Effect of 2-hydroxyethyl methacrylate on Streptococcus spp. biofilms.

AIMS: The effect of different concentrations of 2-hydroxyethyl methacrylate (HEMA) was evaluated on biofilm formation and preformed biofilm of Streptococcus mitis, Streptococcus mutans and Streptococcus oralis, alone or combined to each other. METHODS AND RESULTS: Twofold serial dilution of HEMA ranged from 12 to 0·75 mmol l(-1) was added to Streptococcal broth cultures and mature biofilms in 96-well-microtitre plates to evaluate bacterial biomass and cell viability. HEMA affected the Streptococcal population in a strain-specific way producing few significant effects. A reduction on biofilm formation and a detachment of preformed biofilm was recorded in Strep. mitis ATCC 6249, whereas in mixed cultures, the monomer expressed a general aggregative effect on mature biofilms. A reduction in cell viability was also recorded in an HEMA-concentration-dependent way in each experimental condition studied. CONCLUSIONS: These results suggest that the HEMA prevalent effects are both the reduction of bacterial adhesion to a polystyrene surface and the increase in dead cells also characterized by an aggregative status. SIGNIFICANCE AND IMPACT OF THE STUDY: Understanding the potential effect of HEMA, released from resin-based materials, on oral bacteria may furnish information for surveillance of the risk reduction in secondary caries via hindering biofilm generation.

Subinhibitory concentrations of moxifloxacin decrease adhesion and biofilm formation of Stenotrophomonas maltophilia from cystic fibrosis.

Stenotrophomonas maltophilia is an emerging nosocomial bacterial pathogen that is currently isolated with increasing frequency from the airways of cystic fibrosis (CF) patients. In this study the effect of subinhibitory concentrations (subMICs) of moxifloxacin on adhesion, biofilm formation and cell-surface hydrophobicity of two strains of S. maltophilia isolated from CF patients were evaluated. Adhesion and biofilm formation assays were carried out on polystyrene and quantified by colony counts. Cell-surface hydrophobicity was determined by a test for adhesion to n-hexadecane. Moxifloxacin at 0.03x and 0.06x MIC caused a significant decrease in adhesion and biofilm formation by both strains tested. A significant reduction in cell-surface hydrophobicity following exposure to subMICs of moxifloxacin was observed for one strain only. The results of the present study provide an additional rationale for the use of moxifloxacin in CF patients and more generally in biofilm-related infections involving S. maltophilia.

Quick and reliable galactomannan detection in crude minced lung specimens from haematological patients with suspected invasive fungal infection: results from a case series.

Invasive aspergillosis (IA) is the leading direct or contributory cause of death in patients with haematological malignancies. Early diagnosis remains difficult and often elusive due the heterogeneity of clinical presentations and the low sensitivity of both histological examination and cultures of specimens obtained from patients at risk. We report two cases of IA, both of which lacked both histological and cultural evidence of IA from pulmonary specimens. In both patients, detection of galactomannan (GM) by enzyme immunoassay (EIA) on pulmonary tissue homogenates led to the diagnosis of IA, which was confirmed by Aspergillus DNA (real time PCR). In conclusion, we provide preliminary evidence that lung homogenates may be prepared for GM EIA assays, which may contribute to quick diagnosis of IA on otherwise negative samples. We feel that our results open up the opportunity of a prospective and comparative evaluation of this diagnostic technique.

Influence of temperature on biofilm formation by Listeria monocytogenes on various food-contact surfaces: relationship with motility and cell surface hydrophobicity.

AIMS: To assess the ability of Listeria monocytogenes to form biofilm on different food-contact surfaces with regard to different temperatures, cellular hydrophobicity and motility. METHODS AND RESULTS: Forty-four L. monocytogenes strains from food and food environment were tested for biofilm formation by crystal violet staining. Biofilm levels were significantly higher on glass at 4, 12 and 22 degrees C, as compared with polystyrene and stainless steel. At 37 degrees C, L. monocytogenes produced biofilm at significantly higher levels on glass and stainless steel, as compared with polystyrene. Hydrophobicity was significantly (P < 0.05) higher at 37 degrees C than at 4, 12 and 22 degrees C. Thirty (68.2%) of 44 strains tested showed swimming at 22 degrees C and 4 (9.1%) of those were also motile at 12 degrees C. No correlation was observed between swimming and biofilm production. CONCLUSIONS: L. monocytogenes can adhere to and form biofilms on food-processing surfaces. Biofilm formation is significantly influenced by temperature, probably modifying cell surface hydrophobicity. SIGNIFICANCE AND IMPACTS OF THE STUDY: Biofilm formation creates major problems in the food industry because it may represent an important source of food contamination. Our results are therefore important in finding ways to prevent contamination because they contribute to a better understanding on how L. monocytogenes can establish biofilms in food industry and therefore survive in the processing environment.

