Good Biofilm Formation Research Articles

Prevalence, virulence and antifungal activity of C. albicans isolated from infected root canals

BackgroundThere is limited data on the prevalence of Candida species in infected root canal systems of human teeth. We attempted to investigate the prevalence, genotype, virulence and the antifungal susceptibility of Candida albicans isolated from infected root canals of patients with primary and post-treatment infections in a UAE population.MethodsMicrobiological samples from 71 subjects with infected root canals were aseptically collected, and cultured on Sabouraud dextrose agar, and C. albicans was identified using multiplex polymerase chain reaction, and the isolates were further subtyped using ABC genotyping system. Their relative virulence was compared using further four archival samples of endodontic origin from another geographical region, and four more salivary isolates, as controls. The virulence attributes compared were biofilm formation, and production of phospholipase and haemolysin, and the susceptibility to nystatin, amphotericin B, ketoconazole, and fluoconazole was also tested.Results4 out of 71 samples (5.6%) yielded Candida species. On analysis of variance among the groups, the intracanal isolates, mainly Genotype A, possessed a high degree of phospholipase and haemolysin activity (p < 0.05). The UAE and Finland isolates were stronger biofilm formers and had superior phospholipase production capacity compared with the salivary isolates. All isolates were sensitive to the antifungal chemicals used. The salivary isolates were more sensitive to fluoconazole compared to other groups (p < 0.05).ConclusionThe prevalence of C. albicans in infected root canals of patients attending a dental hospital in UAE is low. The strains isolated were good biofilm formers, possessed strong phospholipase and haemolysin activity and were mostly of the genotype A. The fact that the root canal isolates possessed significant hydrolase activity, imply that they are equipped with virulence attributes necessary for survival within a harsh intracanal ecosystem.

Identification and Morphological Characterization of Biofilms Formed by Strains Causing Infection in Orthopedic Implants.

Objectives: For a better understanding of the mechanisms involved in biofilm formation, we performed a broad identification and characterization of the strains affecting implants by evaluating the morphology of biofilms formed in vitro in correlation with tests of the strains’ antibiotic susceptibility in planktonic form. The ability of the strains to form biofilms in vitro was evaluated by means of colony forming units counting, metabolic activity tests of biofilm cells, and scanning electron microscopy. Methods: A total of 140 strains were isolated from patients with orthopedic implant-related infections during the period of 2015 to 2018. The identification of the isolates was carried out through microbiological cultures and confirmed by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. Antibiotic susceptibility rates of the isolates were accessed according to EUCAST (European Committee on Antimicrobial Susceptibility Testing). The ability of all isolates to form biofilms in vitro was evaluated by counting the colony forming units, by measuring the metabolic activity of biofilm cells, and by analyzing the morphology of the formed biofilms using scanning electron microscopy. Results: From all the isolates, 41.84% (62 strains) were Staphylococcus epidermidis and 15.60% (22 strains) were Staphylococcus aureus. A significant difference in the capacity of biofilm formation was observed among the isolates. When correlating the biofilm forming capacity of the isolates to their antibiotic susceptibility rates, we observed that not all strains that were classified as resistant were biofilm producers in vitro. In other words, bacteria that are not good biofilm formers can show increased tolerance to multiple antibiotic substances. Conclusion: From 2015 until 2018, Staphylococcus epidermidis was the strain that caused most of the orthopedic implant-related infections in our hospital. Not all strains causing infection in orthopedic implants are able to form biofilms under in vitro conditions. Differences were observed in the number of cells and morphology of the biofilms. In addition, antibiotic resistance is not directly related to the capacity of the strains to form biofilms in vitro. Further studies should consider the use of in vitro culture conditions that better reproduce the joint environment and the growth of biofilms in humans.

Efficacy of Commercial Sanitizers Used in Food Processing Facilities for Inactivation of Listeria Monocytogenes, E. Coli O157:H7, and Salmonella Biofilms.

Bacteria entrapped in biofilms are a source of recurring problems in food processing environments. We recently developed a robust, 7-day biofilm microplate protocol for creating biofilms with strongly adherent strains of Listeria monocytogenes, Escherichia coli O157:H7, and Salmonella serovars that could be used to examine the effectiveness of various commercial sanitizers. Listeria monocytogenes 99-38, E.coli O157:H7 F4546, and Salmonella Montevideo FSIS 051 were determined from prior studies to be good biofilm formers and could be recovered and enumerated from biofilms following treatment with trypsin. Extended biofilms were generated by cycles of growth and washing daily, for 7 days, to remove planktonic cells. We examined five different sanitizers (three used at two different concentrations) for efficacy against the three pathogenic biofilms. Quaternary ammonium chloride (QAC) and chlorine-based sanitizers were the least effective, showing partial inhibition of the various biofilms within 2 h (1–2 log reduction). The best performing sanitizer across all three pathogens was a combination of modified QAC, hydrogen peroxide, and diacetin which resulted in ~6–7 log reduction, reaching levels below our limit of detection (LOD) within 1–2.5 min. All treatments were performed in triplicate replication and analyzed by one way repeated measures analysis of variance (RM-ANOVA) to determine significant differences (p < 0.05) in the response to sanitizer treatment over time. Analysis of 7-day biofilms by scanning electron microscopy (SEM) suggests the involvement of extracellular polysaccharides with Salmonella and E. coli, which may make their biofilms more impervious to sanitizers than L. monocytogenes.

