In vitro formation of oropharyngeal biofilms on silicone rubber treated with a palladium/tin salt mixture

Adhesion of yeasts and bacteria to silicone rubber is one of the first steps in the biodeterioration of indwelling, silicone rubber voice prostheses. In this paper, silicone rubber, so-called "Groningen button," voice prostheses were treated with a colloidal palladium/tin solution to form a thin metal coat intended to discourage biofilm formation. First it was demonstrated that this treatment did not negatively affect the airflow resistance of the prostheses or induce any cytotoxicity. Subsequently, palladium/tin-treated voice prostheses were placed in a modified Robbins device together with untreated control prostheses to evaluate biofilm formation. Biofilms were formed by inoculating the device for 3 days with the total cultivable microflora obtained from an explanted, malfunctioning voice prosthesis supplemented with separately isolated yeasts (Candida albicans and Candida tropicalis). After 3 days the device was perfused three times daily with growth medium and phosphate-buffered saline. The device was allowed to drain between perfusions to better mimic the conditions in the oropharynx (moist but not always fully wetted). After 9 days the total number of bacterial and fungal colony-forming units on the prostheses were determined microbiologically, and scanning electron micrographs were taken of the valve sides. Biofilm formation was significantly less on the heavily treated palladium/tin prostheses than it was on the untreated prostheses although some ingrowing microcolonies also were observed on the treated prostheses. The spread of the biofilms was smaller on the treated prostheses than on the untreated ones.     

Effect of probiotic bacteria on prevalence of yeasts in oropharyngeal biofilms on silicone rubber voice prostheses in vitro

The proliferation of yeasts in the mixed bacterial and fungal biofilms colonising silicone rubber voice prostheses in laryngectomised patients is the main cause of malfunctioning of the valve mechanism on the oesophageal side of the prostheses. Indwelling voice prostheses usually have to be replaced every 3-4 months. The consumption of probiotic bacteria is largely motivated by health claims related to the urogenital and lower digestive tract, but not to the upper digestive tract. The present study examined the influence of probiotic bacteria on the prevalence of yeasts in oropharyngeal biofilms on silicone rubber voice prostheses, as formed in a modified Robbins device. Exposure of oropharyngeal biofilms on voice prostheses to suspensions of Bifidobacterium infantis 420 or Enterococcus faecium 603 did not significantly reduce the number of yeasts in the biofilm. However, suspensions of Lactobacillus fermentum B54, L. rhamnosus 744 or L. lactis cremoris SK11 led to a reduction in the number of yeasts harvested from the voice prostheses. Suspensions of L. casei Shirota and Streptococcus thermophilus B significantly reduced the number of yeasts in the biofilm to 39% and 33%, respectively. The reduction brought about in yeast prevalence in the mixed biofilm was greatest by exposure to a suspension of L. lactis 53, with yeast prevalence only 4% of the control. In conclusion, the study demonstrated that the prevalence of yeasts in oropharyngeal biofilms on silicone rubber voice prostheses might be controlled by consumption of probiotic bacteria.     

Biofilm formation and design features of indwelling silicone rubber tracheoesophageal voice prostheses--an electron microscopical study

After total laryngectomy, voice can be restored with a silicone rubber tracheoesophageal voice prosthesis. However, biofilm formation and subsequent deterioration of the silicone material of the prosthesis will limit device life by impairing valve function. To simulate the natural process of biofilm development under dynamic nutrient conditions, a modified Robbins device was used to evaluate the biofilm-related valve dysfunction of the Groningen, Provox2, Blom-Singer indwelling, and VoiceMaster voice prostheses. Obstruction of the semicircular slit-valved Groningen prosthesis leading to increased airway resistance was caused not only by a buildup of deposits on the esophageal flange and valve hat, but also by accumulation of deposits on the semicircular valve seating. The hinged flap valved Provox2 and indwelling Blom-Singer prostheses failed to close sufficiently because of biofilm formation on the valve seating. The esophageal flange of the VoiceMaster prosthesis was affected, but the tripod structure of the ball valve was fully colonized up to the titanium sleeve, which interfered with proper valve opening and closure. These findings on biofilm formation could be used for the further development and modification of critical design features of voice prostheses to facilitate tracheoesophageal speech.     

