Mutations of DNA gyrase and topoisomerase IV in clinical isolates of fluoroquinolone-resistant Proteus mirabilis

The presence of fluoroquinolone resistance-associated mutations within the quinolone resistance-determining region of DNA gyrase and topoisomerase IV was investigated genetically in clinical isolates of Proteus mirabilis recovered from patients with urinay tract infections. Two isolates of fluoroquinolone-resistant P. mirabilis possessed the mutations in GyrA (Ser-83 --> Arg or Ile), GyrB (Ser-464 --> Tyr or Phe) and ParC (Ser-80 --> Ile). A novel mutation with Glu-87 --> Lys in GyrA, where suggested to be responsible for fluoroquinolone resistance, was identified. These results demonstrate that the presence of an additional mutation at Glu-87 in GyrA may contribute to high-level fluoroquinolone resistance, too.     

Molecular surveillance of macrolide, tetracycline and quinolone resistance mechanisms in 1191 clinical European Streptococcus pneumoniae isolates.

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.     

Phenotypic and genotypic analysis of levofloxacin-resistant clinical isolates of Streptococcus pneumoniae collected in 13 countries during 1999-2000

During 1999-2000, 5015 isolates were collected from 13 countries and tested against levofloxacin. Overall, levofloxacin resistance minimum inhibitory concentration (MIC>or =8 mg/l) was found in 40 isolates (0.8%). The highest resistance rates were in Hong Kong (8.0%), China (3.3%) and Spain (1.6%). Levofloxacin retained an MIC(90) of 1 mg/l in all countries. Pulsed-field gel electrophoresis analysis of resistant isolates demonstrated the presence of clones in countries where levofloxacin resistance exceeded 1%, suggesting that the elevated resistance rates could result from resistant clones within participating hospitals. DNA-sequence analysis of the quinolone-resistance-determining regions of gyrA, gyrB, parC and parE genes showed that the most common mutations were in GyrA (Ser81Phe), ParC (Ser79Phe, Lys137Asn) and ParE (Ile460Val), accounting for 40% of the isolates tested. Levofloxacin-resistant isolates were generally non-susceptible to other fluoroquinolones tested. Future studies to characterise resistant isolates by other molecular methods may ensure that the appropriate counter-measures can be taken to control the spread of resistant isolates.     

Activity of moxifloxacin and other quinolones against pneumococci resistant to first-line agents, or with high-level ciprofloxacin resistance

The activity of moxifloxacin and other quinolones was assessed against 288 epidemiologically diverse isolates of Streptococcus pneumoniae, many of them resistant to one or more first-line agents and/or with increased ciprofloxacin resistance (minimum inhibitory concentrations, MICs 16- > 64 mg/l compared with 1-2 mg/l for most isolates). Moxifloxacin and grepafloxacin were the most active quinolone analogues, inhibiting about 90% of the isolates at < or = 1 mg/l, whereas levofloxacin inhibited 64% of isolates at < = 1 mg/l and ciprofloxacin inhibited 42%. Moxifloxacin also was the most active agent against isolates with elevated ciprofloxacin resistance (MIC 16- > 64 mg/l): moxifloxacin MICs of around 4 mg/l were seen for most such isolates, compared with 16-32 mg for levofloxacin and grepafloxacin. The activity of moxifloxacin against pneumococci resistant to one or more first-line agent suggests it will have a useful therapeutic role, although its activity against highly ciprofloxacin resistant isolates seems marginal.     

