Detection of resistance genes from Pseudomonas aeruginosa using immunomagnetic isolation and chemiluminescence

2020 ◽  
Vol 10 (3) ◽  
pp. 412-418
Author(s):  
Fei Xu ◽  
Cheng Chen ◽  
Xing Li ◽  
Bo Zhang
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Resistance Genes ◽  
High Efficiency ◽  
Bacterial Pathogen ◽  
Resistance Mechanisms ◽  
Clinical Samples ◽  
Specific Gene ◽  
Beta Lactamases ◽  
Immunomagnetic Isolation ◽  
Encoding Gene

Pseudomonas aeruginosa (P. aeruginosa) is a common opportunistic and nosocomial bacterial pathogen. Various multi-resistance mechanisms present across numerous P. aeruginosa strains counteract conventional antimicrobial therapy, thereby becoming a great challenge. This study aimed to establish the application of immunomagnetic isolation and chemiluminescence to detect the presence of extended spectra of β-lactamases encoding genes: blaTEM and blaVEB; metallo-beta-lactamases encoding gene: blaVIM; aminoglycoside modifying enzymes encoding gene: aac(6)II, ant(3)I; and the specific gene for P. aeruginosa, gyrB. P. aeruginosa was specifically selected using the immunomagnetic nanoparticles (IMNPs) in the six parallel bacterial plates counting, proving that they are reliable. Then, the high efficiency of IMNPs@Probes in targeting the resistance genes of P. aeruginosa was demonstrated using the results of chemiluminescent intensities of blaTEM, blaVEB, blaVIM aac(6)II, ant(3)I, and gyrB (more than 10 times higher than that of the control). Sixty-eight in situ clinical samples were tested for the presence of these resistance genes, and one more blaTEM and three more blaVIM individuals were detected using this method compared to the traditional PCR. Thus, the application of our method in clinical screening is specific, accurate, and reliable, and it could be useful in the administration of appropriate treatment.

scholarly journals Frequency of MexA gene in pseudomonas aeruginosa isolated from clinical samples of a Tertiary Care Hospital in Pakistan.

2020 ◽  
Vol 27 (11) ◽  
pp. 2389-2393
Author(s):  
Syed Luqman Shuaib ◽  
Amina Gul ◽  
Jawad Ahmed ◽  
Noor Rehman ◽  
Liaqt Ali ◽  
...  
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Resistance Genes ◽  
Efflux Pump ◽  
Tertiary Care Hospital ◽  
Tertiary Care ◽  
Clinical Samples ◽  
Specific Gene ◽  
Care Hospital ◽  
Conventional Pcr ◽  
Isolation And Identification

The drug resistance genes are responsible to preserve the Pseudomonas. This also happens in the case of Mex drug efflux pumps and expression increase of MexA, MexB and OprM genes in Pseudomonas aeruginosa, when grown in sub-inhibitory concentrations of antibiotics. Objectives: This study was designed to detect the MexA gene of Pseudomonas aeruginosa resistant strains in tertiary care hospital, Peshawar. Study Design: Cross-sectional study. Setting: Department of Pathology, Khyber Teaching Hospital, Peshawar, Pakistan. Period: 14 months duration from April 2015 to May 2016. Material & Methods: The specimens including burn wound swabs, pus and urine were obtained from different patients and were processed on blood agar and MacConkey medium for isolation and identification. Conventional PCR was performed for the MexA gene on 50 specimens. Results: The simple conventional PCR was done for MexA (The Mex AB OprM operon resistance genes) and the O-antigen acetylase gene (the species-specific gene) separately, gave positive bands for 49 out of the 50 specimens. Our finding confirms the presence of the MexA gene (and hence most probably MexABOprM operon) in 49 out of 50 specimens of Pseudomonas aeruginosa. Conclusion: Among other resistance mechanisms to antibiotics and disinfectants, the MexABOprM efflux pump might have a role.

