Staphylococcus aureus biofilm formation and antibiotic susceptibility tests on polystyrene and metal surfaces

Aim: We compared the MBEC?‐HTP assay plates made of polystyrene with metal discs composed of TMZF^® and CrCo as substrates for biofilm formation. Methods and Results: Staphylococcus aureus was grown on polystyrene and on metal discs made of titanium and chrome–cobalt. Antibiotic susceptibility was assessed by examining the recovery of cells after antibiotic exposure and by measuring the biofilm inhibitory concentration (BIC). The minimal inhibitory concentration (MIC) was assessed with planktonic cells. Bacterial growth was examined by scanning electron microscopy. The antibiotic concentration for biofilm inhibition (BIC) was higher than the MIC for all antibiotics. Microscopic images showed the biofilm structure characterized by groups of cells covered by a film. Conclusions: All models allowed biofilm formation and testing with several antibiotics in vitro. Gentamicin and rifampicin are the most effective inhibitors of Staph. aureus biofilm‐related infections. We recommend MBEC?‐HTP assay for rapid testing of multiple substances and TMZF^® and CrCo discs for low‐throughput testing of antibiotic susceptibility and for microscopic analysis. Significance and Impact of the Study: In vitro assays can improve the understanding of biofilms and help developing methods to eliminate biofilms from implant surfaces. One advantage of the TMZF^® and CrCo discs as biofilm in vitro assay is that these metals are commonly used for orthopaedic implants. These models are usable for future periprosthetic joint infection studies.     

伊犁黑蜂蜂胶对不同生长状态白假丝酵母菌抑菌作用的体外研究

目的探究伊犁黑蜂蜂胶对白假丝酵母菌游离状态及其生物膜状态的影响。方法采用NCCLS M27-A2酵母微量稀释法结合平板法测定蜂胶对游离状态下白假丝酵母菌的最小抑菌浓度(MIC)和最低杀菌浓度(MBC);玻片法在体外构建白假丝酵母菌生物膜模型,采用MTT法检测蜂胶对白假丝酵母菌生物膜状态下的最低生物膜清除浓度(MBEC)及抑制作用;激光共聚焦显微镜(CLSM)观察不同浓度蜂胶醇提取物对同一时间白假丝酵母菌生物膜的抑菌效果,并进行红蓝荧光染色定量分析。结果伊犁黑蜂蜂胶对浮游白假丝酵母菌的MIC为4mg/mL,MBC为8mg/mL;伊犁黑蜂蜂胶对生物膜状态下白假丝酵母菌的MBEC为16mg/mL,随着药物浓度的增加抑菌性逐渐增大,差异具有统计学意义(P0.05);CLSM观察,伊犁黑蜂蜂胶作用于白假丝酵母菌生物膜后可使生物膜内活菌比例降低,生物膜活性减弱,生物膜内实验菌死亡率随蜂胶浓度增加呈上升趋势,与阴性对照组比较差异有统计学意义(P0.05)。结论伊犁黑蜂蜂胶在体外对不同状态下的白假丝酵母菌均起到了明显的抑制作用。     

Citropin 1.1-treated central venous catheters improve the efficacy of hydrophobic antibiotics in the treatment of experimental staphylococcal catheter-related infection