痰标本中质粒介导喹诺酮耐药基因qnr在肺炎克雷伯菌中的分布特征及耐药分析

目的:了解从痰标本中分离出的肺炎克雷伯菌对16种抗菌药物的耐药性,以及研究由质粒介导的喹诺酮类耐药基因qnr在肺炎克雷伯菌中的存在情况。方法:用PCR及直接测序的方法对135株肺炎克雷伯菌进行qnr基因检测,并用K-B纸片法检测其对16种抗菌药物的体外抗菌活性。另外,用琼脂平皿二倍稀释法检测阳性菌株对左氧氟沙星的MIC值。结果:135株肺炎克雷伯菌中,9株(6.6%)检出qnr基因。阳性菌株均对亚胺培南敏感且对多种抗生素耐药,其中2株qnr阳性菌株对左氧氟沙星敏感。结论:肺炎克雷伯菌中存在质粒介导喹诺酮类耐药基因qnr基因,qnr阳性菌株呈现多重耐药。临床工作中,应加强对耐药基因的监测,降低细菌耐药的发生。     

Alterations in penicillin binding protein gene of Streptococcus pneumoniae and their correlation with susceptibility patterns

Penicillin binding protein (pbp) gene alterations of 328 clinical isolates of Streptococcus pneumoniae were examined for a correlation with their antibiotic-resistance. The frequency of penicillin G (PEN-G) resistance was determined to clarify susceptibility to several antibiotics, namely PEN-G, ampicillin, sulbactam/ampicillin, cefozopram, panipenem (PAPM), clarithromycin (CLR), azithromycin (AZM) and levofloxacin (LVX). Oligonucleotide primers for three pbp genes (pbp1a, pbp2x and pbp2b) were used to detect mutations in pbp. Of the strains, 25.9% were classified as Pen-Gs, 68.0% as Pen-Gir and 6.1% as Pen-Gr. The polymerase chain reaction product for wild-type pbp1a was found in 185 isolates, that for wild-type pbp2x was found in 66 isolates and that for wild-type pbp2b was found in 213 isolates. None of these three genes was detectable in 100 isolates while all of them were detected in 64 isolates (1aw/2xw/2bw). Of those 64 isolates with 1aw/2xw/2bw, the minimum inhibitory concentration (MIC) of PEN-G was < or =0.06 mg/l for 54 isolates and 0.12 mg/l for 10 isolates. Of the 272 strains for which the MIC of PAPM was < or =0.03 mg/l, there were 85 Pen-Gs, 184 Pen-Gir and three Pen-Gr isolates. Three strains for which the MIC of LVX was > or =4.0 mg/l included one Pen-Gs and two Pen-Gir isolates. The MICs of CLR correlated significantly with those of AZM. The MIC of CLR was > or =1 mg/l for 216 isolates, and the MIC of AZM was > or =1 mg/l for 244 of them. These data suggested that PAPM may be effective against S. pneumoniae infection, although acquisition of resistance should be considered. LVX also seemed to be effective against S. pneumoniae.     

In vitro activity of moxifloxacin against recent community-acquired respiratory tract pathogens isolated in France: a national survey

Between February and June 2000, 2345 consecutive strains of Streptococcus pneumoniae, Haemophilus influenzae, Moraxella catarrhalis, Staphylococcus aureus and Klebsiella pneumoniae were isolated from 2088 adult patients suffering from community-acquired respiratory tract infections, in 97 hospital laboratories. Of the 1037 S. pneumoniae isolates, 48.3% were intermediately or highly penicillin resistant. For invasive isolates, the MIC90s of penicillin G, amoxycillin, cefuroxime, ceftriaxone, erythromycin, ofloxacin, ciprofloxacin and moxifloxacin were 2, 2, 4, 0.5, 1024, 2, 2 and 0.25 mg/l, respectively. All but one invasive strain were susceptible to moxifloxacin whereas 97.5% were susceptible to levofloxacin. The MIC90s of clinical isolates with intermediate susceptibility or high resistance to penicillin G, were 2, 2, 4, 1, 1024, 2, 2 and 0.25 mg/l. About 98.1, 97.0, and 83.1% of strains were inhibited by concentrations < or = 1 mg/l of moxifloxacin, levofloxacin and ciprofloxacin, respectively (E-test). Eight of the 1037 S. pneumoniae strains were not susceptible to moxifloxacin and had mutations in gyrA (eight strains), parC (four strains) or parE (three strains). Against H. influenzae (32.7% were beta-lactamase producers) and M. catarrhalis (96.3% were beta-lactamase producers), the MIC90s of moxifloxacin, amoxycillin and co-amoxiclav were 0.094 and 0.125 mg/l, 64 and 8 mg/l, and 1.5 and 0.25 mg/l, respectively. Against oxacillin-susceptible S. aureus and K. pneumoniae, the MIC90s of moxifloxacin were 0.125 and 0.84 mg/l respectively. Moxifloxacin had the highest in vitro activity of all antibiotics tested.     