Strategies for the prevention of microbial biofilm formation on silicone rubber voice prostheses

Total laryngectomy, a surgical treatment for extensive cancer of larynx, which alters swallowing and respiration in patients, is followed up with a surgical voice restoration procedure comprising tracheoesophageal puncture techniques with insertion of a "voice prosthesis" to improve successful voice rehabilitation. However, microbial colonization is a major drawback of these devices. Antimicrobials are usually used to prevent the colonization of silicone rubber voice prostheses by microorganisms. However, long-term medication induces the development of resistant strains with all associated risks and the development of alternative prophylactic and therapeutic agents, including probiotics and biosurfactants, have been suggested. The inhibition of microbial growth on surfaces can also be achieved by several other techniques involving the modification of physicochemical properties of the biomaterial surface or the covalently binding of antimicrobial agents to the biomaterial surface. An overview of the different approaches investigated to date and future perspectives to reduce the frequent replacements of voice prostheses in laryngectomized patients through microbial biofilm retardation is presented and discussed.     

Mechanism of fluconazole resistance in Candida albicans biofilms: phase-specific role of efflux pumps and membrane sterols

Candida albicans biofilms are formed through three distinct developmental phases and are associated with high fluconazole (FLU) resistance. In the present study, we used a set of isogenic Candida strains lacking one or more of the drug efflux pumps Cdr1p, Cdr2p, and Mdr1p to determine their role in FLU resistance of biofilms. Additionally, variation in sterol profile as a possible mechanism of drug resistance was investigated. Our results indicate that parent and mutant strains formed similar biofilms. However, biofilms formed by double and triple mutants were more susceptible to FLU at 6 h (MIC = 64 and 16 microg/ml, respectively) than the wild-type strain (MIC > 256 microg/ml). At later time points (12 and 48 h), all the strains became resistant to this azole (MIC > or = 256 microg/ml), indicating lack of involvement of efflux pumps in resistance at late stages of biofilm formation. Northern blot analyses revealed that Candida biofilms expressed CDR and MDR1 genes in all the developmental phases, while planktonic cells expressed these genes only at the 12- and 48-h time points. Functionality of efflux pumps was assayed by rhodamine (Rh123) efflux assays, which revealed significant differences in Rh123 retention between biofilm and planktonic cells at the early phase (P = 0.0006) but not at later stages (12 and 48 h). Sterol analyses showed that ergosterol levels were significantly decreased (P < 0.001) at intermediate and mature phases, compared to those in early-phase biofilms. These studies suggest that multicomponent, phase-specific mechanisms are operative in antifungal resistance of fungal biofilms.     

Role of dimorphism in the development of Candida albicans biofilms.

Two model biofilm systems, involving growth on disks of catheter material or on cylindrical cellulose filters, were used to investigate the structure of Candida albicans biofilms. To assess the importance of dimorphism in biofilm development, biofilms produced by two wild-type strains were compared with those formed by two morphological mutants, incapable of yeast and hyphal growth, respectively. Scanning electron microscopy and thin sections of biofilms examined by light microscopy revealed that biofilms of the wild-type strains formed on catheter disks consisted of two distinct layers: a thin, basal yeast layer and a thicker, but more open, hyphal layer. The hypha- mutant produced only the basal layer, whereas the yeast- mutant formed a thicker, hyphal biofilm equivalent to the outer zone of the wild-type structures. Biofilms of the yeast- mutant were more easily detached from the catheter surface than the others, suggesting that the basal yeast layer has an important role in anchoring the biofilm to the surface. Biofilms formed on cylindrical cellulose filters were quite different in appearance. The hypha- mutant and both wild types produced exclusively yeast-form biofilms whereas the yeast- mutant generated a dense hyphal mat on the top of the filter. All these biofilms, irrespective of morphological form, were resistant to the antifungal agent, amphotericin B. Overall, these results indicate that the structure of a C. albicans biofilm depends on the nature of the contact surface, but that some surfaces produce biofilms with a layered architecture resembling to that described for bacterial systems.     