Molecular characterization of virulence determinants of Stenotrophomonas maltophilia strains isolated from patients affected by cystic fibrosis.

Stenotrophomonas maltophilia is an emerging nosocomial bacterial pathogen which is currently isolated with increasing frequency from the airways of cystic fibrosis (CF) patients. In this study 13 S. maltophilia strains (11 isolated from the airways of independent CF patients, and two non-CF respiratory reference strains) have been characterized for the expression of several virulence-associated factors. In particular, the ability to form biofilm on abiotic surfaces has been determined and correlated with different features, such as motility, adherence and the ability to invade A549 respiratory epithelial cells. Moreover, the presence of a flagellum-associated gene as well as that of the StmPr1 gene, which encodes an extracellular protease, have been determined by Southern blot hybridization. Our data indicate that the different degree of biofilm formation exhibited by the 11 CF isolates does not correlate with motility, ability to adhere to and invade A549 cells, or with the presence of flagella. On the other hand, among the CF isolates the StmPr1 gene was found only in two strains, both able to establish chronic lung infections in CF patients. Moreover, only four of the strains analyzed show a temperature-independent antibiotic-resistance profile, suggesting either a different origin of these strains or an intervening adaptation to host tissues.

Management and triage of women with human papillomavirus infection in follow-up for low-grade cervical disease: association of HPV-DNA and RNA-based methods.

Type-specific persistent infection with Human Papillomavirus (HPV) is a significant risk factor for the development of cervical diseases. Persistent infection could be further refined by a sequencing approach to detect early cervical lesions that are at high risk of developing an invasive squamous cervical cancer. The aim of the present study is to investigate the clinical utility of detecting mRNA transcripts of HPV oncogenes E6/E7 by using a Real-time NASBA technology (mRNA test) and to identify women with low-grade cytological disease but with an increased risk of developing high-grade cervical abnormalities or invasive squamous cervical cancer. Our preliminary results show that E6/E7 is detected in only a subset of HR-HPV-positive cases. Since viral persistence is considered to be the true precursor of neoplastic progression, only the detection of E6/E7 mRNA can identify the infection which is more likely to persist and induce neoplasia in future. For these reasons we believe that this test would be useful for the characterization of women with HR-HPV DNA positivity who should be effectively treated because at high-risk of developing a high grade cervical lesion or an invasive squamous cervical cancer.

Effect of environmental factors on biofilm formation by clinical Stenotrophomonas maltophilia isolates.

The influence of environmental factors (temperature, aerobiosis-anaerobiosis, static-dynamic conditions, pH) was determined on biofilm formation by 51 S. maltophilia clinical isolates. The strains produced more biofilm at 32 degrees C than at 37 or 18 degrees C. Aerobic and 6% CO2 atmosphere yielded comparable biofilm amounts, higher than under anaerobic conditions. Biofilm production was not affected by static vs. agitated culture conditions. Biofilm production at pH 7.5 and 8.5 was comparable but significantly higher than at pH 5.5. The capacity of individual strains to form biofilm and thus contribute to the severity of some diseases is influenced by host traits and environmental conditions at the site of infection, and play an important role in the pathogenesis of biomaterial-related disease caused by S. maltophilia.

Serum and mucosal cytokine profiles in patients with active Helicobacter pylori and ischemic heart disease: is there a relationship?

This study is designed to investigate, for the first time, circulating and gastric mucosal levels of IL1-alpha, IL-6, IL-8 and TNF-alpha in patients with ischemic heart disease (IHD) and matched controls, according to the presence or absence of active Helicobacter pylori infection. Furthermore, in order to evaluate whether modified lipid profile was associated to an increased cardiovascular risk, this was determined in the same groups. Cytokine levels were measured using ELISA in 58 patients with IHD and 52 controls. Active H. pylori infection was assessed if either culture of H. pylori or rapid urease test gave a positive result. Our findings indicate increasing cytokine mucosal levels in H. pylori-positive patients compared to H. pylori-negative subjects. However, the increase was statistically significant only for IL-6 and TNF-alpha in the gastric mucosa of IHD patients. In H. pylori-positive controls, IL-8 mucosal levels positively correlated with both IL1-alpha (r = 0.98; P = 0.0003) and IL-6 (r = 0.83; P = 0.03) levels. Circulating cytokine levels were comparable in IHD and healthy subjects, regardless of H. pylori status. There were no correlations between mucosal and circulating cytokine levels. Active H. pylori infection was not associated with a modified lipid profile in either controls or IHD patients, although ApoAI levels were significantly higher in H. pylori-positive controls compared to those H. pylori-negative. Taken together, the results of the present study provide evidence that active H. pylori infection may play a role as a trigger factor in the pathophysiology of IHD by inducing an inflammatory cascade concentrated on gastric mucosa.

