model. These data demonstrate that the pyrrolamides are a novel class of DNA gyrase inhibitors with the potential to deliver future antibacterial agents targeting multiple clinical indications.

scholarly journals A Novel Zinc Binding System, ZevAB, Is Critical for Survival of Nontypeable Haemophilus influenzae in a Murine Lung Infection Model

2011 ◽  
Vol 79 (8) ◽  
pp. 3366-3376 ◽  
Author(s):  
Charles V. Rosadini ◽  
Jeffrey D. Gawronski ◽  
Daniel Raimunda ◽  
José M. Argüello ◽  
Brian J. Akerley
Keyword(s):  
Haemophilus Influenzae ◽  
Respiratory Infections ◽  
Bacterial Pathogen ◽  
Lung Infection ◽  
Lung Model ◽  
Mouse Lung ◽  
Infection Model ◽  
Nontypeable Haemophilus Influenzae ◽  
Content Type ◽  
Lung Colonization

ABSTRACTNontypeableHaemophilus influenzae(NTHI) is a Gram-negative bacterial pathogen that causes upper and lower respiratory infections. Factors required for pulmonary infection by NTHI are not well understood. Previously, using high-throughput insertion tracking by deep sequencing (HITS), putative lung colonization factors were identified. Also, previous research indicates that secreted disulfide-dependent factors are important for virulence ofH. influenzae. In the present study, HITS data were compared with an informatics-based list of putative substrates of the periplasmic oxidoreductase DsbA to find and characterize secreted virulence factors. This analysis resulted in identification of the “zinc bindingessential forvirulence” (zev) locus consisting ofzevA(HI1249) andzevB(HI1248). NTHI mutants ofzevAandzevBgrew normally in rich medium but were defective for colonization in a mouse lung model. Mutants also exhibited severe growth defects in medium containing EDTA and were rescued by supplementation with zinc. Additionally, purified recombinant ZevA was found to bind to zinc with high affinity. Together, these data demonstrate thatzevABis a novel virulence factor important for zinc utilization ofH. influenzaeunder conditions where zinc is limiting. Furthermore, evidence presented here suggests that zinc limitation is likely an important mechanism for host defense against pathogens during lung infection.

scholarly journals Mechanism-Based Pharmacokinetic/Pharmacodynamic Modeling of Aerosolized Colistin in a Mouse Lung Infection Model

2017 ◽  
Vol 62 (3) ◽  
Author(s):  
Yu-Wei Lin ◽  
Qi Tony Zhou ◽  
Mei-Ling Han ◽  
Nikolas J. Onufrak ◽  
Ke Chen ◽  
...  
Keyword(s):  
Critically Ill ◽  
Critically Ill Patients ◽  
Pharmacodynamic Modeling ◽  
Mouse Lung ◽  
Infection Model ◽  
Bacterial Reduction ◽  
Bacterial Killing ◽  
Deterministic Simulation ◽  
Dosage Regimens

ABSTRACTOptimized dosage regimens of aerosolized colistin (as colistin methanesulfonate [CMS]) are urgently required to maximize bacterial killing against multidrug-resistant Gram-negative bacteria while minimizing toxicity. This study aimed to develop a mechanism-based pharmacokinetic (PK)/pharmacodynamic (PD) model (MBM) for aerosolized colistin based upon PK/PD data in neutropenic infected mice and to perform a deterministic simulation with the PK of aerosolized colistin (as CMS) in critically ill patients.In vivotime-kill experiments were carried out with three different strains ofPseudomonas aeruginosa. An MBM was developed in S-ADAPT and evaluated by assessing its ability to predict the PK/PD index associated with efficacy in mice. A deterministic simulation with human PK data was undertaken to predict the efficacy of current dosage regimens of aerosolized colistin in critically ill patients. In the final MBM, the total bacterial population for each isolate consisted of colistin-susceptible and -resistant subpopulations. The antimicrobial efficacy of aerosolized colistin was best described by a sigmoidalEmaxmodel whereby colistin enhanced the rate of bacterial death. Deterministic simulation with human PK data predicted that an inhalational dosage regimen of 60 mg colistin base activity (CBA) every 12 h is needed to achieve a ≥2-log10bacterial reduction (as the number of CFU per lung) in critically ill patients at 24 h after commencement of inhaled therapy. In conclusion, the developed MBM is a useful tool for optimizing inhalational dosage regimens of colistin. Clinical studies are warranted to validate and refine our MBM for aerosolized colistin.

