scholarly journals Characterization of Binary Biofilms of Listeria monocytogenes and Lactobacillus and Their Response to Chlorine Treatment

2021 ◽  
Vol 12 ◽  
Author(s):  
Magdalena A. Olszewska ◽  
Francisco Diez-Gonzalez
Keyword(s):  
Listeria Monocytogenes ◽  
Laser Scanning ◽  
Spatial Organization ◽  
Volume Ratio ◽  
Single Species ◽  
Laser Scanning Microscopy ◽  
Confocal Laser ◽  
Interspecies Interactions ◽  
Biofilm Thickness ◽  
Cell Counts

In nature, Listeria may interact competitively and cooperatively with other organisms, resulting in unique spatial organization and functions for cells within the community. This study was undertaken to characterize the biofilm architecture of binary biofilms of Listeria monocytogenes and Lactobacillus species and to assess their effect on the survival of Listeria during exposure to hypochlorite. Three L. monocytogenes strains, ATCC 19115 (Lm5), ATCC 19117 (Lm7), and Coleslaw (LmC), were selected and combined individually with three Lactobacillus strains: L. fermentum (Lf), L. bavaricus (Lb), and L. plantarum (Lp). In binary Lm-Lp biofilms, the Lm cell counts were similar to single-species biofilms (8.5 log CFU/well), and the Lp cell numbers declined by 1.0 log CFU/well. In the presence of Lb, the Lm cell counts were reduced by 1.5 log CFU/well (p < 0.05), whereas the Lf cell counts increased at least by 3.5 log CFU/well. Confocal laser scanning microscopy (CLSM) determined that interspecies interactions significantly affected the spatial organization of three binary biofilms. Biofilm surface-to-volume ratio increased from 0.8 μm2/μm3 for Lm5 in the monoculture to 2.1 μm2/μm3 for Lm5-Lp in the dual-species model (p < 0.05), and was characterized by a thicker structure with a largely increased surface area. Biofilm roughness increased from 0.2 for Lm7 to 1.0 for Lm7-Lb biofilms (p < 0.05), which appeared as interspecific segregation. Biofilm thickness increased from 34.2 μm for LmC to 46.3 μm for LmC–Lf (p < 0.05), which produced flat and compact structures that covered the entire surface available. The biomass of the extracellular matrix was higher in the case of some binary biofilms (p < 0.05); however, this effect was dependent upon the species pair. When treated with hypochlorite, Lm5 in binary biofilms had an approximately 1.5 log CFU/well greater survival than individually. The unique spatial organization and greater protein production may explain the protective effect of Lp after hypochlorite exposure.

scholarly journals Low-Temperature Plasma as an Approach for Inhibiting a Multi-Species Cariogenic Biofilm

2021 ◽  
Vol 11 (2) ◽  
pp. 570
Author(s):  
Leandro W. Figueira ◽  
Beatriz H. D. Panariello ◽  
Cristiane Y. Koga-Ito ◽  
Simone Duarte
Keyword(s):  
Low Temperature ◽  
Laser Scanning ◽  
Single Species ◽  
Laser Scanning Microscopy ◽  
Confocal Laser ◽  
Low Temperature Plasma ◽  
Temperature Plasma ◽  
Plasma Device ◽  
Negative Controls ◽  
Promising Source

This study aimed to determine how low-temperature plasma (LTP) treatment affects single- and multi-species biofilms formed by Streptococcus mutans, Streptococcus sanguinis, and Streptococcus gordonii formed on hydroxyapatite discs. LTP was produced by argon gas using the kINPen09™ (Leibniz Institute for Plasma Science and Technology, INP, Greifswald, Germany). Biofilms were treated at a 10 mm distance from the nozzle of the plasma device to the surface of the biofilm per 30 s, 60 s, and 120 s. A 0.89% saline solution and a 0.12% chlorhexidine solution were used as negative and positive controls, respectively. Argon flow at three exposure times (30 s, 60 s, and 120 s) was also used as control. Biofilm viability was analyzed by colony-forming units (CFU) recovery and confocal laser scanning microscopy. Multispecies biofilms presented a reduction in viability (log10 CFU/mL) for all plasma-treated samples when compared to both positive and negative controls (p < 0.0001). In single-species biofilms formed by either S. mutans or S. sanguinis, a significant reduction in all exposure times was observed when compared to both positive and negative controls (p < 0.0001). For single-species biofilms formed by S. gordonii, the results indicate total elimination of S. gordonii for all exposure times. Low exposure times of LTP affects single- and multi-species cariogenic biofilms, which indicates that the treatment is a promising source for the development of new protocols for the control of dental caries.

