Excellent antibacterial activity of Slovak honeys on bacteria mostly infecting chronic wounds.

INTRODUCTION AND AIM OF STUDY: Chronic wounds are commonly colonized by various bacterial species and colonization frequently turns into wound infection, severely impairing healing process. With increasing antimicrobial resistance, the antimicrobial treatment of chronic wounds may be extremely challenging. Rediscovery of old and forgotten antimicrobial therapeutic options, such as apitherapy, may contribute to solving the problem of incurable chronic wound infections. Aim of this study was to evaluate the antimicrobial properties of four kinds of Slovak honey from ecological beekeeping against the most common bacterial species contaminating and infecting chronic wounds, and to compare these antimicrobial activities with those of the approved medical-grade Manuka honey. The impact of honey sterilisation methods and long-lasting storage on the bactericidal activity was also examined. MATERIAL AND METHODS: Antimicrobial activity of honey was detected against 7 bacterial collection strains by broth microdilution antimicrobial susceptibility test according to EUCAST. The results were statistically analysed by Fisherꞌs exact test. RESULTS AND CONCLUSIONS: Slovak ecologically produced honey samples demonstrated an excellent in vitro antibacterial activity, superior to the monofloral medical-grade Manuka honey activity. Neither the gamma-irradiation, nor the three-year-long storage had impact on the bactericidal activity of the tested honey (Tab. 4, Fig. 2, Ref. 53).

Biofilm-producing potential of urinary pathogens isolated from chronic and recurrent urinary tract infections and impact of biofilm on gentamicin and colistin in vitro efficacy.

AIM: The presented study was to compare in vitro biofilm production by bacterial strains from chronic/recurrent and from acute non-complicated UTIs. The activity of gentamicin and colistin on biofilm form of these strains has also been detected, with goal to predict the gentamicin and colistin therapeutic efficacy in the antimicrobial treatment of patients with a suspected presence of biofilm in urinary tract. MATERIAL AND METHODS: The group of 40 bacterial strains repeatedly isolated from patients with chronic or recurrent UTIs was compared with the group of 40 strains from acute UTIs. Both groups contained comparable number of strains of Escherichia coli, Klebsiella spp., Proteus mirabilis and Pseudomonas aeruginosa. Biofilm production was assessed by method in polystyrene microtiter plate. The MIC and MBC values of gentamicin and colistin were detected by broth microdilution assay. The minimal biofilm inhibitory (MBIC) and biofilm eradication concentrations (MBEC) were tested by microdilution method. Non-inactivated biofilm-associated bacteria were detected after overnight incubation in broth medium free of antimicrobials. The statistical analysis of results was performed by Fisher's exact test and by Student's t-test. RESULTS: Biofilm was produced by 90% strains from chronic UTIs, but only by 52% of strains from acute UTIs (p = 0.0004). In the biofilm producing strains, the MBIC values of gentamicin reached from four to 256 mg/L, the MBIC levels of colistin from two to 64 mg/L. The minimal biofilm eradicating concentrations were even higher: for gentamicin from eight to > 512 mg/L, and for colistin from 32 to > 512 mg/L. The differences between MIC and MBIC/MBEC levels were statistically highly significant (p < 0.0001). Presumably, the therapeutic success of parenterally applied gentamicin or colistin on biofilm-related urinary tract infections would be, without respect to the high concentration of gentamicin or colistin achievable in urine during parenteral application, rather unpredictable. Local intravesical instillation would allow for achieving higher gentamicin and colistin concentrations; however, there is need for interpretation criteria for MBEC values concerning therapy, as well as for clinical studies allowing for application of those values to predict clinical success of therapy. CONCLUSIONS: Laboratory detection of biofilm production and evaluation of the MBIC/MBEC values of antimicrobials for strains producing biofilm might be a valuable complement to the microbiologic diagnostics of chronic and recurrent UTIs. It might provide valuable information for more reliable individualised therapy and so decrease the risk of emergence and selection of multiresistant strains during repeated and non-eradicating therapy of chronic and recurrent UTIs.

[Antibiofilm activity of agents for disinfection of skin, mucosa, and wound on microorganisms isolated from patients with catheter-related infections]. / Vplyv prípravkov urcených na dezinfekciu koze, slizníc a rán na zivotaschopnost biofilmovej formy rastu mikroorganizmov izolovaných od pacientov s katétrovými infekciami.