An in vitro antibiotic susceptibility assay for Staphylococcus aureus biofilms developed on 96-well polystyrene tissue culture plates was performed to elucidate the activity of citropin 1.1, rifampin and minocycline. Efficacy studies were performed in a rat model of staphylococcal CVC infection. Silastic catheters were implanted into the superior cava. Twenty-four hours after implantation the catheters were filled with citropin 1.1 (10 microg/mL). Thirty minutes later the rats were challenged via the CVC with 1.0 x 10(6) CFU of S. aureus strain Smith diffuse. Administration of antibiotics into the CVC (the antibiotic lock technique) began 24 h later. The study included: one control group (no CVC infection), one contaminated group that did not receive any antibiotic prophylaxis, one contaminated group that received citropin 1.1-treated CVC, two contaminated groups that received citropin 1.1-treated CVC plus rifampin and minocycline at concentrations equal to MBCs for adherent cells and 1024 microg/mL in a volume of 0.1 mL that filled the CVC and two contaminated groups that received rifampin or minocycline at the same concentrations. All catheters were explanted 7 days after implantation. Main outcome measures were: minimal inhibitory concentration (MIC), minimal bactericidal concentration (MBC), synergy studies, quantitative culture of the biofilm formed on the catheters and surrounding venous tissues, and quantitative peripheral blood cultures. MICs of conventional antibiotics against the bacteria in a biofilm were at least four-fold higher than against the freely growing planktonic cells. In contrast, when antibiotics were used on citropin 1.1 pre-treated cells they showed comparable activity against both biofilm and planktonic organisms. The in vivo studies show that when CVCs were pre-treated with citropin 1.1 or with a high dose of antibiotics, biofilm bacterial load was reduced from 10(7) to 10(3) CFU/mL and bacteremia reduced from 10(3) to 10(1) CFU/mL. When CVCs were treated both with citropin 1.1 and antibiotics, biofilm bacterial load was further reduced to 10(1) CFU/mL and bacteremia was not detected, suggesting 100% elimination of bacteremia and a log 6 reduction in biofilm load. Citropin 1.1 significantly reduces bacterial load and enhances the effect of hydrophobic antibiotics in the treatment of CVC-associated S. aureus infections.     

引发医院感染表皮葡萄球菌生物被膜的检测

为了解引发医院感染的表皮葡萄球菌中ica操纵元的存在与生物被膜的产生的关系及其对抗生素敏感性的影响,收集了引发医院感染的表葡萄球菌106株,采用定量和定性法检测生物被膜的产生,PCR法检测ica操纵元基因的存在以及测量细菌对红霉素（ERY）、氨苄青霉素（AMP）、头孢西丁（FOX）、头孢曲松（CRO）、替考拉宁（TEC）、环丙沙星（CIP）、四环素（TCY）、复方新诺明（SXT）、万古霉素（VAN）的最小抑菌浓度（MIC）;106株表皮葡萄球菌分离株中,有33株检测出icaABC（31．1％）;ica^＋菌中产膜菌的检出率高于ica^＋菌（P=0．001）;葡萄糖和NaCl可提高产膜菌的检出率;ica^＋浮游菌对红霉素,头孢西丁和头孢曲松的耐药率高于ica^＋浮游菌株,但对氨苄青霉素,环丙沙星,四环素和复方新诺明的耐药率与ica^＋菌相似;ica位点基因的存在与引发表葡菌医院感染密切相关,但生物被膜内菌耐药机制还需进一步研究。     

Macrolides decrease the minimal inhibitory concentrations of anti-pseudomonal agents against Pseudomonas aeruginosa from cystic fibrosis patients in biofilm

ABSTRACT: BACKGROUND: Biofilm production is an important mechanism for bacterial survival and its association with antimicrobial resistance represents a challenge for the patient treatment. In this study we evaluated the in vitro action of macrolides in combination with anti-pseudomonal agents on biofilm-grown Pseudomonas aeruginosa recovered from cystic fibrosis (CF) patient. RESULTS: A total of 64 isolates were analysed. The (BIC) results were consistently higher than those minimal inhibitory concentration (MIC) for most anti-pseudomonal agents tested (ceftazidime: P = 0.001, ciprofloxacin: P = 0.234, tobramycin: P = 0.001, imipenem: P < 0.001, meropenem: P = 0.005). When macrolides were associated with the anti-pseudomonal agents, the BIC values were reduced significantly for ceftazidime (P < 0.001) and tobramycin (P < 0.001), regardless the concentration of macrolides. Strong (IQ) was observed when azithromycin at 8 mg/L was associated with all anti-pseudomonal agents tested in biofilm conditions. CONCLUSIONS: P. aeruginosa from CF patients within biofilms are highly resistant to antibiotics but macrolides proved to augment the in vitro activity of anti-pseudomonal agents.     

