Quinolone-resistant Salmonella Typhi in South Africa, 2003–2007

2009 ◽  
Vol 138 (1) ◽  
pp. 86-90 ◽  
Author(s):  
A. M. SMITH ◽  
N. GOVENDER ◽  
K. H. KEDDY
Keyword(s):  
South Africa ◽  
Amino Acid ◽  
Nalidixic Acid ◽  
Efflux Pump ◽  
Variable Number Tandem Repeat ◽  
Quinolone Resistance ◽  
Efflux Pump Inhibitor ◽  
Pcr Screening ◽  
Active Efflux ◽  
Amino Acid Mutations

SUMMARYIn South Africa, for the years 2003–2007, the Enteric Diseases Reference Unit received 510 human isolates of Salmonella Typhi, of which 27 were nalidixic acid-resistant [minimum inhibitory concentrations (MICs) 128–512 μg/ml] with reduced susceptibility to ciprofloxacin (MICs 0·125–0·5 μg/ml). Pulsed-field gel electrophoresis analysis of 19 available isolates differentiated them into five DNA pattern types; multiple-locus variable-number tandem repeat analysis differentiated the isolates into 10 types. This level of genetic diversity suggested that resistant strains usually emerged independently of one another. A 16- to 32-fold decrease in nalidixic acid MIC and a 2- to 8-fold decrease in ciprofloxacin MIC, was observed in the presence of an efflux pump inhibitor. All isolates were negative by PCR screening for qnr genes. Seven resistant isolates were further analysed for mutations in the quinolone resistance-determining region of gyrA, gyrB, parC and parE. No amino-acid mutations were identified in GyrB and ParE; all isolates showed amino-acid mutations in both GyrA and ParC. We conclude that amino-acid mutations in GyrA and ParC in combination with active efflux of antibiotic out of the bacterial cell are the probable mechanisms conferring quinolone resistance.

scholarly journals In Vitro Activities of Six Quinolones and Mechanisms of Resistance in Staphylococcus aureus and Coagulase-Negative Staphylococci

2001 ◽  
Vol 45 (5) ◽  
pp. 1553-1557 ◽  
Author(s):  
Hans-Jörg Linde ◽  
Mario Schmidt ◽  
Emmi Fuchs ◽  
Udo Reischl ◽  
Hans-Helmut Niller ◽  
...  
Keyword(s):  
Staphylococcus Aureus ◽  
Amino Acid ◽  
Nalidixic Acid ◽  
Quinolone Resistance ◽  
Amino Acid Substitutions ◽  
Mechanisms Of Resistance ◽  
Coagulase Negative Staphylococci ◽  
Active Efflux ◽  
Resistance Phenotype

ABSTRACT Of 94 clinical isolates of Staphylococcus aureus(n = 51) and coagulase-negative staphylococci (CNS) (n = 43), mutations in the quinolone resistance-determining region of topoisomerases GrlA, GrlB, GyrA, and GyrB together with MICs of six quinolones were analyzed. Amino acid substitutions at identical residues (GrlA residues 80 and 84; GyrA residues 84 and 88) were found in S. aureus and CNS. Active efflux, as suggested by blocking by reserpine, contributed substantially to the resistance phenotype in some strains. Among ciprofloxacin, clinafloxacin, levofloxacin, nalidixic acid, trovafloxacin, and sparfloxacin, a 0.5-μg/ml concentration of sparfloxacin discriminated best between strains with two or three mutations and those with no mutations.

The mycobacterial efflux pump EfpA can induce high drug tolerance to many anti-tuberculosis drugs, including moxifloxacin, in Mycobacterium smegmatis

2021 ◽  
Author(s):  
Deepika Rai ◽  
Sarika Mehra
Keyword(s):  
Mycobacterium Smegmatis ◽  
Efflux Pump ◽  
Drug Tolerance ◽  
Inducible Promoter ◽  
Major Facilitator Superfamily ◽  
Expression Level ◽  
Efflux Pump Inhibitor ◽  
Gene Encoding ◽  
Active Efflux ◽  
Major Facilitator

