scholarly journals Identification and Characterization of Chemical Compounds that Inhibit Leucyl-tRNA Synthetase from Pseudomonas aeruginosa

2020 ◽  
Vol 17 (1) ◽  
pp. 119-130 ◽  
Author(s):  
Regina Zamacona ◽  
Pamela N. Chavero ◽  
Eduardo Medellin ◽  
Yanmei Hu ◽  
Casey A. Hughes ◽  
...  
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Human Cell ◽  
Bacterial Infections ◽  
Chemical Compounds ◽  
Human Cell Line ◽  
Trna Synthetase ◽  
Multi Drug Resistance ◽  
Bacterial Strains ◽  
Gene Encoding ◽  
Screening Platform

Background: Pseudomonas aeruginosa is an opportunistic multi-drug resistance pathogen implicated as the causative agent in a high-percentage of nosocomial and community acquired bacterial infections. The gene encoding leucyl-tRNA synthetase (LeuRS) from P. aeruginosa was overexpressed in Escherichia coli and the resulting protein was characterized. Methods: LeuRS was kinetically evaluated and the KM values for interactions with leucine, ATP and tRNA were 6.5, 330, and 3.0 μM, respectively. LeuRS was developed into a screening platform using scintillation proximity assay (SPA) technology and used to screen over 2000 synthetic and natural chemical compounds. Results: The initial screen resulted in the identification of two inhibitory compounds, BT03C09 and BT03E07. IC50s against LeuRS observed for BT03C09 and BT03E07 were 23 and 15 μM, respectively. The minimum inhibitory concentrations (MIC) were determined against nine clinically relevant bacterial strains. In time-kill kinetic analysis, BT03C09 was observed to inhibit bacterial growth in a bacteriostatic manner, while BT03E07 acted as a bactericidal agent. Neither compound competed with leucine or ATP for binding LeuRS. Limited inhibition was observed in aminoacylation assays with the human mitochondrial form of LeuRS, however when tested in cultures of human cell line, BT03C09 was toxic at all concentration whereas BT03E07 only showed toxic effects at elevated concentrations. Conclusion: Two compounds were identified as inhibitors of LeuRS in a screen of over 2000 natural and synthetic compounds. After characterization one compound (BT03E07) exhibited broad spectrum antibacterial activity while maintaining low toxicity against human mitochondrial LeuRS as well as against human cell cultures.

scholarly journals Discovery and Characterization of Chemical Compounds That Inhibit the Function of Aspartyl-tRNA Synthetase from Pseudomonas aeruginosa

2017 ◽  
Vol 23 (3) ◽  
pp. 294-301 ◽  
Author(s):  
Araceli Corona ◽  
Stephanie O. Palmer ◽  
Regina Zamacona ◽  
Benjamin Mendez ◽  
Frank B. Dean ◽  
...  
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Aspartic Acid ◽  
Cell Cultures ◽  
Human Cell ◽  
Efflux Pump ◽  
Chemical Compounds ◽  
Opportunistic Pathogen ◽  
Trna Synthetase ◽  
Bacterial Strains ◽  
Human Cell Cultures

Pseudomonas aeruginosa, an opportunistic pathogen, is highly susceptible to developing resistance to multiple antibiotics. The gene encoding aspartyl-tRNA synthetase (AspRS) from P. aeruginosa was cloned and the resulting protein characterized. AspRS was kinetically evaluated, and the KM values for aspartic acid, ATP, and tRNA were 170, 495, and 0.5 μM, respectively. AspRS was developed into a screening platform using scintillation proximity assay (SPA) technology and used to screen 1690 chemical compounds, resulting in the identification of two inhibitory compounds, BT02A02 and BT02C05. The minimum inhibitory concentrations (MICs) were determined against nine clinically relevant bacterial strains, including efflux pump mutant and hypersensitive strains of P. aeruginosa. The compounds displayed broad-spectrum antibacterial activity and inhibited growth of the efflux and hypersensitive strains with MICs of 16 μg/mL. Growth of wild-type strains were unaffected, indicating that efflux was likely responsible for this lack of activity. BT02A02 did not inhibit growth of human cell cultures at any concentration. However, BT02C05 did inhibit human cell cultures with a cytotoxicity concentration (CC50) of 61.6 μg/mL. The compounds did not compete with either aspartic acid or ATP for binding AspRS, indicating that the mechanism of action of the compound occurs outside the active site of aminoacylation.