Selection of a gyrA mutation and treatment failure with gatifloxacin in a patient with Streptococcus pneumoniae with a preexisting parC mutation

An 81-year-old woman had pneumonia caused by Streptococcus pneumoniae (levofloxacin Etest minimum inhibitory concentration [MIC] 1.5 microg/ml) and was treated with intravenous gatifloxacin 200 mg/day. After 3 days of therapy, repeat sputum cultures were positive for S. pneumoniae, which was resistant to levofloxacin (Etest MIC > 32 microg/ml). The isolate obtained before therapy showed a preexisting parC mutation of aspartic acid-83 to asparagine (Asp83-->Asn), and the isolate obtained during therapy showed an acquired gyrA mutation from serine-81 to phenylalanine (Ser81-->Phe) and a second parC mutation from lysine-137 to Asn (Lys137-->Asn). Both isolates were the same strain, as determined with pulsed-field gel electrophoresis. This case demonstrates the potential for resistance to emerge during 8-methoxy fluoroquinolone therapy for fluoroquinolone-susceptible S. pneumoniae with a preexisting parC mutation. Additional clinical failures with a fluoroquinolone may occur unless these first-step parC mutants can be identified to assist clinicians in selecting appropriate antimicrobial therapy.     

Levofloxacin treatment failure in a patient with fluoroquinolone-resistant Streptococcus pneumoniae pneumonia

The frequency of fluoroquinolone-resistant Streptococcus pneumoniae has increased as fluoroquinolone administration for treatment of respiratory tract infections has increased. Levofloxacin treatment failed in a patient who had pneumococcal pneumonia and had received three previous courses of levofloxacin therapy. Susceptibility testing revealed high-level resistance to levofloxacin (minimum inhibitory concentration [MIC] > 32 microg/ml), and cross-resistance to moxifloxacin (MIC 4 microg/ml), trovafloxacin (6 microg/ml), and gatifloxacin (12 microg/ml). Sequencing of the quinolone-resistance determining region revealed a mutation of serine-81 to phenylalanine (Ser81-->Phe) in the gyrA region of DNA gyrase and a Ser79-->Phe mutation in the parC region of topoisomerase IV The patient was treated successfully with intravenous ceftriaxone followed by oral cefprozil. Clinicians must be aware of local resistance patterns and the potential for fluoroquinolone treatment failures in patients with infections caused by S. pneumoniae.     

Sensitivities of ciprofloxacin-resistant Mycobacterium tuberculosis clinical isolates to fluoroquinolones: role of mutant DNA gyrase subunits in drug resistance

Minimum inhibitory concentrations of sitafloxacin, gatifloxacin, moxifloxacin, sparfloxacin, levofloxacin and ciprofloxacin against 59 ciprofloxacin-resistant clinical isolates of Mycobacterium tuberculosis from Japan were determined. The isolates were most susceptible to sitafloxacin and gatifloxacin. To understand better the basis for drug resistance, nucleotide sequences encoding the gyrA and gyrB quinolone resistance-determining region were determined. Predicted amino acid sequences revealed distinct mutational patterns likely to be responsible for fluoroquinolone resistance. Double gyrA mutations as well as mutations in both gyrA and gyrB correlated with increased resistance to all fluoroquinolones.     

