Significant Role In Biofilm Formation Research Articles

Influence of Fiber Diameter of Polycaprolactone Nanofibrous Materials on Biofilm Formation and Retention of Bacterial Cells.

To develop microbiologically safe nanofibrous materials, it is crucial to understand their interactions with microbial cells. Current research indicates that the morphology of nanofibers, particularly the diameter of the fibers, may play a significant role in biofilm formation and retention. However, it has not yet been determined how the fiber diameter of poly-ε-caprolactone (PCL), one of the most widely used biopolymers, affects these microbial interactions. In this study, two nanofibrous materials electrospun from PCL (PCL45 and PCL80) with different fiber diameter and characteristic distance δ between fibers were compared in terms of their ability to support or inhibit bacterial biofilm formation and retain bacterial cells. Strains of Escherichia coli (ATCC 25922 and ATCC 8739) and Staphylococcus aureus (ATCC 25923 and ATCC 6538) were used as model bacteria. Biofilm formation rate and retention varied significantly between the E.coli and S.aureus strains (p < 0.05) for the tested nanomaterials. In general, PCL showed a lower tendency to be colonized by the tested bacteria compared to the control material (polystyrene). Fiber diameter did not influence the biofilm formation rate of S.aureus strains and E.coli 25922 (p > 0.05), but it did significantly impact the biofilm formation rate of E.coli 8739 and biofilm morphology formed by all of the tested bacterial strains. In PCL45, thick uniform biofilm layers were formed preferably on the surface, while in PCL80 smaller clusters formed preferably inside the structure. Further, fiber diameter significantly influenced the retention of bacterial cells of all the tested strains (p < 0.001). PCL45, with thin fibers (average fiber diameter of 376 nm), retained up to 7 log (CFU mL-1) of staphylococcal cells (100% retention). The overall results indicate PCL45's potential for further research and highlight the nanofibers' morphology influence on bacterial interactions and differences in bacterial strains' behavior in the presence of nanomaterials.

Efficient production of R-2-(4-hydroxyphenoxy) propionic acid by Beauveria bassiana using biofilm-based two-stage fermentation

In this work, a novel biofilm-based fermentation of Beauveria bassiana was employed to convert R-2- phenoxypropionic acid (R-PPA) to R-2-(4-hydroxyphenoxy) propionic acid (R-HPPA). The biofilm culture model of Beauveria bassiana produced a significantly higher R-HPPA titer than the traditional submerged fermentation method. Mannitol dosage, tryptone dosage, and initial pH were the factors that played a significant role in biofilm formation and R-HPPA synthesis. Under the optimal conditions, the maximum R-HPPA titer and productivity approached 22.2 g/L and 3.2 g/(L·d), respectively. A two-stage bioreactor combining agitation and static incubation was developed to further increase R-HPPA production. The process was optimized to achieve 100 % conversion of R-PPA, with a maximum R-HPPA titer of 50 g/L and productivity of 3.8 g/(L·d). This newly developed biofilm-based two-stage fermentation process provides a promising strategy for the industrial production of R-HPPA and related hydroxylated aromatic compounds.

Site-directed mutagenesis of the quorum-sensing transcriptional regulator SinR affects\xa0the biosynthesis of menaquinone in\xa0Bacillus subtilis

