Comparative study of virulence factors among ESβL-producing andnonproducing Pseudomonas aeruginosa clinical isolates

ABSTRACTWe investigated the efficacy and safety of liposomal clarithromycin formulations with different surface charges against clinical isolates ofPseudomonas aeruginosafrom the lungs of cystic fibrosis (CF) patients. The liposomal clarithromycin formulations were prepared by the dehydration-rehydration method, and their sizes were measured using the dynamic-light-scattering technique. Encapsulation efficiency was determined by microbiological assay, and the stabilities of the formulations in biological fluid were evaluated for a period of 48 h. The MICs and minimum bactericidal concentrations (MBCs) of free and liposomal formulations were determined withP. aeruginosastrains isolated from CF patients. Liposomal clarithromycin activity against biofilm-formingP. aeruginosawas compared to that of free antibiotic using the Calgary Biofilm Device (CBD). The effects of subinhibitory concentrations of free and liposomal clarithromycin on bacterial virulence factors and motility on agar were investigated on clinical isolates ofP. aeruginosa. The cytotoxicities of the liposome preparations and free drug were evaluated on a pulmonary epithelial cell line (A549). The average diameter of the formulations was >222 nm, with encapsulation efficiencies ranging from 5.7% to 30.4%. The liposomes retained more than 70% of their drug content during the 48-h time period. The highly resistant strains ofP. aeruginosabecame susceptible to liposome-encapsulated clarithromycin (MIC, 256 mg/liter versus 8 mg/liter;P< 0.001). Liposomal clarithromycin reduced the bacterial growth within the biofilm by 3 to 4 log units (P< 0.001), significantly attenuated virulence factor production, and reduced bacterial twitching, swarming, and swimming motilities. The clarithromycin-entrapped liposomes were less cytotoxic than the free drug (P< 0.001). These data indicate that our novel formulations could be a useful strategy to enhance the efficacy of clarithromycin against resistantP. aeruginosastrains that commonly affect individuals with cystic fibrosis.

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A comparative study of coastal and clinical isolates of Pseudomonas aeruginosa

Brazilian Journal of Microbiology ◽

10.1590/s1517-838246320140502 ◽

2015 ◽

Vol 46 (3) ◽

pp. 725-734 ◽

Cited By ~ 4

Author(s):

Anusree V. Nair ◽

Neetha Joseph ◽

Kiran Krishna ◽

K. G. Sneha ◽

Neenu Tom ◽

...

Keyword(s):

Pseudomonas Aeruginosa ◽

Comparative Study ◽

Clinical Isolates

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Quorum sensing and virulence of Pseudomonas aeruginosa during urinary tract infections

The Journal of Infection in Developing Countries ◽

10.3855/jidc.2543 ◽

2012 ◽

Vol 6 (06) ◽

pp. 501-507 ◽

Cited By ~ 24

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.

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Effect of azithromycin and phenylalanine-arginine beta-naphthylamide on quorum sensing and virulence factors in clinical isolates of Pseudomonas aeruginosa

Iranian Journal of Microbiology ◽

10.18502/ijm.v13i1.5491 ◽

2021 ◽

Author(s):

Amel Elsheredy ◽

Ingy El-Soudany ◽

Eglal Elsherbini ◽

Dalia Metwally ◽

Abeer Ghazal

Keyword(s):

Pseudomonas Aeruginosa ◽

Quorum Sensing ◽

Virulence Factors ◽

Efflux Pump ◽

Cell Communication ◽

Inhibitory Effect ◽

Resistance Rate ◽

Clinical Isolates ◽

Efflux Pump Inhibitor ◽

Bacteria Growth

Background and Objectives: Pseudomonas aeruginosa is a problematic opportunistic pathogen causing several types of nosocomial infections with a high resistance rate to antibiotics. Production of many virulence factors in P. aeruginosa is regulated by quorum sensing (QS), a cell-to-cell communication mechanism. In this study, we aimed to assess and compare the inhibitory effect of azithromycin (AZM) and EPI- PAβN (efflux pump inhibitor- Phenylalanine-Arginine Beta-Naphthylamide) on QS system and QS-dependent virulence factors in P. aeruginosa clinical isolates. Materials and Methods: A total of 50 P. aeruginosa isolates were obtained from different types of clinical specimens. Isolates were investigated for detection of QS system molecules by AHL cross-feeding bioassay and QS-dependent virulence factors; this was also confirmed by detection of QS genes (lasR, lasI, rhlR, and rhlI) using PCR assay. The inhibitory effect of sub-MIC AZM and EPI PAβN on these virulence factors was assessed. Results: All the P. aeruginosa, producing QS signals C4 HSL, failed to produce C4 HSL in the presence of sub-MIC AZM, In the presence of EPI PAβN (20 µg/ml) only 14 isolates were affected, there was a significant reduction in QS-dependent virulence factors production (protease, biofilm, rhamnolipid and pyocyanin) in the presence of either 20 µg/ml EPI or subMIC of AZM with the inhibitory effect of AZM was more observed than PAβN. Conclusion: Anti-QS agents like AZM and EPI (PAβN) are useful therapeutic options for P. aeruginosa due to its inhibitory effect on QS-dependent virulence factors production without selective pressure on bacteria growth, so resistance to these agents is less likely to develop.

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Exoenzyme Y Contributes to End-Organ Dysfunction Caused by Pseudomonas aeruginosa Pneumonia in Critically Ill Patients: An Exploratory Study

Toxins ◽

10.3390/toxins12060369 ◽

2020 ◽

Vol 12 (6) ◽

pp. 369

Author(s):

Brant M. Wagener ◽

Naseem Anjum ◽

Sarah C. Christiaans ◽

Morgan E. Banks ◽

Jordan C. Parker ◽

...

