Diffusion of meticillin-resistant Staphylococcus aureus USA300 strains in central Italy.

Meticillin-resistant Staphylococcus aureus (MRSA) is an outstanding, clonally evolving pathogen that in recent years, under the selective pressure of antibiotics, has acquired the crucial ability to infect people outside of hospitals. MRSA USA300 has progressively become synonymous with severe community-associated staphylococcal disease worldwide. Whilst spreading worldwide, these clones have progressively acquired resistance to several antibiotics and have gained the ability to cause infections in hospital settings. Recently, USA300-related strains showing resistance to several antibiotics have been isolated from community-acquired infections in Italy. This paper reports the high frequency of isolation of USA300-related strains both from community- and hospital-acquired infections in central Italy as well as their genotypic characteristics and antibiotic susceptibility. Analysis of these characteristics by partial least squares discriminant analysis enabled it to be demonstrated that whilst moving from the community to the hospital setting these isolates underwent an adaptive process that generated clones showing distinctive characteristics. These observations further support the hypothesis that the threatening generation of strains combining both resistance and virulence is becoming a reality, and stress the necessity of constant molecular epidemiological surveillance of MRSA.

Microbiological and molecular characterization of nosocomial and community Staphylococcus aureus isolates.

This study aimed to evaluate the incidence of Staphylococcus aureus infections in different departments of Belcolle Hospital in Viterbo and the surrounding area between January 2003 and June 2008. Isolates of methicillin-resistant S. aureus (MRSA) recovered in this time interval were characterized by microbiological and molecular methods to evaluate the reliability of simple criteria to distinguish between hospital-acquired and community-acquired isolates. MRSA accounted for 33% of all S. aureus, with a significantly higher prevalence in isolates from nosocomial infections. MRSA isolates were assayed by PCR for the presence of 13 genes associated with virulence, agr type and SCCmec type. Cumulative data were analysed by partial least square discriminant analysis and a clear correlation was demonstrated between genetic profiles and classification of isolates as hospital or community acquired according to simple temporal criteria. Nosocomial MRSA isolates from blood samples showed significantly higher genetic diversity than other nosocomial isolates. Our data confirm the existence of significant differences between community- and hospital-acquired MRSA isolates.

Genotype and phenotype relationship in MLSb resistance in Streptococcus pyogenes and Streptococcus agalactiae isolated in central Italy.

The aim of this work is to study a correlation between phenotype and genotype in clinical isolates of erythromycin-resistant Streptococcus spp. Among the 25 erythromycin-resistant S. pyogenes, we detected six strains with iMLSB, nine with cMLSB and two with M phenotypes. Among 14 erythromycin-resistant S. agalactiae, we detected five strains with iMLSB, seven with cMLSB and none with an M phenotype. Moreover, 8 S. pyogenes and 2 S. agalactiae showed a phenotype not matching the known ones described in literature, defining an unknown phenotype. Upon examination, the genetic profiles, erm(A), erm(B) and mef(A), of the clinical isolates did not easily correlate with a specific phenotype. Our findings highlighted that the whole matter of phenotypic diversity in macrolide-resistant S. pyogenes and S. agalactiae strains and the correlation with their genetic profiles should be submitted to a more careful analysis of phenotypic and genotypic characterization.

In vitro susceptibility of isolates of Borrelia burgdorferi s.l. to antimicrobial agents.

In the present study, we investigate the in vitro antimicrobial activity of macrolides, beta-lactams and tetracycline against Borrelia burgdorferi s.l. clinical and tick isolates. Minimal inhibitory concentrations (MICs) were determined in normal growth condition and after pre-exposure of the strains to sub-MIC of the founder of each drug family. All the classes of tested antibiotics showed good antibacterial activity against all the borreliae isolates and there were no significant susceptibility differences among clinical and tick isolates. After pre-exposure of the strains to sub-MIC of erythromycin, cefoxitin and tetracycline, we observed that some strains of B. burgdorferi s.l. showed higher MIC values to both the pre-exposed drug and drugs of the same family. The less susceptibility of borreliae, in the last growth condition in vitro, could be one of the justifications of clinical results indicating the limited efficacy of these antibiotics in treatment of B. burgdoferi infections.

Multifactorial aspects of antimicrobial activity of propolis.