Bacterial adhesion and growth on a polymer brush-coating

Biomaterials-related infections pose serious problems in implant surgery, despite the development of non-adhesive coatings. Non-adhesive coatings, like polymer brush-coatings, have so far only been investigated with respect to preventing initial bacterial adhesion, but never with respect to effects on kinetics of bacterial growth. Here, we compare adhesion and 20h growth of three bacterial strains (Staphylococcus aureus, Staphylococcus epidermidis and Pseudomonas aeruginosa) on pristine and brush-coated silicone rubber in a parallel plate flow chamber. Brush-coatings were made using a tri-block copolymer of polyethylene oxide (PEO) and polypropylene oxide (PPO). Brush-coatings prevented adhesion of staphylococci to below 5x10(5)cm(-2) after 30min, which is a 10-fold reduction compared to pristine silicone rubber. Biofilms grew on both brush-coated and pristine silicone rubber, while the viability of biofilms on brush-coatings was higher than on pristine silicone rubber. However, biofilms on brush-coatings developed more slowly and detached almost fully by high fluid shear. Brush-coating remained non-adhesive after S. epidermidis biofilm formation and subsequent removal whereas a part of its functionality was lost after removal of S. aureus biofilms. Adhesion, growth and detachment of P. aeruginosa were not significantly different on brush-coatings as compared with pristine silicone rubber, although here too the viability of biofilms on brush-coatings was higher. We conclude that polymer brush-coatings strongly reduce initial adhesion of staphylococci and delay their biofilm growth. In addition, biofilms on brush-coatings are more viable and easily removed by the application of fluid shear.     

Comparison of Biofilms Formed by Candida albicans and Candida parapsilosis on Bioprosthetic Surfaces

Little is known about fungal biofilms, which may cause infection and antibiotic resistance. In this study, biofilm formation by different Candida species, particularly Candida albicans and C. parapsilosis, was evaluated by using a clinically relevant model of Candida biofilm on medical devices. Candida biofilms were allowed to form on silicone elastomer and were quantified by tetrazolium (XTT) and dry weight (DW) assays. Formed biofilm was visualized by using fluorescence microscopy and confocal scanning laser microscopy with Calcofluor White (Sigma Chemical Co., St. Louis, Mo.), concanavalin A-Alexafluor 488 (Molecular Probes, Eugene, Oreg.), and FUN-1 (Molecular Probes) dyes. Although minimal variations in biofilm production among invasive C. albicans isolates were seen, significant differences between invasive and noninvasive isolates (P < 0.001) were noted. C. albicans isolates produced more biofilm than C. parapsilosis, C. glabrata, and C. tropicalis isolates, as determined by DW assays (P was <0.001 for all comparisons) and microscopy. Interestingly, noninvasive isolates demonstrated a higher level of XTT activity than invasive isolates. On microscopy, C. albicans biofilms had a morphology different from that of other species, consisting of a basal blastospore layer with a dense overlying matrix composed of exopolysaccharides and hyphae. In contrast, C. parapsilosis biofilms had less volume than C. albicans biofilms and were comprised exclusively of clumped blastospores. Unlike planktonically grown cells, Candida biofilms rapidly (within 6 h) developed fluconazole resistance (MIC, >128 microg/ml). Importantly, XTT and FUN-1 activity showed biofilm cells to be metabolically active. In conclusion, our data show that C. albicans produces quantitatively larger and qualitatively more complex biofilms than other species, in particular, C. parapsilosis.     

Biofilm formation by Candida species on the surface of catheter materials in vitro.

