pH-FISH: coupled microscale analysis of microbial identity and acid-base metabolism in complex biofilm samples.

Background: Correlative structural and chemical imaging of biofilms allows for the combined analysis of microbial identity and metabolism at the microscale. Here, we developed pH-FISH, a method that combines pH ratiometry with fluorescence in situ hybridization (FISH) in structurally intact biofilms for the coupled investigation of microbial acid metabolism and biofilm composition. Careful biofilm handling and modified sample preparation procedures for FISH allowed preservation of the three-dimensional biofilm structure throughout all processing and imaging steps.

Cooperation between coagulase and von willebrand factor binding protein in fibrin pseudocapsule formation.

The major human pathogen forms biofilms comprising of a fibrin network that increases attachment to surfaces and shields bacteria from the immune system. It secretes two coagulases, Coagulase (Coa) and von Willebrand factor binding protein (vWbp), which hijack the host coagulation cascade and trigger the formation of this fibrin clot. However, it is unclear how Coa and vWbp contribute differently to the localisation and dynamics of clot assembly in growing biofilms.

Environmental pH and compound structure affect the activity of short-chain carboxylic acids against planktonic growth, biofilm formation, and eradication of the food pathogen .

Short-chain carboxylic acids (SCCAs) that are naturally produced by microbial fermentation play an essential role in delaying microbial spoilage. SCCAs are structurally diverse, but only a few of them are routinely used in food biopreservation. This study investigated the effects of environmental pH and intrinsic properties of 21 structurally different SCCAs on the antimicrobial and antibiofilm activity against .

A high-throughput assay identifies molecules with antimicrobial activity against persister cells.

Persister cells are transiently non-growing antibiotic-tolerant bacteria that cause infection relapse, and there is no effective antibiotic therapy to tackle these infections. High-throughput assays in drug discovery are biased towards detecting drugs that inhibit bacterial growth rather than killing non-growing bacteria. A new and simple assay to discover such drugs is needed.

The efficacy and safety of an enzyme-containing lozenge for dental biofilm control-a randomized controlled pilot trial.

Objectives: To evaluate the effect of daily use of a multiple-enzyme lozenge on de novo plaque formation, on gingivitis development, and on the oral microbiome composition.

Methods: This trial with two parallel arms included 24 healthy adults allocated to the Active (n = 12) or Placebo (n = 12) group. Subjects consumed one lozenge three times daily for seven days, and no oral hygiene procedures were allowed.

Efficacy of rifampicin combination therapy against MRSA prosthetic vascular graft infections in a rat model.

is a major cause of prosthetic vascular graft or endograft infections (VGEIs) and the optimal choice of antibiotics is unclear. We investigated various antibiotic choices as either monotherapy or combination therapy with rifampicin against MRSA and . Fosfomycin, daptomycin and vancomycin alone or in combination with rifampicin was used against MRSA USA300 FPR3757.

Bacterial efflux pumps excrete SYTO™ dyes and lead to false-negative staining results.

Multidrug efflux pumps excrete a range of small molecules from bacterial cells. In this study, we show that bacterial efflux pumps have affinity for a range of SYTO™ dyes that are commonly used to label bacteria. Efflux pump activity will there lead to false negative results from bacterial staining and SYTO™ dyes should be used with caution on live samples.

Extracellular G-quadruplexes and Z-DNA protect biofilms from DNase I, and G-quadruplexes form a DNAzyme with peroxidase activity.

Many bacteria form biofilms to protect themselves from predators or stressful environmental conditions. In the biofilm, bacteria are embedded in a protective extracellular matrix composed of polysaccharides, proteins and extracellular DNA (eDNA). eDNA most often is released from lysed bacteria or host mammalian cells, and it is the only matrix component most biofilms appear to have in common.

Combined pH ratiometry and fluorescence lectin-binding analysis (pH-FLBA) for microscopy-based analyses of biofilm pH and matrix carbohydrates.

Bacterial biofilms have a complex and heterogeneous three-dimensional architecture that is characterized by chemically and structurally distinct microenvironments. Confocal microscopy-based pH ratiometry and fluorescence lectin-binding analysis (FLBA) are well-established methods to characterize pH developments and the carbohydrate matrix architecture of biofilms at the microscale. Here, we developed a combined analysis, pH-FLBA, to concomitantly map biofilm pH and the distribution of matrix carbohydrates in bacterial biofilms while preserving the biofilm microarchitecture.

Considerations on the use of microsensors to profile dissolved H2 concentrations in microbial electrochemical reactors.

Measuring the distribution and dynamics of H2 in microbial electrochemical reactors is valuable to gain insights into the processes behind novel bioelectrochemical technologies, such as microbial electrosynthesis. Here, a microsensor method to measure and profile dissolved H2 concentrations in standard H-cell reactors is described. Graphite cathodes were oriented horizontally to enable the use of a motorized microprofiling system and a stereomicroscope was used to place the H2 microsensor precisely on the cathode surface.

