scholarly journals Three Pseudomonas aeruginosa strains with different protease profiles.

2013 ◽  
Vol 60 (1) ◽  
Author(s):  
Mariola Andrejko ◽  
Agnieszka Zdybicka-Barabas ◽  
Monika Janczarek ◽  
Małgorzata Cytryńska
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Alkaline Protease ◽  
Reference Strain ◽  
Galleria Mellonella ◽  
Proteolytic Enzymes ◽  
Growth Conditions ◽  
Luria Bertani ◽  
Clinical Isolates ◽  
Pcr Analysis ◽  
Growth Media

The proteolytic activity of three Pseudomonas aeruginosa strains, ATCC 27853 - a reference strain, and two clinical isolates was tested. The activity was examined after culturing the bacteria in two different growth media: the minimal M9 medium and rich Luria-Bertani broth (LB). Based on zymograms and protease activity specific assays, it was concluded that the reference strain produced three proteolytic enzymes in the LB medium: protease IV, elastase B and elastase A, while alkaline protease was only produced in the M9 medium. The clinical isolates of P. aeruginosa produced elastase B and alkaline protease when grown in the LB medium and the minimal M9 medium, respectively. PCR analysis confirmed the presence of both the lasB gene encoding elastase B and aprA coding for alkaline protease in the genomes of the three P. aeruginosa strains analyzed. The expression of these genes coding for two important P. aeruginosa virulence factors was dependent on the growth conditions in all the strains studied. The contribution of the extracellular proteinases to the virulence of P. aeruginosa strains used in this study was investigated using an insect model, the greater wax moth Galleria mellonella.

scholarly journals Quorum sensing and virulence of Pseudomonas aeruginosa during urinary tract infections

2012 ◽  
Vol 6 (06) ◽  
pp. 501-507 ◽  
Author(s):  
Sezgi Senturk ◽  
Seyhan Ulusoy ◽  
Gulgun Bosgelmez-Tinaz ◽  
Aysegul Yagci
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Urinary Tract ◽  
Quorum Sensing ◽  
Virulence Factors ◽  
Urinary Tract Infections ◽  
Clinical Isolates ◽  
Pcr Analysis ◽  
Signal Molecules ◽  
Tract Infections

Introduction: In the opportunistic pathogen Pseudomonas aeruginosa, the production of several virulence factors depends on quorum sensing (QS) involving N-acylhomoserine lactone signal molecules. In vitro studies have suggested that the QS system is crucial in the pathogenesis of P. aeruginosa. However, it is unclear whether QS systems of P. aeruginosa play the same role during infections. Methodology:  In this study, to explore the contribution of QS systems to the pathogenesis of P. aeruginosa during urinary tract infections, we collected 82 clinical isolates. Detection of N-acyl-homoserine lactones (C12-HSL and C4-HSL) was performed on agar plates employing biosensor strains C. violaceum. Elastase and biofilm production were determined spectrophotometrically. QS genes were detected by PCR and subsequently underwent sequencing. Results and conclusion:  Six isolates were found to be negative in the production of both C12-HSL and C4-HSL and all virulence factors tested.  PCR analysis of these isolates revealed that four isolates contained all four QS genes while one isolate was negative for lasR gene, and one isolate negative for lasI, lasR and rhlR genes. Sequence analyses of these isolates showed that the lasR, lasI, rhlR and rhlI genes had point mutations. The combination of these mutations probably explains their C12-HSL, C4-HSL and virulence factor deficiencies. Results of this study suggest that QS deficient clinical isolates occur and are still capable of causing clinical infections in humans. 

scholarly journals Phenotypic and genetic analysis of virulence factors in multidrug- sensitive and multidrug-resistant clinical isolates of Pseudomonas aeruginosa

2021 ◽  
Vol 10 (11) ◽  
pp. e457101120032
Author(s):  
Stephanie Targino Silva ◽  
Jailton Lobo da Costa Lima ◽  
Marcelle Aquino Rabelo ◽  
Armando Monteiro Bezerra Neto ◽  
Lílian Rodrigues Alves ◽  
...  
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Phospholipase C ◽  
Virulence Factors ◽  
Alkaline Protease ◽  
Tertiary Hospital ◽  
Multidrug Resistant ◽  
Clinical Isolates ◽  
Protease Production ◽  
High Level ◽  
Almost All