Active efflux of drugs across the membrane is a major survival strategy of bacteria against many drugs. In this work, we characterize an efflux pump EfpA, from the major facilitator superfamily, that is highly conserved among both slow growing and fast-growing mycobacterium species and has been found to be upregulated in many clinical isolates of Mycobacterium tuberculosis . The gene encoding EfpA from Mycobacterium smegmatis was over-expressed under both constitutive and an inducible promoter. Expression of efpA gene under both the promoters resulted in greater than 32-fold increased drug tolerance of M. smegmatis cells to many first-line (rifampicin, isoniazid and streptomycin) and second-line (amikacin) anti-tuberculosis drugs. Notably, drug tolerance of M. smegmatis cells to moxifloxacin increased by more than 180-fold when efpA was over-expressed. The increase in minimum inhibitory concentration (MIC) correlated with the decreased uptake of drugs including norfloxacin, moxifloxacin and ethidium bromide and the high MIC could be reversed in the presence of an efflux pump inhibitor. A correlation was observed between the MIC of drugs and the efflux pump expression level, suggesting that the latter could be modulated by varying the expression level of the efflux pump. The expression of high levels of efpA did not impact the fitness of the cells when supplemented with glucose.The efpA gene is conserved across both pathogenic and non-pathogenic mycobacteria. The efpA gene from the Mycobacterium bovis BCG/ M. tuberculosis , which is 80% identical to efpA from M. smegmatis , also led to decreased antimicrobial efficacy to many drugs, although the fold-change was lower. When over-expressed in M. bovis BCG, an 8-fold higher drug tolerance to moxifloxacin was observed . This is the first report of an efflux pump from mycobacterium species that leads to higher drug tolerance to moxifloxacin, a promising new drug for the treatment of tuberculosis.

scholarly journals Mechanisms of Fluoroquinolone Resistance in Escherichia coli Isolates from Food-Producing Animals

2011 ◽  
Vol 77 (20) ◽  
pp. 7113-7120 ◽  
Author(s):  
Maria Karczmarczyk ◽  
Marta Martins ◽  
Teresa Quinn ◽  
Nola Leonard ◽  
Séamus Fanning
Keyword(s):  
Escherichia Coli ◽  
Nalidixic Acid ◽  
Target Genes ◽  
Efflux Pump ◽  
Multidrug Resistant ◽  
Fluoroquinolone Resistance ◽  
Efflux Pump Inhibitor ◽  
Content Type ◽  
Field Isolates ◽  
K 12

ABSTRACTEleven multidrug-resistantEscherichia coliisolates (comprising 6 porcine and 5 bovine field isolates) displaying fluoroquinolone (FQ) resistance were selected from a collection obtained from the University Veterinary Hospital (Dublin, Ireland). MICs of nalidixic acid and ciprofloxacin were determined by Etest. All showed MICs of nalidixic acid of >256 μg/ml and MICs of ciprofloxacin ranging from 4 to >32 μg/ml. DNA sequencing was used to identify mutations within the quinolone resistance-determining regions of target genes, and quantitative real-time PCR (qRT-PCR) was used to evaluate the expression of the major porin, OmpF, and component genes of the AcrAB-TolC efflux pump and its associated regulatory loci. Decreased MIC values to nalidixic acid and/or ciprofloxacin were observed in the presence of the efflux pump inhibitor phenylalanine-arginine-β-naphthylamide (PAβN) in some but not all isolates. Several mutations were identified in genes coding for quinolone target enzymes (3 to 5 mutations per strain). All isolates harbored GyrA amino acid substitutions at positions 83 and 87. Novel GyrA (Asp87 → Ala), ParC (Ser80 → Trp), and ParE (Glu460 → Val) substitutions were observed. The efflux activity of these isolates was evaluated using a semiautomated ethidium bromide (EB) uptake assay. Compared to wild-typeE. coliK-12 AG100, isolates accumulated less EB, and in the presence of PAβN the accumulation of EB increased. Upregulation of theacrBgene, encoding the pump component of the AcrAB-TolC efflux pump, was observed in 5 of 11 isolates, while 10 isolates showed decreased expression of OmpF. This study identified multiple mechanisms that likely contribute to resistance to quinolone-based drugs in the field isolates studied.

scholarly journals Evidence for Active Efflux as the Primary Mechanism of Resistance to Ciprofloxacin in Salmonella enterica Serovar Typhimurium

2000 ◽  
Vol 44 (5) ◽  
pp. 1223-1228 ◽  
Author(s):  
Etienne Giraud ◽  
Axel Cloeckaert ◽  
Dominique Kerboeuf ◽  
Elisabeth Chaslus-Dancla
Keyword(s):  
Outer Membrane ◽  
Salmonella Enterica ◽  
Efflux Pump ◽  
Enzyme Linked Immunosorbent Assay ◽  
Topoisomerase Iv ◽  
Efflux Pump Inhibitor ◽  
Active Efflux ◽  
Fluorimetric Method ◽  
Serovar Typhimurium ◽  
Acrab Efflux Pump