We investigated the antibacterial activity of sub-inhibitory concentrations of ethanolic extract of propolis (EEP), and its effect on the antibacterial activity of some antibiotics. Some clinically isolated Gram-positive strains were used. Moreover, sub-inhibitory concentrations of EEP were used to value its action on some important virulence factors like lipase and coagulase enzymes, and on biofilm formation in Staphylococcus aureus. Our results indicated that EEP had a significant antimicrobial activity towards all tested clinical strains. Adding EEP to antibacterial tested drugs, it drastically increased the antimicrobial effect of ampicillin, gentamycin and streptomycin, moderately the one of chloramphenicol, ceftriaxon and vancomycin, while there was no effect with erithromycin. Moreover, our results pointed out an inhibitory action of EEP on lipase activity of 18 Staphylococcus spp. strains and an inhibitory effect on coagulase of 11 S. aureus tested strains. The same EEP concentrations showed a negative interaction with adhesion and consequent biofilm formation in S. aureus ATCC 6538P.

Effect of propolis on virulence factors of Candida albicans.

Propolis is a resinous substance collected by honeybees from plant sources. Its antimicrobial activity has been well documented but little is specifically known about its activity on virulence factors of Candida albicans. The aim of this work was therefore to evaluate in vitro the propolis effect on yeast-mycelial conversion (Y-M), extracellular phospholipase activity and fungal adhesion to epithelial cells. The two propolis samples used significantly inhibited the C. albicans strains tested, showing a rapid (between 30 seconds and 15 minutes), dose-dependent cytocidal activity and an inhibitory effect on Y-M conversion at a concentration of 0.22 mg/ml. Moreover, the hyphal length was reduced even at lower propolis concentration. Propolis also caused a dose- and time-dependent inhibition of phospholipase activity. No clear effect was shown on adherence to buccal epithelial cells and surface structure hydrophobicity, but damage to the plasma membrane structure was demonstrated with the Propidium Iodide test.

Antibacterial activity of Hydrastis canadensis extract and its major isolated alkaloids.

The antibacterial activity of extract and isolated major alkaloids (berberine, beta-hydrastine, canadine and canadaline) of Hydrastis canadensis L. (Ranunculaceae) was evaluated against 6 strains of microorganism: Staphylococcus aureus (ATCC 25 993 and ATCC 6538P), Streptococcus sanguis (ATCC 10 556), Escherichia coli (ATCC 25 922), Pseudomonas aeruginosa (ATCC 27 853). Bactericidal activity was evaluated by contact test by measuring the "killing time" on a low density bacterial inoculum, and bacteriostatic activity in liquid medium by M.I.C. values. The results provide a rational basis for the traditional antibacterial use of Hydrastis canadensis.

Effect of slime production on the antibiotic susceptibility of isolates from prosthetic infections.

The antibacterial activity of 6 antibiotics towards 10 gram-positive and 6 gram-negative glycocalyx-producing strains, has been evaluated by employing a method which partially simulates the in vivo colonization of prosthetic devices. The results showed that routine antibiotic sensitivity tests are not predictive about the response of the glycocalyx-embedded bacteria, and that prophylaxis may be useful with ofloxacin and clindamycin, before placing a prosthetic device. Once bacterial colonization had already occurred, however, none of the tested antibiotics was able to eradicate the sessile bacterial form. The minimum bactericidal concentration (MBC) values, indeed, were much higher than those determined on the planktonic form, and were much higher than serum and tissue levels that can be reached in vivo.

In vitro activity of pefloxacin compared with five other quinolones on nalidixic acid-resistant proteae species.

The in vitro activity of pefloxacin, a new fluoroquinolone, was compared with that of 5 other quinolone compounds (nalidixic and pipemidic acids, norfloxacin, ciprofloxacin, and ofloxacin) against 416 strains of Proteae spp. isolated from urine specimens of hospitalized patients with acute urinary tract infections (UTI). Ciprofloxacin was the most active agent. Norfloxacin, ofloxacin, and pefloxacin were similarly active against Proteus strains (MIC90 = 0.39 microgram/ml). Against Providencia spp. pefloxacin and norfloxacin showed similar activity (MIC90 = 3.12 micrograms/ml). There is minimal discrepancy between minimum inhibitory concentrations and minimum bactericidal concentrations exhibited by the quinolones for all urinary tract pathogens tested. Our in vitro studies indicate that pefloxacin is an active antimicrobial agent and suggest that it will prove useful in the treatment of complicated urinary tract infections due to nalidixic acid-resistant Proteae spp.