Priority of Early Colonizers but No Effect on Cohabitants in a Synergistic Biofilm Community.

The arrival order of different species to a habitat can strongly impact community assembly and succession dynamics, thus influencing functionality. In this study, we asked how prior colonization of one community member would influence the assembly of a synergistic multispecies biofilm community grown in vitro. We expected that the prior arrival would confer an advantage, in particular for good biofilm formers. Yet, we did not know if the cohabitants would be impaired or benefit from the pre-colonization of one member, depending on its ability to form biofilm. We used a consortium consisting of four soil bacteria; Stenotrophomonas rhizophila, Xanthomonas retroflexus, Microbacterium oxydans and Paenibacillus amylolyticus. This consortium has been shown to act synergistically when grown together, thus increasing biofilm production. The results showed that the two good biofilm formers gained a fitness advantage (increase in abundance) when allowed prior colonization on an abiotic surface before the arrival of their cohabitants. Interestingly, the significantly higher number of the pre-colonized biofilm formers did not affect the resulting composition in the subsequent biofilm after 24 h.

Prototype Development of the Intelligent Hydrogel Wound Dressing and Its Efficacy in the Detection of Model Pathogenic Wound Biofilms.

The early detection of wound infection in situ can dramatically improve patient care pathways and clinical outcomes. There is increasing evidence that within an infected wound the main bacterial mode of living is a biofilm: a confluent community of adherent bacteria encased in an extracellular polymeric matrix. Here we have reported the development of a prototype wound dressing, which switches on a fluorescent color when in contact with pathogenic wound biofilms. The dressing is made of a hydrated agarose film in which the fluorescent dye containing vesicles were mixed with agarose and dispersed within the hydrogel matrix. The static and dynamic models of wound biofilms, from clinical strains of Escherichia coli, Pseudomonas aeruginosa, Staphylococcus aureus, and Enterococcus faecalis, were established on nanoporous polycarbonate membrane for 24, 48, and 72 h, and the dressing response to the biofilms on the prototype dressing evaluated. The dressing indicated a clear fluorescent/color response within 4 h, only observed when in contact with biofilms produced by a pathogenic strain. The sensitivity of the dressing to biofilms was dependent on the species and strain types of the bacterial pathogens involved, but a relatively higher response was observed in strains considered good biofilm formers. There was a clear difference in the levels of dressing response, when dressings were tested on bacteria grown in biofilm or in planktonic cultures, suggesting that the level of expression of virulence factors is different depending of the growth mode. Colorimetric detection on wound biofilms of prevalent pathogens (S. aureus, P. aeruginosa, and E. faecalis) is also demonstrated using an ex vivo porcine skin model of burn wound infection.

Emerging, Non-PCV13 Serotypes 11A and 35B of Streptococcus pneumoniae Show High Potential for Biofilm Formation In Vitro.

BackgroundSince the use of pneumococcal conjugate vaccines PCV7 and PCV13 in children became widespread, invasive pneumococcal disease (IPD) has dramatically decreased. Nevertheless, there has been a rise in incidence of Streptococcus pneumoniae non-vaccine serotypes (NVT) colonising the human nasopharynx. Nasopharyngeal colonisation, an essential step in the development of S. pneumoniae-induced IPD, is associated with biofilm formation. Although the capsule is the main pneumococcal virulence factor, the formation of pneumococcal biofilms might, in fact, be limited by the presence of capsular polysaccharide (CPS).Methodology/Principal FindingsWe used clinical isolates of 16 emerging, non-PCV13 serotypes as well as isogenic transformants of the same serotypes. The biofilm formation capacity of isogenic transformants expressing CPSs from NVT was evaluated in vitro to ascertain whether this trait can be used to predict the emergence of NVT. Fourteen out of 16 NVT analysed were not good biofilm formers, presumably because of the presence of CPS. In contrast, serotypes 11A and 35B formed ≥45% of the biofilm produced by the non-encapsulated M11 strain.Conclusions/SignificanceThis study suggest that emerging, NVT serotypes 11A and 35B deserve a close surveillance.

Characterization of pre-antibiotic era Klebsiella pneumoniae isolates with respect to antibiotic/disinfectant susceptibility and virulence in Galleria mellonella.