Antibiofilm activity of selected plant essential oils and their major components

The aim of the study was to examine the antibiofilm activity of selected essential oils (EO): Lavandula angustifblia (LEO), Melaleuca alternifolia (TTO), Melissa officinalis (MEO) and some of their major constituents: linalool, linalyl acetate, alpha-terpineol, terpinen-4-ol. Biofilms were formed by Staphylococcus aureus ATCC 29213 and Escherichia coli NCTC 8196 on the surface of medical biomaterials (urinary catheter, infusion tube and surgical mesh). TTC reduction assay was used for the evaluation of mature biofilm eradication from these surfaces. Moreover, time-dependent eradication ofbiofilms preformed in polystyrene 96-well culture microplates was examined and expressed as minimal biofilm eradication concentration (evaluated by MTT reduction assay). TTO, alpha-terpineol and terpinen-4-ol as well as MEO, showed stronger anti-biofilm activity than LEO and linalool or linalyl acetate. Among the biomaterials tested, surgical mesh was the surface most prone to persistent colonization since biofilms formed on it, both by S. aureus and E. coli, were difficult to destroy. The killing rate studies of S. aureus biofilm treated with TTO, LEO, MEO and some of their constituents revealed that partial (50%) destruction of 24-h-old biofilms (MBEC50) was achieved by the concentration 4-8 x MIC after 1 h, whereas 2-4 x MIC was enough to obtain 90% reduction in biomass metabolic activity (MBEC90) after just 4 h of treatment. A similar dose-dependent effect was observed for E. coli biofilm which, however, was more susceptible to the action of phytochemicals than the biofilms of S. aureus. It is noteworthy that an evident decrease in biofilm cells metabolic activity does not always lead to their total destruction and eradication.     

Inhibition and destruction of Pseudomonas aeruginosa biofilms by antibiotics and antimicrobial peptides

Pseudomonas aeruginosa is one of the major nosocomial pathogen that can causes a wide variety of acute and chronic infections P. aeruginosa is a dreaded bacteria not just because of the high intrinsic and acquired antibiotic resistance rates but also the biofilm formation and production of multiple virulence factors. We investigated the in vitro activities of antibiotics (ceftazidime, tobramycin, ciprofloxacin, doripenem, piperacillin and colistin) and antimicrobial cationic peptides (AMPs; LL-37, CAMA: cecropin(1–7)-melittin A(2–9) amide, melittin, defensin and magainin-II) alone or in combination against biofilms of laboratory strain ATCC 27853 and 4 clinical strains of P. aeruginosa. The minimum inhibitory concentrations (MIC), minimum bactericidal concentration (MBC) and minimum biofilm eradication concentrations (MBEC) were determined by microbroth dilution technique. The MBEC values of antibiotics and AMPs were 80–>5120 and 640–>640 mg/L, respectively. When combined with the LL-37 or CAMA at 1/10× MBEC, the MBEC values of antibiotics that active against biofilms, were decreased up to 8-fold. All of the antibiotics, and AMPs were able to inhibit the attachment of bacteria at the 1/10× MIC and biofilm formation at 1× or 1/10× MIC concentrations. Time killing curve studies showed 3-log₁₀ killing against biofilms in 24 h with almost all studied antibiotics and AMPs. Synergism were seen in most of the studied combinations especially CAMA/LL-37 + ciprofloxacin against at least one or two strains’ biofilms. Since biofilms are not affected the antibiotics at therapeutic concentrations, using a combination of antimicrobial agents including AMPs, or inhibition of biofilm formation by blocking the attachment of bacteria to surfaces might be alternative methods to fight with biofilm associated infections.     

Characterization of biofilm growth and biocide susceptibility testing of Mycobacterium phlei using the MBEC assay system

The importance of non-tuberculosis mycobacterial biofilm species in medicine, industry and the environment has recently gained attention. Our objectives were to characterize biofilm growth of Mycobacterium phlei M4, as a model of rapidly growing mycobacteria using the minimal biofilm eradication concentration (MBEC) and to compare biocide susceptibility of planktonic and biofilm organisms. Scanning electron microscopy was also carried out to observe biofilm morphology. With the exception of Sporicidin and Virkon the minimum bactericidal concentration values for all biocides tested were lower than the MBEC values. The MBEC assay system was seen to produce multiple and reproducible biofilms of M. phlei and to be a useful tool for susceptibility studies.     

Biofilm susceptibility to metal toxicity

This study compared bacterial biofilm and planktonic cell susceptibility to metal toxicity by evaluating the minimum inhibitory concentration (MIC), the planktonic minimum bactericidal concentration (MBC), and minimum biofilm eradication concentration (MBEC) using the MBEC device. In total, 17 metal cations and oxyanions, chosen to represent groups VIB to VIA of the periodic table, were each tested on biofilm and planktonic cultures of Escherichia coli JM109, Staphylococcus aureus ATCC 29213, and Pseudomonas aeruginosa ATCC 27853. In contrast to control antibiotic assays, where biofilm cultures were 2 to 64 times less susceptible to killing than logarithmically growing planktonic bacteria, metal compounds killed planktonic and biofilm cultures at the same concentration in the vast majority of combinations. Our data indicate that, under the conditions reported, growth in a biofilm does not provide resistance to bacteria against killing by metal cations or oxyanions.     

