Antimicrobial susceptibilities of Campylobacter strains isolated from Finnish subjects infected domestically or from those infected abroad

The in vitro susceptibilities of 678 Campylobacter jejuni and Campylobacter coli strains isolated from stool samples of the same number of Finnish subjects were studied. A total of 523 patients, representing inhabitants from throughout Finland, had not traveled abroad within the 2 weeks prior to becoming ill, whereas 155 persons had presumably acquired their infections abroad. The antimicrobial agents studied were erythromycin, ciprofloxacin, levofloxacin, trovafloxacin, and moxifloxacin. The MICs of these antimicrobial agents were determined by the agar dilution method. The growth of all domestic isolates was inhibited by erythromycin at concentrations of 4 microg/ml, and for these isolates the fluoroquinolone MICs at which 90% of isolates are inhibited (MIC(90)s) ranged from 0.06 to 0.5 microg/ml. For the foreign isolates, the erythromycin MIC(90) was still low (4 microg/ml), but their susceptibilities to fluoroquinolones were clearly reduced (MIC(90)s, 8 to 64 microg/ml). Of the four different fluoroquinolones studied, ciprofloxacin was the least active (MIC(90), 64 micro g/ml).     

Role of efflux pumps and topoisomerase mutations in fluoroquinolone resistance in Campylobacter jejuni and Campylobacter coli

Point mutations in the topoisomerase (DNA gyrase A) gene are known to be associated with fluoroquinolone resistance in Campylobacter. Recent studies have shown that an efflux pump encoded by cmeABC is also involved in decreased susceptibilities to fluoroquinolones, as well as other antimicrobials. Genome analysis suggests that Campylobacter jejuni contains at least nine other putative efflux pumps. Using insertional inactivation and site-directed mutagenesis, we investigated the potential contributions of these pumps to susceptibilities to chloramphenicol, ciprofloxacin, erythromycin, and tetracycline in C. jejuni and Campylobacter coli. Insertional inactivation of cmeB resulted in 4- to 256-fold decreases in the MICs of chloramphenicol, ciprofloxacin, erythromycin, and tetracycline, with erythromycin being the most significantly affected. In contrast, inactivation of all other putative efflux pumps had no effect on susceptibility to any of the four antimicrobials tested. Mutation of gyrA at codon 86 (Thr-Ile) caused 128- and 64-fold increases in the MICs of ciprofloxacin and nalidixic acid, respectively. The replacement of the mutated gyrA with a wild-type gyrA allele resulted in a 32-fold decrease in the ciprofloxacin MIC and no change in the nalidixic acid MIC. Our findings indicate that CmeABC is the only efflux pump among those tested that influences antimicrobial resistance in Campylobacter and that a point mutation (Thr-86-Ile) in gyrA directly causes fluoroquinolone resistance in Campylobacter. These two mechanisms work synergistically in acquiring and maintaining fluoroquinolone resistance in Campylobacter species.     

Antibiotic resistance of Campylobacter jejuni and Campylobacter coli clinical isolates from Poland

We tested 102 Campylobacter jejuni and 6 Campylobacter coli clinical isolates from Poland. All were susceptible to erythromycin. Among the tested C. jejuni isolates 55.9% and 13.7% were resistant to ciprofloxacin and tetracycline, respectively. Replacement of Thr86 with Ile in GyrA and a plasmid-borne tet(O) gene were the main resistance mechanisms for fluoroquinolones and tetracycline, respectively.     

Susceptibilities of Campylobacter jejuni isolates from Germany to ciprofloxacin,moxifloxacin, erythromycin,clindamycin, and tetracycline

To elucidate Campylobacter jejuni resistance to antibiotics in Germany, MICs of ciprofloxacin, moxifloxacin, erythromycin, clindamycin, and tetracycline were determined (using agar dilution) for 144 clinical isolates. The data indicate a considerable ciprofloxacin resistance (45.1%) without a clonal relationship of the strains and a greater in vitro activity of moxifloxacin, erythromycin, and clindamycin.     

Increasing resistance of Streptococcus pneumoniae to fluoroquinolones: results of a Hong Kong multicentre study in 2000.