scholarly journals Investigation of Ciprofloxacin Resistance and Its Mechanisms in Clinical Pseudomonas aeruginosa Isolates

ANKEM Dergisi ◽  
2021 ◽  
Author(s):  
Nilüfer Uzunbayır Akel ◽  
Yamaç Tekintaş ◽  
Fethiye Ferda Yılmaz ◽  
İsmail Öztürk ◽  
Mustafa Ökeer ◽  
...  
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Efflux Pump ◽  
Resistance Mechanisms ◽  
Regulatory Genes ◽  
Quinolone Resistance ◽  
University Hospital ◽  
Clinical Samples ◽  
Genetic Groups ◽  
Ciprofloxacin Resistance ◽  
Polymerase Chain

Pseudomonas aeruginosa is one of the most important causes of hospital infections. Although different antibiotic groups are used for the treatment of P.aeruginosa infections, quinolone groups are distinguished by the advantages of oral administration. However, in recent years, resistance against members of this group has made treatment more difficult. The aim of this study was to investigate the epidemiological relationship and possible mechanisms of resistance in ciprofloxacin resistant P. aeruginosa isolates from Ege University Hospital. The identification of P.aeruginosa bacteria isolated from clinical samples in Ege University Medical Faculty Medical Microbiology Laboratory was determined by VITEK MS automated systems by VITEK compact, antimicrobial susceptibility. The epidemiological relationships of the ciprofloxacin resistant isolates were determined by Enterobacterial repetitive intergenic consensus-polymerase chain reaction (ERIC-PCR). The presence of qnrA, qnrB, qnrS, qepA genes, the quinolone resistance genes and nfxB, mexR, the regulatory genes of the efflux pump, was determined by PCR. The phenylalanine-arginine β-naphthylamide (PAβN) assay was used to determine the activation of the efflux pump. Twenty-two isolates (26.5 %) were found resistant to ciprofloxacin. According to the ERIC-PCR results, 11 unrelated clones were detected. Ciprofloxacin minimum inhibitory concentration (MIC) values were decreased 2-64 times in 10 isolates in the presence of PAIN. No ciprofloxacin MIC change was detected in one isolate. The presence of pump regulatory genes was determined in 10 of the 11 representative isolates, while only qnrB of the genes associated with quinolone resistance was detected in seven representative isolates. qnrA, qnrS, qepA genes were not detected in any isolate. Ciprofloxacin resistant P.aeruginosa isolates are isolated from our hospital. It is noteworthy that the isolates belonging to different genetic groups are in circulation in clinics. Basic resistance mechanisms are thought to be efflux pumps and qnrB genes.

scholarly journals Evaluation of different phenotypic tests for detection of metallo-β-lactamases in imipenem-resistant Pseudomonas aeruginosa

2017 ◽  
Vol 9 (04) ◽  
pp. 249-253 ◽  
Author(s):  
Rohit Sachdeva ◽  
Babita Sharma ◽  
Rajni Sharma
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Large Scale ◽  
Wide Spectrum ◽  
Carbapenem Resistance ◽  
Resistance Mechanisms ◽  
Clinical Samples ◽  
Resistant Strains ◽  
Synergy Test ◽  
Phenotypic Tests ◽  
Simple Screening

