Laura Gualco

The ARESC study: an international survey on the antimicrobial resistance of pathogens involved in uncomplicated urinary tract infections

The ARESC (Antimicrobial Resistance Epidemiological Survey on Cystitis) study is an international survey to investigate the prevalence and susceptibility of pathogens causing cystitis. Female patients (n=4264) aged 18-65 years with symptoms of uncomplicated cystitis were consecutively enrolled in nine European countries as well as Brazil during 2003-2006. Pathogens were identified and their susceptibility to nine antimicrobials was determined. Escherichia coli accounted for 76.7% of isolates. Among E. coli, 10.3% of the isolates were resistant to at last three different classes of antimicrobial agents. Resistance was most common to ampicillin (48.3%), trimethoprim/sulfamethoxazole (29.4%) and nalidixic acid (18.6%). Fosfomycin, mecillinam and nitrofurantoin were the most active drugs (98.1%, 95.8% and 95.2% susceptible strains, respectively) followed by ciprofloxacin, amoxicillin/clavulanic acid and cefuroxime (91.7%, 82.5% and 82.4%, respectively). Resistance to ciprofloxacin was >10% in Brazil, Spain, Italy and Russia. Overall, Proteus mirabilis were more susceptible to beta-lactams and less susceptible to non-beta-lactams than E. coli, whereas Klebsiella pneumoniae strains, which are intrinsically resistant to ampicillin, were less susceptible to mecillinam (88.8%), fosfomycin (87.9%), cefuroxime (78.6%) and nitrofurantoin (17.7%). Resistance was rare in Staphylococcus saprophyticus, with the exception of ampicillin (36.4%) and trimethoprim/sulfamethoxazole (10.2%). In Italy, Spain, Brazil and Russia, the countries most affected by antimicrobial resistance, extended-spectrum beta-lactamase (ESBL) enzymes (mainly CTX-M type) were detected in 48 strains (39 E. coli, 6 K. pneumoniae and 3 P. mirabilis). Despite wide intercountry variability in bacterial susceptibility rates to the other antimicrobials tested, fosfomycin and mecillinam have preserved their in vitro activity in all countries investigated against the most common uropathogens.

European Emergence of Ciprofloxacin-Resistant Escherichia coli Clonal Groups O25:H4-ST 131 and O15:K52:H1 Causing Community-Acquired Uncomplicated Cystitis

ABSTRACT A total of 148 E. coli strains displaying reduced susceptibility to ciprofloxacin (MIC ≥ 2 μg/ml) and causing uncomplicated urinary tract infections in eight European countries during 2003 to 2006 were studied. Their phylogenetic groups, biochemical profiles, and antibiotic susceptibilities were determined. Determination of the O:H serotype, pulsed-field gel electrophoresis (PFGE), randomly amplified polymorphic DNA (RAPD) PCR, and multilocus sequence typing provided additional discrimination. The majority (82.4%) of the microorganisms (122/148) carried resistance to two or more additional drugs, with the pattern ciprofloxacin-trimethoprim-sufamethoxazole-tetracycline-ampicillin being the most represented (73 strains out of 148; 49.3%). Extended-spectrum beta-lactamase production was detected in 12/148 strains (8.1%), with CTX-M-15 being the most-common enzyme. Six strains out of the whole collection studied (4.0%) contained a qnrB-like gene. Overall, 55 different PFGE or RAPD PCR profiles could be distinguished, indicating a substantial heterogeneity. However, about one-third (51/148) of the strains belonged to two clonal groups: O15:K52:H1 (phylogenetic group B2, lactose-nonfermenting variant, ciprofloxacin MIC of 16 μg/ml) and O25:H4 sequence type 131 (ST-131) (phylogenetic group D, ciprofloxacin MIC of ≥32 μg/ml). With the exception of Poland, strains of these two groups were isolated in samples from all participating countries but more frequently in samples from Spain and Italy. In some representative strains of the two main clonal groups, alterations in GyrA and ParC were the basic mechanism of fluoroquinolone resistance. In some members of the O25:H4 ST-131 group, displaying a ciprofloxacin MIC of >32 μg/ml, additional OmpF loss or pump efflux overexpression was found. In the Mediterranean area, strains belonging to these two clonal groups played a major role in determining the high rate of fluoroquinolone-resistant E. coli strains observed in the community.