Type II topoisomerase mutations in clinical isolates of Enterobacter cloacae and other enterobacterial species harbouring the qnrA gene

The qnr genes are transferable genes that confer low-level quinolone resistance by protection of topoisomerase. The occurrence of mutations in DNA gyrase (gyrA, gyrB) and topoisomerase IV (parC, parE) genes in strains harbouring qnr was investigated in 28 qnrA-positive clinical isolates, among which 7 strains also harboured qnrS. Topoisomerase mutations were found in 25 (89%) of the 28 strains, with at least two mutations (gyrA and parC) in 13 strains and one mutation in 12 strains. Isolates of the Enterobacter cloacae complex were compared with reference strains of the new Enterobacter species. gyrA mutations were found at position 83 (Ser or Thr for Ile, Tyr, Leu or Phe depending on the species), and new gyrB mutations were described (S463A, S464F). qnrA had an additive effect of a 10-fold increase in the minimum inhibitory concentration (MIC) whatever the number of topoisomerase mutations, and qnrS was additive to qnrA with a further 2- to 10-fold increase in the MIC. Comparison of MICs with susceptibility breakpoints showed that strains combining qnrA and topoisomerase mutations were resistant to fluoroquinolones, but the three strains lacking a topoisomerase mutation were susceptible using ciprofloxacin and levofloxacin but not using nalidixic acid or moxifloxacin testing.     

TARGETed surveillance: susceptibility of Streptococcus pneumoniae isolated from community-acquired respiratory tract infections in 2003 to fluoroquinolones and other agents

We assessed antibiotic resistance in Streptococcus pneumoniae collected worldwide in 2003. Resistance to clarithromycin was the highest overall (34.1%) followed by penicillin G (22.1%). Patient age and/or country of origin had the greatest effect on susceptibility. Resistance was highest in children<6 years of age and in patients from South Africa or France. Resistance to penicillin or amoxicillin/clavulanic acid decreased in adults and was low in Germany. Fluoroquinolone resistance was very low overall, but 3.0% levofloxacin resistance (2.6% gatifloxacin and 0.4% moxifloxacin) was observed in Italy. Interestingly, many isolates with minimum inhibitory concentrations (MICs) at the top of the fluoroquinolone susceptibility breakpoints possessed single quinolone resistance-determining region (QRDR) mutations. Care should be taken when treating fluoroquinolone-susceptible isolates with a higher MIC, which are likely to harbour QRDR mutations and may become fully resistant and cause treatment failure. We concur with the conclusions of other recent studies that suggest fluoroquinolone breakpoints should be lowered to ensure these isolates are categorised as resistant. Fluoroquinolones would still remain an important alternative treatment for respiratory tract infections (albeit for adults only), with moxifloxacin being the most potent fluoroquinolone tested in this study.     

High levels of multidrug resistance in clinical isolates of Gram-negative pathogens from Nigeria

In Nigeria, quinolones and β-lactam antibiotics are widely used to treat bacterial infections. This study aimed to identify the prevalence of resistance to these drugs and to determine the mechanisms of resistance to these agents. In total, 134 non-duplicate, Gram-negative enteric isolates of 13 species from different hospitals were investigated for susceptibility to a panel of antibiotics, carriage of plasmid-mediated quinolone and β-lactam resistance genes, production of extended-spectrum β-lactamases (ESBLs), and mutations within topoisomerase genes. The level of resistance to all antibiotics tested was extremely high, with minimum inhibitory concentrations for 90% of the organisms (MIC(90) values) of ≥ 256 μg/mL for all drugs. Of the 134 isolates, 92 had mutations within the quinolone resistance-determining region (QRDR) of gyrA or within gyrA and parC. In addition, the plasmid-mediated quinolone resistance genes qnrA, qnrB, aac(6')-Ib-cr and qepA were identified. The qnrD allele, which has previously only been found in Salmonella isolates from China, was identified in two Proteus isolates and one Pseudomonas isolate. Of the 134 isolates, 23 (17.2%) carried aac(6')-Ib-cr, 11 (8.2%) carried a qnr variant and 5 (3.7%) were positive for qepA. Twenty-eight isolates (20.9%) produced ESBL variants, with a CTX-M variant being carried by 25 isolates (18.7%). In addition, six isolates (4.5%) carried ampC variants [ACT-1 (1 isolate), DHA-1 (4 isolates) and CMY-2 (1 isolate)]. This study demonstrates a very high level of multidrug resistance amongst Gram-negative enteric bacilli isolated from different sites from patients in Nigerian hospitals as well as the presence of a variety of plasmid-associated resistance genes, including some identified from Africa for the first time.     

