Sessile Phases Research Articles

Comparative Evaluation on the effect of Herbal mouthwash on Putative Periodontal Pathogens – In vitro study

Introduction: The architecture of dental plaque biofilms provides a resistant environment for bacteria. Chemical plaque control agents containing antimicrobials (chlorhexidine, herbal mouthwash) are adjunctively used to halt the growth of dental plaque. In order to overcome the adverse effects that result due to the extensive use of synthetic formulation, several herbal preparations have been tested for their effectiveness. In the current study, the effects of chlorhexidine (CHX) and herbal mouthwash in vitro on the planktonic and sessile phases of selected strains of putative periodontal pathogens are compared. Methods: The standard strains of Aggregatibacter actinomycetemcomitans, Porphyromonas gingivalis, and Fusobacterium nucleatum were used for assessing the antimicrobial properties of the two mouthwashes. The methods of broth dilution and agar diffusion were used for assessing the minimum inhibitory concentration (MIC) in the planktonic phase. The antimicrobial effects in the sessile phase were determined by crystal violet microtiter assay. Results: The broth dilution method showed no statistical significant difference in MIC between the two mouthwashes. The agar diffusion method showed a statistically significant difference between the two types of mouthwashes against the F. nucleatum. The crystal violet microtiter assay suggested that the herbal mouthwash exhibited an antimicrobial effect even after 24 hrs on a mixed biofilm. Conclusion: Antimicrobial action exerted by the herbal mouthwash after 24 hrs was higher than that of CHX, against a biofilm constituting the three putative periodontal pathogens.

Investigating natural antibiofilm components: a new therapeutic perspective against candidal vulvovaginitis

The rampant emergence of Candida albicans in the vagina and its ability to thrive as a biofilm has outstood the prevalence of candidal vulvovaginitis (CVV), a gender-based fungal infection approximately affecting 75% of the global female population. The biofilm represents a multidimensional microbial population, which often dictates prominent caveats of CVV such as increased fungal virulence, drug resistance and infection relapse/recurrence. Additionally, the conjugated issues of the ineffectiveness of conventional antifungals (azoles), prolonged treatment durations, compromised patient compliance, economic and social burden, exacerbates CVV complications as well. Henceforth, the current hypothesis narrates an investigational proposal for exploration and combination of naturally derived antibiofilm components with luliconazole (imidazole antifungal agent) as a new therapeutic paradigm against CVV. The purported hypothesis unravels a synergistic approach for fabricating Nanostructured Lipid Carriers, NLCs loaded transvaginal gel with dual APIs of natural (antibiofilm) as well as the synthetic (antifungal) origin to target high therapeutic efficacy, delivery, retention, controlled release and bioadhesion in a vaginal milieu. The multipronged effect of antibiofilm and antifungal agents will expectably enhance drug susceptibility thus, maintaining Minimum Inhibitory Concentration (MIC) against cells of C. albicans and targeting its biofilm in planktonic, adherent, and sessile phases. The effective disruption of a biofilm could further lower infection resistance and recurrence as well. In conclusion, the purported hypothesis could speed up the emergence of novel drug combinations and accelerates new product development with solid, synergistic, and complementary activities against C. albicans and its biofilm, making it amenable for generating pre-clinical and clinical results therebycreating a suitableroadmap for commercialization.

Changes of microbial diversity during pyrite bioleaching

Microorganisms, one of the key factors affecting the bioleaching process, change the components of extracellular polymeric substance (EPS) and community structure to survive in leaching environments. In this work, Fourier transform infrared (FTIR), X-ray powder diffraction (XRD) and 16S rDna high-throughput sequence analyses were used to reveal the microbial changes in planktonic and sessile phases during bioleaching. The results showed the occupation of sessile cells decreased from 66.2% to (10±3)%. After bioleaching, the planktonic and sessile cells have similar EPS, but they are different from the original cells. Pyrite dissolution mainly occurs at the early and late stages with the decreasing of particle diameter, by 50% and 40%, respectively. The 16S rDna gene based sequence analysis results in total of 1117420 Reads across the six samples, presented among 7 phyla, 9 classes, 17 orders, 23 families and 31 genera. Genera Leptospirillum and Sulfobacillus are the main bacteria at the early and middle stages, and Leptospirillum is the main genus at the end of bioleaching. Aquabacterium and Acidovorax are special genera in sessile cells and Weissella is special in planktonic ones.

Community Development between Porphyromonas gingivalis and Candida albicans Mediated by InlJ and Als3.