Persister cells, the biofilm matrix and tolerance to metal cations in biofilm and planktonic Pseudomonas aeruginosa

In this study, we examined Pseudomonas aeruginosa ATCC 27853 biofilm and planktonic cell susceptibility to metal cations. The minimum inhibitory concentration (MIC), the minimum bactericidal concentration (MBC) required to eradicate 100% of the planktonic population (MBC 100), and the minimum biofilm eradication concentration (MBEC) were determined using the MBEC trade mark-high throughput assay. Six metals - Co(2+), Ni(2+), Cu(2+), Zn(2+), Al(3+) and Pb(2+)- were each tested at 2, 4, 6, 8, 10 and 27 h of exposure to biofilm and planktonic cultures grown in rich or minimal media. With 2 or 4 h of exposure, biofilms were approximately 2-25 times more tolerant to killing by metal cations than the corresponding planktonic cultures. However, by 27 h of exposure, biofilm and planktonic bacteria were eradicated at approximately the same concentration in every instance. Viable cell counts evaluated at 2 and 27 h of exposure revealed that at high concentrations, most of the metals assayed had killed greater than 99.9% of biofilm and planktonic cell populations. The surviving cells were propogated in vitro and gave rise to biofilm and planktonic cultures with normal sensitivity to metals. Further, retention of copper by the biofilm matrix was investigated using the chelator sodium diethlydithiocarbamate. Formation of visible brown metal-chelates in biofilms treated with Cu(2+) suggests that the biofilm matrix may coordinate and sequester metal cations from the aqueous surroundings. Overall, our data suggest that both metal sequestration in the biofilm matrix and the presence of a small population of 'persister' cells may be contributing factors in the time-dependent tolerance of both planktonic cells and biofilms to high concentrations of metal cations.     

The assessment of the antibacterial and antifungal activities of aspirin, EDTA and aspirin–EDTA combination and their effectiveness as antibiofilm agents

Aims:  To evaluate the antimicrobial activities of aspirin, EDTA and an aspirin-EDTA (A-EDTA) combination against Pseudomonas aeruginosa , Escherichia coli and Candida albicans in planktonic and biofilm cultures.
Methods and Results:  Minimal inhibitory concentrations (MIC) and minimal biocidal concentrations (MBC) were determined using twofold broth microdilution and viable counting methods, respectively. Aspirin's recorded MIC values ranged from 1·2 to 2·7 mg ml⁻¹. Checkerboard assay demonstrated a synergism in antimicrobial activity upon combination. Aspirin's minimal biofilm eradication concentration values (MBEC) against the established biofilms ranged between 1·35 and 3·83 mg ml⁻¹. A complete eradication of bacterial biofilms was achieved after a 4-h treatment with the A-EDTA combination.
Conclusion:  Both aspirin and EDTA possess broad-spectrum antimicrobial activity for both planktonic and biofilm cultures. Aspirin used at the MBEC for 24 h was successful in eradicating P. aeruginosa , E. coli and C. albicans biofilms established on abiotic surfaces. Moreover, the exposure to the A-EDTA combination (4 h) effected complete bacterial biofilm eradication.
Significance and Impact of the Study:  There is a continuous need for the discovery of new antimicrobial agents. Aspirin and EDTA are 'nonantibiotic drugs', the combination of which can be used successfully to treat and eradicate biofilms established on abiotic surfaces.     

Antibiotic Susceptibility of Biofilm Cells and Molecular Characterisation of Staphylococcus hominis Isolates from Blood

Objectives

We aimed to characterise the staphylococcal cassette chromosome mec (SCCmec) type, genetic relatedness, biofilm formation and composition, icaADBC genes detection, icaD expression, and antibiotic susceptibility of planktonic and biofilm cells of Staphylococcus hominis isolates from blood.