AIM OF THE STUDY: To evaluate the activity of four disinfectious agents used for skin, mucosa and wound disinfection (chlorhexidine digluconate, povidone-iodine, octenidine hydrochloride, super oxygenated water) on the biofilm of Staphylococcus aureus, Escherichia coli and Candida sp. strains, isolated from patients with catheter-related infections. MATERIAL AND METHODS: The tested agents were applied on 24-hours biofilm in the microtiter plate wells. After 20-minutes exposition, the wells were washed, and the microbial vitality was tested by regrowth method after 24-hours cultivation in fresh culture medium. Biofilm formation was confirmed in a parallel microtiter plate; the quantity of produced biofilm was measured after crystal violet staining spectrophotometrically at 570 nm. RESULTS: All four tested disinfectious agents inactivated the biofilm of all S. aureus, E. coli, C. albicans, C. krusei and C. glabrata strains, without respect to the intensity of biofilm production. Three strains of C. tropicalis with intensive biofilm production partially preserved their vitality after exposition to chlorhexidine and povidone-iodine, and 2 strains to octenidine. Super oxygenated water had no effect on yeasts associated with massive biofilm of one C. tropicalis strain, and only partially decreased the vitality of additional two strains. CONCLUSIONS: The tested disinfectious agents proved in-vitro antibiofilm activity on all microbial strains from catheter-related infections, with exception of three C. tropicalis strains with intensive biofilm production. Octenidine was found to be the most active agent. The results enable to assume, that the tested disinfectious agents, when applied to patients, will inactivate not only the individual microorganisms not protected by biofilm, but also the biofilm on the catheter surfaces approachable by local application. However, C. tropicalis strains producing massive biofilm, protecting them partially from effects of disinfectious agents tested in the present study, still remain a challenge.

Enterococcus faecalis and Candida albicans in the dental root canal and periapical infections.

OBJECTIVE: The aim of the present study was to examine the prevalence of Enterococcus faecalis and Candida albicans in endodontic infections. METHODS: Samples for microbiological examination were collected from 32 patients with deep dental caries, infected dental root canal, or periapical infection. RESULTS: Cultivation of the dental samples yielded four strains of Enterococcus faecalis (12.5 %), and three strains of Candida albicans (9.4 %). All Enterococcus faecalis isolates were susceptible to ampicillin, one isolate was resistant to tetracycline, two to erythromycin and azithromycin (additional 2 had intermediate susceptibility), and one strain had intermediate susceptibility to ciprofloxacin and moxifloxacin. CONCLUSION: We conclude that Enterococcus faecalis and Candida albicans can participate in the dental root canal and periapical infections, and the use of effective irrigant solutions and intracanal medicaments active against these microbes is important in order to prevent endodontic therapy failures. Unexpected was the isolation of C. albicans from a nine-year-old child with periodontitis apicalis. This finding must draw attention to the possibility that even at such a young age, this microorganism could be a potential etiological agent in endodontic infections (Tab. 2, Ref. 34). Text in PDF www.elis.sk.

[Susceptibility of Staphylococcus aureus biofilms to vancomycin, gemtamicin and rifampin]. / Citlivost' kmenov staphylococcus aureus rastúcich v biofilme na vankomycín, gentamicín a rifampicin.

STUDY OBJECTIVES: To detect biofilm formation in Staphylococcus aureus strains and to determine the minimal biofilm inhibition concentrations (MBIC) and the minimal biofilm eradicating concentrations (MBEC) of vancomycin, gentamicin and rifampin. To compare the MBIC and MBEC with the minimal inhibition concentration (MIC) and minimal bactericidal concentration (MBC) data for planktonic Staphylococcus aureus forms that are commonly used in antimicrobial susceptibility testing for the purposes of individualized therapy. PATIENTS AND METHODS: Fifteen S. aureus strains isolated from central venous catheters, intratracheal tubes and wound drainage tubes from the patients of the University Hospital, Bratislava-Staré Mesto were included in the study. Selected virulence factors were characterized. The biofilm formation potential was measured by a modified crystal violet micro-assay. The presence of viable cells biofilm in was tested using 3-(4,5-dimethylthiazol--2-yl)-2,5-diphenyl tetrazolium bromide (MTT). The MIC and MBC of vancomycin, gentamicin and rifampin was tested in planktonic S. aureus forms by the broth microdilution method. The MBIC and MBEC of these antimicrobial drugs for biofilm S. aureus forms were determined by a modified microdilution method. Student's t-test was used for statistical analysis of the results. RESULTS: All of the study strains formed biofilm, with only two of them having a low biofilm formation potential. MTT revealed moderate to high metabolic activity of bacteria biofilm in Vancomycin MICs and MBICs were identical in 80 % of the study strains. Vancomycin MBECs are higher than MBCs in all the study strains, are interpreted as resistance according to the criteria of the Clinical and Laboratory Standards Institute (CLSI) and make the drug unsuitable for use in the treatment. In vitro gentamicin MBICs indicated susceptibility according to the CLSI criteria but gentamicin MBECs were interpreted as gentamicin resistance. Rifampin MICs and MBICs of the study strains revealed susceptibility. Rifampin MBCs were interpreted as susceptibility, but based on MBECs, 13% of the study strains were considered as resistant and 13% of the study strains showed intermediate susceptibility. The differences between gentamicin and rifampin MICs and MBICs and those between MBCs and MBECs of all antimicrobials tested were statistically significant. CONCLUSION: The tested biofilm S. aureus forms showed high MBECs of vancomycin, gentamicin and rifampin, with rifampin only being suitable for therapeutic use. To provide reliable results for individualized antibiotic therapy, it will be needed to test in vitro biofilm formation, to determine MBIC and MBEC of antimicrobial drugs using a standardized method, to interpret the test results in relation to biofilm S. aureus forms and to establish the interpretation criteria for MBIC and MBEC similarly to MIC and MBC.

