Mutations in nalc gene of Mex AB-OprM efflux pump in carbapenem resistant Pseudomonas aeruginosa isolated from burn wounds in Yazd, Iran

Background and Objectives: Burn wound infections have emerged as an important cause of morbidity and mortality in patients due to prolonged hospital stay. Pseudomonas aeruginosa, is the second cause of bacterial burn wound infections. Resistance mechanisms among P. aeruginosa are intrinsic or acquired. Intrinsic resistance mechanisms among P. aeruginosa isolates are inducible AmpC cephalosporinase, decrease of specific porin OprD, and overexpression of RND efflux pump. The aim of this study was detection of mutations in nalC gene in carbapenem resistant P. aeruginosa isolated from burn wounds. Materials and Methods: In this cross-sectional study, 180 burn-wound specimens were collected. Suspected lactose-nega- tive colonies were identified by conventional biochemical methods. Kirby-Bauer and Etest methods were used for suscepti- bility testing. PCR and sequencing techniques were used for the detection of nalC mutation. Results: Out of 180 specimens received in the laboratory, 54 of isolates were isolated and identified as P. aeroginosa (30%). Of these isolates 20 (37%) were resistant to at least two carbapenems simultaneously. From these carbapenem resistant iso- lates, 19 (95%), 14 (70%), 14 (70%), 19 (95%) and 16 (80%) were resistant to imipenem, cefepime, piperacillin, ceftizoxime and gentamicin, respectively. Only 1 (2%) isolate was sensitive to all carbapenems and did not has mutation in nalC gene, 20 (37%) isolates were resistant to at least two carbapenems, and had mutations in nalC gene (Gly71►Glu and Ser209►Arg). Conclusion: As the results showed, mutation in efflux pump was observed in carbapenem resistant isolate and this confirmed that the indiscriminate use of antibiotics for treatment or prophylaxis can increase mutation in efflux pump.

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Effect of Human Burn Wound Exudate on Pseudomonas aeruginosa Virulence

mSphere ◽

10.1128/msphere.00111-15 ◽

2016 ◽

Vol 1 (2) ◽

Cited By ~ 17

Author(s):

Manuel R. Gonzalez ◽

Betty Fleuchot ◽

Leonardo Lauciello ◽

Paris Jafari ◽

Lee Ann Applegate ◽

...

Keyword(s):

Pseudomonas Aeruginosa ◽

Chemical Composition ◽

Composition Analysis ◽

Wound Infections ◽

Artificial Medium ◽

Burn Wound ◽

Content Type ◽

Burn Wounds ◽

Chemical Composition Analysis

ABSTRACT Microbial infection of severe burn wounds is currently a major medical challenge. Of the infections by bacteria able to colonize such injuries, those by Pseudomonas aeruginosa are among the most severe, causing major delays in burn patient recovery or leading to fatal issues. In this study, we investigated the growth properties of several burn wound pathogens in biological fluids secreted from human burn wounds. We found that P. aeruginosa strains were able to proliferate but not those of the other pathogens tested. In addition, burn wound exudates (BWEs) stimulate the expression of virulence factors in P. aeruginosa. The chemical composition analysis of BWEs enabled us to determine the major components of these fluids. These data are essential for the development of an artificial medium mimicking the burn wound environment and for in vitro analysis of the initial step in the development of burn wound infections. Burn wound sepsis is currently the main cause of morbidity and mortality after burn trauma. Infections by notorious pathogens such as Pseudomonas aeruginosa, Staphylococcus aureus, and Acinetobacter baumannii impair patient recovery and can even lead to fatality. In this study, we investigated the effect of burn wound exudates (BWEs) on the virulence of those pathogens. BWEs were collected within 7 days after burn trauma from 5 burn patients. We first monitored their effect on pathogen growth. In contrast to A. baumannii and S. aureus, P. aeruginosa was the only pathogen able to grow within these human fluids. Expression of typical virulence factors such as pyocyanin and pyoverdine was even enhanced compared the levels seen with standard laboratory medium. A detailed chemical composition analysis of BWE was performed, which enabled us to determine the major components of BWE and underline the metabolic modifications induced by burn trauma. These data are essential for the development of an artificial medium mimicking the burn wound environment and the establishment of an in vitro system to analyze the initial steps of burn wound infections. IMPORTANCE Microbial infection of severe burn wounds is currently a major medical challenge. Of the infections by bacteria able to colonize such injuries, those by Pseudomonas aeruginosa are among the most severe, causing major delays in burn patient recovery or leading to fatal issues. In this study, we investigated the growth properties of several burn wound pathogens in biological fluids secreted from human burn wounds. We found that P. aeruginosa strains were able to proliferate but not those of the other pathogens tested. In addition, burn wound exudates (BWEs) stimulate the expression of virulence factors in P. aeruginosa. The chemical composition analysis of BWEs enabled us to determine the major components of these fluids. These data are essential for the development of an artificial medium mimicking the burn wound environment and for in vitro analysis of the initial step in the development of burn wound infections.