Methods

The study included 67 S. hominis blood isolates. Methicillin resistance was evaluated with the cefoxitin disk test. mecA gene and SCCmec were detected by multiplex PCR. Genetic relatedness was determined by pulsed-field gel electrophoresis. Biofilm formation and composition were evaluated by staining with crystal violet and by detachment assay, respectively; and the biofilm index (BI) was determined. Detection and expression of icaADBC genes were performed by multiplex PCR and real-time PCR, respectively. Antibiotic susceptibilities of planktonic cells (minimum inhibitory concentration, MIC) and biofilm cells (minimum biofilm eradication concentration, MBEC) were determined by the broth dilution method.

Results

Eighty-five percent (57/67) of isolates were methicillin resistant and mecA positive. Of the mecA-positive isolates, 66.7% (38/57) carried a new putative SCCmec type. Four clones were detected, with two to five isolates each. Among all isolates, 91% (61/67) were categorised as strong biofilm producers. Biofilm biomass composition was heterogeneous (polysaccharides, proteins and DNA). All isolates presented the icaD gene, and 6.66% (1/15) isolates expressed icaD. This isolate presented the five genes of ica operon. Higher BI and MBEC values than the MIC values were observed for amikacin, vancomycin, linezolid, oxacillin, ciprofloxacin, and chloramphenicol.

Conclusions

S. hominis isolates were highly resistant to methicillin and other antimicrobials. Most of the detected SCCmec types were different than those described for S. aureus. Isolates indicated low clonality. The results indicate that S. hominis is a strong biofilm producer with an extracellular matrix with similar composition of proteins, DNA and N-acetylglucosamine; and presents high frequency and low expression of icaD gene. Biofilm production is associated with increased antibiotic resistance.     

Azithromycin Reduces the Production of α-hemolysin and Biofilm Formation in Staphylococcus aureus

Staphylococcus aureus causes a broad range of life-threatening diseases in humans. This bacterium produces a large number of extracellular virulence factors that are closely associated with specific diseases which are controlled by quorum sensing. In this study, we show that azithromycin was active against methicillin-resistant Staphylococcus aureus (MRSA) strains with MICs ranged from 32 to 64 μg/mL. Azithromycin at subinhibitory concentration, markedly reduced the production of α-hemolysin at (1/16MIC, 1/8MIC) and biofilm formation at (1/16MIC, 1/8MIC), respectively. The results indicated that sub-inhibitory concentrations of azithromycin decreased the production of α-hemolysin and biofilm formation in MRSA in a dose-dependent manner. Therefore, azithromycin may be useful in the treatment of α-hemolysin producing and biofilm formation MRSA infections.     

Effectiveness of the antibiotics chloramphenicol and rifampin in the treatment of Streptococcus pneumoniae-induced meningitis and systemic infections

Streptococcus pneumoniae, a common pathogen in pediatric infections, has become resistant to penicillin and make these infections difficult to treat. Rifampin and chloramphenicol have been recommended as alternative therapies, since they are less costly and more accessible to communities with limited resources. However, their use may be restricted by the differing levels of resistance found in target populations. The objective was to determine minimal inhibitory concentration (MIC) and minimal bactericidal concentration (MBC) for chloramphenicol and rifampin in strains of S. pneumoniae. These strains were newly isolated from children under age 5 that had demonstrated systemic infections and meningitis. A subgroup of 107 isolates of S. pneumoniae was selected from 324 strains isolated during a period of 2 years (1994-1996). Among these isolates, 60 were penicillin-resistant and 47 were susceptible; 53 isolates were from children with meningitis. MIC and MBC for chloramphenicol and rifampicin were obtained by standard methods recommended by the National Committee for Clinical Laboratory Standards (NCCLS). S. pneumoniae ATCC strain 49619 served as the control. An isolate was considered susceptible to chloramphenicol when MIC = 4 microg/ml and resistant when MIC = 8 microg/ml. A strain was considered susceptible to rifampin when MIC = 1 microg/ml and resistant when MIC = 4 microg/ml. MBC was determined by recording the lower concentration of the antibiotic that inhibited 99.9% of the initial inoculum. Chloramphenicol resistance was found in 21% of the 107 isolates. In the group susceptible to penicillin, 11% were resistant to chloramphenicol and in the group resistant to penicillin 28% was resistant to chloramphenicol as well. MBC was found > 4 microg/ml in 28% of the isolates susceptible to penicillin and in 60% of the resistant isolates. No isolates were found resistant to rifampin. However, 2 penicillin resistant isolates showed CBM > 1 microg/ml to rifampin, and one with CIM = 1 microg/ml had a MBC to rifampicin of 16 microg/ml. Meningitis isolates showed higher CIM and CBM than the group of total isolates. These data suggest that chloramphenicol is not recommended for invasive infections caused by S. pneumoniae in Colombia. Rifampin is a more effective therapy in combination with other antibiotics for treatment of this kind of infections. Further studies are necessary to clarify the significance of low levels of MBC to rifampin found in some strains, since this may affect the efficacy of therapies that include this antibiotic.     