A model system for studying Candida biofilms growing on the surface of small discs of catheter material is described. Biofilm formation was determined quantitatively by a colorimetric assay involving reduction of a tetrazolium salt or by [3H]leucine incorporation; both methods gave excellent correlation with biofilm dry weight (r = 0.997 and 0.945, respectively). Growth of Candida albicans biofilms in medium containing 500 mM galactose or 50 mM glucose reached a maximum after 48 h and then declined; however, the cell yield was lower in low-glucose medium. Comparison of biofilm formation by 15 different isolates of C. albicans failed to reveal any correlation with pathogenicity within this group, but there was some correlation with pathogenicity when different Candida species were tested. Isolates of C. parapsilosis (Glasgow), C. pseudotropicalis, and C. glabrata all gave significantly less biofilm growth (P < 0.001) than the more pathogenic C. albicans. Evaluation of various catheter materials showed that biofilm formation by C. albicans was slightly increased on latex or silicone elastomer (P < 0.05), compared with polyvinyl chloride, but substantially decreased on polyurethane or 100% silicone (P < 0.001). Scanning electron microscopy demonstrated that after 48 h, C. albicans biofilms consisted of a dense network of yeasts, germ tubes, pseudohyphae, and hyphae; extracellular polymeric material was visible on the surfaces of some of these morphological forms. Our model system is a simple and convenient method for studying Candida biofilms and could be used for testing the efficacy of antifungal agents against biofilm cells.     

Alcohol dehydrogenase restricts the ability of the pathogen Candida albicans to form a biofilm on catheter surfaces through an ethanol-based mechanism

Candida biofilms formed on indwelling medical devices are increasingly associated with severe infections. In this study, we used proteomics and Western and Northern blotting analyses to demonstrate that alcohol dehydrogenase (ADH) is downregulated in Candida biofilms. Disruption of ADH1 significantly (P = 0.0046) enhanced the ability of Candida albicans to form biofilm. Confocal scanning laser microscopy showed that the adh1 mutant formed thicker biofilm than the parent strain (210 microm and 140 microm, respectively). These observations were extended to an engineered human oral mucosa and an in vivo rat model of catheter-associated biofilm. Inhibition of Candida ADH enzyme using disulfiram and 4-methylpyrazole resulted in thicker biofilm (P < 0.05). Moreover, biofilms formed by the adh1 mutant strain produced significantly smaller amounts of ethanol, but larger amounts of acetaldehyde, than biofilms formed by the parent and revertant strains (P < 0.0001), demonstrating that the effect of Adh1p on biofilm formation is mediated by its enzymatic activity. Furthermore, we found that 10% ethanol significantly inhibited biofilm formation in vitro, with complete inhibition of biofilm formation at ethanol concentrations of >/=20%. Similarly, using a clinically relevant rabbit model of catheter-associated biofilm, we found that ethanol treatment inhibited biofilm formation by C. albicans in vivo (P < 0.05) but not by Staphylococcus spp. (P > 0.05), indicating that ethanol specifically inhibits Candida biofilm formation. Taken together, our studies revealed that Adh1p contributes to the ability of C. albicans to form biofilms in vitro and in vivo and that the protein restricts biofilm formation through an ethanol-dependent mechanism. These results are clinically relevant and may suggest novel antibiofilm treatment strategies.     

In vitro leakage susceptibility of tracheoesophageal shunt prostheses in the absence and presence of a biofilm

Although leakage through a tracheoesophageal shunt prosthesis is the main cause of prosthesis failure in a laryngectomy patient, this has never been the subject of in vitro evaluation. The aim of this study was to compare three commercially available voice prostheses by comparison of their in vitro leakage patterns, in absence or presence of a biofilm. To compare in vitro leakage patterns, a model comprised of an artificial throat equipped with a single prosthesis coupled to a water reservoir was developed. By varying the height of the water reservoir, different pressures on the voice prosthesis can be obtained. Both in absence and presence of a biofilm, the Blom Singer voice prosthesis demonstrated the lowest leakage, followed by Groningen Low Resistance. The Provox2 showed significantly the most leakage, however, in presence of a biofilm the leakage of the Provox2 significantly decreased. Regular airflow during biofilm formation significantly increased leakage through the Provox2. Out of 746 clinical replacements, Provox2 showed 76% and Groningen Low Resistance 57% replacements due to leakage. The model used in this study showed significant differences in leakage of the three types of voice prostheses used. Leakage occurred more readily through Provox2 than through Groningen Low Resistance and Blom Singer prostheses, which is in line with clinical observations and enforces the model.     