BackgroundMenaquinone (MK-7) is a highly valuable vitamin K2 produced by Bacillus subtilis. Common static metabolic engineering approaches for promoting the production of MK-7 have been studied previously. However, these approaches caused an accumulation of toxic substances and reduced product yield. Hence, dynamic regulation by the quorum sensing (QS) system is a promising method for achieving a balance between product synthesis and cell growth.ResultsIn this study, the QS transcriptional regulator SinR, which plays a significant role in biofilm formation and MK production simultaneously, was selected, and its site-directed mutants were constructed. Among these mutants, sinR knock out strain (KO-SinR) increased the biofilm biomass by 2.8-fold compared to the wild-type. SinRquad maximized the yield of MK-7 (102.56 ± 2.84 mg/L). To decipher the mechanism of how this mutant regulates MK-7 synthesis and to find additional potential regulators that enhance MK-7 synthesis, RNA-seq was used to analyze expression changes in the QS system, biofilm formation, and MK-7 synthesis pathway. The results showed that the expressions of tapA, tasA and epsE were up-regulated 9.79-, 0.95-, and 4.42-fold, respectively. Therefore, SinRquad formed more wrinkly and smoother biofilms than BS168. The upregulated expressions of glpF, glpk, and glpD in this biofilm morphology facilitated the flow of glycerol through the biofilm. In addition, NADH dehydrogenases especially sdhA, sdhB, sdhC and glpD, increased 1.01-, 3.93-, 1.87-, and 1.11-fold, respectively. The increased expression levels of NADH dehydrogenases indicated that more electrons were produced for the electron transport system. Electrical hyperpolarization stimulated the synthesis of the electron transport chain components, such as cytochrome c and MK, to ensure the efficiency of electron transfer. Wrinkly and smooth biofilms formed a network of interconnected channels with a low resistance to liquid flow, which was beneficial for the uptake of glycerol, and facilitated the metabolic flux of four modules of the MK-7 synthesis pathway.ConclusionsIn this study, we report for the first time that SinRquad has significant effects on MK-7 synthesis by forming wrinkly and smooth biofilms, upregulating the expression level of most NADH dehydrogenases, and providing higher membrane potential to stimulate the accumulation of the components in the electron transport system.

Effect of honey phenolic extract on biofilm formation by Pediococcus pentosaceus and Lactobacillus fermentum

Today, there is an increasing trend to supplement food with phenolic compounds due to the beneficial effect on health. Lactic acid bacteria (LAB) have been shown to form protective biofilms in food industry. However, surrounding condition, such as additives play a significant role in biofilm formation. The aim of this study was to investigate the influence of honey phenolic extract (HPE) on biofilm formation (BF) of Pediococcus pentosaceus and Lactobacillus fermentum. To this end, optimal BF was tested under different environmental conditions, such as pH and cultivation time. Micro-titer plate assay was performed to assess biofilm-forming abilities. In addition, FT-IR spectroscopy and scanning electron microscopy (SEM) were used to monitor the changes in bacterial membrane composition and biofilm morphology. It was observed that the addition of HPE at 0.094 mg/mL reduced BF of L. fermentum and P. pentosaceus by 28% and 15%, respectively. However, BF of P. pentosaceus remained constant as the concentration of phenolic compounds increased. Micrographs of P. pentosaceus revealed a dense production of extracellular polymeric substances that contributed to the bacterial tolerance against phenolic compounds. Band shifts corresponding to COC ring vibrations, confirmed that exopolysaccharides constitute part of P. pentosaceus biofilm. On the other hand, higher concentrations of HPE were found to disrupt biofilm formation by L. fermentum, resulting in loosely associated clusters.

Detection of biofilm formation and assessment of biofilm genes expression in different Pseudomonas aeruginosa clinical isolates

IntroductionBiofilm formation is an important antibiotic resistance mechanism among different bacterial species. AimThis study aimed to determine the ability of different P. aeruginosa clinical isolates to produce biofilm and its association with las R, lec A and pel A genes expression and antibiotic resistance. MethodsIn this study, 100 isolates of P. aeruginosa were obtained from patients with wound infections at Minia University Hospitals. The antibiotic sensitivity of the isolated strains was done using the disc diffusion method. The isolated bacteria were tested for their ability to form biofilm using a microtiter plate assay. The expression of lasR, lecA and pel A genes in isolated strains was detected using real-time rt-PCR. Results: 27 out of 100 P. aeruginosa isolates (27%) were positive biofilm producers: 14% were strong biofilm producers, 7% were moderate biofilm producers and 6% were weak biofilm producers. Resistance to all used antibiotics and multidrug resistance was higher among biofilm producing than non-biofilm producing strains, but the difference was statistically non-significant. The expression of lasR, lecA and pel A genes were detected in 20 (74.1%), 26(96.3%) and 18(66.7%) isolates out of 27 biofilm-producing strains respectively, compared to 23(31.5%), 26(35.6%) and 24(24%) out of 73 non-biofilm producing strains respectively. ConclusionBiofilm producing strains demonstrated higher expression level of lasR, lecA, and pel A genes with a mean fold change of 357.02 ± 248.5, 26.7 ± 23.2 and 420.2 ± 271.3 respectively, therefore las R, lec A, and pel A genes have a significant role in biofilm formation.