scholarly journals Enhanced Bactericidal Efficacy of NaOCl at pH 12 Followed by Acidified NaOCl at pH 6.5 on Enterococcus faecalis Biofilm

2020 ◽  
Vol 10 (17) ◽  
pp. 6096
Author(s):  
Ronald Wigler ◽  
Shlomo Matalon ◽  
Tomer Goldberger ◽  
Anat Or Lerner ◽  
Anda Kfir
Keyword(s):  
Contact Time ◽  
Laser Scanning ◽  
Saline Solution ◽  
Volume Ratio ◽  
Bactericidal Effect ◽  
Laser Scanning Microscopy ◽  
Confocal Laser ◽  
Red Fluorescence ◽  
Bactericidal Efficacy ◽  
Significant Difference

This study aimed to determine the bactericidal efficacy of sequential use of NaOCl pH 12 followed by acidified NaOCl pH 6.5, and compare it to that of either of these NaOCl solutions alone. E. faecalis biofilm was grown on standardized dentine specimens for four weeks. The specimens were randomly divided into four groups: (A) 4 min exposure to 0.9% saline solution (control); (B) 4 min exposure to 4% NaOCl pH 12; (C) 4 min exposure to 4% NaOCl pH 6.5; and (D) 2 min exposure to 4% NaOCl pH 12 followed by 2 min exposure to 4% NaOCl pH 6.5. The bactericidal activity was evaluated after the 4 min of contact time using confocal laser scanning microscopy. The volume ratio of red fluorescence to green and red fluorescence indicated the proportion of dead cells in the biofilm. The percent of dead cells in the saline solution group was significantly lower than those in the other groups. There was no significant difference between NaOCl pH 12 compared to NaOCl pH 6.5. The sequential use of NaOCl pH 12 followed by pH 6.5 significantly increased the percent of dead cells compared to both the samples exposed to either NaOCl pH 12 or pH 6.5. These results show that sequential irrigation protocol had a stronger bactericidal effect than the commonly used NaOCl pH 12.

Effects of Bacteriophage P100 at Different Concentrations on the Structural Parameters of Listeria monocytogenes Biofilms

2018 ◽  
Vol 81 (12) ◽  
pp. 2040-2044 ◽  
Author(s):  
CRISTINA RODRÍGUEZ-MELCÓN ◽  
ROSA CAPITA ◽  
CAMINO GARCÍA-FERNÁNDEZ ◽  
CARLOS ALONSO-CALLEJA
Keyword(s):  
Image Analysis ◽  
Listeria Monocytogenes ◽  
Laser Scanning ◽  
Digital Image Analysis ◽  
Surface Coverage ◽  
Structural Parameters ◽  
Laser Scanning Microscopy ◽  
Confocal Laser ◽  
Foodborne Diseases ◽  
Average Thickness