The EGD Murray collection consists of approximately 500 clinical bacterial isolates, mainly Enterobacteriaceae, isolated from around the world between 1917 and 1949. A number of these "Murray" isolates have subsequently been identified as Klebsiella pneumoniae. Antimicrobial susceptibility testing of these isolates showed that over 30% were resistant to penicillins due to the presence of diverse blaSHV β-lactamase genes. Analysis of susceptibility to skin antiseptics and triclosan showed that while the Murray isolates displayed a range of MIC/minimal bactericidal concentration (MBC) values, the mean MIC value was lower than that for more modern K. pneumoniae isolates tested. All Murray isolates contained the cation efflux gene cepA, which is involved in disinfectant resistance, but those that were more susceptible to chlorhexidine were found to have a 9- or 18-bp insertion in this gene. Susceptibility to other disinfectants, e.g., H2O2, in the Murray isolates was comparable to that in modern K. pneumoniae isolates. The Murray isolates were also less virulent in Galleria and had a different complement of putative virulence factors than the modern isolates, with the exception of an isolate related to the modern lineage CC23. More of the modern isolates (41% compared to 8%) are classified as good/very good biofilm formers, but there was overlap in the two populations. This study demonstrated that a significant proportion of the Murray Klebsiella isolates were resistant to penicillins before their routine use. This collection of pre-antibiotic era isolates may provide significant insights into adaptation in K. pneumoniae in relation to biocide susceptibility.

Culturable bacterial diversity from a feed water of a reverse osmosis system, evaluation of biofilm formation and biocontrol using phages.

Biofilm formation on reverse osmosis (RO) systems represents a drawback in the application of this technology by different industries, including oil refineries. In RO systems the feed water maybe a source of microbial contamination and thus contributes for the formation of biofilm and consequent biofouling. In this study the planktonic culturable bacterial community was characterized from a feed water of a RO system and their capacities were evaluated to form biofilm in vitro. Bacterial motility and biofilm control were also analysed using phages. As results, diverse Protobacteria, Actinobacteria and Bacteroidetes were identified. Alphaproteobacteria was the predominant group and Brevundimonas, Pseudomonas and Mycobacterium the most abundant genera. Among the 30 isolates, 11 showed at least one type of motility and 11 were classified as good biofilm formers. Additionally, the influence of non-specific bacteriophage in the bacterial biofilms formed in vitro was investigated by action of phages enzymes or phage infection. The vB_AspP-UFV1 (Podoviridae) interfered in biofilm formation of most tested bacteria and may represent a good alternative in biofilm control. These findings provide important information about the bacterial community from the feed water of a RO system that may be used for the development of strategies for biofilm prevention and control in such systems.

Characterization of biofilms in different clinical M serotypes of Streptococcus pyogenes

Streptococcus pyogenes is a notorious human pathogen responsible for a wide array of infections. The ability of S. pyogenes to form biofilms is an innate property during the pathogenesis of invasive infections. From the eleven M serotypes tested: M56, M74, M100, M65, M89 and st38 formed dense biofilms in 48 h. The present study is the first of its kind to report about the biofilm formation in the serotypes M56, M65 M74 M100 and st38. XTT reduction assay of the biofilms showed decreased metabolic activity with increase in incubation time. The surface architecture of the biofilms when observed by scanning electron microscopy (SEM) revealed the microcolony formation. Confocal laser scanning microscopy (CLSM) was used to compare the surface topography and thickness of biofilms between the biofilm formers with and without the addition of glucose. Interestingly a non-biofilm former (st2147) was induced to form biofilms with the addition of glucose. On correlating the drug (erythromycin) resistance of the various M serotypes with their biofilm forming ability we noticed that erythromycin sensitive strains were found to be good biofilm formers. We also noticed that biofilm formation in S. pyogenes is independent of sil gene.

Effect of oxygen and growth medium onin vitrobiofilm formation byEscherichia coli

ABSTRACTThe effects of oxygen availability onin vitrobiofilm formation by anEscherichia coliK-12 strain and 13 clinicalE. colistrains were compared. AllE. colistrains were capable of forming monospecies biofilm on polystyrene in aerobic media. The K-12 strain produced biofilm in both aerobic glucose minimal medium (ABTG), and aerobic trypticase soy broth (TSB) whereas the majority of the clinical strains produced significant biofilm only in aerobic TSB (9 of 13). In anaerobic media,E. coliK-12 and 9 of the 13 clinical strains were capable of forming biofilmin vitro. Only three clinical strains formed biofilm in anaerobic TSB whereas six clinical strains produced detectable biofilm in anaerobic ABTG. None of the strains tested were capable of forming biofilm in both anaerobic ABTG and anaerobic TSB. Strains that were good biofilm formers in aerobic ABTG also produced the highest amount of biofilm in anaerobic ABTG (R2= 0.90). Image analysis revealed notable differences in architecture for biofilms grown in the presence and in the absence of oxygen. In aerobic ABTG, the biofilm was dominated by tall, mushroom-shaped microcolonies with pores and channels whereas biofilm in anaerobic ABTG was thinner and less heterogeneous, resulting in reduced maximum thickness and biovolume. Analysis of phospholipid fatty acid (PLFA) profiles fromE. coliK-12 and three clinical strains did not reveal a specific pattern associated with the biofilm phenotypes. Interestingly, the clinicalE. colistrains adjusted their PLFA composition much more than didE. coliK-12 in response to changes in growth regimens. Collectively, the results indicate that oxygen availability may affectE. colibiofilm formation in minimal and complex media. The results confirm thatE. coliK-12 and some clinicalE. colistrains are capable of formingin vitrobiofilm under anaerobic conditions. However, the data also suggest that this attribute is highly strain dependent and may vary significantly among clinical isolates.