This retrospective multicenter study showed a dose-response relationship between antimicrobial use and resistance in uropathogens in older adults. Our results demonstrate, for the first time in human clinical isolates, that the oral route of administration is associated with an increased likelihood of resistance compared to the parenteral route, provided the number of prescriptions (week courses) is below 10 (over one and a half year observation time). This is in full agreement with animal experimental studies in rodents for E. coli exposed to betalactams and tetracyclines , and earlier findings in a randomized control field trial in cattle . The effect of route of administration moreover interacted with the number of preceding prescriptions. Up to 9 prescriptions, when other variables held constant, probability of resistance decreased by increase in proportion of preceding parenteral antibiotic prescriptions. Since, seemingly, resistance gets organized after some threshold, possibly by reorganization of resistance at the molecular level, a different pattern was observed for samples with more than 9 prescriptions.
Comparison with the literature
Recently, a study comparing resistance in faecal E. coli from different groups of children (healthy, cancer, cystic fibrosis), suggested that aminopenicillin administered intravenously had only a modest effect on selection of intestinal resistance in cancer patients and possibly less impact than oral administration, which was the main route of administration of aminopenicillin to children with cystic fibrosis .
Apart from these studies, relatively little attention has been recently given to the route of administration and its particular influence on antimicrobial resistance. One exception is the stimulation to switch from intravenous to oral formulations (IV/PO switch) as soon as possible in acute care hospitals to reduce length of stay, treatment costs, and central line associated infections . Our results should trigger research to examine the influence of this switch on the selection of antimicrobial resistance. A recent investigation in Switzerland has shown that in acute care practice such a switch, leads to a two-step broad spectrum selection pressure, with an oral exposure of predominantly amoxicillin clavulanic acid, and to a lesser extent fluoroquinolones or clindamycine . This impact on resistance in pathogens and commensals thus is substantial and we therefor plea to examine this in defined case control settings and larger at the population level. Infection control intervention studies likewise should include antimicrobial resistance data of pathogens, preferably over consecutive years . Of notice, the IV/PO switch rationale is also fundamentally contradictory to the general mutation prevention theory. This theory states that a short high (loading) dose, followed by regular dosing intervals during an as short as possible time period is able to minimizes the resistance selection pressure while maintaining clinical efficacy [6, 19, 20]. Our study was conducted to assess such dynamics and persistence for E. coli retrieved from urine samples in the older adults (retired population). We assume many of these patients were suspected or confirmed to have a urinary tract infection. For urinary tract infections in women, a Cochrane review published in 2002 has shown that a reduction of treatment duration is feasible without impairing clinical efficacy and therefore should be encouraged to minimize the development and spread of resistance . This is in line with our observations when using the number of week courses as a proxy for treatment duration and should further be stimulated in general and specialized practice.
The urinary tract mostly gets infected with E. coli by retrograde infection from commensal faecal bacteria. Each time an inappropriate antimicrobial therapy is initiated in the individual patient resistant genes can be selected. Other risk factors for developing drug resistant UTI include previous antimicrobial exposure, long-term care residence, older age and comorbidities such as diabetes . Dutch investigators have also identified other medication and diet, including animal derived food, to be a risk factor for resistance in bacteria involved in UTI . Bacteria can obtain antimicrobial resistant genes either by mutation or by acquisition f romneighboring bacteria (horizontal transfer). This has formerly been investigated and well documented for E. coli, both as a commensal [10, 23] and an invasive pathogen organism (e.g. EARS-net). This resistance selection process and maintenance after withdrawal of antibiotic pressure (i.e. persistence) can further be stimulated or driven by unrelated antimicrobial agents (co-selection) . Despite that minimal inhibitory concentration determinations are the golden standard, under routine laboratory conditions, in E. coli and many other fast growing organisms, disk diffusion tests have for long been the method to simultaneously determine susceptibility profiles for a wide variety of antimicrobial agents. For these reasons, the antimicrobial resistance index (ARI, [10, 24]) was used as primary outcome variable. It can theoretically even take subtle changes in the antibiogram into account, and merges selection pressure effects of virtually all antimicrobial agents used including co-selection by unrelated organisms. It has been shown to be strongly correlated with treatment incidences expressed as prescribed or administered daily dosages (PDD & DDD) at different population levels and settings [24,25,26]. Causal relationships between DDD and antimicrobial resistance have been found in single center longitudinal studies .
It was observed that patients who died during the study period were on average, more likely to have strains that were resistant to antibiotics. This is in line with similar observations in other bacteria . The effect of number of days between the sample and the last prescription (log(time)) was significant and negative; indicating that probability of resistance is higher in the days after the treatment and decreases over time, confirming earlier findings in bacteria retrieved from the respiratory tract . We further observed a high variability in ARI across participating laboratories and this demands further research in terms of validation of antimicrobial resistance surveillance.
Strengths and limitations
The study has several limitations, like the voluntary participation of the laboratories, the reliability of the(ir) disk diffusion tests, the lack of information on co-morbidities of the patients, the applied dosage assessment  and the absence of compliance information with regard to the prescribed antibiotics, and the unknown selection criteria related to patients that undergo laboratory examinations of the urinary tract. Selection bias due to inter-laboratory and gender driven differences in the panel of antimicrobial agents tested also could have influenced the analysis. Deviations in dosage regimens that might interfere with the resistance selection could not be identified with the applied methodology. Also patients not receiving antibiotics were excluded in the current study design. The latter information could be used to assess a baseline level of resistance as our study group earlier explored for the respiratory tract system . Since also antimicrobial agents prescribed for indications other that urinary tract infections were included in the analysis it seems reasonable to conclude that resistance selection pressures are not restricted to one organ system, given the potential effect of antimicrobial agents on the digestive tract  and thereby indirectly on organisms shed by stool that can cause urinary tract infections. Also all prescriptions were considered in our analysis because of co-selection due to linked resistance genes as demonstrated for E. coli . An additional confounder that potentially could have driven the selection between oral or injectable administrations is the difference between empirical, prophylactic and microbiologically directed regimes. In a European study in long term care facilities executed in 2009 , empirical treatments were most common (54.4%), followed by prophylactic (28.8%) and microbiologically documented (16.1%) regimes . It is also recommended to repeat the analysis to confirm the finding by prospective randomized and controlled studies and in other study populations. Moreover, undesired effects of the switch of formulations, as recommended by several international guidelines on antimicrobial stewardship, should be considered in further studies.