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Activity of Ceftolozane-Tazobactam against Carbapenem-Resistant, Non-Carbapenemase-Producing Pseudomonas aeruginosa and Associated Resistance Mechanisms

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.01970-17 ◽

2017 ◽

Vol 62 (1) ◽

Cited By ~ 12

Author(s):

Yu Mi Wi ◽

Kerryl E. Greenwood-Quaintance ◽

Audrey N. Schuetz ◽

Kwan Soo Ko ◽

Kyong Ran Peck ◽

...

Keyword(s):

Pseudomonas Aeruginosa ◽

Efflux Pump ◽

Multidrug Resistant ◽

Resistance Mechanisms ◽

Resistance Rate ◽

Content Type ◽

Reverse Transcription Pcr ◽

Carbapenem Resistant ◽

Kill Curve ◽

Time Kill

ABSTRACT Although carbapenems are effective for treating serious multidrug-resistant Pseudomonas aeruginosa infections, carbapenem-resistant P. aeruginosa (CRPA) is now being reported worldwide. Ceftolozane-tazobactam (C/T) demonstrates activity against many multidrug-resistant isolates. We evaluated the activity of C/T and compared its activity to that of ceftazidime-avibactam (C/A) using a well-characterized collection of non-carbapenemase-producing CRPA isolates. Forty-two non-carbapenemase-producing CRPA isolates from a previous study (J. Y. Lee and K. S. Ko, Int J Antimicrob Agents 40:168–172, 2012, https://doi.org/10.1016/j.ijantimicag.2012.04.004) were included. All had been previously shown to be negative for bla IMP, bla VIM, bla SPM, bla GIM, bla SIM, and bla KPC by PCR. In the prior study, expression of oprD, ampC, and several efflux pump genes had been defined by quantitative reverse transcription-PCR. Here, antimicrobial susceptibility was determined by broth microdilution according to Clinical and Laboratory Standards Institute (CLSI) guidelines. Time-kill curve assays were performed using three C/T- and C/A-susceptible CRPA isolates. Among 42 non-carbapenemase-producing CRPA isolates, overall susceptibility to C/T was 95.2%, compared to 71.4%, 42.9%, 23.8%, 21.4%, and 2.4% for C/A, ceftazidime, piperacillin-tazobactam, cefepime, and meropenem, respectively. The C/T resistance rate was significantly lower than that of C/A among isolates showing decreased oprD and increased mexB expression (5.1% versus 25.6%, P = 0.025, and 4.3% versus 34.8%, P = 0.022, respectively). In time-kill curve studies, C/T was less bactericidal than C/A against an isolate with decreased oprD and increased ampC expression. C/T was active against 95.2% of non-carbapenemase-producing CRPA clinical isolates. No apparent correlation of C/T MIC values with specific mutation-driven resistance mechanisms was noted.

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Effects of Topical Antimicrobial Formulations on Pseudomonas aeruginosa Biofilm in an In vivo Porcine Burn Wound Model

10.1101/737338 ◽

2019 ◽

Author(s):

S.C. Davis ◽

M. Solis ◽

J. Gil ◽

J. Valdes ◽

A. Higa ◽

...

Keyword(s):

