scholarly journals Survival and proliferation of an opportunistic pathogen in mixed species biofilms

2021 ◽  
Author(s):  
Mahtab Ghadakpour
Keyword(s):  
Laser Scanning ◽  
Inhibitory Concentration ◽  
Single Cells ◽  
Opportunistic Pathogen ◽  
Laser Scanning Microscopy ◽  
Confocal Laser ◽  
Community Members ◽  
Mixed Species ◽  
Pseudomonas Aeruginosa Pao1 ◽  
Conventional Plate

The purpose of this study was to examine the interaction between an opportunistic pathogen and mixed community biofilms, in terms of integration, proliferation and subsequent release. Pseudomonas aeruginosa PAO1 was used as the test strain in conjuction with community biofilms obtained from sink drains. Confocal laser scanning microscopy (CLSM) analysis showed that PAO1 could successfully incorporate into the community. The relative abundance of PAO1 in the biofilms was dependant on the order of inoculation. Biofilm cell yield was studied using conventional plate counting, CLSM and flow cytometry, which revealed that PAO1 became a dominant community member. Cells were released from the biofilms in the form of single cells, duplets and aggregates of various zies. Detached aggregates were often observed to contain PAO1 and community members. It was also determined that association with communities provided PAO1 with increased protection against EDTA but not against streptomycin, when applied at planktonic minimal inhibitory concentration.

scholarly journals Survival and proliferation of an opportunistic pathogen in mixed species biofilms

2021 ◽  
Author(s):  
Mahtab Ghadakpour
Keyword(s):  
Laser Scanning ◽  
Inhibitory Concentration ◽  
Single Cells ◽  
Opportunistic Pathogen ◽  
Laser Scanning Microscopy ◽  
Confocal Laser ◽  
Community Members ◽  
Mixed Species ◽  
Pseudomonas Aeruginosa Pao1 ◽  
Conventional Plate

The purpose of this study was to examine the interaction between an opportunistic pathogen and mixed community biofilms, in terms of integration, proliferation and subsequent release. Pseudomonas aeruginosa PAO1 was used as the test strain in conjuction with community biofilms obtained from sink drains. Confocal laser scanning microscopy (CLSM) analysis showed that PAO1 could successfully incorporate into the community. The relative abundance of PAO1 in the biofilms was dependant on the order of inoculation. Biofilm cell yield was studied using conventional plate counting, CLSM and flow cytometry, which revealed that PAO1 became a dominant community member. Cells were released from the biofilms in the form of single cells, duplets and aggregates of various zies. Detached aggregates were often observed to contain PAO1 and community members. It was also determined that association with communities provided PAO1 with increased protection against EDTA but not against streptomycin, when applied at planktonic minimal inhibitory concentration.

scholarly journals Cell Adherence and Drug Delivery from Particle Based Mesoporous Silica Films

2019 ◽  
Author(s):  
Emma Björk ◽  
Bernhard Baumann ◽  
Florian Hausladen ◽  
Rainer Wittig ◽  
mika lindén
Keyword(s):  
Drug Delivery ◽  
Laser Scanning ◽  
Single Cells ◽  
Silicon Wafers ◽  
Laser Scanning Microscopy ◽  
Drug Uptake ◽  
C2c12 Cells ◽  
Confocal Laser ◽  
Particle Sizes

Spatially and temporally controlled drug delivery is important for implant and tissue engineering applications, as the efficacy and bioavailability of the drug can be enhanced, and can also allow for drugging stem cells at different stages of development. Long-term drug delivery over weeks to months is however difficult to achieve, and coating of 3D surfaces or creating patterned surfaces is a challenge using coating techniques like spin- and dip-coating. In this study, mesoporous films consisting of SBA-15 particles grown onto silicon wafers using wet processing were evaluated as a scaffold for drug delivery. Films with various particle sizes (100 – 900 nm) and hence thicknesses were grown onto OTS-functionalized silicon wafers using a direct growth method. Precise patterning of the areas for film growth could be obtained by local removal of the OTS functionalization through laser ablation. The films were incubated with the model drug DiO, and murine myoblast cells (C2C12 cells) were seeded onto films with different particle sizes. Confocal laser scanning microscopy (CLSM) was used to study the cell growth, and a vinculin-mediated adherence of C2C12 cells on all films was verified. The successful loading of DiO into the films was confirmed by UV-vis and CLSM. It was observed that the drugs did not desorb from the particles during 24 hours in cell culture. During adherent growth on the films for 4 h, small amounts of DiO and separate particles were observed inside single cells. After 24 h, a larger number of particles and a strong DiO signal were recorded in the cells, indicating a particle mediated drug uptake. A substantial amount of DiO loaded particles were however attached on the substrate after 24 making the films attractive as a long-term reservoir for drugs on e.g. medical implants.<br>

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 Localization and quantification of epidermal growth factor receptors on single cells by confocal laser scanning microscopy.

