Analysis of integrons and associated gene cassettes in clinical isolates of multidrug resistant Pseudomonas aeruginosa from Southwest Nigeria

Molecular detection and surveillance of the resistance genes harboured by Pseudomonas aeruginosa are becoming increasingly important in assessing and controlling spread and colonization in hospitals, and in guiding the treatment of infections. This study analysed the resistance mechanisms of carbapenem-resistant clinical isolates of P. aeruginosa and identified the associated integron-borne metallo-β-lactamase (MBL)-encoding genes. Twenty-seven imipenem (IPM)-resistant clinical isolates of P. aeruginosa were divided into three groups according to their resistance levels to carbapenems. Strains bearing bla IMP-10 showed extremely high-level resistance to IPM, with MICs of 512–2048 μg ml−1. By comparison, strains bearing bla IMP-1, bla IMP-7 and bla VIM-2 showed an intermediate level of resistance, with MICs of 32–256 μg ml−1. The non-MBL-producing strains showed a low level of resistance, with MICs of 8–32 μg ml−1. The same trend in resistance levels was also observed when resistance to other carbapenems, such as meropenem and panipenem, was determined. DNA sequencing showed that the MBL-encoding gene cassettes were carried by class 1 integrons. The bla IMP-1, bla IMP-7 and bla IMP-10 gene cassettes were preceded by a hybrid P ant promoter, TGGACA-N17-TAAACT, and the bla VIM-2 gene cassette was preceded by a weak promoter, TGGACA-N17-TAAGCT. Most of the MBL-encoding genes were linked to one or two resistance genes encoding aminoglycoside-modifying enzymes, such as aac(6′)Iae, aac(6′)II, aacA7, aacC4, aadA1, aadA2 and aadA6, highlighting the multidrug-resistant properties of these clinical isolates.

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Detection of Extended-Spectrum β-Lactamases in Clinical Isolates of Pseudomonas aeruginosa

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.01511-05 ◽

2006 ◽

Vol 50 (9) ◽

pp. 2990-2995 ◽

Cited By ~ 70

Author(s):

Xiaofei Jiang ◽

Zhe Zhang ◽

Min Li ◽

Danqiu Zhou ◽

Feiyi Ruan ◽

...

Keyword(s):

Pseudomonas Aeruginosa ◽

Multidrug Resistant ◽

Clinical Isolates ◽

Screening Methods ◽

Extended Spectrum ◽

Pcr Product ◽

Amoxicillin Clavulanate ◽

Synergy Test ◽

Esbl Producers ◽

Disk Test

ABSTRACT With the occurrence of extended-spectrum β-lactamases (ESBLs) in Pseudomonas aeruginosa being increasingly reported worldwide, there is a need for a reliable test to detect ESBLs in clinical isolates of P. aeruginosa. In our study, a total of 75 clinical isolates of P. aeruginosa were studied. Nitrocefin tests were performed to detect the β-lactamase enzyme; isoelectric focusing electrophoresis, PCR, and PCR product sequencing were designed to further characterize the contained ESBLs. Various ESBL-screening methods were designed to compare the reliabilities of detecting ESBLs in clinical isolates of P. aeruginosa whose β-lactamases were well characterized. Thirty-four of 36 multidrug-resistant P. aeruginosa clinical isolates were positive for ESBLs. bla VEB-3 was the most prevalent ESBL gene in P. aeruginosa in our study. Among the total of 34 isolates that were considered ESBL producers, 20 strains were positive using conventional combined disk tests and 10 strains were positive using a conventional double-disk synergy test (DDST) with amoxicillin-clavulanate, expanded-spectrum cephalosporins, aztreonam, and cefepime. Modifications of the combined disk test and DDST, which consisted of shorter distances between disks (20 mm instead of 30 mm) and the use of three different plates that contained cloxacillin (200 μg/ml) alone, Phe-Arg β-naphthylamide dihydrochloride (MC-207,110; 20 μg/ml) alone, and both cloxacillin (200 μg/ml) and MC-207,110 (20 μg/ml) increased the sensitivity of the tests to 78.8%, 91.18%, 85.29%, and 97.06%.