This study aimed to correlate the pattern of antimicrobial susceptibility, phenotypic production of virulence factors, the occurrence of virulence factors genes and the clonal profile of clinical isolates of Pseudomonas aeruginosa of a tertiary hospital in Recife-PE. The 30 clinical isolates (15 multidrug-sensitive (MDS) and 15 multidrug-resistant (MDR)) were analyzed using phenotypic methods to detect virulence factors (alkaline protease, hemolysin, phospholipase C, lipase, and pigments). The detection of the aprA, lasA, lasB, plcH, and toxA genes was performed through specific PCRs, and the clonal profile was assessed using ERIC-PCR. The results revealed cephalosporins being the class eliciting the highest percentage of resistance; the MDR isolates were all resistant. Among the MDS isolates, all were sensitive to carbapenems and quinolones. The MDR isolates produced less virulence factors such as pyocyanin and lipase, and exhibited lower expression of toxA and lasA genes, whereas the MDS isolates produced less hemolysin and phospholipase C. There was no difference between the groups for alkaline protease production and aprA gene expression. All the isolates produced pyocyanin and expressed lasB and plcH genes. A great genetic diversity was found, and it was possible to observe 28 genetic profiles. Clones were present among the MDR isolates. The occurrence of virulence factors in almost all the isolates studied suggests their high level of pathogenicity, demonstrating that this pathogen is capable of accumulating numerous virulence factors, and in some cases, is associated with multidrug resistance, which makes it difficult to treat these infections.

scholarly journals The effectiveness of Proteolytic bacteria in the leather and detergent industry isolated waste from the Modjo tannery

2021 ◽  
Author(s):  
Tayachew Desalegn ◽  
◽  
Ketema Bacha ◽  
Mesfin Tafesse ◽  
Chandran Masi ◽  
...  
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Bacillus Subtilis ◽  
Proteolytic Enzymes ◽  
Digestive Enzyme ◽  
Total Solid ◽  
Growth Conditions ◽  
Protease Production ◽  
Alkaline Proteases ◽  
Extracellular Proteases ◽  
Protease Enzyme

Protease also called proteinase or peptidase is a digestive enzyme that is categorized under proteolytic enzymes and it has great potential in industrial application. Extracellular proteases are used in a variety of industries because they exhibit practically all of the characteristics needed for biotech applications such as detergent, bioremediation, food, and leather processing. In the synthesis of all three major types of acidic, neutral, and alkaline proteases, microbial sources have dominated an unbeatable area. Alkaline proteases are a large group of industrial enzymes formed by a wide variety of species, including animals, fungi, and bacteria. The fermentation method serves to make bacteria, fungi, and yeast alkaline proteases. Proteases are produced in large quantities by Gram-positive bacteria, especially those belonging to the Bacillus genus. Following standard procedures, the bacterial isolates PMOJ-01 and PMOJ-05 with the prominent zone of clearance and efficient enzyme development were further characterized to the genus level. Moreover, the growth conditions for the highest protease production were optimized with different pH, temperatures, and NaCl concentrations, in the results of PMOJ-01 and PMOJ- 05 pH (7 and 8), temperatures 45oC, and 1% NaCl concentrations both cases respectively. The proteases activities from PMOJ-01, Pseudomonas aeruginosa, and PMOJ-05, Bacillus subtilis were most active at pH 7.0 and pH 8.0 and temperature at 35oC and 45oC, respectively. The enzyme activity and the total solid protease sample of the crude enzyme of Pseudomonas aeruginosa and Bacillus subtilis were 0.299 U/ mL and 0.289 U/ mL, 1.37±0.14 U/mg, and 1.199 U/mg respectively. The effect on dehairing, distaining, and scum removal revealed that the purified protease enzyme of PMOJ-01 and PMOJ-05 can be used in detergent and leather industries.

scholarly journals Phenotypic Adaption of Pseudomonas aeruginosa by Hacking Siderophores Produced by Other Microorganisms