ABSTRACTThe pleiomorphic yeast Candida albicans is a significant pathogen in immunocompromised individuals. In the oral cavity, C. albicans is an inhabitant of polymicrobial communities, and interspecies interactions promote hyphal formation and biofilm formation. C. albicans colonizes the subgingival area, and the frequency of colonization increases in periodontal disease. In this study, we investigated the interactions between C. albicans and the periodontal pathogen Porphyromonas gingivalis. C. albicans and P. gingivalis were found to coadhere in both the planktonic and sessile phases. Loss of the internalin-family protein InlJ abrogated adhesion of P. gingivalis to C. albicans, and recombinant InlJ protein competitively inhibited interspecies binding. A mutant of C. albicans deficient in expression of major hyphal protein Als3 showed diminished binding to P. gingivalis, and InlJ interacted with Als3 heterologously expressed in Saccharomyces cerevisiae. Transcriptional profiling by RNA sequencing (RNA-Seq) established that 57 genes were uniquely upregulated in an InlJ-dependent manner in P. gingivalis-C. albicans communities, with overrepresentation of those corresponding to 31 gene ontology terms, including those associated with growth and division. Of potential relevance to the disease process, C. albicans induced upregulation of components of the type IX secretion apparatus. Collectively, these findings indicate that InlJ-Als3-dependent binding facilitates interdomain community development between C. albicans and P. gingivalis and that P. gingivalis has the potential for increased virulence within such communities.

Comprehensive Evaluation of Biological Growth Control by Chlorine-Based Biocides in Power Plant Cooling Systems Using Tertiary Effluent

Recent studies have shown that treated municipal wastewater can be a reliable cooling water alternative to fresh water. However, elevated nutrient concentration and microbial population in wastewater lead to aggressive biological proliferation in the cooling system. Three chlorine-based biocides were evaluated for the control of biological growth in cooling systems using tertiary treated wastewater as makeup, based on their biocidal efficiency and cost-effectiveness. Optimal chemical regimens for achieving successful biological growth control were elucidated based on batch-, bench-, and pilot-scale experiments. Biocide usage and biological activity in planktonic and sessile phases were carefully monitored to understand biological growth potential and biocidal efficiency of the three disinfectants in this particular environment. Water parameters, such as temperature, cycles of concentration, and ammonia concentration in recirculating water, critically affected the biocide performance in recirculating cooling systems. Bench-scale recirculating tests were shown to adequately predict the biocide residual required for a pilot-scale cooling system. Optimal residuals needed for proper biological growth control were 1, 2-3, and 0.5-1 mg/L as Cl2 for NaOCl, preformed NH2Cl, and ClO2, respectively. Pilot-scale tests also revealed that Legionella pneumophila was absent from these cooling systems when using the disinfectants evaluated in this study. Cost analysis showed that NaOCl is the most cost-effective for controlling biological growth in power plant recirculating cooling systems using tertiary-treated wastewater as makeup.

Ecological behaviour of three serogroups of Legionella pneumophila within a model plumbing system

Three Legionella pneumophila strains isolated from water samples and belonging to serogroups (sgs) 1, 6 and 9 were analysed for their capacity to colonise an experimental model simulating a domestic hot water distribution system. Ecological factors that could influence the persistence of the sgs such as intracellular life within protozoan hosts and bacterial interference by the production of antagonistic compounds were also studied. Viable counts of L. pneumophila increased both in the planktonic and in the sessile phases. Sg 6 showed a marked prevalence during the whole experiment and exhibited the highest host infection efficiency. Sg 1 was significantly less represented, but showed the highest capacity to reproduce in the protozoan hosts. Sg 9 was poorly represented and less adapted to intracellular life. Among the 14 bacteria constantly isolated in the system, five (35.7%) produced antagonistic substances against Legionella, with differences according to the bacterial strain and L. pneumophila sgs.

Using microbial genomics to evaluate the effectiveness of silver to prevent biofilm formation

Increasingly, monovalent silver cations are being considered as an alternative biocide in water distribution systems. The objective of this study was to understand how bacteria respond when challenged with silver. The approach used included cultivation of Escherichia coli in planktonic and sessile phases, exposing biomass to a solution of silver nitrate and evaluating genetic responses using commercial Affymetrix microarrays. Experimental results showed that lower pH enhanced silver toxicity in a dose-dependent manner. Sessile biomass demonstrated resistance to silver, and the microarray results indicated that the genetic mechanism for silver resistance was similar to the mechanism for copper resistance including upregulation of efflux pumps as well as upregulation of metal oxidoreductases. The gene, copA, a P-type ATPase efflux flux, was upregulated in response to silver exposure, and the gene of CusCFBA, a Cu(I) efflux pump, was also upregulated. The gene of CueO, a robust cuprous oxidase, was also upregulated and may have reduced silver toxicity through oxidation of silver ions. This study is significant because it provides baseline data to understand the genetic response of bacteria to silver biocides.

