Lorena Huelves

Correlation between virulence factors and in vitro biofilm formation by Escherichia coli strains.

The ability of 15 Escherichia coli strains to form biofilms on polystirene plates was studied. The strains were serotyped, and their phenotypic expression of surface virulence factors (VFs), and antibiotic susceptibility was also determined. Moreover, 30 VFs-associated genes were analysed, including 15 adhesins (papC, papG and its three alleles, sfa/focDE, sfaS, focG, afa/draBC, iha, bmaE, gafD, nfaE, fimH, fimAvMT78, agn43, F9 fimbriae and type 3 fimbriae-encoding gene clusters), four toxins (hlyA, cnf1, sat and tsh), four siderophore (iron, fyuA, iutA and iucD), five proctetins/invasion-encoding genes (kpsM II, kpsMT III, K1 kps variant- neuC, traT and ibeA), and the pathogenicity island malX and cvaC. Morphological appearance and thickness of biofilms of two strong and three weak biofilm producers were also studied by confocal laser scanning microscopy (CLSM). Seven strains were classified as strong biofilm producers and the remaining eight strains were regarded as weak biofilm producers. Mannose-resistant haemagglutination was the only phenotypically expressed surface virulence factor more frequently found in the strong biofilm group. Five virulence-associated genes were more common (p<0.05) in strong biofilm producers: papC and papG alleles, sfa/focDE, focG, hlyA and cnf1. CLSM images showed irregular biofilms with projections at the top mainly in strong biofilm.

Measurement of biofilm formation by clinical isolates of Escherichia coli is method-dependent

Aims:  In this study, we have evaluated the impact of methodological approaches in the determination of biofilm formation by four clinical isolates of Escherichia coli in static assays.

Effects of human serum albumin, ibuprofen and N-acetyl-l-cysteine against biofilm formation by pathogenic Escherichia coli strains

The aim of this study was to evaluate the effects of human serum albumin (HSA), ibuprofen sodium (IBU) and N-acetyl-L-cysteine (NAC) against biofilm formation by seven biofilm-producing strains of Escherichia coli. Biofilm formation was studied using polystyrene microtitre plates in static conditions. The impact of the three compounds on bacterial growth and biofilm formation was tested by applying each compound in solution and as pre-treatment (coating) of polystyrene wells. When studied in solution, the minimum biofilm inhibitory concentrations of HSA, IBU and NAC were 8 mg/L (all strains), 2-125 mg/L (five strains) and 30-125 mg/L (five strains), respectively. Pre-treatment of polystyrene plates with HSA at 8 and 32,000 mg/L significantly reduced biofilm formation by all strains, whereas coating with 125 mg/L IBU and 1000 mg/L NAC did not. When HSA at 8 and 32,000 mg/L was combined with either 125 mg/L IBU or 1000 mg/L NAC in pre-treatment assays, more potent inhibition of biofilm was observed for some strains. Our results suggest that biofilm formation by E. coli may be prevented by coating medical devices with HSA alone or in combination with IBU or NAC. In addition, IBU and NAC could be useful in the treatment of urinary tract infections caused by E. coli due to their inhibitory effect on both bacterial growth and biofilm formation.

Antimicrobial susceptibility of Streptococcus pyogenes in Central, Eastern, and Baltic European Countries, 2005 to 2006: the cefditoren surveillance program.

The in vitro activity of penicillin, ampicillin, cefditoren, cefotaxime, erythromycin, clarithromycin, and levofloxacin against 763 clinical isolates of Streptococcus pyogenes was determined. Clinically significant isolates collected from November 2005 to December 2006 in the Czech Republic, Slovakia, Hungary, Poland, Romania, Estonia, Latvia, and Lithuania (the latter 3 analyzed as Baltic countries) were studied. No resistance to beta-lactams and levofloxacin was found. The rate of erythromycin resistance in S. pyogenes varied among countries, being low (<10%) in Romania and Baltic countries, intermediate (10-20%) in Poland and Czech Republic, and high (>25%) in Hungary and Slovakia. The predominant (75.0%) erythromycin-resistant phenotype among S. pyogenes isolates was MLS(B). The identification of the prevalence of erythromycin resistance mechanism could have impact on the choice of empiric antibiotic therapy for the clinicians in such countries.

In vitro activity of ciprofloxacin, moxifloxacin, vancomycin and erythromycin against planktonic and biofilm forms of Corynebacterium urealyticum