scholarly journals Novel Lytic Phages Protect Cells and Mice against Pseudomonas aeruginosa Infection

2021 ◽  
Author(s):  
Feng Chen ◽  
Xingjun Cheng ◽  
Jianbo Li ◽  
Xiefang Yuan ◽  
Xiuhua Huang ◽  
...  
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Mouse Models ◽  
Bacterial Infections ◽  
Phage Therapy ◽  
Inflammatory Responses ◽  
Multi Drug Resistance ◽  
Bacterial Strains ◽  
Genetic Traits

With the fast emergence of serious antibiotic resistance and the lagged discovery of novel antibacterial drugs, phage therapy for pathogenic bacterial infections has acquired great attention in the clinics. However, development of therapeutic phages also faces tough challenges, such as laborious screening and time to generate effective phage drugs since each phage may only lyse a narrow scope of bacterial strains. Identifying highly effective phages with broad host ranges is crucial for improving phage therapy. Here, we isolated and characterized several lytic phages from various environments specific for Pseudomonas aeruginosa by testing their growth, invasion, host ranges, and potential for killing targeted bacteria. Importantly, we identified several therapeutic phages (HX1, PPY9, and TH15) with broad host ranges to lyse laboratory strains and clinical isolates of P. aeruginosa with multi-drug resistance (MDR) both in vitro and in mouse models. In addition, we analyzed critical genetic traits related to the high-level broad host coverages by genome sequencing and subsequent computational analysis against known phages. Collectively, our findings establish that these novel phages may have potential for further development as therapeutic options for patients who fail to respond to conventional treatments. IMPORTANCE Novel lytic phages isolated from various environmental settings were systematically characterized for their critical genetic traits, morphology structures, host ranges against laboratory strains and clinical multi-drug resistant (MDR) Pseudomonas aeruginosa, and antibacterial capacity both in vitro and in mouse models. First, we characterized the genetic traits and compared with other existing phages. Furthermore, we utilized acute pneumonia induced by laboratorial strain PAO1, and W19, an MDR clinical isolate and chronic pneumonia by agar beads laden with FDR1, a mucoid phenotype strain isolated from the sputum of a cystic fibrosis (CF) patient. Consequently, we found that these phages not only suppress bacteria in vitro but also significantly reduce the infection symptom and disease progression in vivo, including lowered bug burdens, inflammatory responses and lung injury in mice, suggesting that they may be further developed as therapeutic agents against MDR P. aeruginosa.

scholarly journals Glutaminyl‐tRNA Synthetase from Pseudomonas aeruginosa : Characterization, structure, and development as a screening platform

2019 ◽  
Vol 29 (4) ◽  
pp. 905-918
Author(s):  
Yaritza Escamilla ◽  
Casey A. Hughes ◽  
Jan Abendroth ◽  
David M. Dranow ◽  
Samantha Balboa ◽  
...  
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Trna Synthetase ◽  
Screening Platform

scholarly journals Combined Antibacterial and Anti-Inflammatory Activity of a Cationic Disubstituted Dexamethasone-Spermine Conjugate

2010 ◽  
Vol 54 (6) ◽  
pp. 2525-2533 ◽  
Author(s):  
Robert Bucki ◽  
Katarzyna Leszczyńska ◽  
Fitzroy J. Byfield ◽  
David E. Fein ◽  
Esther Won ◽  
...  
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Bacterial Infections ◽  
Glucocorticoid Receptors ◽  
Fluorescent Protein ◽  
Lipoteichoic Acid ◽  
Bacterial Strains ◽  
Bacterial Killing ◽  
Antibiotic Resistant ◽  
Anti Inflammatory ◽  
Green Fluorescent