Prevalence of plasmid-mediated quinolone resistance determinants qnr and aac(6')-Ib-cr in Escherichia coli and Klebsiella pneumoniae producing extended-spectrum β-lactamases in Spain

The presence of the plasmid-mediated quinolone resistance determinants qnrA, qnrB, qnrS and aac(6')-Ib-cr was evaluated in a collection of 382 isolates of extended-spectrum β-lactamase (ESBL)-producing Escherichia coli and Klebsiella pneumoniae collected between February and March 2006 for the nationwide Spanish GEIH-ESBL 2006 project. In total, 14 isolates (3.7%) were positive for qnr genes (3 qnrA1, 5 qnrB-like and 6 qnrS1) and 62 isolates (16.2%) were positive for the mutant variant of aac(6')-Ib-cr. The Aac(6')-Ib-cr enzyme was the most prevalent plasmid-mediated mechanism of quinolone resistance in Spain. Most of the Aac(6')-Ib-cr-producing E. coli isolates (94.2%) carried two mutations in gyrA and two in parC, whilst only 57.2% of K. pneumoniae harbouring this enzyme were gyrA and/or parC mutants. Most qnr plasmids were transferable, but only four were conjugative. Plasmid incompatibility groups were identified for only four plasmids, belonging to FIA, HI2 and I1γ. The most prevalent ESBLs associated with qnr plasmids belonged to the SHV and CTX-M families. The present study highlights the broad geographical spread of qnr-like determinants in Spain and their association with the SHV-12 and CTX-M-9 ESBLs in human clinical isolates.     

儿科病房肺炎克雷伯菌呼吸道分离株耐药性分析

目的 了解本院儿科病房肺炎克雷伯菌呼吸道分离株耐药性,为临床合理使用抗生素提供依据.方法 应用TDR-300B自动微生物分析系统进行细菌鉴定及药敏试验,采用WHONET5.6对2015年1月至2016年12月本院儿科病房呼吸道标本分离的肺炎克雷伯菌对临床常用的12种抗菌药物的耐药性进行统计分析.结果 儿科病房肺炎克雷伯菌呼吸道分离株对阿米卡星、美罗培南、左氧氟沙星和哌拉西林/他唑巴坦的耐药率低(0％～12.2％);对其余8种抗生素耐药率较高(24.5％ ～ 100.0％);2016年分离株对左氧氟沙星耐药率显著高于2015年,而2016年分离株对头孢西丁的耐药率则显著低于2015年.结论 儿科病房肺炎克雷伯菌呼吸道分离株耐药严重,临床工作者应合理应用抗菌药物,延缓耐药菌的产生,并加强细菌耐药监测和医院感染控制措施以防止耐药菌的播散.     