The MICs of 13 antimicrobial agents including seven fluoroquinolones (ciprofloxacin, levofloxacin, sparfloxacin, grepafloxacin, gatifloxacin, moxifloxacin and clinafloxacin) for Streptococcus pneumoniae isolates obtained from all regions of Hong Kong in the year 2000 were determined by the Etest. Overall, 39.4% of 180 isolates were susceptible to penicillin, 11.7% were intermediate and 48.9% were resistant. The overall prevalence of fluoroquinolone non-susceptibility (levofloxacin MIC > or = 4 mg/L) was 13.3% but increased to 27.3% among the penicillin-resistant isolates. For the fluoroquinolone non-susceptible isolates, within-class cross-resistance was common. For the fluoroquinolone non-susceptible isolates, the median MICs of clinafloxacin, gatifloxacin, moxifloxacin, sparfloxacin and grepafloxacin were, respectively, six-, 24-, 32- 84- and 128-fold higher than those for the susceptible isolates. All fluoroquinolone non-susceptible strains were derived from adults. The prevalence of fluoroquinolone resistance was higher in isolates from older patients (17.1% among those > or = 65 years of age versus 9.1% among those 18-64 years of age, P < 0.001) and from adults with chronic obstructive pulmonary disease (24.6% versus 9.3%, P = 0.01). All fluoroquinolone non-susceptible strains were non-susceptible to penicillin (MIC range 2-4 mg/L), cefotaxime (MIC range 1-4 mg/L) and erythromycin (MIC range 4- > or = 256 mg/L). The fluoroquinolone non-susceptible isolates were genetically related to the Spain(23F)-1 clone when analysed by pulse-field gel electrophoresis and multilocus sequence typing. In conclusion, a rapid increase in the prevalence of fluoroquinolone resistance among S. pneumoniae was found in Hong Kong. Typing analysis suggests that this is due to the pan-regional dissemination of a fluoroquinolone-resistant variant (designated Hong Kong(23F)-1) of the globally distributed Spain(23)F-1 clone.     

Mutant prevention concentrations for single-step fluoroquinolone-resistant mutants of wild-type, efflux-positive, or ParC or GyrA mutation-containing Streptococcus pneumoniae isolates

Three fluoroquinolone-susceptible and five fluoroquinolone-resistant (two with ParC Ser79Phe mutations, one with a GyrA Ser81Phe mutation, and two that were efflux positive) Streptococcus pneumoniae isolates were exposed to one, two, four, eight, and sixteen times the MICs of ciprofloxacin, gatifloxacin, gemifloxacin, levofloxacin, and moxifloxacin. Mutational frequencies were calculated at each multiple of the MIC for which growth was observed. Mutant prevention concentrations (MPCs) and the multiple of the MIC at the MPC (MP(MIC)) were evaluated. All resulting mutants were sequenced for quinolone resistance-determining region changes in GyrA and ParC and were evaluated for reserpine-sensitive efflux. The MPC order was generally ciprofloxacin > levofloxacin > gatifloxacin > moxifloxacin > gemifloxacin. The MP(MIC) order varied depending on the genetic constitution of the original isolates from which the mutants were generated. For those mutants created from fluoroquinolone-susceptible isolates (those that had wild-type ParC and GyrA and were efflux negative), the MP(MIC) order was ciprofloxacin = moxifloxacin > gemifloxacin > levofloxacin > gatifloxacin. The MP(MIC)s of each fluoroquinolone for mutants created from isolates with a ParC mutation (with wild-type GyrA and efflux negative) were similar. A similar occurrence was observed with the mutants created from the efflux-positive isolates (with wild-type ParC and GyrA). The MP(MIC) order for the mutants created from the isolate with a GyrA mutation (with wild-type ParC and efflux negative) was ciprofloxacin = gemifloxacin > levofloxacin = moxifloxacin > gatifloxacin. Gatifloxacin, levofloxacin, and moxifloxacin may be intrinsically more able to prevent the development of resistance by fluoroquinolone-susceptible isolates, isolates that are efflux positive, or isolates that carry a GyrA mutation. However, once a ParC mutation is present, the MPC increases dramatically for all fluoroquinolones.     