Mouse lung infection model to assess Rhodococcus equi virulence and vaccine protection

2014 ◽  
Vol 172 (1-2) ◽  
pp. 256-264 ◽  
Author(s):  
Patricia González-Iglesias ◽  
Mariela Scortti ◽  
Iain MacArthur ◽  
Alexia Hapeshi ◽  
Héctor Rodriguez ◽  
...  
Keyword(s):  
Lung Infection ◽  
Rhodococcus Equi ◽  
Mouse Lung ◽  
Infection Model ◽  
Vaccine Protection

Association of breast cancer risk in women of African ancestry with genetic variants in the TET-related DNA demethylation pathway.

2017 ◽  
Vol 35 (15_suppl) ◽  
pp. e13015-e13015
Author(s):  
Kris M. Lee ◽  
Shengfeng Wang ◽  
Dezheng Huo ◽  
Temidayo O. Ogundiran ◽  
Oladosu Ojengbede ◽  
...  
Keyword(s):  
Breast Cancer ◽  
Breast Cancer Risk ◽  
Cancer Risk ◽  
Genetic Variants ◽  
African Ancestry ◽  
Dna Demethylation ◽  
Increased Risk ◽  
Gene Level ◽  
Demethylation Pathway ◽  
Pathway Level

e13015 Background: Imbalance in DNA methylation/demethylation cycles due to mutations in TET1, TET2, TET3, and TDG has been implicated in the onset and progression of breast cancer. However, few studies have assessed the relationship between genetic variants in the TET-related DNA demethylation pathway and breast cancer risk, particularly among women of African ancestry. Methods: We investigated 4095 single-nucleotide polymorphisms (SNPs) in four TET-related DNA demethylation pathway genes in the ROOT consortium, which includes women of African ancestry, in 1657 cases [403 estrogen receptor-positive (ER+) and 374 ER-negative (ER-)] and 2029 controls. Pathway and gene-level analyses were conducted using the adaptive rank truncated product (ARTP) test for 925 SNPs that were not highly correlated (r2 < 0.8), and SNP-level analyses were conducted with logistic regression, which was used to estimate the odds ratio (OR) and 95 % confidence intervals (CI). Results: The Tet-related DNA demethylation pathway was significantly associated with ER-negative breast cancer (pathway level P = 0.049). Gene-level analyses showed that TET1 was the candidate gene responsible for the association at the pathway level ( P = 0.008). SNP TET1 rs10998376 (OR = 1.21, 95% CI = 1.02-1.42, Padj = 0.0032) was statistically significant even after gene-level correction for multiple comparisons, and was associated with increased risk. There were no SNPs in genes with P > 0.05 in the ARTP tests that were significant after Boferroni correction. Conclusions: In conclusion, specific TET-related DNA demethylation pathway genes may contribute to breast cancer risk, in particular, to risk of ER-negative tumors in women of African ancestry.

scholarly journals The Novel β-Lactam Enhancer Zidebactam Augments theIn VivoPharmacodynamic Activity of Cefepime in a Neutropenic Mouse LungAcinetobacter baumanniiInfection Model

2019 ◽  
Vol 63 (4) ◽  
Author(s):  
S. S. Bhagwat ◽  
H. Periasamy ◽  
S. S. Takalkar ◽  
S. R. Palwe ◽  
H. N. Khande ◽  
...  
Keyword(s):  
Bactericidal Effect ◽  
Multidrug Resistant ◽  
Mouse Lung ◽  
Infection Model ◽  
Bactericidal Action ◽  
Content Type ◽  
Time Kill ◽  
The Impact