ABSTRACT The occurrence of active efflux and cell wall modifications were studied in Salmonella enterica serovar Typhimurium mutants that were selected with enrofloxacin and whose phenotypes of resistance to fluoroquinolones could not be explained only by mutations in the genes coding for gyrase or topoisomerase IV. Mutant BN18/21 exhibited a decreased susceptibility to ciprofloxacin (MIC = 0.125 μg/ml) but did not have a mutation in the gyrA gene. Mutants BN18/41 and BN18/71 had the same substitution, Gly81Cys in GyrA, but exhibited different levels of resistance to ciprofloxacin (MICs = 2 and 8 μg/ml, respectively). None of the mutants had mutations in the parC gene. Evidence for active efflux was provided by a classical fluorimetric method, which revealed a three- to fourfold decrease in ciprofloxacin accumulation in the three mutants compared to that in the parent strain, which was annuled by addition of the efflux pump inhibitor carbonyl cyanide m-chlorophenylhydrazone. In mutant BN18/71, a second fluorimetric method also showed a 50% reduction in the level of accumulation of ethidium bromide, a known efflux pump substrate. Immunoblotting and enzyme-linked immunosorbent assay experiments with an anti-AcrA antibody revealed that the resistance phenotype was strongly correlated with the expression level of the AcrAB efflux pump and suggested that decreased susceptibility to ciprofloxacin due to active efflux probably related to overproduction of this pump could occur before that due to gyrA mutations. Alterations were also found in the outer membrane protein and lipopolysaccharide profiles of the mutants, and these alterations were possibly responsible for the decrease in the permeability of the outer membrane that was observed in the mutants and that could act synergistically with active efflux to decrease the level of ciprofloxacin accumulation.

scholarly journals Genetic determinants of multi-drug resistance in Acinetobacter baumannii at an academic hospital in Pretoria, South Africa

2019 ◽  
Author(s):  
Noel-David Nogbou ◽  
Dikwata Thabiso Phofa ◽  
Maphoshane Nchabeleng ◽  
Andrew Munyalo Musyoki
Keyword(s):  
South Africa ◽  
Antimicrobial Resistance ◽  
Genetic Relatedness ◽  
Efflux Pump ◽  
Multi Drug Resistance ◽  
Drug Resistant ◽  
Genetic Determinants ◽  
Pump System ◽  
Active Efflux ◽  
Almost All

AbstractAntimicrobial resistance is now globally recognised as the greatest threat to human health. Acinetobacter baumanniis’ (A. baumannii) clinical significance has been driven by its ability to obtain and transmit antimicrobial resistance factors. In South Africa, A. baumannii is a leading cause of healthcare associated infections (HAI). In this study, we investigated the genetic determinants of multi-drug resistant A. baumannii (MDRAB) at a teaching hospital in Pretoria, South Africa.One hundred non repetitive isolates of A. baumannii were collected for the study at Dr George Mukhari Tertiary Laboratory (DGMTL). Antimicrobial susceptibility testing was performed using the VITEK2 system (bioMerieux, France). The prevalence of common resistance associated genes and AdeABC efflux pump system associated genes were investigated using conventional PCR. Genetic relatedness of isolates was then determined using rep-PCR.Seventy (70) of 100 isolates collected were confirmed to be multi-drug resistant and were blaOXA51 positive. Phenotypically, the isolates where resistant to almost all tested antibiotics. However, one isolate showed intermediate susceptibility to tigecycline while all were susceptible to colistin. Oxacillinase encoding gene blaOXA-23 was the most detected at 99% and only 1% was positive for blaOXA-40. The PCR results for metallo-betalactamase (MBL) encoding genes showed that MBL blaVIM was the most frequently detected at 86% and blaSIM-1 at 3% was the least detected. Out of 70 isolates, 56 isolates had the required gene combination for an active efflux pump. The most prevalent clone was clone A at 69% of the isolates. Regarding treatment; colistin and tigecycline are the most effective against strains encountered at DGMTL as all tested carbapenems seem to have lost their effectiveness.The major genotypic determinants for drug resistances are oxacillinases: blaOXA-51 (100%) and blaOXA-23 (99%). The study reports for the first time, blaOXA-40 and blaSIM-1 detection in A. baumannii in South Africa.