Clinical pseudomonas aeruginosa: potential factors of pathogenicity and resistance to antimicrobials.

Resistance to 17 antimicrobials, surface hydrophobicity, motility, biofilm, production of N-acylhomoserine lactone signal molecules (N-butyrylhomoserine lactone and N-3-oxolauroylhomoserine lactone) and response to oxidative stress were analyzed in 47 clinical Pseudomonas aeruginosa strains. In addition to natural resistance, the strains demonstrated the greatest level of resistance to cefotaxime (91.5%). Isolates in the range of 44.7-57.4% were resistant to aminoglycosides and ciprofloxacin, of 25.5-36.2% to cephalosporins. On the other hand, 97.9% remained susceptible to meropenem, 93.6% to piperacillin + tazobactam and 87.2% to piperacillin. The majority of the strains (72.3%) manifested their hydrophilic character. Higher zones of motility showed 12 isolates (in average 54.8 mm) as compared to the others (30.2 mm). Approximately 1/3 of the strains (29.8%) produced a higher amount of biofilm quantified by measuring the absorbance of solubilized crystal violet (0.20-0.46) than the rest of isolates (0-0.19). All but two strains produced N-3-oxolauroylhomoserine lactone and in 48.9% of samples N-butyrylhomoserine lactone were detected. Only four isolates with higher biofilm production showed both types of homoserine lactone. Majority of the strains (70.2%) manifested higher resistance to H2O2 than the rest of the strains. The group of strains resistant to aminoglycosides and ciprofloxacin revealed a significantly higher number of hydrophobic strains (compared with the sensitive ones). In contrast, higher number of strains sensitive to aminoglycosides and ciprofloxacin or only to ciprofloxacin produced N-butyrylhomoserine lactone and biofilm (compared to the resistant ones). Such association was not found among the rest of the tested parameters. The results indicate that the resistance to antimicrobials in P. aeruginosa isolates was not generally associated with changes in the production of the pathogenicity factors.

[Influrence of recombinant human procalcitonin on phagocytic and candidacidal ability of polymorphonuclear leukocytes and on killing mechanisms of serum and blood aganist bacteria Staphylococcus aureus and Escherichia coli]. / Vplyv rekombinantného lbdského prokalcitonínu na fagocytovú a kandi- dacídnu schopnost polymorfonukleárnych leukocytov a na cídne mechanizmy séra a krvi voti baktériám Staphylococcus aureus a Escherichia coli.

UNLABELLED: Procalcitonin (PCT) is a highly selective and specific marker for early diagnosis of sepsis. There is enough information confirming that procalcitonin should be considered as an important mediator in the pathophysiology of sepsis. The aim of our work was to evaluate the effect of recombinant human procalcitonin (rhPCT) on phagocytic and candidacidal activity of polymorphonuclear leukocytes (PMNL) in blood, as well as on killing mechanisms of fresh serum and blood against Escherichia coli (E. coli) and Staphylococcus aureus (S. aureus). Our results show that rhPCT dose dependently decreased both phagocytic and candidacidal activity of polymorphonuclear leukocytes (p < 0.001). RhPCT inhibited also the microbicidal activity of both serum and blood on E. coli that was found by inoculation of suspension on culture plates. We found that rhPCT supported the E. coli colony count increase during the incubation in the presence of both serum and blood. The effect of rhPCT on the S. aureus colony count increase was not statistically significant. CONCLUSION: Our results indicate a suppressive effect of rhPCT on phagocytic and microbicidal activity of polymorphonuclear leucocytes.

Antibiotic resistance and virulence factors among clinical and food enterococci isolated in Slovakia.