ABSTRACT Because listeriosis is one of the deadliest foodborne diseases, controlling and eradicating Listeria monocytogenes biofilms is a serious challenge for food safety. Biofilms (24 h old) formed on polystyrene by a L. monocytogenes strain of food origin were exposed for a further 24 h to 12 different concentrations (from 100 to 1011 PFU/mL) of the bacteriophage P100 (Listex P100). The structural parameters of biofilms were studied by using confocal laser scanning microscopy and digital image analysis. The biovolume in the observation field (14,121 μm2) of control (untreated) biofilms was 237,333.1 ± 2,692.6 μm3. The biomass of treated biofilms ranged from 164.7 ± 89.0 μm3 (biofilms exposed to 1010 PFU/mL) to 231,170.5 ± 15,142.0 μm3 (100 PFU/mL). The lowest biomass was achieved after treatment with 108 PFU/mL, with no further decrease in biovolume when higher phage concentrations were used. A strong (P &lt; 0.001) correlation was found between phage concentration (log units) and biovolume (−0.965), surface coverage (−0.939), roughness (0.976), maximum thickness (−0.853), and average thickness (−0.965). Findings from this research suggest that bacteriophage P100 at concentrations equal to or greater than 8 log PFU/mL successfully removes L. monocytogenes biofilms from polystyrene surfaces.

Interspecies Interactions in Dual-Species Biofilms Formed by Psychrotrophic Bacteria and the Tolerance of Sessile Communities to Disinfectants

2020 ◽  
Vol 83 (6) ◽  
pp. 951-958 ◽  
Author(s):  
LEI YUAN ◽  
NI WANG ◽  
FAIZAN A. SADIQ ◽  
GUOQING HE
Keyword(s):  
Hydrogen Peroxide ◽  
Sodium Hypochlorite ◽  
Peracetic Acid ◽  
Laser Scanning ◽  
Structural Changes ◽  
Laser Scanning Microscopy ◽  
Confocal Laser ◽  
Interspecies Interactions ◽  
Mixed Species ◽  
Psychrotrophic Bacteria

ABSTRACT Biofilms on the surface of food processing equipment act as potential reservoirs of microbial contamination. Bacterial interactions are believed to play key roles in both biofilm formation and antimicrobial tolerance. In this study, Aeromonas hydrophila, Chryseobacterium oncorhynchi, and Pseudomonas libanensis, which were previously isolated from Chinese raw milk samples, were selected to establish two dual-species biofilm models (P. libanensis plus A. hydrophila and P. libanensis plus C. oncorhynchi) on stainless steel at 7°C. Subsequently, three disinfectants, hydrogen peroxide (100 ppm), peracetic acid (100 ppm), and sodium hypochlorite (100 ppm), were used to treat the developed sessile communities for 10 min. Structural changes after exposure to disinfectants were analyzed with confocal laser scanning microscopy. The cell numbers of both A. hydrophila and C. oncorhynchi recovered from surfaces increased when grown as dual species biofilms with P. libanensis. Dual-species biofilms were more tolerant of disinfectants than were each single-species biofilm. Peracetic acid was the most effective disinfectant for removing biofilms, followed by hydrogen peroxide and sodium hypochlorite. The results expand the knowledge of mixed-species biofilms formed by psychrotrophic bacteria and will be helpful for developing effective strategies to eliminate bacterial mixed-species biofilms. HIGHLIGHTS

scholarly journals High In Vitro Antibacterial Activity of Pac-525 againstPorphyromonas gingivalisBiofilms Cultured on Titanium

2015 ◽  
Vol 2015 ◽  
pp. 1-8 ◽  
Author(s):  
Ji-yin Li ◽  
Xue-jin Wang ◽  
Li-na Wang ◽  
Xiao-xia Ying ◽  
Xiang Ren ◽  
...  
Keyword(s):  
Laser Scanning ◽  
Pathogenic Bacteria ◽  
Fusobacterium Nucleatum ◽  
Laser Scanning Microscopy ◽  
Confocal Laser ◽  
Average Cell ◽  
Biofilm Thickness ◽  
Live Bacteria ◽  
In Vitro Antibacterial Activity