MOLECULAR STUDY OF BIOFILM PRODUCTION BY METHICILLIN RESISTANT STAPHYLOCOCCUS AUREUS

Background:Staphylococci are a group of bacteria that cause diseases ranging from minor skin infections to life-threatening bacteremia. Biofilm formation was determined by a number of methods and is available to detect the capability of staphylococci to colonize the biomedical devices. The icaA and icaD have been reported to play a significant role in biofilm formation. Objective:To achieve and detect the molecular basis of adhesion properties in respect to methicillin resistant Staphylococcus aureus. Methods:Clinical samples were taken from Burn patients; identified and Methicillin susceptibility was tested. The genes icaA and icaD were amplified in methicillin resistant Staphylococcus aureus and the polymerase chain reaction products were sequenced and aligned with the previous recorded sequences online. Results:There was a great correlation between the presence of icaD genes and the slime production. Methicillin resistant Staphylococcus aureus did not reveal any correlation between icaA and icaD and slime layer production; nonetheless, a correlation was noticed between icaD alone and a biofilm production Conclusion:The present findings indicated that methicillin resistant Staphylococcus aureus was able to form biofilm. None of the methicillin resistant Staphylococcus aureus isolates harboured icaA; while 100% of them contained icaD. Keywords:Methicillin resistant Staphylococcus aureus, icaA, icaD gene Citation:Mohamad DA. Molecular study of biofilm production by methicillin resistant Staphylococcus aureus. Iraqi JMS. 2019; 17(3&amp;4): 191-200. doi: 10.22578/IJMS.17.3&amp;4.5

Role of hypothetical protein YicS in the pathogenicity of Avian Pathogenic Escherichia coli in vivo and in vitro

Avian Pathogenic Escherichia coli (APEC) strains belong to the extra-intestinal pathogenic group of E. coli (ExPEC) that causes colibacillosis in poultry. A variety of putative virulence factors of APEC are recognized as potent causes of pathogenicity, the mechanisms underlying their pathogenicity are still not fully understood. The role of yicS in the virulence of pathogenic E. coli is still unclear. Thus, yicS may be related to biofilm formation, which in some bacteria plays a role in pathogenicity. Therefore, the fact that this gene appears to be under positive selection pressure suggests that yicS may be associated with the pathogenicity of APEC. To better understand the role of yicS protein in APEC biological characteristics and pathogenicity, we deleted yicS in an APEC Swollen Head Syndrome strain (APEC strain SCI-07) and studied its effects by comparing wild type and isogenic mutants through comprehensive in vitro and in vivo assays. We demonstrated that yicS plays a role in pathogenicity of APEC. We suggest that the yicS gene, which encodes an exporter protein, has a significant role in biofilm formation, motility, invasion of CEC-32 and Hep-2 cells and APEC pathogenicity in a day-old chick model.