scholarly journals In VitroSusceptibility of Burkholderia vietnamiensis to Aminoglycosides

2011 ◽  
Vol 55 (5) ◽  
pp. 2256-2264 ◽  
Author(s):  
Agatha N. Jassem ◽  
James E. A. Zlosnik ◽  
Deborah A. Henry ◽  
Robert E. W. Hancock ◽  
Robert K. Ernst ◽  
...  
Keyword(s):  
Burkholderia Cepacia ◽  
Efflux Pump ◽  
Severe Disease ◽  
Resistant Isolate ◽  
Aminoglycoside Resistance ◽  
Efflux Pump Inhibitor ◽  
Content Type ◽  
Active Efflux ◽  
Burkholderia Vietnamiensis

ABSTRACTBurkholderia cepaciacomplex (BCC) bacteria are opportunistic pathogens that can cause severe disease in cystic fibrosis (CF) patients and other immunocompromised individuals and are typically multidrug resistant. Here we observed that unlike other BCC species, most environmental and clinicalBurkholderia vietnamiensisisolates were intrinsically susceptible to aminoglycosides but not to cationic antimicrobial peptides or polymyxin B. Furthermore, strains acquired aminoglycoside resistance during chronic CF infection, a phenomenon that could be induced under tobramycin or azithromycin pressurein vitro. In comparing susceptible and resistantB. vietnamiensisisolates, no gross differences in lipopolysaccharide structure were observed, all had lipid A-associated 4-amino-4-deoxy-l-arabinose residues, and all were resistant to the permeabilizing effects of aminoglycosides, a measure of drug entry via self-promoted uptake. However, susceptible isolates accumulated 5 to 6 times more gentamicin than a resistant isolate, and aminoglycoside susceptibility increased in the presence of an efflux pump inhibitor.B. vietnamiensisis therefore unusual among BCC bacteria in its susceptibility to aminoglycosides and capacity to acquire resistance. Aminoglycoside resistance appears to be due to decreased cellular accumulation as a result of active efflux.

scholarly journals Natural Antibiotic Resistance of Bacteria Isolated from Larvae of the Oil Fly, Helaeomyia petrolei

2000 ◽  
Vol 66 (11) ◽  
pp. 4615-4619 ◽  
Author(s):  
Dana R. Kadavy ◽  
Jacob M. Hornby ◽  
Terry Haverkost ◽  
Kenneth W. Nickerson
Keyword(s):  
Antibiotic Resistance ◽  
Los Angeles ◽  
Nalidixic Acid ◽  
Antimicrobial Agents ◽  
Efflux Pump ◽  
Gut Contents ◽  
Solvent Tolerance ◽  
Bacterial Strains ◽  
Active Efflux ◽  
Mutator Strain

ABSTRACT Helaeomyia petrolei (oil fly) larvae inhabit the asphalt seeps of Rancho La Brea in Los Angeles, Calif. The culturable microbial gut contents of larvae collected from the viscous oil were recently examined, and the majority (9 of 14) of the strains were identified as Providencia spp. Subsequently, 12 of the bacterial strains isolated were tested for their resistance or sensitivity to 23 commonly used antibiotics. All nine strains classified as Providencia rettgeri exhibited dramatic resistance to tetracycline, vancomycin, bacitracin, erythromycin, novobiocin, polymyxin, colistin, and nitrofurantoin. Eight of nineProvidencia strains showed resistance to spectinomycin, six of nine showed resistance to chloramphenicol, and five of nine showed resistance to neomycin. All 12 isolates were sensitive to nalidixic acid, streptomycin, norfloxacin, aztreonam, cipericillin, pipericillin, and cefotaxime, and all but OF008 (Morganella morganii) were sensitive to ampicillin and cefoxitin. The oil fly bacteria were not resistant to multiple antibiotics due to an elevated mutation rate. For each bacterium, the number of resistant mutants per 108cells was determined separately on rifampin, nalidixic acid, and spectinomycin. In each case, the average frequencies of resistant colonies were at least 50-fold lower than those established for known mutator strain ECOR 48. In addition, the oil fly bacteria do not appear to excrete antimicrobial agents. When tested, none of the oil fly bacteria produced detectable zones of inhibition on Pseudomonas aeruginosa, Escherichia coli, Staphylococcus aureus, or Candida albicans cultures. Furthermore, the resistance properties of oil fly bacteria extended to organic solvents as well as antibiotics. When pre-exposed to 20 μg of tetracycline per ml, seven of nine oil fly bacteria tolerated overlays of 100% cyclohexane, six of nine tolerated 10% xylene, benzene, or toluene (10:90 in cyclohexane), and three of nine (OF007, OF010, and OF011) tolerated overlays of 50% xylene–50% cyclohexane. The observed correlation between antibiotic resistance and organic solvent tolerance is likely explained by an active efflux pump that is maintained in oil fly bacteria by the constant selective pressure of La Brea's solvent-rich environment. We suggest that the oil fly bacteria and their genes for solvent tolerance may provide a microbial reservoir of antibiotic resistance genes.