Effects of Fresh Garlic Extract on Candida albicans Biofilms

The effects of fresh garlic extract (FGE) against planktonic Candida spp. have been demonstrated in vitro and have been attributed to the action of allicin (3), a sulfur-containing compound that is formed at levels of approximately 3 to 5 mg/g of fresh clove (8). Biofilm-associated, or sessile, Candida albicans organisms demonstrate increased resistance to traditional antifungal agents that have activity against their planktonic counterparts (5). Therefore, we investigated the activity of FGE against both planktonic and sessile forms of C. albicans. (This data was presented in part at the 44th Interscience Conference on Antimicrobial Agents and Chemotherapy, Washington, D.C., 30 October to 2 November 2004.) FGE was prepared as previously described by Lemar and colleagues (7). Briefly, fresh garlic cloves were crushed in sterile saline at a concentration of 40 mg/ml. FGE was filter sterilized and stored at −70°C until use. We utilized C. albicans GDH 2346, a strain isolated from a denture stomatitis patient that is known to produce an extracellular matrix containing a mannoprotein adhesin, with a previously described method of C. albicans biofilm formation (2, 4). Sterile silicone elastomer disks were immersed in 1 × 107 CFU of C. albicans, incubated at 37°C on an orbital shaker for 90 min, and then rinsed. One-third of the disks were placed in 0, 0.5, or 1.0 mg of FGE/ml in yeast nitrogen base and incubated at 37°C for 48 h prior to evaluation. The other disks were incubated in yeast nitrogen base for 48 h to form mature biofilms. The untreated, mature biofilms were incubated with 0, 2, or 4 mg of FGE/ml in saline for 1 or 48 h prior to evaluation. All biofilms were evaluated with XTT [2,3-bis(2-methoxy-4-nitro-5-sulfophenyl)-2H-tetrazolium-5-carboxanilide] reduction assay, a semiquantitative, colorimetric assay of metabolic activity (2). MICs for planktonic organisms were measured using the NCCLS macrodilution method. All experiments were performed in triplicate on three different days. Comparisons between groups were performed by analysis of variance (ANOVA). P values of <0.05 were considered significant. The mean absorbances at 492 nm for disks treated immediately after adherence, mature biofilms treated for 1 h, and mature biofilms treated for 48 h are presented in Table ​Table1.1. MICs for planktonic yeast were 0.0625 to 0.125 mg/ml. TABLE 1. XTT assay results for adherent and biofilm-associated C. albicans treated with FGE These data demonstrate activity of FGE against C. albicans in its planktonic, adherent, and sessile phases. The in vitro activity decreases as the biofilm phenotype develops, as noted previously with traditional antifungal drugs (1). The superior activity at 1 versus 48 h of treatment probably relates to the half-life of FGE at 37°C and would be an important consideration in the development of in vivo uses (6). Although only one strain of C. albicans was used, the same MIC range and metabolic assay results were demonstrated with a clinical candidemia isolate (data not shown). These results appear promising and merit further investigation for determination of the antifungal activity of FGE against C. albicans biofilms.

Analysis of the functional diversity of the microbial communities in a paper-mill water system

Most microbiological studies of wastewater treatment systems rely on methods that are dependent on culturing. However, due to the limitation of conventional microbiological methods, alternative assays, such as the analysis of the functional diversity of the microbial communities using Biolog substrate utilisation profiles, have been proposed. The Biolog method was thus evaluated in a paper-mill water system. The influence of the production of various paper grades, biocide combinations and monthly maintenance shut-downs on the functional diversity of the microbial communities were determined using the Biolog technique. The communities in the planktonic as well as the sessile phases were analysed approximately every second week for a period of one year. The average well colour development (AWCD) technique was used to transform the data obtained from the Biolog plates prior to multivariate statistical analysis. Our results indicated that different microbial communities developed during the production of the different paper grades. A difference in substrate utilisation was evident after a single day of production of fluting or linerboard in the planktonic communities, while differences in the sessile communities only became significant after an extended period of production. The effect of the application of different biocides was more distinct in the planktonic than in the sessile populations. No clear trends concerning the influence of maintenance shut-downs and cleaning of machines could be observed in the sessile phase, while a shift in the microbial community could be observed in the planktonic phase. Biodiversity indices indicated that a high functional diversity existed in both the planktonic and sessile phases. It was concluded that the substrate utilisation profiles obtained with Biolog plates were a sensitive measure, which enabled the detection of shifts in the microbial community function and possibly structure within the same water system. Water SA Vol.29(1) 2003: 31-34

Thermophilic and mesophilic bacteria in biofilms associated with corrosion in a heat exchanger