ABSTRACT The rising number of antibiotic-resistant bacterial strains represents an emerging health problem that has motivated efforts to develop new antibacterial agents. Endogenous cationic antibacterial peptides (CAPs) that are produced in tissues exposed to the external environment are one model for the design of novel antibacterial compounds. Here, we report evidence that disubstituted dexamethasone-spermine (D2S), a cationic corticosteroid derivative initially identified as a by-product of synthesis of dexamethasone-spermine (DS) for the purpose of improving cellular gene delivery, functions as an antibacterial peptide-mimicking molecule. This moiety exhibits bacterial killing activity against clinical isolates of Staphylococcus aureus, Pseudomonas aeruginosa present in cystic fibrosis (CF) sputa, and Pseudomonas aeruginosa biofilm. Although compromised in the presence of plasma, D2S antibacterial activity resists the proteolytic activity of pepsin and is maintained in ascites, cerebrospinal fluid, saliva, and bronchoalveolar lavage (BAL) fluid. D2S also enhances S. aureus susceptibility to antibiotics, such as amoxicillin (AMC), tetracycline (T), and amikacin (AN). Inhibition of interleukin-6 (IL-6) and IL-8 release from lipopolysaccharide (LPS)- or lipoteichoic acid (LTA)-treated neutrophils in the presence of D2S suggests that this molecule might also prevent systemic inflammation caused by bacterial wall products. D2S-mediated translocation of green fluorescent protein (GFP)-labeled glucocorticoid receptor (GR) in bovine aorta endothelial cells (BAECs) suggests that some of its anti-inflammatory activities involve engagement of glucocorticoid receptors. The combined antibacterial and anti-inflammatory activities of D2S suggest its potential as an alternative to natural CAPs in the prevention and treatment of some bacterial infections.

scholarly journals Characterization and structure determination of prolyl‐tRNA synthetase from Pseudomonas aeruginosa and development as a screening platform

2019 ◽  
Vol 28 (4) ◽  
pp. 727-737 ◽  
Author(s):  
Noah Pena ◽  
David M. Dranow ◽  
Yanmei Hu ◽  
Yaritza Escamilla ◽  
James M. Bullard
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Structure Determination ◽  
Trna Synthetase ◽  
Screening Platform

scholarly journals Optimization of a High-Throughput 384-Well Plate-Based Screening Platform with Staphylococcus aureus ATCC 25923 and Pseudomonas aeruginosa ATCC 15442 Biofilms

2020 ◽  
Vol 21 (9) ◽  
pp. 3034 ◽  
Author(s):  
Shella Gilbert-Girard ◽  
Kirsi Savijoki ◽  
Jari Yli-Kauhaluoma ◽  
Adyary Fallarero
Keyword(s):  
Staphylococcus Aureus ◽  
Pseudomonas Aeruginosa ◽  
High Throughput ◽  
High Throughput Screening ◽  
Bacterial Infections ◽  
Rapid Screening ◽  
Bacterial Biofilms ◽  
Main Concern ◽  
Assay Optimization ◽  
Screening Platform

In recent years, bacterial infections have become a main concern following the spread of antimicrobial resistance. In addition, bacterial biofilms are known for their high tolerance to antimicrobials and they are regarded as a main cause of recalcitrant infections in humans. Many efforts have been deployed in order to find new antibacterial therapeutic options and the high-throughput screening (HTS) of large libraries of compounds is one of the utilized strategies. However, HTS efforts for anti-biofilm discovery remain uncommon. Here, we miniaturized a 96-well plate (96WP) screening platform, into a 384-well plate (384WP) format, based on a sequential viability and biomass measurements for the assessment of anti-biofilm activity. During the assay optimization process, different parameters were evaluated while using Staphylococcus aureus and Pseudomonas aeruginosa as the bacterial models. We compared the performance of the optimized 384WP platform to our previously established 96WP-based platform by carrying out a pilot screening of 100 compounds, followed by the screening of a library of 2000 compounds to identify new repurposed anti-biofilm agents. Our results show that the optimized 384WP platform is well-suited for screening purposes, allowing for the rapid screening of a higher number of compounds in a run in a reliable manner.

scholarly journals Laboratory based antimicrobial resistance surveillance for Pseudomonas aeruginosa blood isolates from South Africa

2018 ◽  
Vol 12 (08) ◽  
pp. 616-624
Author(s):  
Ashika Singh-Moodley ◽  
Adriano Duse ◽  
Preneshni Naicker ◽  
Ranmini Kularatne ◽  
Trusha Nana ◽  
...  
Keyword(s):  
South Africa ◽  
Pseudomonas Aeruginosa ◽  
Antimicrobial Resistance ◽  
Antimicrobial Susceptibility ◽  
Bacterial Infections ◽  
Treatment Guidelines ◽  
Resistance Mechanisms ◽  
Multi Drug Resistance ◽  
Hospital Treatment ◽  
Baseline Information

