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Non-fermenting Gram-negative bacteria (NFGNB) are increasingly cultured in respiratory samples from cystic fibrosis (CF) patients. This study determined the antimicrobial susceptibility of clinical CF respiratory isolates from distinct geographical regions. A total of 286 isolates (106 Stenotrophomonas maltophilia, 100 Burkholderia spp., 59 Achromobacter spp., 12 Pandoraea spp., 9 Ralstonia spp.) from the Netherlands, Northern Ireland, Spain, USA and Australia were tested. MIC 50/90 values and susceptibility categorisation were determined. Trimethoprim/sulfamethoxazole (SXT) was the most active compound for all micro-organisms (MIC 50 , 0.12-4 mg/L; MIC 90 , 1-16 mg/L). For S. maltophilia, 47% and 62% of isolates were susceptible to SXT according to CLSI and EUCAST breakpoints, respectively. Ceftazidime presented lower susceptibility (35%; MIC 50 , 32 mg/L; MIC 90 , 256 mg/L). MIC 90 values for tobramycin and colistin were >128 mg/L and >16 mg/L, respectively. Regarding Burkholderia, 72%, 56% and 44% were susceptible to SXT, ceftazidime and meropenem, respectively. For both ceftazidime and meropenem, MIC 50 and MIC 90 values were within the intermediate or resistant category. The most active antibiotics for Achromobacter spp. were SXT (MIC 50 , 0.5 mg/L; MIC 90 , 8 mg/L) and imipenem (MIC 50 , 2 mg/L; MIC 90 , 8 mg/L). SXT, imipenem and ciprofloxacin were active against 12 Pandoraea spp. (MIC 50 , 0.12-4 mg/L; MIC 90 , 1-8 mg/L). Ciprofloxacin (MIC 50 , 4 mg/L) and SXT (MIC 50 , 1 mg/L) were the only active antibiotics for Ralstonia spp. There were no statistically significant differences in susceptibility rates between countries. NFGNB other than Pseudomonas aeruginosa are potential pathogens in CF. SXT was demonstrated to be the most active compound against these isolates., (Copyright © 2018 Elsevier B.V. and International Society of Chemotherapy. All rights reserved.)

Worldwide, resistance of Gram-negative micro-organisms to third-generation cephalosporins and carbapenems owing to β-lactamases is an increasing problem. Although the CTX-M, TEM and SHV extended-spectrum β-lactamases (ESBLs) are most widely disseminated, other β-lactamase families have also recently emerged, such as plasmid-mediated AmpC β-lactamases and carbapenemases. Here we describe a new set of multiplex polymerase chain reactions (PCRs) with one amplification protocol enabling detection of 25 prevalent β-lactamase families, including ESBLs, carbapenemases, plasmid-mediated AmpC β-lactamases and OXA β-lactamases., (Copyright © 2011 Elsevier B.V. and the International Society of Chemotherapy. All rights reserved.)

The objective of this study was to investigate the antimicrobial resistance patterns, integron characteristics and gene cassettes as well as the presence of Salmonella genomic island 1 (SGI1) in non-typhoidal Salmonella (NTS) isolates from human and animal origin. Epidemiologically unrelated Dutch NTS strains (n=237) originating from food-producing animals and human cases of salmonellosis were tested for their susceptibility to 15 antimicrobial agents. Resistance to 14 of these antimicrobials, including the third-generation cephalosporins, was detected. Resistance to sulphonamides, ampicillin, tetracycline, streptomycin, trimethoprim and nalidixic acid was common (>/=10% of the strains were resistant). Resistance against three or more antimicrobials was observed in 57 isolates. The same 237 strains were studied for the prevalence of class 1 integrons, their gene cassettes and the presence of SGI1. Thirty-six isolates (15.2%) carried class 1 integrons. These integrons had ten distinct profiles based on the size of the integron and restriction fragment length polymorphism analysis. Integrons were detected for the first time in serovars Indiana and Senftenberg. Multidrug resistance was strongly associated with the presence of class 1 integrons in which the aadA2, aadA1, bla(PSE-1), dfrA1, dfrA5, dfrA14 or sat genes were present, as determined by nucleotide sequence determination. The presence of gene cassettes or combinations of gene cassettes not previously found in integrons in Salmonella was observed. SGI1 or its variants (SGI-B, -C and -F) were present in 16 isolates belonging to either serovar Typhimurium, Derby or Albany. Regardless of whether the isolate was of human or animal origin, the same resistance phenotype, integron profile and SGI1 structure could be observed.