Abstract PURPOSE: Pseudomonas aeruginosa causes a wide spectrum of infections including bacteremia, pneumonia, urinary tract infection, etc., Metallo-beta-lactamase (MBL) producing P. aeruginosa is an emerging threat and cause of concern as they have emerged as one of the most feared resistance mechanisms. This study was designed to know the prevalence of MBL production in P. aeruginosa and to evaluate the four phenotypic tests for detection of MBL production in imipenem-resistant clinical isolates of P. aeruginosa. METHODS: Totally, 800 isolates of P. aeruginosa isolated from various clinical samples were evaluated for carbapenem resistance and MBL production. All imipenem-resistant strains were tested for carabapenemase production by modified Hodge test. Screening for MBL production was done by double-disc synergy test and combined disc test (CDT). Confirmation of MBL production was done by the E-test (Ab BioDisk, Solna, Sweden). RESULTS: Out of the 800 isolates of P. aeruginosa, 250 isolates were found resistant to imipenem. Based on the results of E-test, 147 (18.37%) isolates of P. aeruginosa were positive for MBL production. The CDT has the highest sensitivity and specificity for the detection of MBL production as compared to other tests. CONCLUSION: The results of this study are indicative that MBL production is an important mechanism of carbapenem resistance among P. aeruginosa. Use of simple screening test like CDT will be crucial step toward large-scale monitoring of these emerging resistant determinants. Phenotypic test for MBL production has to be standardized, and all the isolates should be routinely screened for MBL production.

scholarly journals Role of biofilm-specific antibiotic resistance genes PA0756-0757, PA5033 and PA2070 in Pseudomonas aeruginosa

2018 ◽  
Author(s):  
Prasanth Manohar ◽  
Thamaraiselvan Shanthini ◽  
Reethu Ann Philip ◽  
Subramani Ramkumar ◽  
Manali Kale ◽  
...  
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Antibiotic Resistance ◽  
Resistance Genes ◽  
Biofilm Formation ◽  
Tamil Nadu ◽  
Antibiotic Resistance Genes ◽  
Clinical Samples ◽  
Cross Sectional ◽  
Antibiotic Resistant ◽  
Resistant Genes

AbstractTo evaluate the presence of biofilm-specific antibiotic-resistant genes, PA0756-0757, PA5033 and PA2070 in Pseudomonas aeruginosa isolated from clinical samples in Tamil Nadu. For this cross-sectional study, 24 clinical isolates (included pus, urine, wound, and blood) were collected from two diagnostic centers in Chennai from May 2015 to February 2016. Biofilm formation was assessed using microtiter dish biofilm formation assay and minimal inhibitory concentration (MIC) and minimal bactericidal concentrations (MBC) were determined for planktonic and biofilm cells (MBC assay). Further, PCR amplification of biofilm-specific antibiotic resistance genes PA0756-0757, PA5033 and PA2070 were performed. Biofilm formation was found to be moderate/strong in 16 strains. MBC for planktonic cells showed that 4, 7, 10 and 14 strains were susceptible to gentamicin, ciprofloxacin, meropenem and colistin respectively. In MBC assay for biofilm cells (MBC-B), all the 16 biofilm producing strains were resistant to ciprofloxacin and gentamicin whereas nine and four were resistant to meropenem, and colistin respectively. The biofilm-specific antibiotic-resistant genes PA0756-0757 was found in 10 strains, 6 strains with PA5033 and 9 strains with PA2070 that were found to be resistant phenotypically. This study highlighted the importance of biofilm-specific antibiotic resistance genes PA0756-0757, PA5033, and PA2070 in biofilm-forming P. aeruginosa.

scholarly journals Detection of antibiotic resistance genes among multiple drug resistant Pseudomonas aeruginosa strains isolated from clinical sources in selected health institutions in Kwara State.

2021 ◽  
Vol 10 (1) ◽  
pp. 40-48
Author(s):  
O.C. Adekunle ◽  
A. Mustapha ◽  
G. Odewale ◽  
R.O. Ojedele
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Antibiotic Resistance ◽  
Resistance Genes ◽  
Multiple Drug Resistance ◽  
Antibiotic Resistance Genes ◽  
Clinical Samples ◽  
Multiple Drug ◽  
Antibiotic Resistant ◽  
Clinical Specimens ◽  
Health Institutions