The resistance to antibiotics and the distribution of virulence factors in enterococci isolated from traditional Slovak sheep cheese bryndza was compared with strains from human infections. The occurrence of 4 enterococcal species was observed in 117 bryndza-cheese isolates. The majority of strains were identified as E. faecium (76 %) and E. faecalis (23 %). Several strains of E. durans and 1 strain of E. hirae were also present. More than 90 % of strains isolated from 109 clinical enterococci were E. faecalis, the rest belonged to E. faecium. The resistance to 6 antimicrobial substances (ampicillin, ciprofloxacin, higher concentration of gentamicin, nitrofurantoin, tetracycline and vancomycin) was tested in clinical and food enterococci. A higher level of resistance was found in clinical than in food strains and E. faecium had a higher resistance than E. faecalis; no resistance to vancomycin was detected. The occurrence of 3 virulence-associated genes, cylA (coding for hemolysin), gelE (coding for gelatinase) and esp (coding for surface protein) was monitored. Differences were found in the distribution of cylA gene between clinical and bryndza-cheese E. faecalis strains; in contrast to clinical strains (45 %), cylA gene was detected in 22 % of food isolates. The distribution of 2 other virulence factors, gelE and esp, was not significantly different in the two groups of E. faecalis strains. cylA and gelE genes were not detected in E. faecium but more than 70 % of clinical E. faecium were positive for esp, even thought none of the 79 E. faecium cheese isolates contained this gene.

Susceptibility of Streptococcus pneumoniae isolated from the respiratory tract of hospitalized children with respiratory tract infections.

The most frequent nasopharyngeal carriers of Streptococcus pneumoniae are young children. Frequent use of antimicrobial therapy in children facilitates the selection of penicillin-resistant strains in this population. These strains, especially if highly resistant, may cause serious therapeutic problems. Aim of the study was to monitor penicillin- and multidrug-resistant S. pneumoniae strains in hospitalized children with respiratory tract infections. Hospitalized children up to five years were examined for S. pneumoniae presence in their upper respiratory tract. Susceptibility to penicillin, erythromycin, trimethoprim/sulfamethoxazole, tetracycline, and chloramphenicol was determined by the disk-diffusion method. The minimal inhibitory concentrations (MIC) of penicillin, erythromycin and trimethoprim/sulfamethoxazole were measured by the E-test. S. pneumoniae strain was isolated from 60 (34.7%) out of 173 microbiologically examined children; 2 different strains were isolated in 9 cases. Nine strains (13.0%) were penicillin resistant with MICs ranging from 1.5 to 8 mg/L, and 17 strains (24.6%) had intermediate susceptibility. Seventeen (24.6%) strains were erythromycin resistant (MIC > or = 1 mg/L). Eighteen strains (26.1%) were resistant and 7 strains (10.1%) were intermediately susceptible to trimethoprim/sulfamethoxazole. Ten strains (14.5%) were not susceptible to tetracycline, and 11 (15.9%) to chloramphenicol. Non-susceptibility (resistance or intermediate susceptibility) to the tested antimicrobials was more prevalent in penicillin-nonsusceptible strains. The current level of S. pneumoniae resistant to antimicrobial drugs in children with respiratory tract infections in the hospital department monitored in our study do not cause problems in the choice of antibacterial therapy. Penicillins still can remain the drug of choice in cases when typical bacterial causing agents of respiratory tract infections are suspected. (Tab. 3, Fig. 2, Ref. 31.)

[Staphylococcus aureus and the human immune system]. / Staphylococcus aureus a imunitný systém cloveka.

The author characterizes factors of virulence of Staphylococcus aureus and its different interactions with immune systems of the host. She describes the immune mechanisms which play a key role in the elimination of staphylococcal infection and where inadequate function leads to the development of staphylococcal disease. The author draws attention to the complicated interrelationship of the complex of different virulence factors of staphylococcal strains involved in inhibition and dysregulation or in stimulation of effector mechanisms of immunity.

Staphylococcus aureus in chronic and recurrent infections.

Ninety-four Staphylococcus aureus strains isolated from chronic and recurrent skin and respiratory tract infections were investigated for several virulence factor expressions. Production of protein A was noticed in all of the tested strains in amounts from less than 0.1 to more than 2.5 ng per 10(6) bacterial cells. The percentage of the extracellularly produced protein A was found to lie between 4.5 and 27.8%. Two strains (both from the respiratory tract) produced more than 50% of protein A in the extracellular form and one strain did not produce any detectable amount of the extracellular protein A; 99% of the tested strains produced the clumping factor, 96% staphylocoagulase, 79% staphylokinase and 90% gelatinolytic activity; 79% produced alpha-toxin exclusively or in combination with delta- or beta-toxin; 8% of strains produced beta-toxin. There were differences in beta-toxin production between strains from the respiratory tract (5%) and skin infections (25%). delta-Toxin was produced by 53% of the strains. In each of the tested strains a complex of virulence factors was detected. The importance of inactivated extracellular products (especially alpha- and delta-toxin and in the case of skin infections also beta-toxin) as components of staphylococcal whole-cell vaccine was suggested.