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Objectives Our aim was to investigate the emergence and spread of ciprofloxacin resistance in clinical Neisseria gonorrhoeae isolates in New South Wales, Australia, from the first reported case in 1991 until ciprofloxacin resistance was sustained at or above the WHO threshold for treatment change of 5% (1999), to inform future strategies for controlling gonococcal antimicrobial resistance. Methods The index isolate and all subsequent clinical isolates of ciprofloxacin-resistant N. gonorrhoeae in New South Wales from 1991 to 1999 were genotyped using a previously described method on the Agena MassARRAY iPLEX platform. Region of acquisition data, where available, were used to determine whether cases were travel associated. Results In New South Wales, of the 325 ciprofloxacin-resistant N. gonorrhoeae isolates reported from 1991 to 1999, 98% (320/325) were able to be recovered and 100% (320/320) were genotyped. There were 66 different genotypes, comprising 1–99 isolates each. Notably no single clone was found to account for ciprofloxacin resistance being sustained in the population, with considerable variability in genotype prevalence observed throughout the study period. A total of 65% (209/320) of genotyped isolates had information regarding the likely place of acquisition; of these, 44% (93/209) were associated with overseas travel or sexual contact with an overseas visitor. The first ciprofloxacin-resistant N. gonorrhoeae in New South Wales was associated with travel to Thailand. Index cases of each resistant genotype were significantly more likely to have been acquired overseas (51.5%), predominantly in Asia (45%, 30/66). Conclusions The continued importation of multiple genotypes, rather than the expansion of a single genotype, led to ciprofloxacin-resistant N. gonorrhoeae being established in New South Wales.

A patient's prior urine cultures are often considered when choosing empiric antibiotic therapy for a suspected urinary tract infection. We sought to evaluate how well previous urine cultures predict the identity and susceptibility of organisms in a patient's subsequent urine cultures.We conducted a multinational, multicenter, retrospective cohort study, including 22 019 pairs of positive urine cultures from 4351 patients across 2 healthcare systems in Toronto, Ontario, and Chicago, Illinois. We examined the probability of the same organism being identified from the same patient's positive urine culture as a function of time elapsed from the previous positive urine specimen; in cases where the same organism was identified we also examined the likelihood of the organism exhibiting the same or better antimicrobial susceptibility profile.At 4-8 weeks between cultures, the correspondence in isolate identity was 57% (95% confidence interval [CI], 55%-59%), and at32 weeks it was 49% (95% CI, 48%-50%), still greater than expected by chance (P.001). The susceptibility profile was the same or better in 83% (95% CI, 81%-85%) of isolate pairs at 4-8 weeks, and 75% (95% CI, 73%-77%) at32 weeks, still greater than expected by chance (P.001). Despite high local rates of ciprofloxacin resistance in urine isolates across all patients (40%; 95% CI, 39.5%-40.5%), ciprofloxacin resistance was20% among patients with a prior ciprofloxacin sensitive organism and no subsequent fluoroquinolone exposure.A patient's prior urine culture results are useful in predicting the identity and susceptibility of a current positive urine culture. In areas of high fluoroquinolone resistance, ciprofloxacin can be used empirically when prior urine culture results indicate a ciprofloxacin-susceptible organism and there has been no history of intervening fluoroquinolone use.

Aims: There has been an increasing number of pathogens becoming resistant to multiple classes of antibiotics. The study on how mutation emerges is therefore crucial to promote further understanding in this area. Conventional methods for such studies involve the monitoring of growth by standard plate count and biomolecular sequencing. This is however tedious and not cost effective. The aim of this paper is thus to introduce a novel system that enables real-time monitoring of bacterial ‘mutation-in-progress’. Methods and Results: This system provides real-time data, thus enabling confirmatory and further work to be performed at the important points when mutation is initiated. The system integrates spectroscopic techniques as the detection system and various supporting systems, such as a nutrient replenishing system, a pH control system and a waste system to allow for extended monitoring. In this paper, the feasibility of monitoring the emergence of ciprofloxacin resistance in Staphylococcus aureus was demonstrated as an initial example. The integrated system was found to require significantly less material resource and manpower compared with conventional techniques. Conclusions: The novel system to monitor bacterial mutation-in-progress is presented. The work reported herein demonstrates such a system to be effective and efficient in performing real-time monitoring of mutation-in-progress, especially in extended time frames for mutation into the weeks and months. Significance and Impact of the Study: With the successful optimization of this system, researchers can learn about the dynamics of antibiotic resistance and further understand how the mutation of bacteria occurs.