Plasmid "curing" by some recently synthetized 4-quinolone compounds.

Nalidixic acid and two recently synthetized 4-quinolones eliminated F'lac and R-plasmids from E. coli at concentrations of one half or one quarter of the minimum inhibitory concentration (MIC). Two of the three plasmids tested were cured by all derivatives, although with different frequencies. Pefloxacin was the most effective compound compared with the other quinolones and nalidixic acid the least active.

Inhibition of plasmid conjugation by some recently synthetized 4-quinolone compounds.

Four fluoroquinolones (norfloxacin, ciprofloxacin, ofloxacin, and pefloxacin) were compared with nalidixic acid for their inhibitory effect on conjugal plasmid transfer. The inhibition was observed in mating experiments using various combinations of drugs at subinhibitory concentrations and 3 different plasmids in the E. coli k12 genetic background. Fluoroquinolones inhibited plasmid transfer to a greater extent than nalidixic acid. Ofloxacin and pefloxacin were consistently the most active agents, causing 90 to 100% inhibition of plasmid transfer in all mating systems studied.

On the formation of a dicloxacillin-p-hydroxymercuribenzoate suicide complex mediated by beta-lactamase I from Bacillus cereus.

p-Hydroxymercuribenzoate is a non-competitive inhibitor of beta-lactamase I from Bacillus cereus and also, after preliminary preincubation, an inactivator of the enzyme. Submitted to the simultaneous action of PCMB plus dicloxacillin, the enzyme completely loses its activity. Extensive dialysis can restore the enzymatic activity only if preincubation had been carried out with either PCMB or dicloxacillin but not if both inhibitors had been simultaneously present. Mercaptoethanol protects the enzyme from the action of PCMB, but not from the severe inactivation caused by dicloxacillin-PCMB mixtures. All these data suggest the formation of a complex between PCMB and the acyl-enzyme intermediate generated upon hydrolysis of the beta-lactam bond of dicloxacillin.

Inhibition of beta-lactamase production in gram-positive and gram-negative strains by sub-inhibitory concentrations of nalidixic acid.

We found it of interest to try to determine a microbiological parameter for the effects that sub-inhibitory doses of a chemotherapeutic agent may have on bacteria. To this end, the antibacterial activity of nalidixic acid was analyzed and our attention was directed to evidence of the production of beta-lactamase by 60 strains. The tests carried out showed that sub-inhibitory concentrations of nalidixic acid, from 2 to 64 times lower than the MIC (minimum inhibitory concentrations), were able to inhibit production of penicillinases in the 14 strains of Staphylococcus spp. tested, while only in 7 of the 21 strains of Gram-negative bacteria. Strains from our collection of beta-lactamase producing bacteria were analyzed, and our results confirmed that nalidixic acid inhibits plasmidic and chromosomic beta-lactamases.

[Antibiotic and metal resistance in "Escherichia coli" strains isolated from the environment and from patients (author's transl)]. / Resistenza ad antibiotici e metalli in ceppi di "Escherichia coli" isolati dall'ambiente e da pazienti.

The incidence and the patterns of the antibiotic and metal resistance in 106 strains of Escherichia coli isolated from ground waters, used also as drinking water supply (sample A), was studied in comparison with the resistance behaviour in the 104 strains of the same microorganism isolated from non hospitalized patients (sample P). Significant differences between the percentage of resistant strains in the two examined samples were found for some of the antibiotics and the metals tested (ampicillin, streptomycin, kanamycin, mercury and zinc) while non statistically significant differences were found for gentamicin, tetracyclin, nalidixic acid and cadmium. From the high percentages of the resistant strains in the environmental sample (up to 44.3% for tetracyclin) we may deduce that also the ground waters, especially if used as drinking water, contribute to the spread of the resistant bacteria. The patterns of the antibiotic multiresistances in the strains isolated from patients and from ground waters do not differ greatly and this strengthens the hypothesis that resistance to antibiotics has been acquired by Escherichia coli strains before reaching the ground waters.