In order to investigate the potential of short antimicrobial peptides (AMPs) as alternative antibacterial agents during the treatment of peri-implantitis, the cytotoxic activity of three short AMPs, that is, Pac-525, KSL-W, and KSL, was determined using the MTT assay. The antimicrobial activity of these AMPs, ranging in concentration from 0.0039 mg/mL to 0.5 mg/mL, against the predominant planktonic pathogens, includingStreptococcus sanguis, Fusobacterium nucleatum, andPorphyromonas gingivalis, involved in peri-implantitis was investigated. Furthermore, 2-day-oldP. gingivalisbiofilms cultured on titanium surfaces were treated with Pac-525 and subsequently observed and analysed using confocal laser scanning microscopy (CLSM). The average cell proliferation curve indicated that there was no cytotoxicity due to the three short AMPs. The minimum inhibitory concentration and minimum bactericidal concentration values of Pac-525 were 0.0625 mg/mL and 0.125 mg/mL, respectively, forP. gingivalisand 0.0078 mg/mL and 0.0156 mg/mL, respectively, forF. nucleatum. Using CLSM, we confirmed that compared to 0.1% chlorhexidine, 0.5 mg/mL of Pac-525 caused a significant decrease in biofilm thickness and a decline in the percentage of live bacteria. These data indicate that Pac-525 has unique properties that might make it suitable for the inhibition the growth of pathogenic bacteria around dental implants.

scholarly journals Individual or Combined Effects of Meropenem, Imipenem, Sulbactam, Colistin, and Tigecycline on Biofilm-Embedded Acinetobacter baumannii and Biofilm Architecture

2016 ◽  
Vol 60 (8) ◽  
pp. 4670-4676 ◽  
Author(s):  
Yung-Chih Wang ◽  
Shu-Chen Kuo ◽  
Ya-Sung Yang ◽  
Yi-Tzu Lee ◽  
Chun-Hsiang Chiu ◽  
...  
Keyword(s):  
Acinetobacter Baumannii ◽  
Laser Scanning ◽  
Bactericidal Effect ◽  
Laser Scanning Microscopy ◽  
Confocal Laser ◽  
Roughness Coefficient ◽  
Content Type ◽  
Biofilm Thickness ◽  
Carbapenem Resistant ◽  
Biofilm Architecture

ABSTRACTAcinetobacter baumanniibiofilms are difficult to eradicate. We investigated the effects of meropenem (2 mg/liter), imipenem (2 mg/liter), sulbactam (4 mg/liter), colistin (2 mg/liter), and tigecycline (2 mg/liter), alone or in combination, on biofilm-embedded carbapenem-resistant and carbapenem-susceptibleA. baumannii(CRAb and CSAb, respectively) cells, as well as on the architecture of the biofilms.A. baumanniiATCC 15151 (Ab15151) and its OXA-82-overproducing transformant, along with two clinical CSAb and two clinical CRAb isolates of differing clonalities, were used. The minimal bactericidal concentrations for biofilm-embedded cells of the six tested isolates were >50-fold those of their planktonic cells. When used individually, meropenem exhibited a higher killing effect than the other four antimicrobials on biofilm-embedded CSAb cells in the colony biofilm assay. For two clinical CRAb isolates, meropenem plus sulbactam or sulbactam plus tigecycline showed >100-fold the bactericidal effect exhibited by these agents used alone after 48 h of treatment. The effect of antimicrobials on the architecture of Ab15151 biofilm emitting green fluorescence was determined by confocal laser scanning microscopy using COMSTAT software. Significant decreases in the maximum biofilm thickness were observed after exposure to meropenem and imipenem. Meropenem plus sulbactam significantly decreased the biomass and mean thickness and increased the roughness coefficient of biofilms, but sulbactam plus tigecycline only decreased the maximum and mean biofilm thickness compared to any of these agents used alone. Meropenem was active against biofilm-embedded CSAb, whereas meropenem plus sulbactam exhibited synergism against biofilm-embedded CRAb and caused significantly more damage to the biofilm architecture than did any of the agents used alone.

scholarly journals Bacteria on Catheters in Patients Undergoing Peritoneal Dialysis