Diversity of Terrestrial Cyanobacteria Colonizing Selected Stone Monuments in Serbia

ABSTRACTThe presence of biofilms on stone monuments can lead to biodeterioration processes that significantly lower not only the aesthetic value of cultural heritage sites but also cause considerable physical and chemical damage to stone surfaces. In terrestrial environments, cyanobacteria have a significant role in biofilm formation on a variety of natural and artificial stone substrata, including granite, sandstone, limestone, lime mortar, etc. Studies on cyanobacterial diversity and estimation of biodeteriorative activity on stone cultural heritage objects can be very important for conservators and restorers, and cyanobacteria removal from stone monuments is currently a great challenge for conservation science. In that sense, the diversity of terrestrial cyanobacteria was investigated at six localities in Serbia. Samples for cyanobacterial analyses were taken from surface biofilms covering different substrata: granite and lime mortar (The Monument to the Unknown Hero), sandstone (Branko's Bridge, Eiffel’s Lock) and carbonate rock (Medieval tombstones from Mramorje and Rastište, Gradac Monastery). A total of 18 cyanobacterial taxa were identified. The highest diversity was found on carbonate rock, followed by sandstone. Cyanobacteria were also recorded on an artificial substratum – lime mortar, while cyanobacteria were not found on granite. All detected cyanobacteria seriously impaired the structural and aesthetic integrity of the studied monuments via mechanical and chemical actions, so their presence must not be overlooked.

Computational insights into promoter architecture of toxin-antitoxin systems of Mycobacterium tuberculosis

Toxin-antitoxin (TA) systems are two component genetic modules widespread in many bacterial genomes, including Mycobacterium tuberculosis (Mtb). The TA systems play a significant role in biofilm formation, antibiotic tolerance and persistence of pathogen inside the host cells. Deciphering regulatory motifs of Mtb TA systems is the first essential step to understand their transcriptional regulation. In this study, in silico approaches, that is, the knowledge based motif discovery and de novo motif discovery were used to identify the regulatory motifs of 79 Mtb TA systems. The knowledge based motif discovery approach was used to design a Perl based bio-tool Mtb-sig-miner available at (https://github.com/zoozeal/Mtb-sig-miner), which could successfully detect sigma (σ) factor specific regulatory motifs in the promoter region of Mtb TA modules. The manual curation of Mtb-sig-miner output hits revealed that the majority of them possessed σB regulatory motif in their promoter region. On the other hand, de novo approach resulted in the identification of a novel conserved motif [(T/A)(G/T)NTA(G/C)(C/A)AT(C/A)] within the promoter region of 14 Mtb TA systems. The identified conserved motif was also validated for its activity as conserved core region of operator sequence of corresponding TA system by molecular docking studies. The strong binding of respective antitoxin/toxin with the identified novel conserved motif reflected the validation of identified motif as the core region of operator sequence of respective TA systems. These findings provide computational insight to understand the transcriptional regulation of Mtb TA systems.

Virulence factor in Candida albicans infections and their clinical relevance

Most of fungal infections are superficial and relatively easy to treat. Nevertheless, millions of invasive fungal infections have also been reported. Considering that most of the time the presence of this fungus represents colonization, the challenge is to differentiate between colonization and infection. The objective is to summarize the Candida spp. virulence factors and to highlight their clinical relevance. We have selected the most pertinent articles from the recent medical literature regarding this subject using Google Academic and Pubmed databases. The first virulence factors involved in the infection are adhesion proteins (e.g., the fungal protein gpm1 interacts with vibronectin, leading to fungal adhesion to keratinocytes, endothelial cells and monocytes). In order to invade, the fungus penetrates the epithelium/endothelium using hydrolytic enzymes - aspartyl proteases (encoded by SAP genes), phospholipases and lipases. Candida albicans could induce its own endocytosis by the epithelial cells, process governed by Efg1p and Tpk2p proteins. Penetration is favored by the yeast-hyphae conversion, pleomorphism being essential in pathogenicity. It is triggered by various factors including pH and temperature changes, hormonal variations etc. The fungus is very versatile, having the capacity to survive within a wide range of environmental conditions. It can cause co-infections with different bacteria such as Pseudomonas aeruginosa, Klebsiella pneumoniae etc. The virulence factor with the most important clinical significance is the capacity of biofilm formation. A significant role in biofilm formation is played by the adhesion proteins codified by ALS and EAP Candida albicans genes. Biofilm presence on medical instruments leads to antifungal resistance. The study of the Candida spp. virulence factors is a continually developing field. Understanding these mechanisms has a major clinical importance when comes to correct evaluation of a positive Candida spp. sample and correct treatment administration. It could also lead to new therapeutic approaches if we consider the increase of antifungal resistance in the last years. We highlight the importance of collaboration between the clinician and the laboratory physician.