Effect of fosfomycin alone and in combination with N-acetylcysteine on E. coli biofilms

Four slime-producing uropathogenic Escherichia coli strains were used to investigate the activity of fosfomycin and N-acetylcysteine (NAC) against biofilms developed on 96-well polystyrene tissue culture plates. Biofilms aged, respectively, 5 (initial) and 48 h (mature) and two fosfomycin concentrations (128 and 2000 mg/l) were used. The effect of various levels (0.007-8 mg/ml) of NAC alone and in combination with fosfomycin on the formation or disruption of biofilms was assessed. Following exposure to the drugs, the percentage of residual slime relative to the control, ranged from 62.5-100 to 26.2-64.1% in the presence of 0.007 and 8 mg/ml of NAC. After treatment of pre-formed biofilms with NAC at the highest concentrations used, the remaining exopolysaccharide matrix was reduced to 25-68% of the amount found with the untreated control. Exposure to fosfomycin at 2000 mg/l reduced biofilms 40-57 and 41-49% for the initial and mature forms, respectively. Fosfomycin was more active at 2000 mg/l combined with NAC 2 mg/ml. Under these conditions initial and mature biofilms were reduced 66-80 and 60-73%, respectively. NAC, when used in combination, enhanced fosfomycin bactericidal activity producing a 99-99.9% reduction in viable cells. Fosfomycin and NAC at concentrations achievable in urine displayed a synergistic effect promoting both the formation of biofilms and reduction of sessile cell viability.

In vitro activity of fosfomycin against Gram-negative urinary pathogens and the biological cost of fosfomycin resistance

The aim of this study was to reassess the activity of fosfomycin against recently isolated uropathogens circulating in Italy and to evaluate the effect of fosfomycin resistance on the expression of several virulence traits using the rare mutant strains. In vitro activity of fosfomycin was evaluated using 441 Gram-negative organisms isolated from patients with uncomplicated urinary tract infections (UTIs). Fosfomycin was the most active antibiotic against Escherichia coli (99% susceptibility). The activity against Proteus mirabilis was more potent than that of co-trimoxazole and nitrofurantoin (87.5, 67 and 0% susceptibility, respectively). The other microorganisms, accounting for about 7% of all pathogens tested, showed variable susceptibilities to fosfomycin. Compared with susceptible strains, fosfomycin-resistant mutants showed a reduced rate of growth and were impaired in their ability to adhere to uroepithelial cells and to urinary catheters. They were also more resistant to UV irradiation and to phage T7 and showed diminished rates of colicin synthesis and transfer of plasmids.

Molecular analysis and susceptibility patterns of meticillin-resistant Staphylococcus aureus (MRSA) strains circulating in the community in the Ligurian area, a northern region of Italy: emergence of USA300 and EMRSA-15 clones

For many years meticillin-resistant Staphylococcus aureus (MRSA) has been considered a typical nosocomial pathogen. Recently, MRSA has emerged as a frequent cause of infections in the community. A multicentre surveillance study was carried out in the Ligurian area of Italy to evaluate the incidence, molecular nature and susceptibility patterns of MRSA strains circulating among outpatients. The genetic background of MRSA strains was analysed by pulsed-field gel electrophoresis (PFGE) and multilocus sequence typing (MLST). Determination of antimicrobial susceptibility patterns, staphylococcal cassette chromosome mec (SCCmec) type, accessory gene regulator (agr) group and Panton-Valentine leukocidin (PVL) production was also performed. In total, 12 (6.4%) of 188 S. aureus isolates collected during 2006-2007 were found to be MRSA by phenotypic and genotypic methods. Analysis of isolates by PFGE showed that the majority of strains (11/12) belonged to two well-known international clones (EMRSA-15 and USA300) and their variations. High variability regarding SCCmec IV subtypes, susceptibility patterns and PVL toxin production were found among members of the USA300 clonal group, even when displaying the same PFGE profiles. The remaining MRSA strain belonged to sequence type (ST) 8, agr group I and carried SCCmec type I. Both community-associated MRSA and healthcare-associated MRSA epidemic international clones circulate among outpatients in our region. It is alarming that members of the most represented clonal group in our collection (USA300) can acquire multiresistance as well as PVL genes. Infection control measures in our area should be improved to avoid the selection of microorganisms displaying both traits simultaneously as well as the spread of these epidemic international clones.

Epidemiology of Major Respiratory Pathogens

Abstract A vast literature attests to the fact that Streptococcus pneumoniae, Haemophilus influenzae and Moraxella catarrhalis represent the prevailing bacterial pathogens of community-acquired lower respiratory tract infections. Their specific incidence as causative agents of the more common syndromes is known to vary even profoundly, depending on geographic area, and the same holds true for their rates of resistance to antimicrobial drugs. Europe does not escape the threat posed by the present pandemic spread of penicillin resistance in S. pneumoniae although, as expected, some countries like Spain and France are highly affected and others including Germany, Italy, the Netherlands and the Scandinavian region, are relatively spared. In several sites multiple resistance has been described in S. pneumoniae with the most affected drugs being penicillin, the macrolides, co-trimoxazole and tetracycline. In H. influenzae synthesis of β-lactamases is the main resistance trait expressed. Lack of susceptibility to β-lactams dictated by a different mechanism remains extremely rare. Large variations in the incidence of this character are apparent when considering European countries. France and Spain are again widely affected while Germany, the Netherlands and Italy display rates of β-lactamase-positive H. influenzae of about 16%. M. catarrhalis must be considered generally resistant to non-protected aminopenicillins since over 90% of these organisms produce β-lactamases.