In vitro susceptibility to 15 beta-lactam antibiotics was evaluated using Enterobacteriaceae isolated during the SENTRY Antimicrobial Surveillance Program. Piperacillin/tazobactam was the most active penicillin against Escherichia coli, Proteus mirabilis, Klebsiella oxytoca and Klebsiella pneumoniae (94.9%, 98.3%, 87.4% and 82.9% of isolates susceptible). Of the cephalosporins, cefepime was most effective against Escherichia coli, Proteus mirabilis and Enterobacter cloacae (99.2%, 96.3% and 95.2% of isolates susceptible, respectively) and cefoxitin against Klebsiella oxytoca and Klebsiella pneumoniae (98.6% and 95.6% of isolates susceptible). Carbapenems had excellent activity (> or =99.5% of all isolates). ESBL-production was confirmed with the ESBL-Etest and disk diffusion test in 1.3% of Escherichia coli isolates, 18.4% of Klebsiella pneumoniae, 12.6% of Klebsiella oxytoca and 5.3% of Proteus mirabilis isolates.

Bacterial spread between patients may contribute to the high prevalence of antibiotic-resistant pathogens within ICUs. The aim of this study was to evaluate the fate of Pseudomonas aeruginosa during the different antibiotic regimens. Susceptibility patterns and genotyping were performed to determine whether there was a predominant clone and to track the spread of resistant strains within the unit. Twenty-eight different ribotypes were found among 82 Pseudomonas isolates. Four ribotypes accounted for 42 (51%) isolates and were designated the "major clones" occurring throughout multiple cycles. The ribotypes with multiple occurrences were more resistant to antibiotics than ribotypes that appeared only once. The correlation of antibiotic use with antibiotic resistance and the finding of a large number of ribotypes suggested that de novo development of antibiotic resistance is a likely event in P. aeruginosa. In addition, ribotypes associated with antibiotic resistance appeared to have a survival advantage and can become frequent colonizers in the ICU.

Antibiotic resistance is a major and well-known problem in intensive care units (ICUs) world-wide and previously susceptible isolates become resistant through the acquisition of resistance determinants from other bacteria or the development of mutations, as is the case in beta-lactam resistance. We evaluated the presence of resistance determinants involved in beta-lactam resistance and multi-resistance in order to establish the contribution of horizontal gene transfer to the spread of resistance in a surgical ICU during an antibiotic rotation study. Pseudomonas aeruginosa and Enterobacteriaceae isolates were selected and iso-electric focusing (IEF), DNA-typing methods such as specific beta-lactamase and specific integron PCRs were performed to determine the presence of beta-lactamases. The PCRs specific for IMP-1, OXA-1, and VIM-type beta-lactamases performed on the selected P. aeruginosa and Enterobacteriaceae isolates with MICs for cephalosporins >1 mg/l did not demonstrate any of these beta-lactamases. IEF for 14 pseudomonads, representing 7 genotypes from 9 patients, showed a beta-lactamase with a pI larger than 8.5 in 13 of the isolates. The integrase PCR was positive for only five isolates from three patients and conserved segment PCR showed integrons of variable sizes (700, 900, 1,400 and 1,500 bp). Each patient had its own integron types. It can be concluded that integrons and associated resistance determinants played only a minor role in the surgical ICU and beta-lactam resistance among P. aeruginosa isolates was most likely due to the derepression of its AmpC gene.

The objective of this study was to determine the activity of daptomycin, a novel lipopeptide, against European Gram-positive isolates (n = 1539). The MIC(90)-values of daptomycin against Staphylococcus aureus isolates was 0.25 mg/L, against Enterococcus faecalis 4 mg/L, against Enterococcus faecium 8 mg/L, 0.25 mg/L against Staphylococcus epidermidis, and 0.25mg/L against Streptococcus pneumoniae. Daptomycin was equally potent against antibiotic-susceptible and resistant strains within a particular species. Based on a breakpoint of 1 mg/L for S. aureus and group A streptococci, all isolates tested were susceptible to daptomycin. Based on a breakpoint of 4 mg/L for vancomcyin-susceptible E. faecalis 99.7% of these isolates were susceptible to daptomycin.