Introduction: Pseudomonas aeruginosa (P. aeruginosa) is a frequent nosocomial pathogen that causes severe diseases in many clinical and community settings. The objectives were to investigate the occurrence of multiple antibiotic resistant P. aeruginosa strains among clinical samples and to detect the presence of antibiotic resistance genes in the DNA molecules of the strains.Methods: Clinical specimens were collected aseptically from various human anatomical sites in five selected health institutions within Kwara State, Nigeria. Multiple drug resistance patterns of isolated micro-organisms to different antibiotics were determined using the Bauer Kirby disc diffusion technique. The DNA samples of the multiple resistant P. aeruginosa strains were extracted and subjected to Polymerase Chain Reaction (PCR) for resistance gene determination.Results: A total of 145 isolates were identified as P. aeruginosa from the clinical samples. Absolute resistance to ceftazidime, gentamicin and ceftriaxone was observed while low resistance to ciprofloxacin, piperacillin and imipenem was documented. The prevalence of bla VIM , ,bla CTX-M and blaTEM were 34.4 %, 46.7 % and 16.7 % respectively.Conclusion: This study has shown that there is a high occurrence of metallo â-lactamase- producing and antibiotic-resistant strains of P. aeruginosa in clinical specimens from the studied area. Keywords: Metallo â-lactamase enzyme, P. aeruginosa, clinical samples, antibiotic-resistance genes

scholarly journals In Vitro Activity of the Ultra-Broad-Spectrum Beta-lactamase Inhibitor QPX7728 in Combination with Multiple Beta-Lactam Antibiotics against Pseudomonas aeruginosa

2021 ◽  
Author(s):  
Olga Lomovskaya ◽  
Debora Rubio-Aparicio ◽  
Kirk Nelson ◽  
Dongxu Sun ◽  
Ruslan Tsivkovski ◽  
...  
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Resistance Mechanisms ◽  
Clinical Isolates ◽  
Beta Lactamase ◽  
In Vitro Activity ◽  
Beta Lactam ◽  
Beta Lactams ◽  
Beta Lactamases ◽  
Lactamase Inhibitor

QPX7728 is an ultra-broad-spectrum beta-lactamase inhibitor with potent inhibition of key serine and metallo beta-lactamases. QPX7728 enhances the potency of multiple beta-lactams in beta-lactamase producing Enterobacterales and Acinetobacter spp. In this study we evaluated the in vitro activity of QPX7728 (8 μg/ml) combined with multiple beta-lactams against clinical isolates of Pseudomonas aeruginosa with varying beta-lactam resistance mechanisms. Seven-hundred-ninety clinical isolates were included in this study; 500 isolates, termed a “representative panel”, were selected to be representative the MIC distribution of meropenem (MEM), ceftazidime-avibactam (CAZ-AVI), and ceftolozane-tazobactam (TOL-TAZ) resistance for clinical isolates according to 2017 SENTRY surveillance data (representative panel). An additional 290 selected isolates (“challenge panel”), that were either non-susceptible to MEM or were resistant to TOL-TAZ or CAZ-AVI were also tested; 61 strains carried metallo beta-lactamases (MBLs), 211 strains were defective in the carbapenem porin OprD and 185 strains had the MexAB-OprM efflux pump overproduced based on a phenotypic test. Against the representative panel, susceptibility for all QPX7728/beta-lactam combinations was >90%. For the challenge panel, QPX-ceftolozane (TOL) was the most active combination (78.6% susceptible) followed by equipotent QPX-piperacillin (PIP) and QPX-cefepime (FEP), restoring susceptibility in 70.3% of strains (CLSI breakpoints for the beta-lactam compound alone). For MBL-negative strains, QPX-TOL and QPX-FEP restored the MIC values to susceptibility rates in ∼90% and ∼80% of strains, respectively, vs 68-70% for QPX-MEM and QPX-PIP and 63-65% for TOL-TAZ and CAZ-AVI. For MBL-positive strains, QPX-PIP restored the MIC to susceptibility values for ∼70% of strains vs 2-40% for other combinations. Increased efflux and impaired OprD had varying effect on QPX7728 combination depending on the partner beta-lactam tested. QPX7728 enhanced the potency of multiple beta-lactams against P. aeruginosa, with varying results according to the beta-lactamase production and other intrinsic resistance mechanisms.