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Enhancement of pyocyanin production by subinhibitory concentration of royal jelly in Pseudomonas aeruginosa

F1000Research ◽

10.12688/f1000research.27915.3 ◽

2021 ◽

Vol 10 ◽

pp. 14

Author(s):

Dina Auliya Amly ◽

Puspita Hajardhini ◽

Alma Linggar Jonarta ◽

Heribertus Dedy Kusuma Yulianto ◽

Heni Susilowati

Keyword(s):

Pseudomonas Aeruginosa ◽

Bacterial Growth ◽

Biofilm Formation ◽

Royal Jelly ◽

Microtiter Plate ◽

Multidrug Resistant ◽

Clinical Isolates ◽

Antibiotic Tolerance ◽

Subinhibitory Concentration ◽

Pyocyanin Production

Background: Pseudomonas aeruginosa, a multidrug-resistant Gram-negative bacterium, produces pyocyanin, a virulence factor associated with antibiotic tolerance. High concentrations of royal jelly have an antibacterial effect, which may potentially overcome antibacterial resistance. However, in some cases, antibiotic tolerance can occur due to prolonged stress of low-dose antibacterial agents. This study aimed to investigate the effect of subinhibitory concentrations of royal jelly on bacterial growth, pyocyanin production, and biofilm formation of P. aeruginosa. Methods: Pseudomonas aeruginosa ATCC 10145 and clinical isolates were cultured in a royal jelly-containing medium to test the antibacterial activity. Pyocyanin production was observed by measuring the absorbance at 690 nm after 36 h culture and determined using extinction coefficient 4310 M-1 cm-1. Static microtiter plate biofilm assay performed to detect the biofilm formation, followed by scanning electron microscopy. Results: Royal jelly effectively inhibited the viability of both strains from a concentration of 25%. The highest production of pyocyanin was observed in the subinhibitory concentration group 6.25%, which gradually decreased along with the decrease of royal jelly concentration. Results of one-way ANOVA tests differed significantly in pyocyanin production of the two strains between the royal jelly groups. Tukey HSD test showed concentrations of 12.5%, 6.25%, and 3.125% significantly increased pyocyanin production of ATCC 10145, and the concentrations of 12.5% and 6.25% significantly increased production of the clinical isolates. Concentrations of 12.5% and 6.125% significantly induced biofilm formation of P. aeruginosa ATCC 10145, in line with the results of the SEM analysis. Conclusions: The royal jelly concentration of 25% or higher inhibits bacterial growth; however, the subinhibitory concentration increases pyocyanin production and biofilm formation in P. aeruginosa. It is advisable to determine the appropriate concentration of royal jelly to obtain beneficial virulence inhibiting activity.

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DNA microarray for genotyping multidrug-resistant Pseudomonas aeruginosa clinical isolates

Diagnostic Microbiology and Infectious Disease ◽

10.1016/j.diagmicrobio.2007.06.005 ◽

2007 ◽

Vol 59 (3) ◽

pp. 325-338 ◽

Cited By ~ 11

Author(s):

Jan Weile ◽

Rolf D. Schmid ◽

Till T. Bachmann ◽

Milorad Susa ◽

Cornelius Knabbe

Keyword(s):

Pseudomonas Aeruginosa ◽

Dna Microarray ◽

Multidrug Resistant ◽

Clinical Isolates

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Molecular mechanisms of efflux pump mediated resistance in clinical isolates of multidrug resistant pseudomonas aeruginosa

International Journal of Infectious Diseases ◽

10.1016/j.ijid.2016.02.269 ◽

2016 ◽

Vol 45 ◽

pp. 104 ◽

Cited By ~ 1

Author(s):

L. Mohanam ◽

L. Priya ◽

E.M. Selvam ◽

S.S. Shivekar ◽

T. Menon

Keyword(s):

Pseudomonas Aeruginosa ◽

Molecular Mechanisms ◽

Efflux Pump ◽

Multidrug Resistant ◽

Clinical Isolates

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A Novel Cecropin D-Derived Short Cationic Antimicrobial Peptide Exhibits Antibacterial Activity Against Wild-Type and Multidrug-Resistant Strains of Klebsiella pneumoniae and Pseudomonas aeruginosa

Evolutionary Bioinformatics ◽

10.1177/1176934320936266 ◽

2020 ◽

Vol 16 ◽

pp. 117693432093626

Author(s):

Iván Darío Ocampo-Ibáñez ◽

Yamil Liscano ◽

Sandra Patricia Rivera-Sánchez ◽

José Oñate-Garzón ◽

Ashley Dayan Lugo-Guevara ◽

...