Introduction: Antimicrobial resistant bacterial infections are widespread globally and increases in antimicrobial resistance presents a major threat to public health. Pseudomonas aeruginosa is an opportunistic healthcare-associated pathogen with high rates of morbidity and mortality and an extensive range of resistance mechanisms. This study describes the antibiotic susceptibility profiles of P. aeruginosa isolates from patients with bacteraemia submitted by sentinel laboratories in South Africa from 2014 to 2015. Methodology: Organism identification and antimicrobial susceptibility testing were done using automated systems. Molecular methods were used to detect common resistance genes and mechanisms. Results: Overall the susceptibility was high for all antibiotics tested with a decrease over the two-year period. There was no change in the MIC50 and MIC90 breakpoints for all antibiotics from 2014 to 2015. The MIC50 was within the susceptible breakpoint range for most antibiotics and the MIC90 was within the susceptible breakpoint range for colistin only. Phenotypically carbapenem non-susceptible isolates harboured the following plasmid-mediated genes: blaVIM (n = 81, 12%) and blaGES (n = 6, 0.9%); blaNDM (n = 4, 0.6%) and blaOXA-48 and variants (n = 3, 0.45%). Porin deletions were observed in one meropenem non-susceptible isolate only, and multi-drug resistance efflux pumps were expressed in the majority of the non-susceptible isolates investigated. BlaVEB-1, blaIMP and blaKPC were not detected. Conclusion: The prevalence of resistance to commonly used antibacterial agents was low for P. aeruginosa isolates and similarly, tested resistance mechanisms were detected in a relatively small proportion of isolates. Findings in this study represent baseline information for understanding antimicrobial susceptibility patterns in P. aeruginosa isolates from blood. Our surveillance report may assist in contributing to hospital treatment guidelines.

scholarly journals Bacterial contamination of table eggs in Babylon, Iraq

2014 ◽  
Vol 38 (1) ◽  
pp. 124-128
Author(s):  
Muna Sabbar Al-Rubiae
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Bacterial Contamination ◽  
Antibiotic Sensitivity ◽  
Multi Drug Resistance ◽  
Bacterial Strains ◽  
Salmonella Spp ◽  
Sensitivity Tests ◽  
Stool Samples ◽  
Different Sources ◽  
Growth Of Bacteria

To study the bacterial contamination of table eggs in Babylon city, a total of 214 eggs collected from different sources, including 100 from farms and 114 from supermarkets, all samples were cultured for the bacteria on Salmonella Shigella Agar(SS Agar) and nutrient agar. The results of the farms samples showed that there are no growth of bacteria in all samples under study whereas the results of supermarkets samples showed that about 21.05% of supermarkets eggs were contaminated with bacterial strains, and the results showed the presence of Stahylococcus aureus in 10.52% of the samples, Pseudomonas aeruginosa in 7.89%, a Proteus mirabilis in 3.50%, S. epidermidis in 0.87% and Bacillus subtilis in 0.87%. Also, the antibiotic sensitivity tests were tested for all isolates and the result showed that the sensitivity was 100% for ciprofloxacin, 85.18% for gentamicin, 85.18% for Amikacin, 59.25% for rifampin, 48.14% for cefotaxime, 44.44% for chloramphenicol, 28.5% for clarithromycin and 0 % for cephalexin. The results showed there was not Salmonella spp. strains in all eggs samples so that present work tried to check the presence of Salmonella spp in farm chickens in farms, 213 chicken stool samples were collected from four farms, the samples were cultured on SS Agar, the results showed presence of Salmonella spp. in 10.37% of stool samples and the antibiotic sensitivity tested, also,the result showed that the sensitivity was 63.6% for ciprofloxacin, 86.36% for gentamicin, 86.36% for amikacin, 36.36% for rifampin, 27.27% for cefotaxime, 4.54% for chloramphenicol and 0% for cephalexin. The results indicated that there were 90% of the isolate of Salmonella spp. isolate have Multi-drug resistance phenomenon.

scholarly journals Antimicrobial susceptibility patterns of respiratory Gram-negative bacterial isolates from COVID-19 patients in Switzerland

2021 ◽  
Vol 20 (1) ◽  
Author(s):  
Marina Gysin ◽  
Claudio Tirso Acevedo ◽  
Klara Haldimann ◽  
Elias Bodendoerfer ◽  
Frank Imkamp ◽  
...  
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Antimicrobial Resistance ◽  
Bacterial Infections ◽  
Drug Exposure ◽  
Bacterial Strains ◽  
Gram Negative ◽  
Genotypic Analysis ◽  
Susceptibility Profiles ◽  
Phenotypic Shift ◽  
Efficacious Drug