Role of OmpA2 surface regions of Porphyromonas gingivalis in host-pathogen interactions with oral epithelial cells.

Outer membrane protein A (OmpA) is a key outer membrane protein found in Gram‐negative bacteria that contributes to several crucial processes in bacterial virulence. In Porphyromonas gingivalis, OmpA is predicted as a heterotrimer of OmpA1 and OmpA2 subunits encoded by adjacent genes. Here we describe the role of OmpA and its individual subunits in the interaction of P. gingivalis with oral cells. Using knockout mutagenesis, we show that OmpA2 plays a significant role in biofilm formation and interaction with human epithelial cells. We used protein structure prediction software to identify extracellular loops of OmpA2, and determined these are involved in interactions with epithelial cells as evidenced by inhibition of adherence and invasion of P. gingivalis by synthetic extracellular loop peptides and the ability of the peptides to mediate interaction of latex beads with human cells. In particular, we observe that OmpA2‐loop 4 plays an important role in the interaction with host cells. These data demonstrate for the first time the important role of P. gingivalis OmpA2 extracellular loops in interaction with epithelial cells, which may help design novel peptide‐based antimicrobial therapies for periodontal disease.

The comER Gene Plays an Important Role in Biofilm Formation and Sporulation in both Bacillus subtilis and Bacillus cereus.

Bacteria adopt alternative cell fates during development. In Bacillus subtilis, the transition from planktonic growth to biofilm formation and sporulation is controlled by a complex regulatory circuit, in which the most important event is activation of Spo0A, a transcription factor and a master regulator for genes involved in both biofilm formation and sporulation. In B. cereus, the regulatory pathway controlling biofilm formation and cell differentiation is much less clear. In this study, we show that a novel gene, comER, plays a significant role in biofilm formation as well as sporulation in both B. subtilis and B. cereus. Mutations in the comER gene result in defects in biofilm formation and a delay in spore formation in the two Bacillus species. Our evidence supports the idea that comER may be part of the regulatory circuit that controls Spo0A activation. comER likely acts upstream of sda, a gene encoding a small checkpoint protein for both sporulation and biofilm formation, by blocking the phosphor-relay and thereby Spo0A activation. In summary, our studies outlined a conserved, positive role for comER, a gene whose function was previously uncharacterized, in the regulation of biofilm formation and sporulation in the two Bacillus species.

Prevalence of mecA, PVL and ica genes in Staphylococcus aureus strains isolated from urinary tract infections patients

The prevalence of Staphylococcus aureus among urinary tract infections (UTIs) patients has been increasing worldwide. The aim of this study was to determine the occurrence of the mecA, PVL, ica genes in a collection of MRSA urinary isolates by PCR. Methicillin resistance S. aureus (MRSA) is considered to have emerged from S. aureus through the acquisition of staphylococcal cassette chromosome (SCCmec), which carries the mecA gene for methicillin resistance. Panton-Valentine leukocidin (PVL)-producing strains of MRSA appear to be associated with increased risk of transmission, complications and hospitalization. IcaA and icaD genes have been reported to play a significant role in biofilm formation in S. aureus. Out of 50 isolates of S. aureus from UTI patients, 36 (72%) were found to be MRSA by oxacillin screen agar. All these MRSA strains were found to be positive for mecA genes, 9 (25%) were found to be positive for PVL and 23 (64%) were positive for both icaA and icaD genes. MRSA isolated from UTI patients show the presence of mecA, PVL, ica genes, which may have consequences for the treatment of UTIs especially in catheter-associated and nosocomial infections. Key words: Urinary tract infections (UTIs), mecA gene, PVL gene, ica gene, MRSA.