2020 ◽  
Vol 19 (4) ◽  
pp. 589-607 ◽  
Author(s):  
Quentin Perraud ◽  
Paola Cantero ◽  
Béatrice Roche ◽  
Véronique Gasser ◽  
Vincent P. Normant ◽  
...  
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Phenotypic Plasticity ◽  
Iron Uptake ◽  
Opportunistic Pathogen ◽  
Growth Conditions ◽  
Growth Media ◽  
Cell Infection ◽  
Uptake Pathway ◽  
Pathway Expression ◽  
Strong Competition

Bacteria secrete siderophores to access iron, a key nutrient poorly bioavailable and the source of strong competition between microorganisms in most biotopes. Many bacteria also use siderophores produced by other microorganisms (exosiderophores) in a piracy strategy. Pseudomonas aeruginosa, an opportunistic pathogen, produces two siderophores, pyoverdine and pyochelin, and is also able to use a panel of exosiderophores. We first investigated expression of the various iron-uptake pathways of P. aeruginosa in three different growth media using proteomic and RT-qPCR approaches and observed three different phenotypic patterns, indicating complex phenotypic plasticity in the expression of the various iron-uptake pathways. We then investigated the phenotypic plasticity of iron-uptake pathway expression in the presence of various exosiderophores (present individually or as a mixture) under planktonic growth conditions, as well as in an epithelial cell infection assay. In all growth conditions tested, catechol-type exosiderophores were clearly more efficient in inducing the expression of their corresponding transporters than the others, showing that bacteria opt for the use of catechol siderophores to access iron when they are present in the environment. In parallel, expression of the proteins of the pyochelin pathway was significantly repressed under most conditions tested, as well as that of proteins of the pyoverdine pathway, but to a lesser extent. There was no effect on the expression of the heme and ferrous uptake pathways. Overall, these data provide precise insights on how P. aeruginosa adjusts the expression of its various iron-uptake pathways (phenotypic plasticity and switching) to match varying levels of iron and competition.

scholarly journals Antioxidant Enzyme Expression in Clinical Isolates of Pseudomonas aeruginosa: Identification of an Atypical Form of Manganese Superoxide Dismutase

2001 ◽  
Vol 69 (12) ◽  
pp. 7396-7401 ◽  
Author(s):  
Bradley E. Britigan ◽  
Rachel A. Miller ◽  
Daniel J. Hassett ◽  
Michael A. Pfaller ◽  
Michael L. McCormick ◽  
...  
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Antioxidant Enzyme ◽  
Catalase Activity ◽  
Stop Codon ◽  
Premature Stop Codon ◽  
Growth Conditions ◽  
Clinical Isolates ◽  
Missense Mutations ◽  
Enzyme Expression

ABSTRACT Expression of superoxide dismutases (FeSOD and MnSOD) and catalases by laboratory strains of Pseudomonas aeruginosa is modulated by exogenous factors. Whether clinical isolates behave similarly and whether antioxidant enzyme expression influencesP. aeruginosa virulence remain unclear. Fifty-sevenP. aeruginosa blood culture isolates, plus seven pairs of blood and local-site isolates, were examined for FeSOD, MnSOD, and catalase production in vitro. Under iron-replete growth conditions FeSOD and catalase activities were maximized. MnSOD was not detected. FeSOD and catalase activity decreased under iron-limited growth conditions, whereas MnSOD activity appeared. SOD and catalase activity did not change with site of isolation or by patient. MnSOD could not be expressed by one isolate due to a missense mutation insodA that produced a premature stop codon. Eleven percent of the isolates expressed a novel, rapidly migrating MnSOD that was associated with missense mutations in the normal stop codon ofsodA. We conclude that clinical P. aeruginosa isolates vary little in FeSOD and catalase expression. Some strains produce a newly described MnSOD variant, whereas one is deficient in MnSOD production. The absence of MnSOD expression in a P. aeruginosa strain causing invasive human disease indicates that MnSOD is probably not essential forP. aeruginosa virulence.

scholarly journals The Core Proteome of Biofilm-Grown Clinical Pseudomonas aeruginosa Isolates