Keyword(s):

Pseudomonas Aeruginosa ◽

Organ Dysfunction ◽

Virulence Factors ◽

Opportunistic Pathogen ◽

Lavage Fluid ◽

Host Cells ◽

Clinical Isolates ◽

Icu Patients ◽

Edema Factor ◽

Host Infection

Pseudomonas aeruginosa is an opportunistic pathogen that causes pneumonia in immunocompromised and intensive care unit (ICU) patients. During host infection, P. aeruginosa upregulates the type III secretion system (T3SS), which is used to intoxicate host cells with exoenzyme (Exo) virulence factors. Of the four known Exo virulence factors (U, S, T and Y), ExoU has been shown in prior studies to associate with high mortality rates. Preclinical studies have shown that ExoY is an important edema factor in lung infection caused by P. aeruginosa, although its importance in clinical isolates of P. aeruginosa is unknown. We hypothesized that expression of ExoY would be highly prevalent in clinical isolates and would significantly contribute to patient morbidity secondary to P. aeruginosa pneumonia. A single-center, prospective observational study was conducted at the University of Alabama at Birmingham Hospital. Mechanically ventilated ICU patients with a bronchoalveolar lavage fluid culture positive for P. aeruginosa were included. Enrolled patients were followed from ICU admission to discharge and clinical P. aeruginosa isolates were genotyped for the presence of exoenzyme genes. Ninety-nine patients were enrolled in the study. ExoY was present in 93% of P. aeruginosa clinical isolates. Moreover, ExoY alone (ExoY+/ExoU−) was present in 75% of P. aeruginosa isolates, compared to 2% ExoU alone (ExoY−/ExoU+). We found that bacteria isolated from human samples expressed active ExoY and ExoU, and the presence of ExoY in clinical isolates was associated with end-organ dysfunction. This is the first study we are aware of that demonstrates that ExoY is important in clinical outcomes secondary to nosocomial pneumonia.

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Antibiotic Resistance Profiles and Quorum Sensing-Dependent Virulence Factors in Clinical Isolates of Pseudomonas Aeruginosa

Indian Journal of Microbiology ◽

10.1007/s12088-013-0370-7 ◽

2013 ◽

Vol 53 (2) ◽

pp. 163-167 ◽

Cited By ~ 14

Author(s):

Huafu Wang ◽

Faping Tu ◽

Zhihong Gui ◽

Xianghong Lu ◽

Weihua Chu

Keyword(s):

Pseudomonas Aeruginosa ◽

Antibiotic Resistance ◽

Quorum Sensing ◽

Virulence Factors ◽

Clinical Isolates

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Priming with intranasal lactobacilli prevents Pseudomonas aeruginosa acute pneumonia in mice.

10.21203/rs.3.rs-47604/v1 ◽

2020 ◽

Author(s):

Marie-Sarah FANGOUS ◽

Philippe Gosset ◽

Nicolas Galakhoff ◽

Stéphanie Gouriou ◽

Charles-Antoine Guilloux ◽

...

Keyword(s):

Pseudomonas Aeruginosa ◽

Virulence Factors ◽

Saline Solution ◽

Clinical Isolates ◽

Control Groups ◽

Strain Pao1 ◽

Log Reduction ◽

Animal Survival

Abstract Background : Increasing resistance to antibiotics of Pseudomonas aeruginosa leads to therapeutic deadlock and alternative therapies are needed. We aimed to evaluate the effects of Lactobacillus clinical isolates in vivo, through intranasal administration on a murine model of Pseudomonas aeruginosa pneumonia.Results : We screened in vitro 50 pulmonary clinical isolates of Lactobacillus for their ability to decrease the synthesis of two QS dependent-virulence factors (elastase and pyocyanin) produced by Pseudomonas aeruginosa strain PAO1.Two blends of three Lactobacillus isolates were then tested in vivo: one with highly effective anti-PAO1 virulence factors properties (blend named L.rff for L. rhamnosus, two L. fermentum strains), and the second with no properties (blend named L.psb, for L. paracasei, L. salivarius and L. brevis). Each blend was administered intranasally to mice 18h prior to PAO1 pulmonary infection. Animal survival, bacterial loads, cytological analysis, and cytokines secretion in the lungs were evaluated at 6 or 24h post infection with PAO1. Intranasal priming with both lactobacilli blends significantly improved 7-day mice survival from 12% for the control PAO1 group to 71% and 100% for the two groups receiving L.rff and L.psb respectively. No mortality was observed for both control groups receiving either L.rff or L.psb. Additionally, the PAO1 lung clearance was significantly enhanced at 24h. A 2-log and 4-log reduction was observed in the L.rff+PAO1 and L.psb+PAO1 groups respectively, compared to the control PAO1 group. Significant reductions in neutrophil recruitment and proinflammatory cytokine and chemokine secretion were observed after lactobacilli administration compared to saline solution, whereas IL-10 production was increased. Conclusions : These results demonstrate that intranasal priming with lactobacilli acts as a prophylaxis, and avoids fatal complications caused by Pseudomonas aeruginosa pneumonia in mice. These results were independent of in vitro anti-Pseudomonas aeruginosa activity on QS-dependent virulence factors. Further experiments are required to identify the immune mechanism before initiating clinical trials.

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

Research Society and Development ◽

10.33448/rsd-v10i11.20032 ◽

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.

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