Antibiotic resistance in Campylobacter jejuni and Campylobacter coli isolated from poultry in the South-East Queensland region

OBJECTIVES: The aim of this study was to determine the antimicrobial resistance patterns of 125 Campylobacter jejuni and 27 Campylobacter coli isolates from 39 Queensland broiler farms. METHODS: Two methods, a disc diffusion assay and an agar-based MIC assay, were used. The disc diffusion was performed and interpreted as previously described (Huysmans MB, Turnidge JD. Disc susceptibility testing for thermophilic campylobacters. Pathology 1997; 29: 209-16), whereas the MIC assay was performed according to CLSI (formerly NCCLS) methods and interpreted using DANMAP criteria. RESULTS: In both assays, no C. jejuni or C. coli isolates were resistant to ciprofloxacin or chloramphenicol, no C. coli were resistant to nalidixic acid, and no C. jejuni were resistant to erythromycin. In the MIC assay, no C. jejuni isolate was resistant to nalidixic acid, whereas three isolates (2.4%) were resistant in the disc assay. The highest levels of resistance of the C. jejuni isolates were recorded for tetracycline (19.2% by MIC and 18.4% by disc) and ampicillin (19.2% by MIC and 17.6% by disc). The C. coli isolates gave very similar results (tetracycline resistance 14.8% by both MIC and disc; ampicillin resistance 7.4% by MIC and 14.8% by disc). CONCLUSIONS: This work has shown that the majority of C. jejuni and C. coli isolates were susceptible to the six antibiotics tested by both disc diffusion and MIC methods. Disc diffusion represents a suitable alternative methodology to agar-based MIC methods for poultry Campylobacter isolates.     

Effect of fluoroquinolone treatment on growth of and toxin production by epidemic and nonepidemic clostridium difficile strains in the cecal contents of mice

Several recent outbreaks of Clostridium difficile-associated disease (CDAD) have been attributed to the emergence of an epidemic strain with increased resistance to fluoroquinolone antibiotics. Some clinical studies have suggested that fluoroquinolones with enhanced antianaerobic activity (i.e., gatifloxacin and moxifloxacin) may have a greater propensity to induce CDAD than ciprofloxacin and levofloxacin do. We examined the effects of subcutaneous fluoroquinolone treatment on in vitro growth of and toxin production by epidemic and nonepidemic C. difficile isolates in cecal contents of mice and evaluated the potential for these agents to inhibit fluoroquinolone-susceptible isolates during treatment. When C. difficile isolates were inoculated into cecal contents collected 2 days after the final antibiotic dose, gatifloxacin and moxifloxacin promoted significantly more growth and toxin production than ciprofloxacin and levofloxacin did. During treatment, gatifloxacin and moxifloxacin inhibited growth of fluoroquinolone-susceptible but not fluoroquinolone-resistant isolates. Ciprofloxacin and levofloxacin promoted growth of C. difficile when administered at higher doses (i.e., 20 times the human dose in mg/kg of body weight), and levofloxacin inhibited growth of fluoroquinolone-susceptible, but not fluoroquinolone-resistant, C. difficile isolates when administered in combination with ceftriaxone. Thus, fluoroquinolones with enhanced antianaerobic activity (i.e., gatifloxacin and moxifloxacin) promoted C. difficile growth to a greater extent than did ciprofloxacin and levofloxacin in this model. However, our findings suggest that fluoroquinolones may exert selective pressure favoring the emergence of epidemic fluoroquinolone-resistant C. difficile strains by inhibiting fluoroquinolone-susceptible but not fluoroquinolone-resistant isolates during treatment and that agents such as levofloxacin or ciprofloxacin can exert such selective pressure when administered in combination with antibiotics that disrupt the anaerobic microflora.     