1992 ◽  
Vol 40 (9) ◽  
pp. 1353-1361 ◽  
Author(s):  
M J Good ◽  
W J Hage ◽  
C L Mummery ◽  
S W De Laat ◽  
J Boonstra
Keyword(s):  
Growth Factor ◽  
Epidermal Growth Factor ◽  
Laser Scanning ◽  
Single Cells ◽  
Laser Scanning Microscopy ◽  
Confocal Laser Scanning ◽  
Confocal Laser ◽  
Scatchard Analysis ◽  
Epidermal Growth

We have established a method for quantifying binding of fluorescence-labeled growth factors to their receptors on single cells in situ with the confocal laser scanning microscope (CLSM). Biotinylated epidermal growth factor (EGF) coupled to phycoerythrin-labeled anti-biotin was used to compare the levels of fluorescence on three different cell types for which the number of EGF factors was known from Scatchard analysis of [125I]-EGF binding. The results showed that as few as 10,000 receptors/cell were detectable above back-ground. This method will provide a rapid and quantifiable alternative to autoradiography for ligand binding to single cells in situ.

Confocal Laser Microscopy of Physiologically Characterized Fluorescently Labeled Central Nervous System Neurons

1988 ◽  
Vol 46 ◽  
pp. 274-275
Author(s):  
J.N. Turner ◽  
J. Swann ◽  
K. Smith ◽  
M. Siemens ◽  
D. Szarowski ◽  
...  
Keyword(s):  
Central Nervous System ◽  
Nervous System ◽  
Laser Scanning ◽  
Tissue Sample ◽  
Lucifer Yellow ◽  
Single Cells ◽  
Brain Slices ◽  
Laser Scanning Microscopy ◽  
Confocal Laser ◽  
Efficient Detection

Confocal laser scanning microscopy (CLSM) is capable of three-dimensional imaging of fluorescently labeled single cells. Efficient detection via a photomultiplier and optical sectioning with high rejection of light from other specimen levels make it possible to image cells surrounded by either labeled or unlabeled tissue. It is no longer necessary to restrict high resolution light microscopy to cultured cells or those near the surface of a tissue sample. Cells can be observed üin situ” in a physiologically characterized environment. Central nervous system neurons can be electrophysiologically characterized and then injected with a fluorescent dye such as lucifer yellow. The CLSM can excite the dye and image the fluorescent emission in thick tissue preparations (hundreds of micrometers) making possible a new approach to the correlation of physiology and anatomy.Brain slices 350 μm thick were obtained from hippocampus and inferior colliculus of immature rats and incubated in oxygenated artificial cerebrospinal fluid. Cells were penetrated with micropipets, characterized electrophysiologically and ionophoretically injected with 5% lucifer yellow in LiAc.

scholarly journals Sanguinarine Inhibits Mono- and Dual-Species Biofilm Formation by Candida albicans and Staphylococcus aureus and Induces Mature Hypha Transition of C. albicans

2020 ◽  
Vol 13 (1) ◽  
pp. 13 ◽  
Author(s):  
Weidong Qian ◽  
Wenjing Wang ◽  
Jianing Zhang ◽  
Miao Liu ◽  
Yuting Fu ◽  
...  
Keyword(s):  
Biofilm Formation ◽  
Laser Scanning ◽  
Inhibitory Concentration ◽  
Fluorescent Dyes ◽  
Laser Scanning Microscopy ◽  
Confocal Laser ◽  
Matrix Component ◽  
New Agents ◽  
Diffusion Assay ◽  
And Staphylococcus Aureus