Promethazine improves antibiotic efficacy and disrupts biofilms of Burkholderia pseudomallei

Efflux pumps are important defense mechanisms against antimicrobial drugs and maintenance of Burkholderia pseudomallei biofilms. This study evaluated the effect of the efflux pump inhibitor promethazine on the structure and antimicrobial susceptibility of B. pseudomallei biofilms. Susceptibility of planktonic cells and biofilms to promethazine alone and combined with antimicrobials was assessed by the broth microdilution test and biofilm metabolic activity was determined with resazurin. The effect of promethazine on 48 h-grown biofilms was also evaluated through confocal and electronic microscopy. The minimum inhibitory concentration (MIC) of promethazine was 780 mg l^?1, while the minimum biofilm elimination concentration (MBEC) was 780–3,120 mg l^?1. Promethazine reduced the MIC values for erythromycin, trimethoprim/sulfamethoxazole, gentamicin and ciprofloxacin and reduced the MBEC values for all tested drugs (p<0.05). Microscopic analyses demonstrated that promethazine altered the biofilm structure of B. pseudomallei, even at subinhibitory concentrations, possibly facilitating antibiotic penetration. Promethazine improves antibiotics efficacy against B. pseudomallei biofilms, by disrupting biofilm structure.     

Antimicrobial and antibiofilm potential of injectable platelet rich fibrin—a second-generation platelet concentrate—against biofilm producing oral staphylococcus isolates

Injectable Platelet rich fibrin (i-PRF) is a platelet concentrate that has been extensively used for multiple medical purposes and is a valuable adjunct for the regeneration of damaged tissues in surgical procedures. The enriched bioactive substances in i-PRF are responsible for speeding the wound healing process. Infection of biofilm producing bacteria in surgical wounds is becoming a serious threat. Research in this field is focused on new strategies to fight infections and to reduce the healing time. The present study was aimed to evaluate the in vitro antimicrobial and antibiofilm effects of i-PRF against oral pathogenic biofilm producing staphylococcus bacteria isolated from patient with dental and oral abscess. The antibacterial activity of i-PRF, was determined through broth microdilution as minimal inhibitory concentration (MIC) and minimal bactericidal concentration (MBC). i-PRF exhibited bactericidal activity against both non biofilm and biofilm producing bacteria. i-PRF could be potential antimicrobial peptide used to combat postoperative infections caused by biofilm producing staphylococcus.     

Growing Burkholderia pseudomallei in Biofilm Stimulating Conditions Significantly Induces Antimicrobial Resistance

Background

Burkholderia pseudomallei, a Gram-negative bacterium that causes melioidosis, was reported to produce biofilm. As the disease causes high relapse rate when compared to other bacterial infections, it therefore might be due to the reactivation of the biofilm forming bacteria which also provided resistance to antimicrobial agents. However, the mechanism on how biofilm can provide tolerance to antimicrobials is still unclear.

Methodology/Principal Findings

The change in resistance of B. pseudomallei to doxycycline, ceftazidime, imipenem, and trimethoprim/sulfamethoxazole during biofilm formation were measured as minimum biofilm elimination concentration (MBEC) in 50 soil and clinical isolates and also in capsule, flagellin, LPS and biofilm mutants. Almost all planktonic isolates were susceptible to all agents studied. In contrast, when they were grown in the condition that induced biofilm formation, they were markedly resistant to all antimicrobial agents even though the amount of biofilm production was not the same. The capsule and O-side chains of LPS mutants had no effect on biofilm formation whereas the flagellin-defective mutant markedly reduced in biofilm production. No alteration of LPS profiles was observed when susceptible form was changed to resistance. The higher amount of N-acyl homoserine lactones (AHLs) was detected in the high biofilm-producing isolates. Interestingly, the biofilm mutant which produced a very low amount of biofilm and was sensitive to antimicrobial agents significantly resisted those agents when grown in biofilm inducing condition.