Molecular characterization of fluoroquinolone resistant Streptococcus pneumoniae clinical isolates obtained from across Canada

There is little published data detailing fluoroquinolone resistance in clinical isolates of S. pneumoniae. The purpose of this study was to characterize the resistance mechanisms of 34 fluoroquinolone-resistant S. pneumoniae clinical isolates obtained from medical centers in 8 of 10 Canadian provinces between 1997 and 2000. The quinolone resistance determining regions of gyrA, parC, and parE from the isolates were sequenced. The isolates were evaluated for reserpine-sensitive efflux of ciprofloxacin and the new fluoroquinolones: gatifloxacin, gemifloxacin, levofloxacin and moxifloxacin. The isolates were typed using pulsed field gel electrophoresis. The majority of the isolates were genetically unrelated. Lower level fluoroquinolone resistance (ciprofloxacin MIC 4-8 microg/ml) was associated with amino acid substitutions in ParC, while higher level resistance (ciprofloxacin MIC > or = 16 microg/ml) was associated with amino acid substitutions in both ParC and GyrA. ParE substitutions were not associated with clinical resistance. Twelve of 34 (35%) isolates demonstrated reserpine-sensitive efflux of ciprofloxacin. Efflux alone conferred low level ciprofloxacin resistance in 3 isolates. Significant reserpine-sensitive efflux of the new fluoroquinolones was not observed.     

Relationships among ciprofloxacin, gatifloxacin, levofloxacin, and norfloxacin MICs for fluoroquinolone-resistant Escherichia coli clinical isolates

Fluoroquinolones are some of the most prescribed antibiotics in the United States. Previously, we and others showed that the fluoroquinolones exhibit a class effect with regard to the CLSI-established breakpoints for resistance, such that decreased susceptibility (i.e., an increased MIC) to one fluoroquinolone means a simultaneously decreased susceptibility to all. For defined strains, however, clear differences exist in the pharmacodynamic properties of each fluoroquinolone and the extent to which resistance-associated genotypes affect the MICs of each fluoroquinolone. In a pilot study of 920 clinical Escherichia coli isolates, we uncovered tremendous variation in norfloxacin MICs. The MICs for all of the fluoroquinolone-resistant isolates exceeded the resistance breakpoint, reaching 1,000 microg/ml. Approximately 25% of the isolates (n = 214), representing the full range of resistant norfloxacin MICs, were selected for the simultaneous determinations of ciprofloxacin, gatifloxacin, levofloxacin, and norfloxacin MICs. We found that (i) great MIC variation existed for all four fluoroquinolones, (ii) the ciprofloxacin and levofloxacin MICs of >90% of the fluoroquinolone-resistant isolates were higher than the resistance breakpoints, (iii) ciprofloxacin and levofloxacin MICs were distributed into two distinct groups, (iv) the MICs of two drug pairs (ciprofloxacin and norfloxacin by Kendall's Tau-b test and gatifloxacin and levofloxacin by paired t test) were similar with statistical significance but were different from each other, and (v) approximately 2% of isolates had unprecedented fluoroquinolone MIC relationships. Thus, although the fluoroquinolones can be considered equivalent with regard to clinical susceptibility or resistance, fluoroquinolone MICs differ dramatically for fluoroquinolone-resistant clinical isolates, likely because of differences in drug structure.     

Antimicrobial activity of moxifloxacin, gatifloxacin and six fluoroquinolones against Streptococcus pneumoniae

The in vitro and pharmacodynamic effects of moxifloxacin and gatifloxacin against Streptococcus pneumoniae were compared with six other fluoroquinolones. Organisms included penicillin-susceptible (54) and penicillin-non-susceptible (145) isolates from 1998-1999. Moxifloxacin and clinafloxacin demonstrated the greatest in vitro activity, with MIC90s of 0.13 mg/L, followed by trovafloxacin, grepafloxacin > gatifloxacin, sparfloxacin > levofloxacin > ciprofloxacin. There was no difference in fluoroquinolone activity between penicillin-susceptible and -non-susceptible strains. Pharmacodynamic analysis using published pharmacokinetic information indicates that all the agents tested except ciprofloxacin had an AUC/MIC90 > 30, with moxifloxacin having the greatest free-drug availability.     