Formation of Propionibacterium acnes biofilms on orthopaedic biomaterials and their susceptibility to antimicrobials

Failure to treat and eradicate prosthetic hip infection with systemic antibiotic regimens is usually due to the fact that the infection is associated with biofilm formation and that bacterial cells growing within a biofilm exhibit increased resistance to antimicrobial agents. In this in vitro study, we investigated the susceptibility of prosthetic hip Propionibacterium acnes and Staphylococcus spp. isolates growing within biofilms on polymethylmethacrylate (PMMA) bone cement to a range of antibiotics. All P. acnes isolates in the biofilm mode of growth demonstrated considerably greater resistance to cefamandole, ciprofloxacin and vancomycin. In contrast, only four of the eight P. acnes isolates demonstrated an increase in resistance to gentamicin. All ten Staphylococcus spp. isolates in the biofilm mode of growth exhibited large increases in resistance to gentamicin and cefamandole with eight of the ten isolates also exhibiting an increase in resistance to vancomycin. However, only three of the ten Staphylococcus spp. isolates exhibited an increase in resistance to ciprofloxacin. Biofilms were also formed on three different titanium alloys and on PMMA bone cement using P. acnes, Staphylococcus epidermidis and Staphylococcus aureus strains to determine if the underlying biomaterial surface had an effect on biofilm formation and the antimicrobial susceptibility of the bacteria growing within biofilms. Although differences in the rate at which the three strains adhered to the different biomaterials were apparent, no differences in biofilm antibiotic resistance between the biomaterials were observed. In the light of these results, it is important that the efficacy of other antibiotics against P. acnes and Staphylococcus spp. prosthetic hip isolates growing within biofilms on orthopaedic biomaterials be determined to ensure optimal treatment of orthopaedic implant infection.     

Characterization of biofilms formed by Candida parapsilosis, C. metapsilosis, and C. orthopsilosis

Infections due to Candida parapsilosis have been associated with the ability of this fungus to form biofilms on indwelling medical devices. Recently, C. parapsilosis isolates were reclassified into 3 genetically non-identical classes: C. parapsilosis, C. orthopsilosis, and C. metapsilosis. Little information is available regarding the ability of these newly reclassified species to form biofilms on biomedical substrates. In this study, we characterized biofilm formation by 10 clinical isolates each of C. parapsilosis, C. orthopsilosis, and C. metapsilosis. Biofilms were allowed to form on silicone elastomer discs to early (6 h) or mature (48 h) phases and quantified by tetrazolium (XTT) and dry weight assays. Surface topography and three-dimensional architecture of the biofilms were visualized using scanning electron microscopy (SEM) and confocal laser scanning microscopy (CLSM), respectively. Metabolic activity assay revealed strain-dependent biofilm forming ability of the 3 species tested, while biomass determination revealed that all 3 species formed equivalent biofilms (P>0.05 for all comparisons). SEM analyses of representative isolates of these species showed biofilms with clusters of yeast cells adherent to the catheter surface. Additionally, confocal microscopy analyses showed the presence of cells embedded in biofilms ranging in thickness between 62 and 85 μm. These results demonstrate that similar to C. parapsilosis, the 2 newly identified Candida species (C. orthopsilosis and C. metapsilosis) were able to form biofilms.     