OBJECTIVES To study the ability of Corynebacterium urealyticum to produce biofilms and to compare the in vitro activity of antimicrobials against planktonic and biofilm-associated organisms. METHODS Biofilm formation on polystyrene plates by three C. urealyticum strains was studied in artificial urine under static conditions. The bactericidal activities of ciprofloxacin, moxifloxacin, vancomycin and erythromycin were studied against biofilm-associated organisms, and the results were compared with those obtained against planktonic organisms. Persister biofilm-associated organisms of each strain exposed to antibiotics were retested to determine the MIC of the same antibiotic. RESULTS The three strains tested consistently produced biofilms. Planktonic organisms was susceptible to ciprofloxacin, moxifloxacin and vancomycin, and their MBC values were two to eight times higher than their corresponding MICs. Bactericidal effect on biofilm-associated organisms required very high antibiotic concentrations; the minimum biofilm bactericidal concentrations for ciprofloxacin, moxifloxacin and vancomycin ranged from 128 to > or =1024 times their respective MBCs for planktonic organisms. Erythromycin was not bactericidal against either planktonic or biofilm-associated organisms for the single susceptible strain tested. Persister biofilm-associated organisms exposed to erythromycin increased their MIC by a factor >8000, but no changes in susceptibility were observed with the other compounds. CONCLUSIONS This work demonstrates that C. urealyticum produces biofilms on polystyrene plates and biofilm-associated organisms are much less susceptible to the bactericidal effect of the antibiotics; and the exposure of C. urealyticum to erythromycin may favour resistance selection. Overall, these results may explain the difficulties for bacterial eradication in chronic infections caused by C. urealyticum.

Pneumococcal LytA Autolysin, a Potent Therapeutic Agent in Experimental Peritonitis-Sepsis Caused by Highly β-Lactam-Resistant Streptococcus pneumoniae

ABSTRACT The in vitro and in vivo antipneumococcal activities of the main pneumococcal autolysin (LytA) and Cpl-1, a lysozyme encoded by phage Cp-1, were studied. Intraperitoneal therapy with LytA or high-dose Cpl-1 remarkably reduced peritoneal bacterial counts (>5 log10 CFU/ml) compared with those for the controls. After intravenous injection, LytA was the most effective treatment.

Antimicrobial susceptibility of Haemophilus influenzae and Moraxella catarrhalis isolates in eight Central, East and Baltic European countries in 2005–06: results of the Cefditoren Surveillance Study

Sir, Cefditoren pivoxil is an aminothiazolyl oral cephalosporin with a broad spectrum of activity against Gram-positive and Gram-negative pathogens and is stable to hydrolysis by many common b-lactamases. It has been shown to be active in vitro against many respiratory tract pathogens including Haemophilus influenzae and Moraxella catarrhalis. Resistance of both organisms to b-lactam antibiotics is generally conferred by either the presence of b-lactamase or as a result of alterations in penicillinbinding protein 3. In this study, we compare the in vitro activity of cefditoren with that of nine other antibiotics against 665 and 133 clinical isolates of H. influenzae and M. catarrhalis strains collected from eight Central, East and Baltic European countries. The isolates were prospectively collected from 1 November 2005 to 31 December 2006, in 25 centres from the Czech Republic, Hungary, Poland, Romania, Slovakia, Estonia, Latvia and Lithuania. Isolates from Estonia, Latvia and Lithuania were analysed all together as ‘Baltic countries’. No isolates of M. catarrhalis were received from Romania. Strain isolates were from patients with upper (ear and sinus exudates) and lower (sputum, bronchial washes and bronchial aspirates) respiratory tract infections and bacteraemia. Duplicate isolates from the same patient were not accepted. The identity of the isolates was confirmed by morphology of the colonies, growth on blood and chocolate agar, and API-NH (bioMerieux, Marcy l’Etoile, France). The following data were collected for each isolate: age group (adults/children), sex, status (outpatients/inpatients) and sample origin (upper respiratory tract/lower respiratory tract/blood). Susceptibility testing was carried out by the CLSI broth microdilution method, using dried commercial plates (Sensititre, Trek Diagnostic Systems Ltd, West Sussex, UK) reconstituted with Haemophilus test medium for H. influenzae and cation-adjusted Mueller–Hinton broth for M. catarrhalis. The antimicrobials tested were ampicillin, amoxicillin, amoxicillin/ clavulanic acid, cefditoren, cefixime, cefpodoxime, cefuroxime, cefotaxime, clarithromycin and levofloxacin. Breakpoint concentrations published by the CLSI (document M100-S16, 2006) to interpret MIC data qualitatively for H. influenzae were also applied to M. catarrhalis. b-Lactamase production was determined using the chromogenic cephalosporin Cefinase test (bioMerieux). The results of the susceptibility testing of H. influenzae isolates are shown in Table 1. Of the 665 isolates tested, 7 (1.1%) were intermediate and 62 (9.3%) resistant to ampicillin. Among ampicillin-resistant isolates, 95.2% were b-lactamase producers. Susceptibility to ampicillin was higher in isolates from the Czech Republic, Slovakia, Hungary, Poland and Baltic countries and lower in those from Romania, the difference being statistically significant (P 0.005) when compared with any of the other countries. Three (4.8%) ampicillin-resistant isolates were categorized as b-lactamase-negative, ampicillin-resistant (BLNAR); all of them having an MIC of 4 mg/L. No isolate was b-lactamase-positive, amoxicillin/clavulanic-resistant (BLPACR). All isolates were susceptible to amoxicillin/clavulanic acid, cefixime, cefpodoxime, cefuroxime, cefotaxime and levofloxacin. All H. influenzae isolates were inhibited by 0.06 mg/L cefditoren. More than 98% of the isolates were susceptible to clarithromycin and none was categorized as intermediate. No differences in susceptibility to the antibiotics tested were found comparing source, localization of the patient, age or gender. One hundred and twenty-seven of the 133 (95.5%) M. catarrhalis isolates produced b-lactamase, but only 94 (70.7%) were non-susceptible to ampicillin using CLSI breakpoints. Susceptibility to ampicillin was higher in isolates from Poland than from any other country, but the difference was not statistically significant. All isolates were susceptible to amoxicillin/ clavulanic acid, cefixime, cefpodoxime, cefuroxime, cefotaxime, clarithromycin and levofloxacin. All M. catarrhalis isolates were inhibited by 0.5 mg/L cefditoren and the MIC50/MIC90 values were 0.12/0.5 mg/L. No differences in susceptibility to the antibiotics tested were found comparing source, localization of the patient, age or gender. The prevalence of H. influenzae isolates producing b-lactamase in Europe varies from low (,6%) in Germany, The Netherlands and Italy to high (.20%) in the UK, Spain and France. – 4 Reports on surveillance studies in our screened countries are scarce but give the following results: 7.9% to 13% in the Czech Republic, 4.8% to 6.5% in Slovakia, 0% to 3.3% in Hungary, 6% to 24% in Poland – 5 and 3.0% in Estonia. Data on the prevalence of BLNAR H. influenzae isolates are variable but prevalence is usually low (,10%), – 4 although some reports have given figures of up to 20% in Poland. No BLPACR isolates were detected in the present study, and the prevalence of such organisms being very low. A high rate (95.5%) of b-lactamase-producing M. catarrhalis isolates was found in our study, with no significant differences Journal of Antimicrobial Chemotherapy doi:10.1093/jac/dkn083