ABSTRACTWCK 5222 is a combination of cefepime and the high-affinity PBP2-binding β-lactam enhancer zidebactam. The cefepime-zidebactam combination is active against multidrug-resistant Gram-negative bacteria, including carbapenemase-expressingAcinetobacter baumannii. The mechanism of action of the combination involves concurrent multiple penicillin binding protein inhibition, leading to the enhanced bactericidal action of cefepime. The aim of the present study was to assess the impact of the zidebactam-mediated enhancedin vitrobactericidal action in modulating the percentage of the time that the free drug concentration remains above the MIC (percentfT>MIC) for cefepime required for thein vivokilling ofA. baumannii. Cefepime and cefepime-zidebactam MICs were comparable and ranged from 2 to 16 mg/liter for theA. baumanniistrains (n = 5) employed in the study. Time-kill studies revealed the improved killing of these strains by the cefepime-zidebactam combination compared to that by the constituents alone. Employing a neutropenic mouse lung infection model, exposure-response analyses for all theA. baumanniistrains showed that the cefepimefT>MIC required for 1-log10kill was 38.9%. In the presence of a noneffective dose of zidebactam, the cefepimefT>MIC requirement dropped significantly to 15.5%, but it still rendered a 1-log10kill effect. Thus, zidebactam mediated the improvement in cefepime’s bactericidal effect observed in time-kill studies, manifestedin vivothrough the lowering of cefepime’s pharmacodynamic requirement. This is a first-ever study demonstrating a β-lactam enhancer role of zidebactam that helps augment thein vivoactivity of cefepime by reducing the magnitude of its pharmacodynamically relevant exposures againstA. baumannii.

scholarly journals In Vivo Efficacies and Pharmacokinetics of DX-619, a Novel Des-Fluoro(6) Quinolone, against Streptococcus pneumoniae in a Mouse Lung Infection Model

2006 ◽  
Vol 50 (1) ◽  
pp. 121-125 ◽  
Author(s):  
Yuichi Fukuda ◽  
Katsunori Yanagihara ◽  
Hideaki Ohno ◽  
Yasuhito Higashiyama ◽  
Yoshitsugu Miyazaki ◽  
...  
Keyword(s):  
Streptococcus Pneumoniae ◽  
Mouse Lung ◽  
Infection Model ◽  
Time Curve ◽  
Concentration Time ◽  
Viable Bacteria ◽  
Effective Doses ◽  
The Mean ◽  
Lung Infections

ABSTRACT DX-619 is a novel des-fluoro(6) quinolone with potent activity against gram-positive pathogens. The in vivo activity of DX-619 against Streptococcus pneumoniae was compared with those of fluoro(6) quinolones, sitafloxacin, and ciprofloxacin in a mouse model. Two strains of S. pneumoniae were used: a penicillin-sensitive S. pneumoniae (PSSP) strain and a penicillin-resistant S. pneumoniae (PRSP) strain. Furthermore, these strains showed intermediate susceptibilities to ciprofloxacin. In murine lung infections caused by PSSP, the 50% effective doses (ED50s) of DX-619, sitafloxacin, and ciprofloxacin were 9.15, 11.1, and 127.6 mg/kg of body weight, respectively. Against PRSP-mediated pneumonia in mice, the ED50s of DX-619, sitafloxacin, and ciprofloxacin were 0.69, 4.84, and 38.75 mg/kg, respectively. The mean ± standard error of the mean viable bacterial counts in murine lungs infected with PSSP and treated with DX-619, sitafloxacin, ciprofloxacin (10 mg/kg twice daily), and saline (twice daily) were 1.75 ± 0.06, 1.92 ± 0.23, 6.48 ± 0.28, and 7.57 ± 0.13 log10 CFU/ml, respectively. Furthermore, the numbers of viable bacteria in lungs infected with PRSP and treated with the three agents and not treated (control) were 1.73 ± 0.04, 2.28 ± 0.17, 4.61 ± 0.59, and 5.54 ± 0.72 log10 CFU/ml, respectively. DX-619 and sitafloxacin significantly decreased the numbers of viable bacteria in the lungs compared to the numbers in the lungs of ciprofloxacin-treated and untreated mice. The pharmacokinetic parameter of the area under the concentration-time curve (AUC)/MIC ratio in the lungs for DX-619, sitafloxacin, and ciprofloxacin were 171.0, 21.92, and 1.22, respectively. The AUC/MIC ratio in the lungs was significantly higher for DX-619 than for sitafloxacin and ciprofloxacin. Our results suggest that DX-619 and sitafloxacin are potent against both PSSP and PRSP in our mouse pneumonia model.