Previous studies have reported that sanguinarine possesses inhibitory activities against several microorganisms, but its effects on mono- and dual-species biofilms of C. albicans and S. aureus have not been fully elucidated. In this study, we aimed to evaluate the efficacy of sanguinarine for mono- and dual-species biofilms and explore its ability to induce the hypha-to-yeast transition of C. albicans. The results showed that the minimum inhibitory concentration (MIC) and minimum biofilm inhibitory concentration (MBIC90) of sanguinarine against C. albicans and S. aureus mono-species biofilms was 4, and 2 μg/mL, respectively, while the MIC and MBIC90 of sanguinarine against dual-species biofilms was 8, and 4 μg/mL, respectively. The decrease in the levels of matrix component and tolerance to antibiotics of sanguinarine-treated mono- and dual-species biofilms was revealed by confocal laser scanning microscopy combined with fluorescent dyes, and the gatifloxacin diffusion assay, respectively. Meanwhile, sanguinarine at 128 and 256 μg/mL could efficiently eradicate the preformed 24-h biofilms by mono- and dual-species, respectively. Moreover, sanguinarine at 8 μg/mL could result in the transition of C. albicans from the mature hypha form to the unicellular yeast form. Hence, this study provides useful information for the development of new agents to combat mono- and dual-species biofilm-associated infections, caused by C. albicans and S. aureus.

Sorption and metabolism of selected herbicides in river biofilm communities

2001 ◽  
Vol 47 (7) ◽  
pp. 634-641 ◽  
Author(s):  
J R Lawrence ◽  
G Kopf ◽  
J V Headley ◽  
T R Neu
Keyword(s):  
Laser Scanning ◽  
Maximum Level ◽  
Sole Source ◽  
Bulk Water ◽  
Laser Scanning Microscopy ◽  
Water Phase ◽  
Confocal Laser ◽  
Community Members ◽  
River Biofilm ◽  
Biofilm Structure

In the present study, biofilms were grown in rotating annular bioreactors with river water as inoculum and sole source of nutrients. The herbicides atrazine and diclofop methyl were applied to the bioreactors, while an identical reactor acted as a control. Biofilm structure was visualized using specific fluorescent probes in conjunction with confocal laser scanning microscopy. The concentration of both herbicides in the bulk water phase followed the pattern of application. Atrazine and metabolites were detected in biofilm samples using direct insertion probe tandem mass spectrometry (DIP-MS/MS) and only trace levels were detected after the addition phase. Monoclonal antibody (MAb) studies indicated that sorption of atrazine was associated with a unique microcolony type. In contrast, diclofop and metabolites reached a maximum level in the biofilm at the end of the addition phase and persisted in the biofilm. Experiments with14C-labeled atrazine and diclofop methyl indicated that mineralization of these compounds to CO2(<1%) occurred in the river biofilms. Thus, both herbicides were sorbed and metabolized by the river biofilm community and detected in biofilms when they were not detected in the bulk water phase. These results indicate that biofilms and specific community members may act as a sink for herbicides, and that this should be taken into account in terms of both sampling and studies of the environmental chemodynamics of contaminants.Key words: biofilm, confocal, image analyses, herbicide, sorption.

scholarly journals Two-Sided Antibacterial Cellulose Combining Probiotics and Silver Nanoparticles

Molecules ◽  
2021 ◽  
Vol 26 (10) ◽  
pp. 2848
Author(s):  
Laura Sabio ◽  
Andrea Sosa ◽  
José M. Delgado-López ◽  
José M. Dominguez-Vera
Keyword(s):  
Silver Nanoparticles ◽  
Bacterial Infections ◽  
Laser Scanning ◽  
Opportunistic Pathogen ◽  
Laser Scanning Microscopy ◽  
Confocal Laser ◽  
Resistant Bacteria ◽  
Skin Infections ◽  
Antibiotic Resistant ◽  
Promising Strategy

The constant increase of antibiotic-resistant bacteria demands the design of novel antibiotic-free materials. The combination of antibacterials in a biocompatible biomaterial is a very promising strategy to treat infections caused by a broader spectrum of resistant pathogens. Here, we combined two antibacterials, silver nanoparticles (AgNPs) and living probiotics (Lactobacillus fermentum, Lf), using bacterial cellulose (BC) as scaffold. By controlling the loading of each antibacterial at opposite BC sides, we obtained a two-sided biomaterial (AgNP-BC-Lf) with a high density of alive and metabolically active probiotics on one surface and AgNPs on the opposite one, being probiotics well preserved from the killer effect of AgNPs. The resulting two-sided biomaterial was characterized by Field-Emission Scanning Electron Microscopy (FESEM) and Confocal Laser Scanning Microscopy (CLSM). The antibacterial capacity against Pseudomonas aeruginosa (PA), an opportunistic pathogen responsible for a broad range of skin infections, was also assessed by agar diffusion tests in pathogen-favorable media. Results showed an enhanced activity against PA when both antibacterials were combined into BC (AgNP-BC-Lf) with respect to BC containing only one of the antibacterials, BC-Lf or AgNP-BC. Therefore, AgNP-BC-Lf is an antibiotic-free biomaterial that can be useful for the therapy of topical bacterial infections.