2013 ◽  
Vol 33 (1) ◽  
pp. 51-59 ◽  
Author(s):  
Maria Pihl ◽  
Julia R. Davies ◽  
Ann-Cathrine Johansson ◽  
Gunnel Svensäter
Keyword(s):  
Peritoneal Dialysis ◽  
Laser Scanning ◽  
Close Association ◽  
Bacterial Species ◽  
Clinical Signs ◽  
Single Species ◽  
Laser Scanning Microscopy ◽  
Confocal Laser ◽  
Rrna Gene ◽  
Micro Organisms

♦BackgroundPeritonitis is the leading cause of morbidity for peritoneal dialysis (PD) patients, and microbial biofilms have previously been identified on catheters from infected patients. However, few studies of catheters from patients without clinical signs of infection have been undertaken. The aim of the present study was to investigate the extent to which bacteria are present on catheters from PD patients with no symptoms of infection.♦MethodsMicrobiologic culturing under aerobic and anaerobic conditions and confocal laser scanning microscopy were used to determine the distribution of bacteria on PD catheters from 15 patients without clinical signs of infection and on catheters from 2 infected patients. The 16S rRNA gene sequencing technique was used to identify cultured bacteria.♦ResultsBacteria were detected on 12 of the 15 catheters from patients without signs of infection and on the 2 catheters from infected patients. Single-species and mixed-microbial communities containing up to 5 species were present on both the inside and the outside along the whole length of the colonized catheters. The bacterial species most commonly found were the skin commensals Staphylococcus epidermidis and Propionibacterium acnes, followed by S. warneri and S. lugdunensis. The strains of these micro-organisms, particularly those of S. epidermidis, varied in phenotype with respect to their tolerance of the major classes of antibiotics.♦ConclusionsBacteria were common on catheters from patients without symptoms of infection. Up to 4 different bacterial species were found in close association and may represent a risk factor for the future development of peritonitis in patients hosting such micro-organisms.

Impact of Hydrodynamics on Oral Biofilm Strength

2009 ◽  
Vol 88 (10) ◽  
pp. 922-926 ◽  
Author(s):  
E. Paramonova ◽  
O.J. Kalmykowa ◽  
H.C. van der Mei ◽  
H.J. Busscher ◽  
P.K. Sharma
Keyword(s):  
Laser Scanning ◽  
Periodontal Diseases ◽  
Single Species ◽  
Laser Scanning Microscopy ◽  
Confocal Laser ◽  
Oral Biofilm ◽  
Actinomyces Naeslundii ◽  
Mechanical Removal ◽  
Hydrodynamic Shear ◽  
Oral Biofilms

Mechanical removal of oral biofilms is ubiquitously accepted as the best way to prevent caries and periodontal diseases. Removal effectiveness strongly depends on biofilm strength. To investigate the influence of hydrodynamics on oral biofilm strength, we grew single- and multi-species biofilms of Streptococcus oralis J22, Actinomyces naeslundii TV14-J1, and full dental plaque at shear rates ranging from 0.1 to 50 1/sec and measured their compressive strength. Subsequently, biofilm architecture was evaluated by confocal laser scanning microscopy. Multi-species biofilms were stronger than single-species biofilms, with strength values ranging from 6 to 51 Pa and from 5 to 17 Pa, respectively. In response to increased hydrodynamic shear, biofilm strength decreased, and architecture changed from uniform carpet-like to more “fluffy” with higher thickness. S. oralis biofilms grown under variable shear of 7 and 50 1/sec possessed properties intermediate of those measured at the respective single shears.

scholarly journals Genetic Features of Resident Biofilms Determine Attachment of Listeria monocytogenes

2009 ◽  
Vol 75 (24) ◽  
pp. 7814-7821 ◽  
Author(s):  
Olivier Habimana ◽  
Mickael Meyrand ◽  
Thierry Meylheuc ◽  
Saulius Kulakauskas ◽  
Romain Briandet
Keyword(s):  
Listeria Monocytogenes ◽  
Laser Scanning ◽  
Time Course ◽  
Surface Hydrophobicity ◽  
Flow Cell ◽  
Laser Scanning Microscopy ◽  
Confocal Laser ◽  
Microscopy Imaging ◽  
Initial Fixation ◽  
Genetic Features