E-test method for detecting antibiotic synergy against Pseudomonas aeruginosa from neutropenic patients: a cost-effective approach.

The aim of this study was to evaluate the accuracy of E-test for the detection of synergy or antagonism of antibiotic combinations against Pseudomonas aeruginosa isolates from neutropenic patients. The activity of levofloxacin or grepafloxacin combined with ceftriaxone or cefotaxime against 20 P. aeruginosa clinical strains was assessed by checkerboard technique in comparison with results performed by E-test. The combination grepafloxacin + ceftriaxone appeared to be most effective (synergy, 55%) by checkerboard technique. The agreement between checkerboard and E-test results was 71.2%. Synergy was detected by checkerboard and E-test methods in 35 (43.8%) and 23 (31.3%) of 80 possible combinations, respectively. Antagonism was detected once (1.2%) by checkerboard method only. No major errors were recorded. E-test was preferable to checkerboard method for the total cost (reagent cost + cost of technologist time) (8,60 vs 21,80 euros/test, respectively). E-test appeared a promising alternative for testing antibiotic combinations although further testing should be performed to better refine this metodology.

Slime production by clinical isolates of Blastoschizomyces capitatus from patients with hematological malignancies and catheter-related fungemia.

In order to expand the present knowledge of the pathogenic potential of Blastoschizomyces capitatus in central venous catheter (CVC)-related bloodstream infections, six strains of the organism recovered from three leukemic patients with CVC-related fungemia in different years were investigated. Isolates and control strains were tested for their genetic relatedness and for their ability to produce slime in glucose-containing solutions. DNA restriction enzyme analysis revealed that all strains of B. capitatus were identical, whereas slime production assays and examination of ex vivo material showed that they were able to produce large amounts of slime. Slime production may therefore play a relevant pathogenic role in cases of CVC-related fungemia caused by B. capitatus.

Addition of teicoplanin or vancomycin for the treatment of documented bacteremia due to gram-positive cocci in neutropenic patients with hematological malignancies: microbiological, clinical and economic evaluation.

A prospective, randomized, double-blind trial was conducted on 124 febrile patients with hematological malignancies to compare teicoplanin with vancomycin as an addition to the initial empiric amikacin-ceftazidime regimen after documented bacteremia due to gram-positive cocci. At enrollment, patients in both groups were comparable with respect to age, sex, underlying hematologic disorders and duration of neutropenia. Rates of therapeutic success were 55/63 (87.3%) in the teicoplanin group and 56/61 (91.8%) in the vancomycin group (p = 0.560). The mean duration of treatment was similar, being 12.2 and 11.4 days, respectively (p = 0.216). Patients treated with teicoplanin remained febrile for slightly longer than those treated with vancomycin (4.9 vs. 4.0 days) (p = 0.013). Thirteen patients experienced an adverse drug reaction, but without any significant difference in the two arms. Isolated staphylococci showed a progressive and significant decrease in susceptibility to both glycopeptides during the 8 study years. The economic analysis performed showed that the addition of vancomycin is cost-saving.

Bacterial and viral DNA in periodontal disease: a study using multiplex PCR.

Recent studies have suggested an association between periodontal disease and the presence of Herpesviruses, in particular: Epstein-Barr virus (EBV) and Human Cytomegalovirus (CMV) (Contreras et al., 1999--Contreras et al., 2000--Slots et al., 2000--Ting et al., 2000). In the work reported in this paper, we use a multiplex Polymerase Chain Reaction (PCR) to compare the presence of Herpesviruses and putative bacterial pathogens in patients with periodontal disease and in healthy individuals. Direct detection of microorganisms with PCR is shown to offer significant advantages in terms of time, effort and cost. The study detected no statistically significant differences between the prevalence of EBV and CMV in patients and controls. The failure to replicate previous findings may be due to differences in the age composition and the geographical and social origins of the study groups. The study detected a significant excess of HSV-1 in periodontal patients. This suggests that the role of Herpesviruses in the pathogenesis of periodontal disease deserves further investigation. The bacterial assay confirmed the results of previous studies showing a strong association between periodontitis and the presence of A. actinomycetemcomitans, P. gingivalis and P. intermedia.