Cells ◽  
2019 ◽  
Vol 8 (10) ◽  
pp. 1129 ◽  
Author(s):  
Jelena Erdmann ◽  
Janne G. Thöming ◽  
Sarah Pohl ◽  
Andreas Pich ◽  
Christof Lenz ◽  
...  
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Bacterial Species ◽  
Protein Profile ◽  
Opportunistic Pathogen ◽  
Growth Conditions ◽  
Protein Quantification ◽  
Clinical Isolates ◽  
Biofilm Growth ◽  
Regulatory Pathways ◽  
Data Independent Acquisition

Comparative genomics has greatly facilitated the identification of shared as well as unique features among individual cells or tissues, and thus offers the potential to find disease markers. While proteomics is recognized for its potential to generate quantitative maps of protein expression, comparative proteomics in bacteria has been largely restricted to the comparison of single cell lines or mutant strains. In this study, we used a data independent acquisition (DIA) technique, which enables global protein quantification of large sample cohorts, to record the proteome profiles of overall 27 whole genome sequenced and transcriptionally profiled clinical isolates of the opportunistic pathogen Pseudomonas aeruginosa. Analysis of the proteome profiles across the 27 clinical isolates grown under planktonic and biofilm growth conditions led to the identification of a core biofilm-associated protein profile. Furthermore, we found that protein-to-mRNA ratios between different P. aeruginosa strains are well correlated, indicating conserved patterns of post-transcriptional regulation. Uncovering core regulatory pathways, which drive biofilm formation and associated antibiotic tolerance in bacterial pathogens, promise to give clues to interactions between bacterial species and their environment and could provide useful targets for new clinical interventions to combat biofilm-associated infections.

scholarly journals Cephalosporin nitric oxide-donor prodrug DEA-C3D disperses biofilms formed by clinical cystic fibrosis isolates of Pseudomonas aeruginosa

2019 ◽  
Vol 75 (1) ◽  
pp. 117-125 ◽  
Author(s):  
Odel Soren ◽  
Ardeshir Rineh ◽  
Diogo G Silva ◽  
Yuming Cai ◽  
Robert P Howlin ◽  
...  
Keyword(s):  
Nitric Oxide ◽  
Cystic Fibrosis ◽  
Pseudomonas Aeruginosa ◽  
Laser Scanning ◽  
Laser Scanning Microscopy ◽  
Clinical Isolates ◽  
Nitric Oxide Donor ◽  
Confocal Laser ◽  
Broth Microdilution Method

Abstract Objectives The cephalosporin nitric oxide (NO)-donor prodrug DEA-C3D (‘DiEthylAmin-Cephalosporin-3′-Diazeniumdiolate’) has been shown to initiate the dispersal of biofilms formed by the Pseudomonas aeruginosa laboratory strain PAO1. In this study, we investigated whether DEA-C3D disperses biofilms formed by clinical cystic fibrosis (CF) isolates of P. aeruginosa and its effect in combination with two antipseudomonal antibiotics, tobramycin and colistin, in vitro. Methods β-Lactamase-triggered release of NO from DEA-C3D was confirmed using a gas-phase chemiluminescence detector. MICs for P. aeruginosa clinical isolates were determined using the broth microdilution method. A crystal violet staining technique and confocal laser scanning microscopy were used to evaluate the effects of DEA-C3D on P. aeruginosa biofilms alone and in combination with tobramycin and colistin. Results DEA-C3D was confirmed to selectively release NO in response to contact with bacterial β-lactamase. Despite lacking direct, cephalosporin/β-lactam-based antibacterial activity, DEA-C3D was able to disperse biofilms formed by three P. aeruginosa clinical isolates. Confocal microscopy revealed that DEA-C3D in combination with tobramycin produces similar reductions in biofilm to DEA-C3D alone, whereas the combination with colistin causes near complete eradication of P. aeruginosa biofilms in vitro. Conclusions DEA-C3D is effective in dispersing biofilms formed by multiple clinical isolates of P. aeruginosa and could hold promise as a new adjunctive therapy to patients with CF.

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