Keyword(s):

Pseudomonas Aeruginosa ◽

Antibacterial Activity ◽

Klebsiella Pneumoniae ◽

Multidrug Resistant ◽

Clinical Isolates ◽

Health Concern ◽

Wild Type ◽

Cationic Antimicrobial Peptide ◽

Promising Alternative ◽

Resistant Strains

Infections caused by multidrug-resistant (MDR) Pseudomonas aeruginosa and Klebsiella pneumoniae are a serious worldwide public health concern due to the ineffectiveness of empirical antibiotic therapy. Therefore, research and the development of new antibiotic alternatives are urgently needed to control these bacteria. The use of cationic antimicrobial peptides (CAMPs) is a promising candidate alternative therapeutic strategy to antibiotics because they exhibit antibacterial activity against both antibiotic susceptible and MDR strains. In this study, we aimed to investigate the in vitro antibacterial effect of a short synthetic CAMP derived from the ΔM2 analog of Cec D-like (CAMP-CecD) against clinical isolates of K pneumoniae (n = 30) and P aeruginosa (n = 30), as well as its hemolytic activity. Minimal inhibitory concentrations (MICs) and minimal bactericidal concentrations (MBCs) of CAMP-CecD against wild-type and MDR strains were determined by the broth microdilution test. In addition, an in silico molecular dynamic simulation was performed to predict the interaction between CAMP-CecD and membrane models of K pneumoniae and P aeruginosa. The results revealed a bactericidal effect of CAMP-CecD against both wild-type and resistant strains, but MDR P aeruginosa showed higher susceptibility to this peptide with MIC values between 32 and >256 μg/mL. CAMP-CecD showed higher stability in the P aeruginosa membrane model compared with the K pneumoniae model due to the greater number of noncovalent interactions with phospholipid 1-Palmitoyl-2-oleyl-sn-glycero-3-(phospho-rac-(1-glycerol)) (POPG). This may be related to the boosted effectiveness of the peptide against P aeruginosa clinical isolates. Given the antibacterial activity of CAMP-CecD against wild-type and MDR clinical isolates of P aeruginosa and K pneumoniae and its nonhemolytic effects on human erythrocytes, CAMP-CecD may be a promising alternative to conventional antibiotics.

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Integrated Genome-Wide Analysis of an Isogenic Pair of Pseudomonas aeruginosa Clinical Isolates with Differential Antimicrobial Resistance to Ceftolozane/Tazobactam, Ceftazidime/Avibactam, and Piperacillin/Tazobactam

International Journal of Molecular Sciences ◽

10.3390/ijms21031026 ◽

2020 ◽

Vol 21 (3) ◽

pp. 1026 ◽

Cited By ~ 1

Author(s):

Weihua Huang ◽

Joelle El Hamouche ◽

Guiqing Wang ◽

Melissa Smith ◽

Changhong Yin ◽

...

Keyword(s):