scholarly journals Cell Adherence and Drug Delivery from Particle Based Mesoporous Silica Films

2019 ◽  
Author(s):  
Emma Björk ◽  
Bernhard Baumann ◽  
Florian Hausladen ◽  
Rainer Wittig ◽  
mika lindén
Keyword(s):  
Drug Delivery ◽  
Laser Scanning ◽  
Single Cells ◽  
Silicon Wafers ◽  
Laser Scanning Microscopy ◽  
Drug Uptake ◽  
C2c12 Cells ◽  
Confocal Laser ◽  
Particle Sizes

Spatially and temporally controlled drug delivery is important for implant and tissue engineering applications, as the efficacy and bioavailability of the drug can be enhanced, and can also allow for drugging stem cells at different stages of development. Long-term drug delivery over weeks to months is however difficult to achieve, and coating of 3D surfaces or creating patterned surfaces is a challenge using coating techniques like spin- and dip-coating. In this study, mesoporous films consisting of SBA-15 particles grown onto silicon wafers using wet processing were evaluated as a scaffold for drug delivery. Films with various particle sizes (100 – 900 nm) and hence thicknesses were grown onto OTS-functionalized silicon wafers using a direct growth method. Precise patterning of the areas for film growth could be obtained by local removal of the OTS functionalization through laser ablation. The films were incubated with the model drug DiO, and murine myoblast cells (C2C12 cells) were seeded onto films with different particle sizes. Confocal laser scanning microscopy (CLSM) was used to study the cell growth, and a vinculin-mediated adherence of C2C12 cells on all films was verified. The successful loading of DiO into the films was confirmed by UV-vis and CLSM. It was observed that the drugs did not desorb from the particles during 24 hours in cell culture. During adherent growth on the films for 4 h, small amounts of DiO and separate particles were observed inside single cells. After 24 h, a larger number of particles and a strong DiO signal were recorded in the cells, indicating a particle mediated drug uptake. A substantial amount of DiO loaded particles were however attached on the substrate after 24 making the films attractive as a long-term reservoir for drugs on e.g. medical implants.<br>

scholarly journals Effects of lipid emulsions on the formation of Escherichia coli–Candida albicans mixed-species biofilms on PVC

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Shanshan Li ◽  
Wanshi Duan ◽  
Yujie Lei ◽  
Zhonghui Wang ◽  
Chaojiang Fu ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Candida Albicans ◽  
Laser Scanning ◽  
Bloodstream Infections ◽  
Laser Scanning Microscopy ◽  
Confocal Laser ◽  
Lipid Emulsions ◽  
Mixed Species ◽  
E Coli ◽  
Increased Risk

AbstractPatients receiving lipid emulsions are at increased risk of contracting catheter-related bloodstream infections (CRBSIs) in the clinic. More than 15% of CRBSIs are polymicrobial. The objective of this study was to explore the effects of lipid emulsions on the formation of Escherichia coli (E. coli)–Candida albicans (C. albicans) mixed-species biofilms (BFs) on polyvinyl chloride (PVC) surfaces and the underlying mechanism. Mixed-species BFs were produced by coculturing E. coli and C. albicans with PVC in various concentrations of lipid emulsions. Crystal violet staining and XTT assays were performed to test the mixed-species BF biomass and the viability of microbes in the BFs. The microstructures of the BFs were observed by an approach that combined confocal laser scanning microscopy, fluorescence in situ hybridization, and scanning electron microscopy. The study found that lipid emulsions could promote the formation of E. coli–C. albicans mixed-species BFs, especially with 10% lipid emulsions. The mechanism by which lipid emulsions promote mixed-species BF formation may involve significant upregulation of the expression of the flhDC, iha, HTA1, and HWP1 genes, which are associated with bacterial motility, adhesion, and BF formation. The results derived from this study necessitate strict aseptic precautions when handling lipid emulsions and avoiding the use of high concentrations of lipid emulsions for as long as possible.

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