Antipneumococcal activity of DK-507k, a new quinolone, compared with the activities of 10 other agents

Agar dilution MIC determination was used to compare the activity of DK-507k with those of ciprofloxacin, levofloxacin, gatifloxacin, moxifloxacin, sitafloxacin, amoxicillin, cefuroxime, erythromycin, azithromycin, and clarithromycin against 113 penicillin-susceptible, 81 penicillin-intermediate, and 67 penicillin-resistant pneumococci (all quinolone susceptible). DK-507k and sitafloxacin had the lowest MICs of all quinolones against quinolone-susceptible strains (MIC at which 50% of isolates were inhibited [MIC50] and MIC90 of both, 0.06 and 0.125 microg/ml, respectively), followed by moxifloxacin, gatifloxacin, levofloxacin, and ciprofloxacin. MICs of beta-lactams and macrolides rose with those of penicillin G. Against 26 quinolone-resistant pneumococci with known resistance mechanisms, DK-507k and sitafloxacin were also the most active quinolones (MICs, 0.125 to 1.0 microg/ml), followed by moxifloxacin, gatifloxacin, levofloxacin, and ciprofloxacin. Mutations in quinolone resistance-determining regions of quinolone-resistant strains were in the usual regions of the parC and gyrA genes. Time-kill testing showed that both DK-507k and sitafloxacin were bactericidal against all 12 quinolone-susceptible and -resistant strains tested at twice the MIC at 24 h. Serial broth passages in subinhibitory concentrations of 10 strains for a minimum of 14 days showed that development of resistant mutants (fourfold or greater increase in the original MIC) occurred most rapidly for ciprofloxacin, followed by moxifloxacin, DK-507k, gatifloxacin, sitafloxacin, and levofloxacin. All parent strains demonstrated a fourfold or greater increase in initial MIC in <50 days. MICs of DK-507k against resistant mutants were lowest, followed by those of sitafloxacin, moxifloxacin, gatifloxacin, ciprofloxacin, and levofloxacin. Four strains were subcultured in subinhibitory concentrations of each drug for 50 days: MICs of DK-507k against resistant mutants were lowest, followed by those of sitafloxacin, moxifloxacin, gatifloxacin, levofloxacin, and ciprofloxacin. Exposure to DK-507k and sitafloxacin resulted in mutations, mostly in gyrA.     

In vitro activity of ABT-773 versus macrolides and quinolones against resistant respiratory tract pathogens.

ABT-773, a novel ketolide, was compared to erythromycin, azithromycin, clarithromycin, ciprofloxacin, ofloxacin, levofloxacin, moxifloxacin, gatifloxacin, and gemifloxacin against antibiotic-resistant strains recently isolated from patients with respiratory tract infections. MICs were determined by agar dilution using standard NCCLS methodology. ABT-773 (MIC(90) 0.06 mg/L) was more active than the macrolides (MIC(90) > or = 2 mg/L) and fluoroquinolones (MIC(90) > or = 0.5 mg/L) against penicillin-resistant Streptococcus pneumoniae. The fluoroquinolones were the most active agents tested against beta-lactamase-positive Haemophilus influenzae (MIC(90) < or = 0.01-0.06 mg/L), against which ABT-773 (MIC(90) 4 mg/L) was comparable to azithromycin and two- and four-fold more active than erythromycin and clarithromycin, respectively. Against beta-lactamase positive Moraxella catarrhalis, the activity of ABT-773 (MIC(90) 0.06 mg/L) was comparable to gemifloxacin, trovafloxacin, levofloxacin, and ciprofloxacin (MIC(90) 0.03-0.06 mg/L) and 4- to eightfold greater than that of clarithromycin, gatifloxacin, and erythromycin. These data suggest ABT-773 could be a valuable compound for the treatment of respiratory tract infections, including those resistant to usual oral therapy.     