Abstract Background Bacterial superinfections associated with COVID-19 are common in ventilated ICU patients and impact morbidity and lethality. However, the contribution of antimicrobial resistance to the manifestation of bacterial infections in these patients has yet to be elucidated. Methods We collected 70 Gram-negative bacterial strains, isolated from the lower respiratory tract of ventilated COVID-19 patients in Zurich, Switzerland between March and May 2020. Species identification was performed using MALDI-TOF; antibiotic susceptibility profiles were determined by EUCAST disk diffusion and CLSI broth microdilution assays. Selected Pseudomonas aeruginosa isolates were analyzed by whole-genome sequencing. Results Pseudomonas aeruginosa (46%) and Enterobacterales (36%) comprised the two largest etiologic groups. Drug resistance in P. aeruginosa isolates was high for piperacillin/tazobactam (65.6%), cefepime (56.3%), ceftazidime (46.9%) and meropenem (50.0%). Enterobacterales isolates showed slightly lower levels of resistance to piperacillin/tazobactam (32%), ceftriaxone (32%), and ceftazidime (36%). All P. aeruginosa isolates and 96% of Enterobacterales isolates were susceptible to aminoglycosides, with apramycin found to provide best-in-class coverage. Genotypic analysis of consecutive P. aeruginosa isolates in one patient revealed a frameshift mutation in the transcriptional regulator nalC that coincided with a phenotypic shift in susceptibility to β-lactams and quinolones. Conclusions Considerable levels of antimicrobial resistance may have contributed to the manifestation of bacterial superinfections in ventilated COVID-19 patients, and may in some cases mandate consecutive adaptation of antibiotic therapy. High susceptibility to amikacin and apramycin suggests that aminoglycosides may remain an effective second-line treatment of ventilator-associated bacterial pneumonia, provided efficacious drug exposure in lungs can be achieved.

scholarly journals Surveillance of bacteria Pseudomonas aeruginosa and MRSA associated with acute suppurative otitis media (ASOM)

2017 ◽  
Vol 6 (1) ◽  
pp. 69 ◽  
Author(s):  
Kundan K. Sahu ◽  
Siba N. Rath ◽  
Rabindra N. Padhy ◽  
Rajashree Panigrahi
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Otitis Media ◽  
Bacterial Infections ◽  
Multidrug Resistant ◽  
Coagulase Negative Staphylococcus ◽  
Bacterial Strains ◽  
Suppurative Otitis Media ◽  
Intracranial Complications ◽  
Susceptibility Rate ◽  
Middle Ear Cleft

Background: Otitis media particularly with suppuration is a critical disease-causing perforation of the tympanic membrane associated with changes of the mucoperiosteum of the middle ear cleft. This surveillance includes isolation and antibiotic profiles of causative bacteria from ear discharges of patients in 3years attending outpatients of a hospital.Methods: Bacterial strains were grown in suitable media and were subjected to antibiotic profiling by the Kirby-Bauer’s method with most antibiotics of the day.Results: In total there were 1164 colonies with 1043 bacterial and 121 fungal isolates from 1230 ear discharge samples. Among 371 Pseudomonas aeruginosa isolates, tobramycin 30 µg/disk had the highest susceptibility rate as 93.2%, followed by ceftazidime 30µg/disk 91.5% and amikacin 10µg/disk 64.4%. From 359 Staphylococcus isolates, there were 236 coagulase negative Staphylococcus (CONS) + methicillin sensitive S. aureus (MSSA) and 123 methicillin resistant S. aureus (MRSA). Staphylococcus including MRSA isolates were most susceptibility to cloxacillin 15µg/disk 95.2%, followed by erythromycin 15µg/disk 83.3% and gentamicin 30µg/disk 78.5%. Of 1164, 49 patients presented post aural abscess, 12 patients had intracranial complications, 9 patients had facial palsy and 3 patients had labyrinthitis.Conclusions: Isolated bacteria, P. aeruginosa and MRSA were multidrug resistant. P. aeruginosa was most common followed by S. aureus. More than 90% P. aeruginosa and 90% S. aureus isolates were sensitive to tobramycin 30 µg/disk and cloxacillin 30 µg/disk, respectively. Therefore, these two antibiotics may be included in the formulary regimen to overcome bacterial infections involved in ASOM.

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