Pseudomonas Aeruginosa ◽

Wound Healing ◽

Post Infection ◽

Wound Dressings ◽

Wound Infections ◽

Burn Wound ◽

Bacterial Counts ◽

Burn Wounds ◽

Wound Model

AbstractSilver has been incorporated into a variety of wound dressings and topical agents to prevent and combat wound infections. Pseudomonas aeruginosa is a common cause of burn wound infections and well-known biofilm producer. The objective of this study was to evaluate the effects of a panel of wound dressings containing different silver formulations on P. aeruginosa biofilms using an in vivo porcine burn wound model. Second-degree burns were created on the skin of specific pathogen-free pigs (n = 3) and inoculated with 2.14 × 105 cfu P. aeruginosa per wound. Biofilms were allowed to develop for 24 h, and then each wound was treated with one of 6 treatments: silver oxynitrate dressing (OXY), silver oxynitrate powder (POWD), nanocrystalline silver dressing (NANO), silver chloride dressing (AGCL), silver sulfadiazine (SSD), or a negative control polyurethane film with no silver-based formulation (NEG). Wounds were cultured at D3 post-infection (n = 3 per pig per treatment) and at D6 post-infection (n = 3 per pig per treatment) for quantification of bacteria. On D6, biopsies (n = 3 per treatment) were taken from POWD, SSD, and NEG wounds and wound healing progress was evaluated histologically. At the time of treatment initiation, 24 h post-infection, 8.71 log cfu P. aeruginosa were present in burn wounds. On D3 and D6, all treatments significantly reduced bacterial counts in wounds as compared to NEG, but POWD caused an approximately 7-log reduction in bacterial counts on both days and was the only treatment to reduce the bacterial counts to below the threshold for detecting bacteria. The OXY, NANO, and SSD treatments had similar reductions in bacterial recovery on D3 and D6 of approximately 2.5-4 log. The histological healing metrics of reepithelialization percentage, epithelial thickness, white cell infiltration, angiogenesis, and granulation tissue formation were similar among wounds from POWD, SSD, and NEG groups at 6 days post-infection. Silver oxynitrate powder reduced P. aeruginosa growth in burn wounds more effectively than other silver-based dressings but did not impact wound healing.

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Antagonistic Interactions of Pseudomonas aeruginosa Antibiotic Resistance Mechanisms in Planktonic but Not Biofilm Growth

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.00519-11 ◽

2011 ◽

Vol 55 (10) ◽

pp. 4560-4568 ◽

Cited By ~ 34

Author(s):

Xavier Mulet ◽

Bartolomé Moyá ◽

Carlos Juan ◽

María D. Macià ◽

José L. Pérez ◽

...

Keyword(s):

Pseudomonas Aeruginosa ◽

Antibiotic Resistance ◽

Efflux Pump ◽

Resistance Mechanisms ◽

Negative Regulator ◽

Intrinsic Resistance ◽

Major Drawback ◽

Biofilm Growth ◽

Content Type ◽

Bacterial Physiology

ABSTRACTPseudomonas aeruginosahas an extraordinary capacity to evade the activity of antibiotics through a complex interplay of intrinsic and mutation-driven resistance pathways, which are, unfortunately, often additive or synergistic, leading to multidrug (or even pandrug) resistance. However, we show that one of these mechanisms, overexpression of the MexCD-OprJ efflux pump (driven by inactivation of its negative regulator NfxB), causes major changes in the cell envelope physiology, impairing the backbone ofP. aeruginosaintrinsic resistance, including the major constitutive (MexAB-OprM) and inducible (MexXY-OprM) efflux pumps and the inducible AmpC β-lactamase. Moreover, it also impaired the most relevant mutation-driven β-lactam resistance mechanism (constitutive AmpC overexpression), through a dramatic decrease in periplasmic β-lactamase activity, apparently produced by an abnormal permeation of AmpC out of the cell. While these results could delineate future strategies for combating antibiotic resistance in cases of acute nosocomial infections, a major drawback for the potential exploitation of the described antagonistic interaction between resistance mechanisms came from the differential bacterial physiology characteristics of biofilm growth, a hallmark of chronic infections. Although the failure to concentrate AmpC activity in the periplasm dramatically limits the protection of the targets (penicillin-binding proteins [PBPs]) of β-lactams at the individual cell level, the expected outcome for cells growing as biofilm communities, which are surrounded by a thick extracellular matrix, was less obvious. Indeed, our results showed that AmpC produced bynfxBmutants is protective in biofilm growth, suggesting that the permeation of AmpC into the matrix protects biofilm communities against β-lactams.

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Molecular Epidemiology and Characterization of Carbapenem-Resistant Klebsiella pneumoniae Isolated from Urine at a Teaching Hospital in Taiwan

Microorganisms ◽

10.3390/microorganisms9020271 ◽

2021 ◽

Vol 9 (2) ◽

pp. 271

Author(s):

Yuarn-Jang Lee ◽

Chih-Hung Huang ◽

Noor Andryan Ilsan ◽

I-Hui Lee ◽

Tzu-Wen Huang

Keyword(s):