Conclusions/Significance

The possible drug resistance mechanism of biofilm mutants and other isolates is not by having biofilm but rather from some factors that up-regulated when biofilm formation genes were stimulated. The understanding of genes related to this situation may lead us to prevent B. pseudomallei biofilms leading to the relapse of melioidosis.     

Susceptibility of Candida glabrata biofilms to echinocandins: alterations in the matrix composition

Candidiases are the most recurrent fungal infections, especially among immunosuppressed patients. Although Candida albicans is still the most widespread isolated species, non-Candida albicans Candida species have been increasing. The goal of this work was to determine the susceptibility of C. glabrata biofilms to echinocandins and to evaluate their effect on the biofilm matrix composition, comparing the results with other Candida species. Drug susceptibilities were assessed through the determination of minimum inhibitory concentration (MIC), minimum fungicidal concentration (MFC) and minimum biofilm eradication concentration (MBEC) of caspofungin (Csf) and micafugin (Mcf). The β-1,3 glucans content of the matrices was assessed after contact with the drugs. The data suggest that, generally, after contact with echinocandins, the concentration of β-1,3 glucans increased. These adjustments in the matrix composition of C. glabrata biofilms and the chemical differences between Csf and Mcf, seem responsible and may determine the effectivity of the drug responses.     

Fimbria targeting superparamagnetic iron oxide nanoparticles enhance the antimicrobial and antibiofilm activity of ciprofloxacin against quinolone-resistant E. coli

High quinolone resistance of Escherichia coli limits the therapy options for urinary tract infection (UTI). In response to the urgent need for efficient treatment of multidrug-resistant infections, we designed a fimbriae targeting superparamagnetic iron oxide nanoparticle (SPION) delivering ciprofloxacin to ciprofloxacin-resistant E. coli. Bovine serum albumin (BSA) conjugated poly(acrylic acid) (PAA) coated SPIONs (BSA@PAA@SPION) were developed for encapsulation of ciprofloxacin and the nanoparticles were tagged with 4-aminophenyl-α-D-mannopyrannoside (mannoside, Man) to target E. coli fimbriae. Ciprofloxacin-loaded mannoside tagged nanoparticles (Cip-Man-BSA@PAA@SPION) provided high antibacterial activity (97.1 and 97.5%, respectively) with a dose of 32 μg/mL ciprofloxacin against two ciprofloxacin-resistant E. coli isolates. Furthermore, a strong biofilm inhibition (86.9% and 98.5%, respectively) was achieved in the isolates at a dose 16 and 8 times lower than the minimum biofilm eradication concentration (MBEC) of ciprofloxacin. Weaker growth inhibition was observed with untargeted nanoparticles, Cip-BSA@PAA@SPIONs, confirming that targeting E. coli fimbria with mannoside-tagged nanoparticles increases the ciprofloxacin efficiency to treat ciprofloxacin-resistant E. coli. Enhanced killing activity against ciprofloxacin-resistant E. coli planktonic cells and strong growth inhibition of their biofilms suggest that Cip-Man-BSA@PAA@SPION system might be an alternative and/or complementary therapeutic option for the treatment of quinolone-resistant E. coli infections.     

Comparison of Antifungal Activities of Gentian Violet and Povidone-Iodine Against Clinical Isolates of Candida Species and Other Yeasts: A Framework to Establish Topical Disinfectant Activities

We evaluated antifungal activity as assessed by the contact time in topical use of gentian violet (GV) and povidone-iodine (PI) against Candida strains. A total of 102 yeast isolates were used in this study. A markedly lower minimal inhibitory concentration (MIC)₉₀ of GV than of PI was detected for all yeast isolates. No remarkable difference in the MICs was observed among the identical strains isolated from different clinical sites for both GV and PI. Although the minimal fungicidal activities (MFCs) of PI were identical for all tested time points, the fungicidal activity of GV decreased during the time course of incubation. These results indicate that, whereas GV is more effective than PI, the topical disinfectant efficacy of GV should be estimated using the MFC_5 min and not the MIC or the MFC_24 h for overall prevention of catheter-related bloodstream infections and oral infections.