In Vitro Activity of Thiamphenicol Against Multiresistant Streptococcus pneumoniae, Haemophilus influenzae and Staphylococcus aureus in Italy

Abstract Thiamphenicol is a derivative of chloramphenicol characterized by a spectrum comparable to that of the parent compound against multiresistant pathogens but showing satisfactory tolerability. The In Vitro activity of thiamphenicol and of 11 comparative drugs against 397 recently isolated antibiotic-resistant and/or invasive pneumococci and 52 multiply-resistant MRSA including 2 VISA strains was determined. Bactericidal activity against Haemophilus influenzae and the post-antibiotic effect on Streptococcus pneumoniae, H. influenzae, Staphylococcus aureus and Escherichia coli were also assessed. Against invasive pneumococci, thiamphenicol and chloramphenicol were the most potent non-β-lactam molecules together with vancomycin and rifampin. Against high-level penicillin-resistant strains phenicol activities were superior to those of cefotaxime, ceftriaxone and imipenem. Against MRSA thiamphenicol and chloramphenicol were second only to the glycopetides and also inhibited the VISA strains. Thiamphenicol showed a significant PAE (0.33 to 2.9h) on all pathogens studied and a powerful bactericidal effect against β-lactamase-positive and -negative H. influenzae. These results indicate a good In Vitro activity of thiamphenicol against difficult-to-treat multiply resistant pathogens.

Do Different Susceptibility Breakpoints Affect the Selection of Antimicrobials for Treatment of Uncomplicated Cystitis

Abstract Because of increasing antibiotic resistance in Escherichia coli, the main uropathogen of uncomplicated urinary tract infections (UTIs), updated susceptibility data are vital in guiding the selection of first-line treatment agents. interpretation of these data depends on the breakpoints adopted, that may vary among different guidelines. In this study we report the minimum inhibitory concentrations (MICs) of eight antibiotics and compare antimicrobial susceptibility results obtained in 2315 E. coli strains recently collected during the ARESC survey using EUCAST and CLSI breakpoints. We have also evaluated the clinical impact of breakpoint discrepancies on the overall susceptibility patterns. Fosfomycin, nitrofurantoin and mecillinam showed the highest susceptibility rates in all countries (>92%) according to both CLSI and EUCAST criteria. Minor category shifts were observed for ciprofloxacin, amoxicillin-clavulanic acid, ampicillin and trimethoprim/sulfamethoxazole. A large number of strains classified as intermediate resistant to cefuroxime according to CLSI are included by the EUCAST in the susceptible category. In conclusion, fosfomycin, mecillinam, and nitrofurantoin have preserved their in vitro activity in all countries investigated, regardless of the criteria adopted. They continue to represent effective options for the empiric therapy of female patients with uncomplicated cystitis. The use of different interpretative criteria for E. coli responsible for Utis therefore has no influence on the decision to be taken by the physicians managing the patients.

In vitro activity of ceftibuten at sub-inhibitory concentrations in comparison with other antibiotics against respiratory and urinary tract pathogens.

Abstract Some new features of the In Vitro activity of ceftibuten, an oral third generation cephalosporin, have been studied in reference to respiratory and urinary tract pathogens included in its antibacterial spectrum. At 0.25XMIC (minimum inhibitory concentration) and 0.5XMIC levels, ceftibuten was able to affect the biofilm production in 2/3 of both Escherichia coli and Proteus mirabilis strains, and reduced the number of strains capable of adhering to epithelial cells by about 35% in comparison to the control. Surface hydrophobicity was also influenced by ceftibuten and the other drugs at 0.25-0.5XMIC. In general, no marked variation in the virulence traits of the pathogens studied were found by exposing bacteria to sub-MICs of ceftibuten. Plasmid loss (from 1.8 to 37.2%), and Flac transfer inhibition (about 30-50% reduction in the number of recombinants) were detected under the experimental conditions used. This study confirms the excellent antibacterial properties of ceftibuten by adding new information about the effects of this antibiotic against pathogens often involved in respiratory and urinary tract infections that may be treated with this compound, supporting the appropriate use of this cephalosporin.

Microbial Epidemiology Patterns of Surgical Infection Pathogens

Abstract Resistance, as assessed in vitro, has a number of serious consequences in clinical situations. Treatment failures are common when an inappropriate drug has been prescribed and this, in turn, may lead to hospitalization of patients who normally would have been treated on an outpatient basis, as well as to longer hospital stay for inpatients and to the use of alternative drugs, which may be more expensive and more toxic. These factors all contribute to increased health care costs, morbidity and mortality. Microbiological procedures may identify the causative pathogen and provide the appropriate susceptibility pattern to the physician, thus reducing the chances of therapeutic failures. However, for a number of reasons including cost—even in hospitals—not to mention general practice, infections are seldom diagnosed on an etiological basis. From what has been stated, the knowledge of bacterial epidemiology and resistance represents basic support for correct therapeutic decision-making.