Characterization of the tumor immune microenvironment of triple-negative breast cancer (TNBC) patients who self-identify as African American (AA) or non-African American (NonAA).

2021 ◽  
Vol 39 (15_suppl) ◽  
pp. 564-564
Author(s):  
Kim Blenman ◽  
Michal Marczyk ◽  
Tao Qing ◽  
Tess O'Meara ◽  
Vesal Yaghoobi ◽  
...  
Keyword(s):  
Breast Cancer ◽  
Gene Expression ◽  
African American ◽  
Triple Negative Breast Cancer ◽  
Somatic Mutations ◽  
Triple Negative ◽  
Immune Microenvironment ◽  
Gene Level ◽  
Tumor Immune Microenvironment ◽  
Pathway Level

564 Background: What tumor biological differences, if any, contribute to the higher incidence and worse prognosis of triple negative breast cancer (TNBC) in African American (AA) compared to NonAA patients are unknown. We hypothesized that differences in the tumor immune microenvironment may contribute to the outcome disparities. The purpose of this study was to characterize and compare the immune microenvironment of TNBC between patients self-identified as NonAA or AA. Methods: Formalin fixed paraffin embedded surgically resected cancer and paired normal tissues collected before any systemic therapy and the corresponding clinical data were collected for NonAA (n = 56) and AA (n = 54) stage I-III TNBC treated at Yale Cancer Center between 2000-2017. The two cohorts were matched for clinical stage, age of diagnosis, and year of diagnosis. We performed somatic and germline whole exome sequencing (WES), bulk RNA sequencing, and immunohistochemistry to assess PD-L1 expression (SP142). Stromal tumor infiltrating lymphocytes (sTILs) were assessed on H&E slides. Mutation load, mutation frequencies, and gene expression differences were compared at gene and pathway level. Immune cell composition was estimated through gene expression deconvolution analyses (TIDE). Results: Tumor mutational burden was similar between the two cohorts. At gene level, few genes had significantly different somatic mutation frequencies, or differential mRNA expression between AA and NonAA samples. Pathway level alterations showed inflammation, immunity (adaptive; innate), antigen presentation, and allograft rejection pathways were more affected by somatic mutations in AA samples. The affected genes differed from cancer to cancer and were not recurrent and therefore were missed at gene level analysis. Gene set enrichment and co-expression analysis also showed higher immune related pathway expression in AA samples. Unsupervised co-expression cluster analysis confirmed coordinated overexpression of genes involved in immunity, inflammation, and cytokine/chemokine signaling in AA patients. Two immunotherapy response predictive signatures, immune inflamed and the IFNG as well as sTILs score and PD-L1 positivity were also higher in AA samples. These findings raise the possibility that immune checkpoint inhibitors might be particularly effective in AA patients. In NonAA samples, the EMT transition, angiogenesis, adipogenesis, myogenesis, fatty acid metabolism, TGFβ signaling, UV-response, and hypoxia pathways were overexpressed. TIDE analysis suggested higher levels of TAM M2, overall TIDE score, and the Immune Exclusion score in NonAA samples. Conclusions: TNBC in AA patients more frequently harbor somatic mutations in genes involved with immune functions and overexpress immune and inflammatory genes compared to NonAA patients.

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