Klebsiella Pneumoniae ◽

Teaching Hospital ◽

Efflux Pump ◽

Resistance Mechanisms ◽

Pcr Analysis ◽

Carbapenem Resistant ◽

High Prevalence ◽

Antimicrobial Susceptibility Tests ◽

Susceptibility Tests ◽

Tract Infections

Urinary tract infections (UTIs) are common in clinics and hospitals and are associated with a high economic burden. Enterobacterium Klebsiella pneumoniae is a prevalent agent causing UTIs. A high prevalence of carbapenem-resistant K. pneumoniae (CRKP) has emerged recently and is continuing to increase. Seventeen urinary CRKP isolates collected at a teaching hospital in Taiwan from December 2016 to September 2017 were analyzed to elucidate their drug resistance mechanisms. Two-thirds of the isolates were obtained from outpatients. Antimicrobial susceptibility tests demonstrated multidrug resistance in all the isolates. Multilocus sequence typing analysis showed high diversity among the isolates. PCR analysis demonstrated the presence of carbapenemases in three isolates. All isolates carried at least one other extended-spectrum β-lactamase, including TEM, DHA, and CTX-M. Fifteen isolates contained mutations in one of the outer membrane porins that were assessed. The expression levels of the acrB and/or oqxB efflux pump genes, as determined by qRT-PCR, were upregulated in 11 isolates. Six isolates might have utilized other efflux pumps or antimicrobial resistance mechanisms. These analyses demonstrated a highly diverse population and the presence of complex resistance mechanisms in urinary isolates of K. pneumoniae.

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Characterization of resistance mechanisms of Enterobacter cloacae Complex co-resistant to carbapenem and colistin

BMC Microbiology ◽

10.1186/s12866-021-02250-x ◽

2021 ◽

Vol 21 (1) ◽

Author(s):

Shixing Liu ◽

Renchi Fang ◽

Ying Zhang ◽

Lijiang Chen ◽

Na Huang ◽

...

Keyword(s):

Lipid A ◽

Efflux Pump ◽

Resistance Mechanism ◽

Carbapenem Resistance ◽

Enterobacter Cloacae ◽

Resistance Mechanisms ◽

Colistin Resistance ◽

Carbapenem Resistant ◽

Horizontal Spread

Abstract Background The emergence of carbapenem-resistant and colistin-resistant ECC pose a huge challenge to infection control. The purpose of this study was to clarify the mechanism of the carbapenems and colistin co-resistance in Enterobacter cloacae Complex (ECC) strains. Results This study showed that the mechanisms of carbapenem resistance in this study are: 1. Generating carbapenemase (7 of 19); 2. The production of AmpC or ESBLs combined with decreased expression of out membrane protein (12 of 19). hsp60 sequence analysis suggested 10 of 19 the strains belong to colistin hetero-resistant clusters and the mechanism of colistin resistance is increasing expression of acrA in the efflux pump AcrAB-TolC alone (18 of 19) or accompanied by a decrease of affinity between colistin and outer membrane caused by the modification of lipid A (14 of 19). Moreover, an ECC strain co-harboring plasmid-mediated mcr-4.3 and blaNDM-1 has been found. Conclusions This study suggested that there is no overlap between the resistance mechanism of co-resistant ECC strains to carbapenem and colistin. However, the emergence of strain co-harboring plasmid-mediated resistance genes indicated that ECC is a potential carrier for the horizontal spread of carbapenems and colistin resistance.

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Resistance Mechanisms of Multiresistant Pseudomonas aeruginosa Strains from Germany and Correlation with Hypermutation

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.00148-07 ◽

2007 ◽

Vol 51 (11) ◽

pp. 4062-4070 ◽

Cited By ~ 105

Author(s):

B. Henrichfreise ◽

I. Wiegand ◽

W. Pfister ◽

B. Wiedemann

Keyword(s):