Detection of Helicobacter pylori by PCR on gastric biopsy specimens taken for CP test: comparison with histopathological analysis.

The aims of the present study were: (i) to assess whether H. pylori could be successfully detected by PCR from the same biopsy sample used for CPtest; and ii) to evaluate CPtest comparatively to both PCR and histology for detection of H. pylori infection in dyspeptic patients. Three antral gastric biopsies were collected from each of 80 consecutive dyspeptic patients undergoing oesophago-gastroduodenoscopy. Two biopsies were for histology (gold standard), one for CPtest, scored at 20min, 1h and 24h for the presence of urease activity. Gastric biopsy was then removed from CPtest and used for ureC-targeted PCR. Fifty-five (68.7%) patients were positive for H. pylori infection by histology. CPtest yielded an overall diagnostic accuracy of 93.8% (95% CI: 91-96.4%), regardless of observation period. No erroneous categorization of H. pylori status occurred using PCR, yielding sensitivity, specificity, positive and negative predictive values, and overall diagnostic accuracy of 100%. Our results suggest that H. pylori can be detected by PCR in gastric biopsies previously taken for CPtest, so reducing the workload of the endoscopist by saving additional biopsies for culture analysis and susceptibility tests.

Microbiological study and scanning electron microscopic analysis of root canal wall dentin following pumped Diodium Nd:YAG laser irradiation.

The capability of Nd:YAG laser in sterilizing root canals and the alterations of dentinal walls induced by laser treatment were investigated. Thirty root canals were infected by P. aeruginosa ATCC 27853 and thirty canals by A. naeslundii CH-12. Within each infection, 4 groups were selected on the basis of the treatment. Among them, 2 test groups (TGs) were treated by Nd:YAG laser at 15 Hz for 15 s, using 2 different settings: 1 Watt/70 Joule and 1.5 Watt/100 Joule, respectively (n = 10 each). The other 2 groups, used as controls (CGs), were: untreated (positive control, n = 5) and sterilized by 5.25% NaClO group (negative control, n = 5). Observations under scanning electron microscope (SEM) and quantitative bacterial counts were performed. These analyses were performed once per group after infections and treatments. Laser treatments significantly reduced the number of both bacteria. SEM investigation showed melting and crystallization of canal dentin over 1.5 W/100 J. Laser irradiation has a bactericidal effect but it does not completely sterilize the root canal as NaClO 5.25% solution does if the goal of treatment is also to avoid alterations of dentinal walls.

Role of antibiotic sensitivity testing before first-line Helicobacter pylori eradication treatments.

BACKGROUND: The resistance of Helicobacter pylori to antibiotics has been advocated as a major cause of treatment failure, and antimicrobial sensitivity testing has been proposed to improve efficacy; however, its role before first-line therapy has not been investigated in detail. AIM: To assess whether antimicrobial sensitivity testing improves the eradication rate of first-line anti-Helicobacter treatments and to compare the effectiveness of ranitidine bismuth citrate and omeprazole in the presence of H. pylori resistance to antibiotics. METHODS: Two hundred and forty-two patients were assigned to either empirical or antimicrobial sensitivity testing-based treatment; within each group, subjects were further randomized to receive ranitidine bismuth citrate, 400 mg b.d., tinidazole, 500 mg b.d., and clarithromycin, 500 mg b.d., or omeprazole, 20 mg b.d., clarithromycin, 500 mg b.d., and amoxicillin, 1 g b.d., for 1 week, with substitution of the resistant antibiotic in the antimicrobial sensitivity testing-based treatment group. RESULTS: Eradication rates were 67% [confidence interval (CI), 55-79%] in the empirical treatment group and 76% (CI, 65-87%) in the antimicrobial sensitivity testing-based group (P=N.S.). The overall success rate was 60% (CI, 51-69%) with omeprazole and 82% (CI, 73-91%) with ranitidine bismuth citrate (P<0.03); the latter overcame antibiotic resistance in 12 of 15 strains vs. zero of eight strains by omeprazole. CONCLUSIONS: Antimicrobial sensitivity testing before first-line treatment does not improve the eradication rate, which is greater when ranitidine bismuth citrate is included in the treatment.

Comparative in vitro activity of levofloxacin and ciprofloxacin against bacterial isolates from neutropenic patients.