Molecular Characterization of Fluoroquinolone Resistance Mechanisms of Campylobacter Isolates from Duck Meats

2017 ◽  
Vol 80 (12) ◽  
pp. 2056-2059
Author(s):  
Min Kang ◽  
Bai Wei ◽  
Sung-Woon Choi ◽  
Se-Yeoun Cha ◽  
Hyung-Kwan Jang
Keyword(s):  
Nalidixic Acid ◽  
Efflux Pump ◽  
Resistance Mechanisms ◽  
Quinolone Resistance ◽  
Fluoroquinolone Resistance ◽  
Gyra Gene ◽  
Meat Samples ◽  
High Level ◽  
Level Resistance

ABSTRACT The purpose of this study was to identify the molecular basis of quinolone resistance of Campylobacter isolates recovered from duck meats. Sixty-one isolates from duck meat samples were studied using sequence analysis of the gyrA gene, and PCR assays were used to identify the presence of the CmeABC efflux pump and its restored sensitivity in the presence of efflux-pump inhibitors. High-level resistance to nalidixic acid and ciprofloxacin was attributed to amino acid substitutions Thr-86-Ile in some isolates. The PCR assay confirmed the presence of the cmeB gene in 29 (47.5%) of the 61 Campylobacter isolates. Phenylalanine arginine β-naphthylamide reduced the MICs of ciprofloxacin and nalidixic acid in 16 (55.2%) and 26 (89.7%) isolates, respectively. The Thr-86-Ile substitution in the gyrA was the primary contributor to the high-level quinolone resistance in Campylobacter isolates from duck meats.

Prevalence and characterization of quinolone resistance in Laribacter hongkongensis from grass carp and Chinese tiger frog

2013 ◽  
Vol 62 (10) ◽  
pp. 1559-1564 ◽  
Author(s):  
Ding-Qiang Chen ◽  
Ling Yang ◽  
Yu-Ting Luo ◽  
Min-Jie Mao ◽  
Yong-Ping Lin ◽  
...  
Keyword(s):  
Molecular Mechanisms ◽  
Efflux Pump ◽  
Quinolone Resistance ◽  
Efflux Pump Inhibitor ◽  
Electrophoretic Patterns ◽  
Food Borne ◽  
Laribacter Hongkongensis ◽  
Resistant Strains

Laribacter hongkongensis is a food-borne bacterium associated with community-acquired gastroenteritis and diarrhoea. Quinolone resistance was recently reported in bacterial isolates from aquatic products, but the molecular mechanisms for resistance were still unknown. In this study, a total of 157 L. hongkongensis strains were isolated from grass carps (n = 443) and Chinese tiger frogs (n = 171). Twenty-one ciprofloxacin-resistant strains were analysed for mutations in quinolone resistance-determining regions (QRDR), acquired quinolone resistance (AQR) genes and the role of efflux pumps in resistance. All QRDR mutations in gyrA (codons 85 and 89) and parC (codons 83 and 231) were found to be closely associated with ciprofloxacin resistance. The AQR gene aac(6′)-Ib-cr was found in 42.9 % (9/21) of the resistant strains, but qnrA, qnrB, qnrC, qnrD, qnrS and qepA were not detected. No significant change of MICs to ciprofloxacin was observed in the presence of an efflux pump inhibitor, indicating the role of efflux pump was probably absent. All 21 ciprofloxacin-resistant strains showed different electrophoretic patterns, which suggested they were not genetically related. These data highlight the importance of QRDR mutations and the AQR gene aac(6′)-Ib-cr during the development of quinolone resistance in a heterogeneous population of L. hongkongensis.

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