Biofilm development on AISI-1020 carbon steel coupons installed at the outlet of a heat exchanger was evaluated at the thirtieth and the sixtieth days of exposure. Water temperature varied between 41 and 60 °C. The most probable number technique (MPN) was applied to quantify mesophilic and thermophilic species of aerobic, anaerobic, and sulphate-reducing bacteria (SRB) in planktonic and sessile phases. The results showed predominance of thermophilic aerobic bacteria in both phases, corresponding to 9.5 ± 0.8 × 106 cells/ml in the planktonic phase. In biofilms, maximal aerobic cell concentration, 7.8 ± 0.6 × 108 cells/cm2, was registered at the sixtieth day. An increase in the number of thermophilic anaerobic and SRB with elapsed time was also observed. The results obtained after 60 days were 5.8 ± 0.4 × 107 and 8.9 ± 0.9 × 104 cells/cm2 for anaerobic and sulphate-reducing bacteria, respectively. Scanning electron microscopy showed a varied composition of species in the biofilms and corrosion on the carbon steel surfaces after biofilm removal.

Biofilm formation on brass coupons exposed to a cooling system of an oil refinery

Brass coupons (70% Cu 30% Zn) were exposed to a cooling freshwater system of an oil refinery, in order to investigate susceptibility of the metal to biofilm formation. The coupons were fixed on bypasses at points which allowed the circulation of makeup, cooling and return water. The number of aerobic, anaerobic and sulfate-reducing bacteria was determined in both the planktonic and the sessile phases. Maximum bacterial concentrations were detected in the cooling water, corresponding to 2.1 ± 0.1 × 106 CFU ml−1 (planktonic phase) and 1.3 ± 0.2 × 105 CFU cm−2 (sessile phase) for aerobic bacteria and to 3.2 ± 0.3 × 105 cells ml−1 (planktonic phase) and 6.2 ± 0.7 × 105 cells cm−2 (sessile phase) for anaerobic bacteria. Sulfate-reducing bacteria (SRB) were observed only in the planktonic phase, being found in greater numbers in the return water. Scanning electron microscopy (SEM) analysis indicated that biofilm formation occurred at the three monitored sites and showed a diversity in cell morphology. Nonetheless, no evidence of corrosion was observed on the brass coupons during the experimental period.

Changes in the distribution of melanophores and xanthophores inTriturus alpestris embryos during their transition from the uniform to banded pattern.

The change in distribution of melanophores from stage 28+ (uniform melanophore pattern) to stage 34 (banded melanophore pattern) and the participation of xanthophores in these changes has been investigated inTriturus alpestris embryos by studying the social behaviour of single cells. While melanophores are clearly visible from outside the embryo at stage 28+, xanthophores cannot be recognized from the outside until after stage 34. In ultrathin sections of stage 34 embryos, xanthophores are observed alternating with melanophores in a zonal distribution (Epperlein 1982). Using detached pieces of dorsolateral trunk skin, which retain their chromatophores after detachment, the entire distribution of melanophores and xanthophores can be visualized in a scanning electron microscope (SEM). In ambiguous cases (early stages), cells were reprocessed for transmission electron microscopy (TEM) and the presence of the characteristic pigment organelles was assessed. In addition, xanthophores were specifically identified by pteridine fluorescence with dilute ammonia. Pteridines were also identified chromatographically in skin homogenates. The combination of these methods allowed precise identification and quantitative determination of melanophores and xanthophores. Both cell types were present as codistributed, biochemically differentiated cells at stage 28+. Changes in the pattern up to stage 34 were due to the rearrangement at the epidermal-mesodermal interface of a relatively fixed number of melanophores which became preferentially localised at the dorsal somite edge and at the lateral plate mesoderm, and to the distribution of an increasing number of xanthophores to subepidermal locations in the dorsal fin and between the melanophore bands. Concomitant was an increase in the thickness of the epidermal basement membrane and a change in shape of chromatophores from elongate via stellate to rosette shaped, which may be correlated with a shift from migratory to sessile phases.

The zonation phenomenon and structure of the swarm colony in Proteus mirabilis.

SUMMARY Zonation of the swarm colonies of six strains of Proteus mirabilis did not appear to be caused by an alternationof swarming and sessile phases. The margin was constantly composed of active swarmers; and, although a thickened ring could be caused by a check, it was more often due to the rapid advance of the edge, leaving a thinner area which permitted a new, interior edge to form behind it. Motility ceased when the bacteria became crowded; the change to the non-swarming phase took place later. The structure of the swarm colonies was lobate, and the zonation of the separate lobes was synchronised. Reduction of nutrients in the medium reduced the speed of advance of the swarm, but the timing of the zone formation was unaltered. Lobes, at the edges of old colonies, in which swarming had ceased, also showed synchronised zonation. These appearances are suggestive of genetic programming.