Pseudomonas Aeruginosa ◽

Efflux Pump ◽

Acquired Resistance ◽

Resistance Mechanisms ◽

Type Ii ◽

Mechanisms Of Resistance ◽

Resistance Determinants ◽

Class 1 ◽

Mutator Strains ◽

Multiresistant Strains

ABSTRACT In this study, we analyzed the mechanisms of multiresistance for 22 clinical multiresistant and clonally different Pseudomonas aeruginosa strains from Germany. Twelve and 10 strains originated from cystic fibrosis (CF) and non-CF patients, respectively. Overproduction of the efflux systems MexAB-OprM, MexCD-OprJ, MexEF-OprN, and MexXY-OprM was studied. Furthermore, loss of OprD, alterations in type II topoisomerases, AmpC overproduction, and the presence of 25 acquired resistance determinants were investigated. The presence of a hypermutation phenotype was also taken into account. Besides modifications in GyrA (91%), the most frequent mechanisms of resistance were MexXY-OprM overproduction (82%), OprD loss (82%), and AmpC overproduction (73%). Clear differences between strains from CF and non-CF patients were found: numerous genes coding for aminoglycoside-modifying enzymes and located, partially in combination with β-lactamase genes, in class 1 integrons were found only in strains from non-CF patients. Furthermore, multiple modifications in type II topoisomerases conferring high quinolone resistance levels and overexpression of MexAB-OprM were exclusively detected in multiresistant strains from non-CF patients. Correlations of the detected phenotypes and resistance mechanisms revealed a great impact of efflux pump overproduction on multiresistance in P. aeruginosa. Confirming previous studies, we found that additional, unknown chromosomally mediated resistance mechanisms remain to be determined. In our study, 11 out of 12 strains and 3 out of 10 strains from CF patients and non-CF patients, respectively, were hypermutable. This extremely high proportion of mutator strains should be taken into consideration for the treatment of multiresistant P. aeruginosa.

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Phenotypic and Molecular Characteristics of Pseudomonas Aeruginosa Isolated from Burn Unit

10.51429/ejmm30103 ◽

2021 ◽

Vol 30 (1) ◽

pp. 19-28

Author(s):

Yasser M. Ismail ◽

Sahar M. Fayed ◽

Fatma M. Elesawy ◽

Nora Z Abd El-Halim ◽

Ola S. El-Shimi

Keyword(s):

Pseudomonas Aeruginosa ◽

Virulence Factors ◽

Antimicrobial Susceptibility ◽

Virulence Gene ◽

Molecular Characteristics ◽

Drug Resistant ◽

Burn Wound ◽

Burn Wounds ◽

Related Data ◽

Genes Encoding

Background: The biggest concern for a burn team is a nosocomial infection in burn patients, which is a significant health issue. Pseudomonas aeruginosa is an extremely troublesome drug-resistant bacterium in the world today. We are now faced with rising P. aeruginosa pan-drug-resistant clones in hospital settings. Objectives: To evaluate the distribution of different virulence factors generated by P. aeruginosa isolated from burn wound infections, together with its antimicrobial susceptibility. Methodology: The isolates reported as P. aeruginosa were further tested for the presence of various phenotypic and genotypic virulence factors including (Biofilm formation, lipase, protease, gelatinase, DNase, bile esculin hydrolysis & hemolysin). Also, genes encoding (nan 1 and Exo A) were investigated by PCR using specific primers. All the isolates were tested for their antimicrobial susceptibility patterns. Results: The study reported that toxins and enzymes were expressed by the tested strains in varying proportions; (92.0%) were producing β-hemolysin, lipase (86%), and protease (86%). The formation of biofilm was observed in 84%. Exo A (70%) was the main virulence gene found in the tested strains. Nan 1 gene was identified in 30% of the samples. 82% of MDRPA isolates were found. There is indeed a relationship between biofilm production and drug resistance, as well as the presence of virulence genes (nan 1 and Exo A) were associated with certain patients and burn wounds characteristics as burn size, burn wound depth, length of hospital stays, and socioeconomic status. Conclusions: Correlation of Pseudomonas aeruginosa virulence profiles with burn wounds and patient-related data can be useful in establishing of an appropriate preventive protocol for hospitalized patients with P. aeruginosa burn serious infections. The targeting of these bacterial virulence arsenals is also a promising approach to developing alternative drugs, which act by attenuating the aggressiveness of the pathogen and reducing its potential to cause vigorous infection.

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Complete Genome Sequence of Pseudomonas aeruginosa K34-7, a Carbapenem-Resistant Isolate of the High-Risk Sequence Type 233

Microbiology Resource Announcements ◽

10.1128/mra.00886-18 ◽

2018 ◽

Vol 7 (4) ◽

Cited By ~ 2

Author(s):

George Taiaroa ◽

Ørjan Samuelsen ◽

Tom Kristensen ◽

Ole Andreas Løchen Økstad ◽

Adam Heikal

Keyword(s):

Pseudomonas Aeruginosa ◽

High Risk ◽

Genome Sequence ◽

Complete Genome Sequence ◽

Complete Genome ◽

Sequence Type ◽

Resistant Isolate ◽

Content Type ◽

Carbapenem Resistant ◽

New Antibiotics

Carbapenem-resistant Pseudomonas aeruginosa is defined as a “critical” priority pathogen for the development of new antibiotics. Here we report the complete genome sequence of an extensively drug-resistant, Verona integron-encoded metallo-β-lactamase-expressing isolate belonging to the high-risk sequence type 233.

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