Multidrug resistance in Campylobacter jejuni strains collected from Finnish patients during 1995-2000

OBJECTIVES: The resistance of Campylobacter jejuni to fluoroquinolones is increasing globally. This study was performed to delineate those antimicrobial agents that are effective in vitro against ciprofloxacin-resistant C. jejuni isolates and potentially suitable for the treatment of severe disease when fluoroquinolone resistance or multidrug resistance is known or suspected. METHODS: During 1995-2000 we collected 376 C. jejuni strains, of which 354 were of foreign origin from multiple countries and 22 were of domestic origin. The MICs of 12 antimicrobial agents against the isolates were determined. RESULTS: Of the 376 strains, 174 (46%) were resistant to ciprofloxacin. Among other antimicrobials, resistance was most common to tetracycline (46%) and ampicillin (17%). Of the ciprofloxacin-resistant strains, 68% and 25%, respectively, were resistant to tetracycline and ampicillin, and 3% were resistant to erythromycin, gentamicin or clindamycin. One (0.6%) ciprofloxacin-resistant isolate was resistant to co-amoxiclav and none was resistant to imipenem. Resistance to three or more antimicrobial groups was detected in 22% of the isolates. Multidrug resistance was significantly associated with ciprofloxacin resistance (33% versus 12%; P<0.01). Eight (2%) strains were resistant to macrolides, of which 75% were also resistant to ciprofloxacin, but none was resistant to co-amoxiclav or imipenem. CONCLUSIONS: Macrolides still appear to be the first-choice alternative for suspected C. jejuni enteritis, if antimicrobial treatment is needed. The in vitro susceptibilities suggest that clinical trials to treat enteritis caused by multidrug-resistant C. jejuni with co-amoxiclav, and life-threatening infections with a carbapenem, may be valuable.     

Comparison of the in vitro activities of several new fluoroquinolones against respiratory pathogens and their abilities to select fluoroquinolone resistance

In this study the in vitro activities and pharmacodynamic properties of moxifloxacin, levofloxacin, gatifloxacin and gemifloxacin were compared on recently isolated respiratory pathogens and strains of Streptococcus pneumoniae with known mechanisms of fluoroquinolone resistance. In addition, the resistance selection frequencies of moxifloxacin and levofloxacin on three recently isolated respiratory pathogens and four strains of S. pneumoniae with known mechanisms of fluoroquinolone resistance were investigated. The four fluoroquinolones had similar activities against both Moraxella catarrhalis (MIC(90)s 0.015-0.06 mg/L) and Haemophilus influenzae (MIC(90)s 0.008-0.03 mg/L). More marked differences in activity were noted with S. pneumoniae, with MIC(90)s of 0.25, 1, 0.5 and 0.03 mg/L for moxifloxacin, levofloxacin, gatifloxacin and gemifloxacin, respectively. With the S. pneumoniae strains, the four fluoroquinolones exhibited similar concentration-dependent time-kill kinetics. The resistance selection frequencies of levofloxacin were higher than those of moxifloxacin at concentrations equivalent to those at the end of the dosing interval. Therefore moxifloxacin may have less of an impact on the development of resistance than levofloxacin.     

In vitro activities of 11 fluoroquinolones against 226 Campylobacter jejuni strains isolated from Finnish patients, with special reference to ciprofloxacin resistance

OBJECTIVES: Although Campylobacter jejuni is naturally susceptible to fluoroquinolones, the resistance to these antimicrobials has increased rapidly during the recent years. The aim of this study was to compare the activities of various older and newer fluoroquinolones towards C. jejuni, with special attention on ciprofloxacin-resistant strains. METHODS: We analysed the in vitro activities of 11 fluoroquinolones against 226 C. jejuni strains collected from Finnish patients between 1995 and 2000. RESULTS: Of all 226 C. jejuni strains, 134 (59.3%) were resistant to ciprofloxacin (MIC > or = 4 mg/L), 1 (0.4%) was intermediately resistant (MIC = 2 mg/L) and 91 (40.3%) were ciprofloxacin-susceptible (MIC < or = 1 mg/L). The MIC50 and MIC90 values of ciprofloxacin for the 91 ciprofloxacin-susceptible strains were 0.25 and 0.5 mg/L, respectively. The corresponding MIC50 and MIC90 values of levofloxacin were 0.25 and 0.5 mg/L, and those of moxifloxacin were 0.125 and 0.25 mg/L, these being lower than those of norfloxacin and ofloxacin. The two newer fluoroquinolones, sitafloxacin and clinafloxacin, exhibited the lowest MIC50 and MIC90 values: 0.016 and 0.064 mg/L of sitafloxacin and 0.032 and 0.125 mg/L of clinafloxacin, respectively. Sitafloxacin and clinafloxacin exhibited the lowest MIC50 and MIC90 values also for the 134 ciprofloxacin-resistant C. jejuni strains: 0.25 and 1 mg/L of sitafloxacin and 1 and 4 mg/L of clinafloxacin, respectively. CONCLUSIONS: Of the newer fluoroquinolones presently under development, sitafloxacin is in vitro highly effective towards C. jejuni, with low MIC values also for the ciprofloxacin-resistant strains. Sitafloxacin might be a candidate for clinical trials on campylobacteriosis.     