不同剂量左氧氟沙星使用频度与细菌耐药率相关性研究

目的：了解不同荆量左氧氟沙星使用频度与8种细菌耐药率的相关性,为临床合理用药提供依据。方法：分别统计2002～2009年0．3g,0．4g、0．5g3种剂量左氧氟沙星、总左氧氟沙星及喹诺酮类抗菌药的用药频度与8种细菌对左氧氟沙星的耐药率,采用SPSS13．0统计软件进行相关性分析。结果：8种细菌中肺炎克雷伯菌对左氧氟沙星的耐药率最低（17．3％-35．4％）,耐甲氧西林金黄色葡萄球菌和产ESBL大肠埃希菌对左氧氟沙星耐药率较高,分别为75．4％～95．6％和81．1％～92．6％。左氧氟沙星（0．5g）的用药频度与大肠埃希菌、铜绿假单胞菌耐药率呈负相关（r=-0．9974,P〈0．01;r=-1．0000,P〈0．01）;左氧氟沙星（0．3g）、总左氧氟沙星、喹诺酮类抗菌药的用药频度与产ESBL肺炎克雷伯菌耐药率呈负相关（r=-0．8678,P〈0．05;r=-0．8465,P〈0．05;r=-0．7857,P〈0．05）;左氧氟沙星（0．3g）和总左氧氟沙星的用药频度分别与金黄色葡萄球菌和鲍曼不动杆菌的耐药率呈正相关（r=0．7256,P〈0．05,r=0．8214,P〈0．05）。结论：不同剂量左氧氟沙星、总左氧氟沙星、喹诺酮类抗菌药用药频度与细菌耐药率存在相关性,临床应注意合理使用不同剂量的左氧氟沙星。     

Single and multi-step resistance selection study in Streptococcus pneumoniae comparing ceftriaxone with levofloxacin,gatifloxacin and moxifloxacin

Attempts were made to select resistant pneumococcal mutants by sequential subculturing of 12 clinically isolated pneumococci, [four were penicillin sensitive (MIC) 0.03-0.06 mg/l, four penicillin intermediate (MIC 0.25-0.5 mg/l) and four penicillin resistant (MIC 2-4 mg/l)] in sub-inhibitory concentrations of ceftriaxone, levofloxacin, gatifloxacin and moxifloxacin. Subculturing in gatifloxacin, levofloxacin, moxifloxacin and ceftriaxone selected 12 mutants (12/12), 10 mutants (10/12), 10 mutants (10/12) and three mutants (3/12), respectively. DNA sequencing of the quinolone-resistant mutants showed that most strains had mutations in GyrA at E85 or S81. This in vitro mutation study demonstrates a clear distinction between the low frequency of development of resistance with ceftriaxone exposure as opposed to the high frequency with quinolone exposure.     

质粒介导的喹诺酮耐药基因在中国大陆尿液分离的大肠埃希菌中的广泛分布

目的分析中国大陆20家三甲医院尿来源大肠埃希菌的耐药特点并调查质粒介导的喹诺酮类耐药基因的分布情况和流行特点。方法收集卫生部全国耐药监测网2007年1月至2008年3月非重复298株尿液分离大肠埃希菌;琼脂稀释法测定其对20种抗菌药物的敏感性,多聚酶链反应和DNA测序分析qn-rA,qnrB,qnrS,aac(6’)-ib和qepA基因的流行性;接合实验分析质粒的转移性;Eric-PCR分析喹诺酮基因阳性菌株之间的遗传相关性;卡方检验用于分析耐药基因与氟喹诺酮耐药之间的相关性。结果 298株大肠埃希菌对20种抗菌药物耐药现象严重,其中对环丙沙星和左氧氟沙星有很高的耐药性,耐药率高达78.5%和74.2%。经基因比对分析,62株(20.8%)细菌携带aac(6’)-Ib基因;45株(15.1%)细菌携带喹诺酮耐药基因,1株(0.3%)检测出qnrA基因,3株(11.4%)检出qnrB基因,5株(1.7%)检出qnrS基因,25株(8.4%)确定为aac(6’)-Ib-cr基因,12株(4.7%)检出qepA基因;此外,有3株细菌分别发现aac(6’)-Ib-cr和qepA1基因aac(6’)-Ib-cr和qnrB1基因,qepA和qnrS1基因共存。45株喹诺酮基因阳性菌株之间具有很大的遗传差异,并且其中有16株细菌携带的基因具有可转移性。aac(6’)-Ib的流行性与细菌的环丙沙星和左氧氟沙星不敏感性相关(P<0.05);喹诺酮耐药基因的流行性与细菌的氟喹诺酮不敏感性相关(P<0.05)。结论尿液分离的大肠埃希菌耐药严重,质粒介导的喹诺酮耐药基因主要以aac(6’)-ib-cr为主,qepA1次之,这些潜在播散的喹诺酮耐药基因对于临床尿路感染的治疗有很大的挑战。     