Structure/Function studies on Glucan Binding Protein C of Streptococcus mutans

Streptococcus mutans is a known etiological agent in dental caries. In the past several years, we have taken a concerted effort toward understanding the adhesion mechanisms adopted by the surface proteins S. mutans. The Glucan Binding Protein C (GBPC) is a LPXTG anchored surface protein on S. mutans that has been widely implicated to play a significant role in biofilm formation. GBPC displays limited homology to the V-region of Antigen I/II (AgI/II), another surface protein of S. mutans (1,2). We undertook to crystallize and resolve the structure of GBPC. Recombinant GbpC111-552 (residues encompassing 111-552) was cloned into a pET23d vector, and thereafter expressed in E.coli BL21(DE3). GbpC111-552 purified initially using affinity chromatography (his-tag), followed by anion exchange (MonoQ) and finally polished with size exclusion (Superdex 75). GbpC111-552 was crystallized using the vapor diffusion method by scanning various commercial crystallization kits on a 96 well plate format through the Art Robbins Gryphon robot. Large crystals were obtained from a refined droplet condition that contained 1 μl of protein (43.3 mg/ml) mixed with 1 μl of reservoir made of 100 mM Bis-Tris pH 6.5 and 25% (w/v) PEG3350. The crystal structure could not be resolved by molecular replacement. Therefore crystals were soaked in Sodium Iodide (NaI) and thereafter flash frozen with 9% glycerol as crytoprotectant. MAD data sets were collected and the crystal structure was resolved. We will present the crystal structure, and how based on its structural homology we discovered similar functionalities among the surface proteins of S. mutans. The structural and functional studies of these surface proteins we hope would allow the identification of the adherence motifs, which would aid in development of inhibitors. The development of such anti-adhesive inhibitors would aid in disease preventative therapies.

Beta-hexosaminidase activity of the oral pathogen Tannerella forsythia influences biofilm formation on glycoprotein substrates.

Tannerella forsythia is an important pathogen in periodontal disease. Previously, we showed that its sialidase activity is key to utilization of sialic acid from a range of human glycoproteins for biofilm growth and initial adhesion. Removal of terminal sialic acid residues often exposes β-linked glucosamine or galactosamine, which may also be important adhesive molecules. In turn, these residues are often removed by a group of enzymes known as β-hexosaminidases. We show here that T. forsythia has the ability to cleave glucosamine and galactosamine from model substrates and that this activity can be inhibited by the hexosaminidase inhibitor PugNAc (O-(2-acetamido-2-deoxy-d-glucopyranosylidene)amino N-phenyl carbamate). We now demonstrate for the first time that β-hexosaminidase activity plays a role in biofilm growth on glycoprotein-coated surfaces because biofilm growth and initial cell adhesion are inhibited by PugNAc. In contrast, adhesion to siallo-glycoprotein-coated surfaces is unaltered by PugNAc in the absence of sialidase activity (using a sialidase-deficient mutant) or surprisingly on the clinically relevant substrates saliva or serum. These data indicate that β-hexosaminidase activity has a significant role in biofilm formation in combination with sialidase activity in the biofilm lifestyle of T. forsythia.

Virulence Regulator PrfA is Essential for Biofilm Formation in Listeria monocytogenes but not in Listeria innocua

The ability of the foodborne pathogen Listeria monocytogenes to develop biofilm in food-processing environment is a major concern for the food safety, because biofilms allow bacteria to better resist environmental stresses. PrfA is a key transcriptional activator that positively regulates most of the known listerial virulence gene expression. In order to explore the role of PrfA on Listeria biofilm development, we compared the abilities of biofilm formation by L. monocytogenes wild type strains (EGD and EGDe) and their prfA deletion mutants (EGD∆prfA and EGDe∆prfA), nonpathogenic Listeria innocua, as well as the recombinant strains that express constitutively active mutant PrfA (PrfA*) in L. innocua (LI-pERL3-prfA*) and in EGDe∆prfA (EGDe∆prfA-pERL3-prfA*) at 37°C in brain heart infusion (BHI) medium using the polyvinyl chloride (PVC) microtiter plate assay and microscopic examination. Our results showed that the wild types of L. monocytogenes had strong abilities to develop biofilm with meshwork of bacterial aggregates, while biofilm with sparse small clumps were observed in L. innocua. The biofilm production of strains EGD∆prfA and EGDe∆prfA that lack funtional PrfA was reduced and could be recovered by the introduction of the PrfA*, however, the PrfA* had no impact on the biofilm forming ability of L. innocua. Our results suggest that PrfA plays a significant role in biofilm formation in L. monocytogenes but not in L. innocua, thus may reflect differences in the molecular mechanisms of biofilm formation by these two closely related species.