Fluoroquinolone resistance in Streptococcus pneumoniae in United States since 1994-1995

The in vitro activities of ciprofloxacin, levofloxacin, gatifloxacin, and moxifloxacin against a large collection of clinical isolates of Streptococcus pneumoniae (n = 4,650) obtained over a 5-year period, 1994-1995 through 1999-2000, were assessed as part of a longitudinal multicenter U.S. surveillance study of antimicrobial resistance. Three sampling periods were used during this investigation, the winter seasons of 1994-1995, 1997-1998, and 1999-2000; and 1,523, 1,596 and 1,531 isolates were collected during these three periods, respectively. The overall rank order of activity of the four fluoroquinolones examined in this study was moxifloxacin > gatifloxacin > levofloxacin = ciprofloxacin, in which moxifloxacin (MIC at which 90% of isolates are inhibited [MIC(90)], 0.25 microg/ml; modal MIC, 0.12 microg/ml) was twofold more active than gatifloxacin (MIC(90), 0.5 microg/ml; modal MIC, 0.25 microg/ml), which in turn was fourfold more active than either levofloxacin (MIC(90), 1 microg/ml; modal MIC, 1 microg/ml) or ciprofloxacin (MIC(90), 2 microg/ml; modal MIC, 1 microg/ml). Changes in the in vitro activities of fluoroquinolones against S. pneumoniae strains in the United States over the 5-year period of the survey were assessed by comparing the MIC frequency distributions of the study drugs against the isolates obtained during the three sampling periods encompassing this investigation. These comparisons revealed no evidence of changes in the in vitro activities of the fluoroquinolones. In addition, the percentages of isolates in the three sampling periods for which MICs were above the resistance breakpoints were compared. Low percentages of resistant strains were detected, and there was no evidence of resistance rate changes over time. For example, by use of a ciprofloxacin MIC of > or = 4 microg/ml to define resistance, the proportions of isolates from the three sampling periods for which MICs were at or above this breakpoint were 1.2, 1.6, and 1.4%, respectively. A total of 164 unique isolates (n = 58 from 1994-1995, 65 from 1997-1998, and 42 from 1999-2000) were examined for evidence of mutations in the quinolone resistance-determining regions (QRDRs) of the parC and the gyrA genes. Forty-nine isolates harbored at least one mutation in the QRDRs of one or both genes (1994-1995, n = 15; 1997-1998, n = 19; 1999-2000, n = 15). Among the 4,650 isolates of S. pneumoniae examined in the study, we estimated that 0.3% had mutations in both the parC and gyrA loci. The majority of mutations (67.3% of the mutations in 49 isolates with mutations) were amino acid substitutions in the parC locus only. Four isolates had a mutation in the gyrA locus only, and 12 isolates had mutations in both genes (8.2 and 24.5% of isolates with mutations, respectively). There was no significant difference in the number of isolates with parC and/or gyrA mutations detected during each study period. Finally, because of the magnitude of the study, we had reasonably large numbers of pneumococcal isolates with genotypically defined mechanisms of fluoroquinolone resistance and were thus able to determine the effects of specific resistance mutations on the activities of different fluoroquinolones. In general, isolates with mutations in parC only were resistant to ciprofloxacin but remained susceptible to levofloxacin, gatifloxacin, and moxifloxacin, whereas isolates with mutations in gyrA only and isolates with mutations in both parC and gyrA were resistant to all four fluoroquinolones tested.     