The target enzymes GyrA and ParC and two efflux pump regulatory genes mexR and nfxB were analysed to determine changes associated with fluoroquinolone resistance in Pseudomonas aeruginosa. Both low- and high-level ciprofloxacin resistance was associated with a Thr-83Ile substitution in GyrA. A ParC Ser-80Leu substitution was found in highly resistant isolates in tandem with the Thr-83Ile substitution in GyrA. Mutations in the efflux regulatory genes were associated with resistance only when in tandem with a mutation in GyrA or ParC. These data show that the main mechanism of fluoroquinolone resistance in P. aeruginosa is mediated primarily through mutations in GyrA, and that mutations in ParC and the efflux regulatory genes are secondary.

Streptococcus pneumoniae isolates (n=1191) were collected during a 1997-1999 European surveillance study. In addition to susceptibility data, a molecular epidemiological survey of their mechanisms of resistance to macrolides, tetracyclines, and quinolones was provided. Of the isolates tested, 72.6% were penicillin-susceptible, 19.9% penicillin-intermediate and 7.5% penicillin-resistant. There was an obvious relationship between resistance to penicillin and resistance to erythromycin (19% of all isolates), clindamycin (14%) and tetracycline (23%). Only one isolate was resistant to levofloxacin. Seventy-three percent of the European S. pneumoniae isolates resistant to erythromycin (n=229) carried the erm(B) gene, while the remaining 27% possessed the mef(A) gene. No mutations were detected in 23S rRNA or in ribosomal proteins L4 and L22. All tetracycline-resistant isolates (n=277) carried the tet(M) gene; none carried the tet(O) gene. Classical mutations in gyrA (Ser 81-Phe or Tyr) and parC (Ser 79-Phe and Asp 83-Asn) and efflux contributed to the decreased quinolone susceptibility. This study of recent European S. pneumoniae isolates can be used to recognize any changes in susceptibility patterns and resistance mechanisms that may occur in the future.

Cross-resistance and multi-resistance to selected antibiotics was determined for Escherichia coli, Klebsiella pneumoniae, Enterobacter cloacae, Pseudomonas aeruginosa, Staphylococcus aureus, Staphylococcus epidermidis, Enterococcus faecalis, Streptococcus pneumoniae, Haemophilus influenzae and Moraxella catarrhalis. Amikacin-resistant Enterobacteriaceae often showed cross-resistance to ss-lactam antibiotics. Only 1% of the Escherichia coli isolates showed resistance to more than four antibiotics from a set of seven. This rate was higher for other Enterobacteriaceae and there were high levels of cross-resistance for P. aeruginosa. The cross-resistance of oxacillin with other antibiotics is well known in staphylococci. Penicillin-resistant pneumococcal isolates were cross-resistant to macrolides. Cross-resistance was only a minor problem in H. influenzae and M. catarrhalis. Cross- and multi-resistance are important problems for Gram-negative and Gram-positive bacteria but not for fastidious bacteria with the exception of penicillin-resistant S. pneumoniae.

The in-vitro activities of gatifloxacin, trovafloxacin, levofloxacin, sparfloxacin, ofloxacin, and ciprofloxacin were tested against 9,682 clinical bacterial isolates from 20 European university hospitals participating in the European SENTRY surveillance programme. Gatifloxacin and trovafloxacin exhibited the highest activities against gram-positive cocci, while levofloxacin, ofloxacin, ciprofloxacin, and gatifloxacin were the most active against Enterobacteriaceae. Ciprofloxacin and levofloxacin showed the highest antimicrobial activities against Pseudomonas spp., while gatifloxacin and trovafloxacin were the most active against Acinetobacter spp. and Stenotrophomonas maltophilia. All Haemophilus spp. and Moraxella catarrhalis isolates were fully susceptible to all quinolones tested. Overall, the new quinolones, showed improved activity against gram-positive cocci and gram-negative non-fermenters while retaining their broad-spectrum activity against gram-negative bacilli.

The past 15 years the field of molecular biology and especially DNA technology has developed rapidly. This did not leave microbiology unaffected. DNA sequencing and the use of DNA probes led to new insights in the evolution and spread of antibiotic resistance genes. It became clear that resistance determinants often show partial homology even when species are not closely related. DNA probes have established their value as epidemiological tools and currently efforts are being made to introduce them into routine diagnostics.