ABSTRACT Planktonic Listeria monocytogenes cells in food-processing environments tend most frequently to adhere to solid surfaces. Under these conditions, they are likely to encounter resident biofilms rather than a raw solid surface. Although metabolic interactions between L. monocytogenes and resident microflora have been widely studied, little is known about the biofilm properties that influence the initial fixation of L. monocytogenes to the biofilm interface. To study these properties, we created a set of model resident Lactococcus lactis biofilms with various architectures, types of matrices, and individual cell surface properties. This was achieved using cell wall mutants that affect bacterial chain formation, exopolysaccharide (EPS) synthesis and surface hydrophobicity. The dynamics of the formation of these biofilm structures were analyzed in flow cell chambers using in situ time course confocal laser scanning microscopy imaging. All the L. lactis biofilms tested reduced the initial immobilization of L. monocytogenes compared to the glass substratum of the flow cell. Significant differences were seen in L. monocytogenes settlement as a function of the genetic background of resident lactococcal biofilm cells. In particular, biofilms of the L. lactis chain-forming mutant resulted in a marked increase in L. monocytogenes settlement, while biofilms of the EPS-secreting mutant efficiently prevented pathogen fixation. These results offer new insights into the role of resident biofilms in governing the settlement of pathogens on food chain surfaces and could be of relevance in the field of food safety controls.

scholarly journals Microscopic and Spectroscopic Analyses of Chlorhexidine Tolerance in Delftia acidovorans Biofilms

2014 ◽  
Vol 58 (10) ◽  
pp. 5673-5686 ◽  
Author(s):  
Tara Rema ◽  
John R. Lawrence ◽  
James J. Dynes ◽  
Adam P. Hitchcock ◽  
Darren R. Korber
Keyword(s):  
Laser Scanning ◽  
Cell Types ◽  
Laser Scanning Microscopy ◽  
Confocal Laser ◽  
Content Type ◽  
Flow Cells ◽  
Biofilm Thickness ◽  
Delftia Acidovorans ◽  
Scanning Transmission ◽  
Negative Effect

ABSTRACTThe physicochemical responses ofDelftia acidovoransbiofilms exposed to the commonly used antimicrobial chlorhexidine (CHX) were examined in this study. A CHX-sensitive mutant (MIC, 1.0 μg ml−1) was derived from a CHX-tolerant (MIC, 15.0 μg ml−1)D. acidovoransparent strain using transposon mutagenesis.D. acidovoransmutant (MT51) and wild-type (WT15) strain biofilms were cultivated in flow cells and then treated with CHX at sub-MIC and inhibitory concentrations and examined by confocal laser scanning microscopy (CLSM), scanning transmission X-ray microscopy (STXM), and infrared (IR) spectroscopy. Specific morphological, structural, and chemical compositional differences between the CHX-treated and -untreated biofilms of both strains were observed. Apart from architectural differences, CLSM revealed a negative effect of CHX on biofilm thickness in the CHX-sensitive MT51 biofilms relative to those of the WT15 strain. STXM analyses showed that the WT15 biofilms contained two morphochemical cell variants, whereas only one type was detected in the MT51 biofilms. The cells in the MT51 biofilms bioaccumulated CHX to a similar extent as one of the cell types found in the WT15 biofilms, whereas the other cell type in the WT15 biofilms did not bioaccumulate CHX. STXM and IR spectral analyses revealed that CHX-sensitive MT51 cells accumulated the highest levels of CHX. Pretreating biofilms with EDTA promoted the accumulation of CHX in all cells. Thus, it is suggested that a subpopulation of cells that do not accumulate CHX appear to be responsible for greater CHX resistance inD. acidovoransWT15 biofilm in conjunction with the possible involvement of bacterial membrane stability.

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