scholarly journals Analysis of Class 1 Integrons and Antibiotic Resistance Genes in Pseudomonas aeruginosa Strains from Benin City, Nigeria

2020 ◽  
Vol 24 (4) ◽  
pp. 633-637
Author(s):  
B.O. Isichei-Ukah ◽  
O.I. Enabulele
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Antibiotic Resistance ◽  
Resistance Genes ◽  
Antibiotic Resistance Genes ◽  
Environmental Isolates ◽  
Significance Level ◽  
Beta Lactamases ◽  
Class 1 Integrons ◽  
Benin City ◽  
Class 1

The presence of integrons and antibiotic resistance genes in the genome of Pseudomonas aeruginosa pose a serious problem in the treatment and control of infections caused by this pathogen in hospitals. This study was carried to analyse the presence of class 1 integrons and some antibiotic resistance genes on selected clinical and environmental strains of Pseudomonas aeruginosa. A total of 120 strains were employed for this study.The strains were confirmed using molecular method and species-specific primers targeting the 16S ribosomal ribonucleic acid (rRNA). Polymerase chain reaction (PCR) was used to detect the presence of class 1 integrons and resistance genes using appropriate primers and conditions. The strains were analysed for the presence of the following antibiotic resistance genes - aadA, blaPSE, blaAMPC, blaIMP and tetC encoding  aminoglycosides, betalactamases, metallo-beta-lactamases (MBL) and tetracylines resistance respectively. On screening the isolates for the presence of class 1 integrons, 50/60 (83.3 %) clinical isolates and 46/60 (76.7 %) environmental isolates showed positive results (P > 0.05). In both clinical and environmental isolates, the highest occurring resistance genes were blaAMPC and tetC (encoding beta-lactamases and tetracylines respectively), while the least was observed in blaIMP (encoding metallo-beta-lactamases). In comparison, there was high significance difference (at P<0.01 significance level) in the resistance gene blaPSE between the clinical and environmental strains. The high prevalence of these resistance genes is a great threat in the treatment of Pseudomonas infections. Keywords: Pseudomonas aeruginosa, Resistance genes, Integrons, Beta-lactamases.

scholarly journals Antimicrobial Susceptibility of Bacteroides fragilis Group Isolates to Clindamycin, Tetracycline and Tigecycline, and Their Possession of tet and ermF Genes, which are Responsible for Resistance

2021 ◽  
Author(s):  
Bermal Tekeş ◽  
Semra Eminoğlu ◽  
Elvam Sayın ◽  
Nurver Ülger Toprak
Keyword(s):  
Resistance Genes ◽  
Bacteroides Fragilis ◽  
Resistance Mechanisms ◽  
Clinical Samples ◽  
Starting Point ◽  
Resistance Transfer ◽  
Bacteroides Fragilis Group ◽  
Agar Dilution ◽  
Resistance Rates ◽  
Sterile Body Fluids

Objective: We aimed to determine the resistance of Bacteroides fragilis group (BFG) bacteria to clindamycin, tetracycline and tigecycline and establish the distribution of related resistance genes. Method: In total 82 BFG strains, isolated from different clinical samples between January 2017 and December 2018, were identified by MALDI-TOF MS. Their antimicrobial sensitivities to were determined using agar dilution methodology (CLSI; M11-A7). The tetM, tetQ, tetX, tetX1, tet36, and ermF genes were investigated by PCR. Results: Eighty-two strains of BFG bacteria, isolated from intra-abdominal abscess (n=36), tissue biopsy (n=16), blood (n=14) and other sterile body fluids (n=12), were identified as Bacteroides fragilis (n=48), Bacteroides thetaiotaomicron (n=17), Bacteroides vulgatus (n=5), Bacteroides ovatus (n=4), Bacteroides caccae (n=1), Bacteroides uniformis (n=1) and Parabacteroides distasonis (n=6). The resistance rates to clindamycin, tetracycline and tigecycline were 54.9%, 84.1%, 4.9%, respectively. Non-B. fragilis isolates were more resistant than B. fragilis strains. In total, 57.3% of the isolates were ermF gene positive, while B. thetaiotaomicron had the highest rate (70.6%). The tet gene positivity ranged from 18.8% to 66.7% among species. The tetQ gene positivity was higher than other tet genes. The 92.7% of the isolates were resistant to at least one antibiotic, while 94% had at least one resistance gene. Conclusion: This study provided data on antimicrobial resistance of our BFG isolates to clindamycin, tetracycline and tigecycline and the related resistance genes. However, our information obtained could also be a starting point for further investigation of the antibiotic resistance mechanisms of Bacteroides species, as well as, resistance transfer among BFG isolates and other bacteria.