Adherence of Streptococcus pneumoniae to polystyrene plates and epithelial cells and the antiadhesive potential of albumin and xylitol.

Aimed to prevent Streptococcus pneumoniae biofilm infections, we studied the adherence of nine pneumococcal strains to polystyrene plates and on epithelial cells and the antiadhesive effect of albumin and xylitol. The adherence was variable among strains, but there was a good correlation between their adherent ability and binding to abiotic material and cells. Strains of serotypes 6B and 23F were the most adherent organisms, whereas serotype 3 strains were the least adherent. Human serum albumin (HSA) enhanced bacterial growth at low concentrations (0.5–2.5%) but inhibited it at 10%. Xylitol inhibited bacterial growth of all strains at concentrations ranging from 5 to 15%. Exposure to 0.5–5% HSA in solubilized form and to 5% HSA precoating of plates diminished adherence to polystyrene >80% for all strains, except for serotype 3 strains. Contrarily, 0.5 and 5% xylitol did not diminish significantly pneumococcal adherence to polystyrene plates or on epithelial cells. Our results suggest that 1) the potential application of HSA coatings on medical devices to inhibit pneumococcal adherence and 2) the possible beneficial effect of xylitol in preventing some pneumococcal infections could be because of its antimicrobial activity rather than to an antiadhesive effect.

Impact of Ibuprofen Therapy in the Outcome of Experimental Pneumococcal Acute Otitis Media Treated With Amoxicillin or Erythromycin

The impact of ibuprofen combined with amoxicillin or erythromycin for therapy of penicillin-resistant pneumococcal acute otitis media (AOM) was evaluated in a gerbil model. Ibuprofen (at 2.5 or 7.5 mg/kg, orally) and/or amoxicillin or erythromycin (5 mg/kg each, s.c.) were administered at 5 h (early therapy, as single-dose regimen) or at 18 h (delayed therapy, five doses) postinoculation (PI). Each antibiotic alone and combined with ibuprofen was more effective administered as early regimen than as delayed treatment when evaluating the presence of otorrhea, otoscopic aspect, culture-positive and bacterial counts in middle ear (ME) samples, and loss of body weight. There was a trend for a better bacteriological outcome in animals receiving amoxicillin or erythromycin and ibuprofen, especially with the high dose. Such a dose of ibuprofen, associated with each antibiotic regimen, also preserved the animal well-being, avoiding a great weight loss in comparison to those receiving the antibiotic alone but a statistically significant difference was only observed for animals receiving delayed therapy with erythromycin and high-dose ibuprofen. In conclusion, ibuprofen combined with antibiotics seemed to improve the outcome of this experimental pneumococcal AOM.

Antimicrobial susceptibility of multidrug-resistant Streptococcus pneumoniae strains with penicillin MICs of 8 to 32 mg/L

The in vitro activity of 22 antibiotics (including novobiocin) and beta-lactam/gentamicin combinations was assessed against 11 multidrug-resistant pneumococcal strains. Among orally administered drugs, only telithromycin, levofloxacin, and linezolid were active against all isolates, but their use is not indicated in pediatrics. Novobiocin could be a potential therapeutic alternative.