Activities of newer fluoroquinolones against ciprofloxacin-resistant Streptococcus pneumoniae

The incidence of ciprofloxacin resistance in Streptococcus pneumoniae is low but steadily increasing, which raises concerns regarding the clinical impact of potential cross-resistance with newer fluoroquinolones. To investigate this problem, we utilized an in vitro pharmacodynamic model and compared the activities of gatifloxacin, grepafloxacin, levofloxacin, moxifloxacin, and trovafloxacin to that of ciprofloxacin against two laboratory-derived, ciprofloxacin-resistant derivatives of S. pneumoniae (strains R919 and R921). Ciprofloxacin resistance in these strains involved the activity of a multidrug efflux pump and possibly, for R919, a mutation resulting in an amino acid substitution in GyrA. Gatifloxacin, grepafloxacin, levofloxacin, moxifloxacin, and trovafloxacin achieved 99.9% killing of both R919 and R921 in < or =28 h. With respect to levofloxacin, significant regrowth of both mutants was observed at 48 h (P < 0.05). For gatifloxacin, grepafloxacin, moxifloxacin, and trovafloxacin, regrowth was minimal at 48 h, with each maintaining 99.9% killing against both mutants. No killing of either R919 or R921 was observed with exposure to ciprofloxacin. During model experiments, resistance to gatifloxacin, grepafloxacin, moxifloxacin, and trovafloxacin did not develop but the MICs of ciprofloxacin and levofloxacin increased 1 to 2 dilutions for both R919 and R921. Although specific area under the concentration-time curve from 0 to 24 h (AUC(0--24))/MIC and maximum concentration of drug in serum (C(max))/MIC ratios have not been defined for the fluoroquinolones with respect to gram-positive organisms, our study revealed that significant regrowth and/or resistance was associated with AUC(0-24)/MIC ratios of < or =31.7 and C(max)/MIC ratios of < or =3.1. It is evident that the newer fluoroquinolones tested possess improved activity against S. pneumoniae, including strains for which ciprofloxacin MICs were elevated.     

Antimicrobial resistance of Campylobacter jejuni and Campylobacter coli with special reference to plasmid profiles of Japanese clinical isolates.

A total of 111 clinical isolates of Campylobacter jejuni and 10 clinical isolates of Campylobacter coli were characterized by their susceptibility to nine antimicrobial agents and by their plasmid profiles on agarose gel electrophoresis. All of the C. jejuni isolates were susceptible to chloramphenicol, ciprofloxacin, erythromycin, kanamycin, and nalidixic acid, but 55% were tetracycline resistant. In the 10 C. coli isolates, a high prevalence of multiple-antibiotic resistance was noted. Plasmids were found in 82% of the tetracycline-resistant and 15% of the tetracycline-susceptible C. jejuni isolates. Tetracycline resistance in six randomly selected C. jejuni isolates, which contained 50- or 135-kilobase (kb) plasmids, was transferred by conjugation to a Campylobacter fetus subsp. fetus recipient with recovery of a 50- or a 45-kb plasmid from transconjugants. From one multiple-antibiotic-resistant C. coli isolate, resistance to tetracycline, kanamycin, and chloramphenicol was transferred concomitantly with a 58-kb plasmid, pNR9589. Nonconjugative 98-kb plasmids, pNR9131 and pNR9581, from C. coli isolates with resistance to tetracycline, kanamycin, and erythromycin were shown by cloning experiments to code for at least kanamycin resistance. Restriction digests revealed that 50-kb plasmids from tetracycline-resistant C. jejuni isolates were identical, although plasmids from multiple-antibiotic-resistant C. coli isolates shared partial DNA homology to each other. Cloning of the kanamycin and chloramphenicol resistance genes of pNR9589 into Escherichia coli showed that the two genes are closely linked or clustered. Double-digestion analysis of the fragments encoding the kanamycin resistance of pNR9131, pNR9581, and pNR9589 showed that these three plasmids contain a common fragment related to kanamycin resistance.