Dna Methylation ◽

Pseudomonas Aeruginosa ◽

Antimicrobial Resistance ◽

Multidrug Resistant ◽

Clinical Isolates ◽

Epigenetic Mechanism ◽

Phenotypic Change ◽

Genome Wide ◽

Family Protein ◽

Significant Difference

Multidrug-resistant (MDR) Pseudomonas aeruginosa is one of the main causes of morbidity and mortality in hospitalized patients and the leading cause of nosocomial infections. We investigated, here, two MDR P. aeruginosa clinical isolates from a hospitalized patient with differential antimicrobial resistance to ceftazidime/avibactam (CZA), ceftolozane/tazobactam (C/T), and piperacillin/tazobactam (P/T). Their assembled complete genomes revealed they belonged to ST235, a widespread MDR clone; and were isogenic with only a single nucleotide variant, causing G183D mutation in AmpC β-lactamase, responsible for a phenotypic change from susceptible to resistant to CZA and C/T. Further epigenomic profiling uncovered two conserved DNA methylation motifs targeted by two distinct putative methyltransferase-containing restriction-modification systems, respectively; more intriguingly, there was a significant difference between the paired isolates in the pattern of genomic DNA methylation and modifications. Moreover, genome-wide gene expression profiling demonstrated the inheritable genomic methylation and modification induced 14 genes being differentially regulated, of which only toxR (downregulated), a regulatory transcription factor, had its promoter region differentially methylate and modified. Since highly expressed opdQ encodes an OprD porin family protein, therefore, we proposed an epigenetic regulation of opdQ expression pertinent to the phenotypic change of P. aeruginosa from resistant to susceptible to P/T. The disclosed epigenetic mechanism controlling phenotypic antimicrobial resistance deserves further experimental investigation.

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The Resistome of Pseudomonas aeruginosa in Relationship to Phenotypic Susceptibility

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.03954-14 ◽

2014 ◽

Vol 59 (1) ◽

pp. 427-436 ◽

Cited By ~ 135

Author(s):

Veronica N. Kos ◽

Maxime Déraspe ◽

Robert E. McLaughlin ◽

James D. Whiteaker ◽

Paul H. Roy ◽

...

Keyword(s):

Pseudomonas Aeruginosa ◽

Multidrug Resistant ◽

Clinical Isolates ◽

Genetic Determinants ◽

Content Type ◽

Susceptibility Data ◽

Next Generation Sequencing Ngs ◽

Sequence Types ◽

Generation Sequencing ◽

Phenotypic Susceptibility

ABSTRACTMany clinical isolates ofPseudomonas aeruginosacause infections that are difficult to eradicate due to their resistance to a wide variety of antibiotics. Key genetic determinants of resistance were identified through genome sequences of 390 clinical isolates ofP. aeruginosa, obtained from diverse geographic locations collected between 2003 and 2012 and were related to microbiological susceptibility data for meropenem, levofloxacin, and amikacin. β-Lactamases and integron cassette arrangements were enriched in the established multidrug-resistant lineages of sequence types ST111 (predominantly O12) and ST235 (O11). This study demonstrates the utility of next-generation sequencing (NGS) in defining relevant resistance elements and highlights the diversity of resistance determinants withinP. aeruginosa. This information is valuable in furthering the design of diagnostics and therapeutics for the treatment ofP. aeruginosainfections.

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Inactivation of the Pseudomonas -Derived Cephalosporinase-3 (PDC-3) by Relebactam

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.02406-17 ◽

2018 ◽

Vol 62 (5) ◽

Cited By ~ 12

Author(s):

Melissa D. Barnes ◽

Christopher R. Bethel ◽

Jim Alsop ◽

Scott A. Becka ◽

Joseph D. Rutter ◽

...

Keyword(s):

Pseudomonas Aeruginosa ◽

Multidrug Resistant ◽

Clinical Isolates ◽

Gram Negative ◽

Content Type ◽

Life Threatening ◽

Lactamase Inhibitor

ABSTRACT Pseudomonas aeruginosa is a prevalent and life-threatening Gram-negative pathogen. Pseudomonas -derived cephlosporinase (PDC) is the major inducible cephalosporinase in P. aeruginosa . In this investigation, we show that relebactam, a diazabicyclooctane β-lactamase inhibitor, potently inactivates PDC-3, with a k 2 / K of 41,400 M −1 s −1 and a k off of 0.00095 s −1 . Relebactam restored susceptibility to imipenem in 62% of multidrug-resistant P. aeruginosa clinical isolates, while only 21% of isolates were susceptible to imipenem-cilastatin alone. Relebactam promises to increase the efficacy of imipenem-cilastatin against P. aeruginosa .

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