Biofilm on dental implants: a review of the literature.

The aim of this article was to review the current literature with regard to biofilm formation on dental implants and the influence of surface characteristics (chemistry, surface free energy, and roughness) of dental implant and abutment materials and their design features on biofilm formation and its sequelae. An electronic MEDLINE literature search was conducted of studies published between 1966 and June 2007. The following search terms were used: biofilm and dental implants, biofilm formation/plaque bacterial adhesion and implants, plaque/biofilm and surface characteristics/roughness/surface free energy of titanium dental implants, implant-abutment interface and plaque/biofilm, biofilm and supragingival/subgingival plaque microbiology, biofilm/plaque and implant infection, antibacterial/bacteriostatic titanium, titanium nanocoating/nanopatterning, antimicrobial drug/titanium implant. Both in vitro and in vivo studies were included in this review. Fifty-three articles were identified in this review process. The articles were categorized with respect to their context on biofilm formation on teeth and dental implant surfaces and with regard to the influence of surface characteristics of implant biomaterials (especially titanium) and design features of implant and abutment components on biofilm formation. The current state of literature is more descriptive, rather than providing strong data that could be analyzed through meta-analysis. Basic research articles on surface modification of titanium were also included in the review to analyze the applications of such studies on the fabrication of implant surfaces that could possibly decrease early bacterial colonization and biofilm formation. Increase in surface roughness and surface free energy facilitates biofilm formation on dental implant and abutment surfaces, although this conclusion is derived from largely descriptive literature. Surface chemistry and the design features of the implant-abutment configuration also play a significant role in biofilm formation.

Biofilm formation by Pseudomonas aeruginosa: role of the C4-HSL cell-to-cell signal and inhibition by azithromycin.

In Pseudomonas aeruginosa, biofilm formation is controlled by a cell-to-cell signalling circuit relying on the secretion of 3-oxo-C12-HSL and C4-HSL. Previous studies suggested that C4-HSL plays no significant role in biofilm formation. However the wild-type PAO1 strain PAO-BI, used as a control in these studies is itself impaired in the production of C4-HSL. We wondered therefore whether the role of C4-HSL in biofilm formation might have been underestimated, and whether azithromycin inhibits biofilm formation by interfering with cell-to-cell signalling. We used isogenic mutants of wild-type PAO1 strains PAO-BI and PT5 in a static biofilm model. Biofilm formation was quantified using Crystal Violet staining and exopolysaccharide measurements. Wild-type strain PAO-BI, as a result of its reduced C4-HSL secretion, produced 40% less biofilm compared with the wild-type PAO1 strain PT5. Using isogenic mutants of strain PT5 we have shown that whereas a lasI mutant (deficient in 3-oxo-C12-HSL) produced similar amounts of biofilm to the wild-type, a rhlI mutant (deficient in C4-HSL) produced 70% less biofilm. In the latter strain, biofilm formation could be restored by addition of exogenous C4-HSL. Azithromycin, known to reduce the production of both 3-oxo-C12-HSL and C4-HSL, inhibited biofilm formation of wild-type PT5 by 45%. This inhibition could be reversed by the addition of both cell-to-cell signals. Our results indicate that C4-HSL also plays a significant role in biofilm formation. Furthermore, we demonstrate the potential of using cell-to-cell signalling blocking agents such as azithromycin to interfere with biofilm formation.