In vitro activity of gemifloxacin and contemporary oral antimicrobial agents against 27247 Gram-positive and Gram-negative aerobic isolates: a global surveillance study

This study was a multi-centre, multi-country surveillance of 27247 Gram-positive and Gram-negative isolates collected from 131 study centres in 44 countries from 1997 to 2000. MICs of gemifloxacin were compared with penicillin, amoxicillin-clavulanic acid, cefuroxime, azithromycin, clarithromycin, trimethoprim-sulphamethoxazole, ciprofloxacin, grepafloxacin and levofloxacin by broth microdilution. Penicillin resistance in Streptococcus pneumoniae was extremely high in the Middle East (65.6%), Africa (64.0%) and Asia (60.4%) and lower in North America (40.3%), Europe (36.9%) and the South Pacific (31.8%). Macrolide resistance in S. pneumoniae was highest in Asia (51.7%) but varied widely between laboratories in Europe (26.0%), North America (21.6%), the Middle East (13.7%), the South Pacific (10.6%) and Africa (10.0%). All the study quinolones were highly active against penicillin-resistant and macrolide-resistant S. pneumoniae. Overall, gemifloxacin had the lowest MIC(90) at 0.06 mg/l with MICs 4-64-fold lower than ciprofloxacin, levofloxacin and grepafloxacin against S. pneumoniae. Gemifloxacin MICs were more potent than grepafloxacin > levoflaxacin > ciproflaxin against the Gram-positive aerobes and shared comparable Gram-negative activity with ciprofloxacin and levofloxacin.     

Linkage of ciprofloxacin resistance with a single genotypic cluster of Klebsiella pneumoniae

The objective of this study was to examine the epidemiology of ciprofloxacin-resistant, extended-spectrum beta-lactamase (ESBL)-producing Klebsiella pneumoniae strains. Sixty-nine unique patient isolates of K. pneumoniae isolated from a variety of clinical specimens submitted to the clinical bacteriology laboratories of The Royal Infirmary of Edinburgh and associated General Practices were identified and susceptibility testing was performed with the Vitek system. Strains flagged as ESBL-positive by the Vitek system were subjected to isoelectric focusing. The results suggested that all 69 isolates harboured at least one ESBL, which was later confirmed by polymerase chain reaction (PCR) with bla(TEM) and/or bla(SHV) primers. The purified PCR product was subjected to automated sequencing and the results were compared with the BLAST online search engine. Of the 69 isolates, 32 (46.4%) were found to be resistant to ciprofloxacin, 11 (15.9%) were intermediate and 26 (37.7%) were sensitive. To investigate the epidemiological relationship between the ciprofloxacin-resistant ESBL-positive strains, pulsed-field gel electrophoresis (PFGE) was performed. Rapidest software was used to calculate the genetic distance by the Nei distance method. PFGE analysis indicated that the clinical isolates belonged to four distinct genotype clusters (Groups A, B, C and D); each group or cluster was homogeneous or compact with respect to certain characteristics. Group A consisted of 25 isolates, group B of 3 isolates and Groups C and D of 2 isolates each. These results indicate that the spread of resistance is largely as a result of the dissemination of a single clonal strain. PCR was used to amplify the gyrA and parC genes from genomic DNA of the ciprofloxacin-resistant isolates. The amplified product was sent for analysis by automated DNA sequencing and the resulting DNA sequences were compared with the gyrA gene of K. pneumoniae. The sequencing results demonstrated that alteration of the GyrA subunit of DNA gyrase at amino acid 83 and/or amino acid 87 plays a central role in conferring high-level quinolone resistance in K. pneumoniae possessing ESBLs.