BACKGROUND: Novel fluoroquinolones have been recently introduced in the management of neutropenic patients because of their increased activity against gram-positive and gram-negative micro-organisms. METHODS: The activities of levofloxacin and ciprofloxacin were determined by the E test against 223 bacterial isolates from patients with haematological malignancies. RESULTS: In general, the activity of levofloxacin was comparable to that of ciprofloxacin. Levofloxacin was somewhat more active against methicillin-resistant Staphylococcus aureus isolates. All methicillin-susceptible S. aureus isolates were inhibited by ciprofloxacin and levofloxacin at a concentration of < or =0.5 and < or =0.25 microg/ml, respectively. Among gram-negative isolates tested, levofloxacin was significantly (p < 0.001) more active than ciprofloxacin against Stenotrophomonas maltophilia, inhibiting 68 and 53% of these isolates, respectively. CONCLUSIONS: Levofloxacin was not superior to ciprofloxacin in its overall antibacterial activity, although small differences between these agents were seen depending on the species tested. In particular, our data suggested that levofloxacin may potentially be used for the management of S. maltophilia infections in neutropenic patients.

[In vitro antimicrobial activity of glutaraldehyde plus O-phenylphenol association (ratio 2:1)]. / Attività antimicrobica in vitro dell'associazione tra glutaraldeide e ofenilfenolo (ratio 2:1).

BACKGROUND: The correct use of antiseptics, disinfectants and sterilization processes to inactivate or remove micro-organisms is an essential component of an effective infection control program. To reach this result the use of phenolic detergent-disinfectants which work well as a presoak has always been suggested in lieu of sterilization involving submerging instruments in a properly prepared glutaraldehyde solution for about 6 to 10 hrs. This study investigated the in vitro effectiveness of disinfectant solution containing 2% glutaraldehyde and 1% o-phenylphenol. METHODS: The organisms used were purchased from the American Type Culture Collection (Escherichia coli ATCC 25922, Candida albicans ATCC 10231, Pseudomonas aeruginosa ATCC 27853, Staphylococcus aureus ATCC 25923, Staphylococcus epidermidis ATCC 12228), and from our clinical collection (Proteus mirabilis CH 14, Klebsiella oxytoca CH 22, Serratia liquefaciens CH 90). MIC and MBC were evaluated as testing solution for antimicrobial activity. The contact time studied between disinfectant and bacteria were 30 sec, 1, 2, 5, 10, 20 and 30 min. RESULTS: The results showed that MIC values for most bacteria were 3.75 mg/ml. The MBC values were similar or higher than the MIC. The disinfectant solution killed E. coli and C. albicans already after 10 min, but for S. aureus 30 min was necessary. CONCLUSIONS: This study proved that this solution attained the sterilization of surgical instruments in good timing and at low cost.

Fluids and microbial penetration in the internal part of cement-retained versus screw-retained implant-abutment connections.

BACKGROUND: It has been recently observed that in implants with screw-retained abutments, in in vitro as well as in vivo conditions, bacteria can penetrate inside the internal cavity of the implant as a consequence of leakage at the implant-abutment interface. An alternative to screw-retained abutments is represented by implants that can receive cemented abutments. In this case, the abutment goes through a transmucosal friction implant extension (collar) and is cemented inside the internal hexagonal portion of the implant. The aim of the present research was to compare fluids and bacterial penetration in 2 different implant systems, one with cement-retained abutments (CRA) and the other with screw-retained abutments (SRA). METHODS: Twelve CRA dental implants and 12 SRA implants were used in this study. The research was done in 3 steps: scanning electron microscopic (SEM) analysis, fluid penetration analysis, and bacterial penetration analysis. RESULTS: 1) Under SEM it was possible to observe in the SRA implants a mean 2 to 7 micron gap between implant and abutment, while in the CRA implants, the gap was 7 micron. In the latter group, however, the gap was always completely filled by the fixation cement. All the spaces between abutment and implant were filled by the cement. 2) With SRA implants, it was possible to observe the presence of toluidine blue at the level of the fixture-abutment interface and the internal threads; the absorbent paper was stained in all cases. With CRA implants, the absorbent paper inside the hollow portion of the implants was never stained by toluidine blue. No penetration of toluidine blue was observed at the implant-abutment interface and inside the hollow portion of the implants. 3) In all the SRA implant assemblies, bacterial penetration was observed at the implant-abutment interface. No bacteria were detected in the hollow portion of the CRA implants. CONCLUSION: On the basis of the results obtained in the present study using 2 different implant systems, we conclude that CRA implants offer better results relating to fluid and bacterial permeability compared to SRA implants.