The plasmid-mediated quinolone resistance qnr genes in clinical isolates in adults have been described in different countries; however, the frequency of their occurrence has not been detected in pediatric patients. A total of 410 clinical isolates of Klebsiella pneumoniae, identified as producers of an extended-spectrum beta-lactamase (ESBL), or AmpC beta-lactamase, were collected from five children's hospitals in China during 2005-2006. The isolates were screened for the presence of the qnrA, qnrB, and qnrS genes, and then the harboring qnr gene isolates were detected for a bla gene coding for the TEM, SHV, CTX-M, and plasmid-mediated ampC gene by a PCR experiment. Ninety-two isolates (22.7%) were positive for the qnr gene, including 10 of qnrA (2.4%), 25 of qnrB (6.1%), and 62 of qnrS (15.1%). Eighty-one of the 92 (88.0%) qnr-positive isolates carried at least one bla gene for TEM, SHV, CTX-M, or DHA-1. The ciprofloxacin resistance increased 16-256-fold and oflaxacin resistance increased 2-32-fold in transconjugants, respectively. These results indicated that the plasmid-mediated qnr quinolone resistance gene was qnrS, followed by qnrB and qnrA. Most of the isolates also carried a bla gene coding ESBL or ampC gene coding DHA-1 among Klebsiella pneumoniae isolated from Chinese pediatric patients.

Sir, The increase in resistance of Gram-negative bacilli is a serious public health problem and our aim was to determine the effect of different duplicate elimination criteria on the evolution of fluoroquinolone resistance in Escherichia coli and Klebsiella pneumoniae in our setting over a period of 18 years. The criteria used are described in Figure 1. Antibiotic susceptibility was studied using microdilution with the Wider semiautomatic system (Soria Melguizo, Spain) and Etest (AB Biodisk, Solna, Sweden), following CLSI (formerly NCCLS) recommendations. In E. coli, the mean percentage of fluoroquinolone-resistant strains according to the EARS-Net criterion was greater than that detected using the CLSI criterion, both when the total number of strains and when the number of extended-spectrum b-lactamase (ESBL)-producing strains was considered (28.4% versus 22.4% and 65.2% versus 60.5%, respectively). With regard to K. pneumoniae, we obtained 7.2% versus 6.3% and 52.2% versus 40.0%. Analysis of the criterion of time showed that when the period considered for elimination of duplicates increased, the ciprofloxacin resistance rate also increased compared with the CLSI criterion. When the EARS-Net criterion was applied, the prevalence of these microorganisms increased in both hospitalized and non-hospitalized patients, although this phenomenon was more important in the former than in the latter. All data are shown in Table 1. The increase in fluoroquinolone resistance is a serious public health problem, especially in ESBL-producing strains, for which the therapeutic options are far more limited, and the local epidemiology of resistance has been considered a key tool in the control of this process. It is essential to apply a duplicate elimination criterion, since analysis of the data based on the total criterion does not reflect the real situation of resistance. What has not yet been definitively established is which criterion is the best to use in each case. The first isolate from each patient is an objective criterion, but has serious limitations since the same patient may frequently be infected by different strains of the same species during the course of their disease and hospital stay, or a strain may become resistant during treatment of the patient due to the selection of resistant mutants during antibiotic therapy. The EARS-Net criterion reflects more faithfully the real situation of resistance, but suggests that it is necessary to apply strict quality control in phenotypic microbiological studies to detect resistance. Our study suggests that the new EARS-Net method of studying duplicates should be applied in order to detect changes in resistance patterns that may occur during the course of disease due to mutations of the microorganism causing the infection or superinfections by more resistant microorganisms.

UK clinical laboratories overestimate ciprofloxacin resistance amongst Pseudomonas aeruginosa isolates, relative to the MIC breakpoint of 1 mg/L. Most tests leading to this overestimation use 1 microg discs and are by Stokes' method with the breakpoint taken as the zone radius for P. aeruginosa NCTC 10662 minus 3 mm. Aiming to reduce this error rate, we examined alternative disc breakpoints. Tests were performed for 100 P. aeruginosa isolates on three media, with breakpoints selected (i) as the zone for P. aeruginosa NCTC 10662 minus 7 mm, as recommended for ciprofloxacin by the BSAC; (ii) with reference to MIC/zone correlation lines; (iii) from natural divisions in zone distribution histograms; and (iv) so as to minimize categorization errors. Breakpoints from regression lines, and those optimized to the susceptibility distribution, reduced the proportion of susceptible organisms misreported as resistant, but the improvement was not significant (P0.05, chi2 test). The breakpoint of the zone radius for P. aeruginosa NCTC 10662 minus 7 mm significantly reduced (P0.05) the number of susceptible organisms reported as resistant, but led to 50-75% of those with low level resistance (MIC 2-4 mg/L) and 4-10% of those with high-level resistance (MIC4 mg/L) being classed as susceptible. Irrespective of the medium and the basis of choosing breakpoints, 5 microg ciprofloxacin discs gave a lower rate of susceptible organisms being reported as resistant than did 1 microg discs; however, the improvement was not significant (P0.05, chi2 test) and the 5 microg discs had the disadvantages of forming very large zones for susceptible isolates and giving some--albeit small--zones for highly resistant organisms. In conclusion, the over-reporting of resistance could be reduced by use of zone breakpoints optimized to the MIC distribution and by the use of 5 microg discs, but the case for these changes is not overwhelming; taking the breakpoint as the zone for NCTC 10662 minus 7 mm led to unacceptable numbers of resistant organisms being reported as susceptible. More fundamentally, ciprofloxacin zones and MICs are continuously distributed for P. aeruginosa isolates, so susceptibility tests cannot divide the species into discrete populations. In these circumstances, it is optimistic to expect disc and MIC categorizations to agree perfectly.

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