scholarly journals Prevalence, Risk Factors, and Molecular Epidemiology of Intestinal Carbapenem-Resistant Pseudomonas aeruginosa

2021 ◽  
Author(s):  
Yanyan Hu ◽  
Yan Qing ◽  
Jiawei Chen ◽  
Congcong Liu ◽  
Jiayue Lu ◽  
...  
Keyword(s):  
Risk Factors ◽  
Pseudomonas Aeruginosa ◽  
Gastrointestinal Tract ◽  
Resistance Genes ◽  
Acquired Resistance ◽  
Multidrug Resistant ◽  
Resistance Mechanisms ◽  
Human Gastrointestinal Tract ◽  
Content Type ◽  
Carbapenem Resistant

Pseudomonas aeruginosa may become multidrug-resistant (MDR) due to multiple inherited and acquired resistance mechanisms. The human gastrointestinal tract is known as a reservoir of P. aeruginosa and its resistance genes.

scholarly journals Detection of carbapenem resistance genes in Pseudomonas aeruginosa isolates with several phenotypic susceptibility profiles

CES Medicina ◽  
2018 ◽  
Vol 32 (3) ◽  
pp. 203-214 ◽  
Author(s):  
Sara Morales ◽  
Marlon A Gallego ◽  
Johanna M Vanegas ◽  
J Natalia Jiménez
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Resistance Genes ◽  
Molecular Mechanisms ◽  
Efflux Pump ◽  
Carbapenem Resistance ◽  
Resistance Mechanisms ◽  
Gene Encoding ◽  
Resistant Group ◽  
Susceptibility Profiles ◽  
Phenotypic Susceptibility

Introduction: Pseudomonas aeruginosa display several resistance mechanisms to carbapenems and such variety makes it difficult to infer from the antibiogram. The aim of this study was to determine the carbapenem resistance genes in P. aeruginosa isolates with different profiles of phenotypic susceptibility to these antibiotics. Materials and methods: From a microbial collection of P. aeruginosa isolates from infected patients, 40 isolates with different carbapenem resistance profiles were selected. The carbapenemases genes, and expression of the OprD porin, the MexAB-OprM efflux pump and the β-lactamase AmpC were determined. Results: From a total of 40 isolates evaluted, in 21 (52.5%) any mechanism of resistance evaluated were detected. In the meropenem-resistant group, overexpression of AmpC (n = 1) and decreased expression of MexAB-OprM (n = 2) and OprD (n = 1) were found. A decrease in the expression of MexAB-OprM was observed in imipenem-resistant group (n = 3) and mutations in the gene encoding the OprD porin (n = 1). Finally, the presence of carbapenemases (VIM, n= 3, KPC-2 / VIM, n = 1) was detected in imipenem-meropenem resistant isolates. Conclusion: The phenotypic susceptibility profiles in P. aeruginosa isolates were not explained by the molecular mechanisms explored, with the exception of carbapenemase-producing isolates. These results evidence the complexity of the antibiotic resistance mechanisms involved in this bacterium.

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