Biochemical and genetic characteristics of Cronobacter sakazakii biofilm formation

Cronobacter sakazakii is a wide-spread opportunistic foodborne pathogen that can form biofilms on a number of different substances, creating food safety risk. However, there is little information about biofilm characteristics for this species. In this study, biofilm formation of 14 foodborne C. sakazakii strains was examined. Transposon mutants of the strain (IQCC10423), the isolate with the greatest biofilm activity, were prepared. A total of 12 mutants were developed with >40% reduction in biofilm formation ability. Eight of these mutants were successfully sequenced with genes putatively identified for: biofilm formation, fundamental cellular processes, phage tail complete protein and uncertain functional protein. The morphology of the biofilm showed that the wild type strain formed a thick biofilm and mutants formed less extracellular polymeric substances (EPSs). Raman spectroscopy was employed to confirm less biofilm formation by different bacterial mutants and demonstrate a similar chemical composition, but different contents of EPS. Wild type biofilms contained a high level of carotenoids, with the distribution of carotenoids mapped using confocal Raman imaging. We demonstrate that various selective functional genes are responsible for the forming ability of C. sakazakii biofilms, which may have the potential to cause risks to food safety.     

Biofilm formation by Candida dubliniensis

Candida dubliniensis is an opportunistic yeast closely related to Candida albicans that has been recently implicated in oropharyngeal candidiasis in human immunodeficiency virus-infected patients. Most manifestations of candidiasis are associated with biofilm formation, with cells in biofilms displaying properties dramatically different from free-living cells grown under normal laboratory conditions. Here, we report on the development of in vitro models of C. dubliniensis biofilms on the surfaces of biomaterials (polystyrene and acrylic) and on the characteristics associated with biofilm formation by this newly described species. Time course analysis using a formazan salt reduction assay to monitor metabolic activities of cells within the biofilm, together with microscopy studies, revealed that biofilm formation by C. dubliniensis occurred after initial focal adherence, followed by growth, proliferation, and maturation over 24 to 48 h. Serum and saliva preconditioning films enhanced the initial attachment of C. dubliniensis and subsequent biofilm formation. Scanning electron microscopy and confocal scanning laser microscopy were used to further characterize C. dubliniensis biofilms. Mature C. dubliniensis biofilms consisted of a dense network of yeasts cells and hyphal elements embedded within exopolymeric material. C. dubliniensis biofilms displayed spatial heterogeneity and an architecture showing microcolonies with ramifying water channels. Antifungal susceptibility testing demonstrated the increased resistance of sessile C. dubliniensis cells, including the type strain and eight different clinical isolates, against fluconazole and amphotericin B compared to their planktonic counterparts. C. dubliniensis biofilm formation may allow this species to maintain its ecological niche as a commensal and during infection with important clinical repercussions.     

Interspecies variation in Candida biofilm formation studied using the Calgary biofilm device

An in vitro assay to study multiple Candida biofilms, in parallel, has been carried out using the Calgary biofilm device (CBD). We here report: i) standardization of the CBD for Candida albicans biofilm formation, ii) kinetics of C. albicans biofilm formation, iii) biofilm formation by five Candida species, and iv) effect of dietary carbohydrates on biofilm formation. The biofilm metabolic activity on all CBD pegs was similar (p=0.6693) and C. albicans biofilm formation revealed slow growth up to 36 h and significantly higher growth up to 48 h (p<0.001). Significant differences in total biofilm metabolic activity were seen for glucose, fructose and lactose grown C. albicans compared with sucrose and maltose grown yeasts. Candida krusei developed the largest biofilm mass (p<0.05) relative to C. albicans, C. glabrata, C. dubliniensis and C. tropicalis. Scanning electron microscopy revealed that C. krusei produced a thick multilayered biofilm of pseudohyphal forms embedded within the polymer matrix, whereas C. albicans, C. dubliniensis and C. tropicalis biofilms consisted of clusters or chains of cells with sparse extracellular matrix material. We conclude that CBD is a useful, simple, low cost miniature device for parallel study of Candida biofilms and factors modulating this phenomenon.     

Involvement of amyloid proteins in the formation of biofilms in the pathogenic yeast Candida albicans

Candida species represent a major fungal threat for human health. Within the Candida genus, the yeast Candida albicans is the most frequently incriminated species during episodes of candidiasis or candidemia. Biofilm formation is used by C. albicans to produce a microbial community that is important in an infectious context. The cell wall, the most superficial cellular compartment, is of paramount importance regarding the establishment of biofilms. C. albicans cell wall contains proteins with amyloid properties that are necessary for biofilm formation due to their adhesion properties. This review focuses on these amyloid proteins during biofilm formation in the yeast C. albicans.     

Dietary sugars, serum and the biocide chlorhexidine digluconate modify the population and structural dynamics of mixed Candida albicans and Escherichia coli biofilms

Despite the increasing recognition of the role played by mixed species biofilms in health and disease, the behavior and factors modulating these biofilms remain elusive. We therefore compared the effect of serum, two dietary sugars (sucrose and galactose) and a biocide, chlorhexidine digluconate, on a dual species biofilm (DSB) of Candida albicans and Escherichia coli and, their single species biofilm (SSB) counterparts. Both modes of biofilm growth on polystyrene plastic surfaces were quantified using a viable cell count method and visualized using confocal scanning laser microscopy (CSLM). Present data indicate that co-culture of C. albicans with varying initial concentrations of E. coli leads to a significant inhibition of yeast growth (r=-0.964; p<0.001). Parallel ultrastructural studies using CSLM and a Live/Dead stain confirmed that E. coli growth rendered blastospores and hyphal yeasts non-viable in DSB. SSB of C. albicans showed pronounced growth when its growth surface was pretreated with serum and by sugar supplements in the incubating medium (p<0.05). Intriguingly, C. albicans in DSB was more resistant to the antiseptic effect of chlorhexidine digluconate. Taken together, the current data elucidate some features of the colonization resistance offered by bacteria in mixed bacterial/fungal habitats and how such phenomena may contribute to the development of fungal superinfection during antimicrobial therapy.     

The role of extracellular DNA in the establishment,maintenance and perpetuation of bacterial biofilms

Abstract

The significance of extracellular DNA (eDNA) in biofilms was overlooked until researchers added DNAse to a Pseudomonas aeruginosa biofilm and watched the biofilm disappear. Now, a decade later, the widespread importance of eDNA in biofilm formation is undisputed, but detailed knowledge about how it promotes biofilm formation and conveys antimicrobial resistance is only just starting to emerge. In this review, we discuss how eDNA is produced, how it aids bacterial adhesion, secures the structural stability of biofilms and contributes to antimicrobial resistance. The appearance of eDNA in biofilms is no accident: It is produced by active secretion or controlled cell lysis – sometimes linked to competence development. eDNA adsorbs to and extends from the cell surface, promoting adhesion to abiotic surfaces through acid–base interactions. In the biofilm, is it less clear how eDNA interacts with cells and matrix components. A few eDNA-binding biomolecules have been identified, revealing new concepts in biofilm formation. Being anionic, eDNA chelates cations and restricts diffusion of cationic antimicrobials. Furthermore, chelation of Mg²⁺ triggers a genetic response that further increases resistance. The multifaceted role of eDNA makes it an attractive target to sensitize biofilms to conventional antimicrobial treatment or development of new strategies to combat biofilms.     

Candida albicans and Candida dubliniensis: comparison of biofilm formation in terms of biomass and activity

Candida albicans and C. dubliniensis are two species responsible for oral candidiasis, especially in immunocompromised patients. Microbial infection is preceded by adherence and biofilm formation. Biofilm formation represents the most common form of C. albicans in the oral cavity and is considered to be one of the most important virulence factors. In this study, the biofilm formation ability of C. dubliniensis was compared with that of C. albicans in terms of biomass (quantified using crystal violet) and activity (assessed by formazan salts formation). Both species formed heterogeneous biofilms; however, species and strain variations were seen in the quantification of biomass and activity. There was no correlation between pseudohyphae formation and biofilm formation capability.