The Effect of Enzymatic Treatment with Mutanase, Beta-Glucanase, and DNase on a Saliva-Derived Biofilm Model.

Introduction: The dental biofilm matrix is an important determinant of virulence for caries development and comprises a variety of extracellular polymeric substances that contribute to biofilm stability. Enzymes that break down matrix components may be a promising approach to caries control, and in light of the compositional complexity of the dental biofilm matrix, treatment with multiple enzymes may enhance the reduction of biofilm formation compared to single enzyme therapy. The present study investigated the effect of the three matrix-degrading enzymes mutanase, beta-glucanase, and DNase, applied separately or in combinations, on biofilm prevention and removal in a saliva-derived in vitro-grown model.

A New Device for Dental Biofilm Collection Additively Manufactured by Direct Metal Laser Sintering and Vat Photopolymerization.

Dental biofilms are complex medical biofilms that cause caries, the most prevalent disease of humankind. They are typically collected using handcrafted intraoral devices with mounted carriers for biofilm growth. As the geometry of handcrafted devices is not standardized, the shear forces acting on the biofilms and the access to salivary nutrients differ between carriers.

Fibrinolytic and antibiotic treatment of prosthetic vascular graft infections in a novel rat model.

Objectives: We developed a rat model of prosthetic vascular graft infection to assess, whether the fibrinolytic tissue plasminogen activator (tPA) could increase the efficacy of antibiotic therapy.

Materials And Methods: Rats were implanted a polyethylene graft in the common carotid artery, pre-inoculated with approx. 6 log10 colony forming units (CFU) of methicillin resistant Staphylococcus aureus.

Protein ligand and nanotopography separately drive the phenotype of mouse embryonic stem cells.

Biochemical and biomechanical signals regulate stem cell function in the niche environments in vivo. Current in vitro culture of mouse embryonic stem cells (mESC) uses laminin (LN-511) to provide mimetic biochemical signaling (LN-521 for human systems) to maintain stemness. Alternative approaches propose topographical cues to provide biomechanical cues, however combined biochemical and topographic cues may better mimic the in vivo environment, but are largely unexplored for in vitro stem cell expansion.

Polymyxin B complexation enhances the antimicrobial potential of graphene oxide.

Introduction: The antibacterial activity of graphene oxide (GO) has been widely explored and tested against various pathogenic bacterial strains. Although antimicrobial activity of GO against planktonic bacterial cells was demonstrated, its bacteriostatic and bactericidal effect alone is not sufficient to damage sedentary and well protected bacterial cells inside biofilms. Thus, to be utilized as an effective antibacterial agent, it is necessary to improve the antibacterial activity of GO either by integration with other nanomaterials or by attachment of antimicrobial agents.

GFP fusions of Sec-routed extracellular proteins in reveal surface-associated coagulase in biofilms.

is a major human pathogen that utilises many surface-associated and secreted proteins to form biofilms and cause disease. However, our understanding of these processes is limited by challenges of using fluorescent protein reporters in their native environment, because they must be exported and fold correctly to become fluorescent. Here, we demonstrate the feasibility of using the monomeric superfolder GFP (msfGFP) exported from By fusing msfGFP to signal peptides for the Secretory (Sec) and Twin Arginine Translocation (Tat) pathways, the two major secretion pathways in we quantified msfGFP fluorescence in bacterial cultures and cell-free supernatant from the cultures.

Antibody-Drug Conjugates to Treat Bacterial Biofilms via Targeting and Extracellular Drug Release.

The treatment of implant-associated bacterial infections and biofilms is an urgent medical need and a grand challenge because biofilms protect bacteria from the immune system and harbor antibiotic-tolerant persister cells. This need is addressed herein through an engineering of antibody-drug conjugates (ADCs) that contain an anti-neoplastic drug mitomycin C, which is also a potent antimicrobial against biofilms. The ADCs designed herein release the conjugated drug without cell entry, via a novel mechanism of drug release which likely involves an interaction of ADC with the thiols on the bacterial cell surface.

Polyether Ionophore Antibiotics Target Drug-Resistant Clinical Isolates, Persister Cells, and Biofilms.

Polyether ionophores are complex natural products known to transport various cations across biological membranes. While several members of this family are used in agriculture (e.g.

Biofilm formation and inflammatory potential of A possible cause of orthopedic implant-associated infections.

a coagulase-negative staphylococcal species, has some unusual characteristics for human-associated staphylococci, such as slow growth and its preference for anoxic culture conditions. This species is a relatively abundant member of the human skin microbiota, but its microbiological properties, as well as the pathogenic potential, have scarcely been investigated so far, despite being occasionally isolated from different types of infections including orthopedic implant-associated infections. Here, we investigated the growth and biofilm properties of clinical isolates of and determined host cell responses.

JMM Profile: .

is the most abundant commensal bacterium of human skin. Despite protecting us from foreign invaders, itself exploits human vulnerability when given the opportunity. Such opportunities arise when patients are immunocompromised or when biomedical implants present an opportunity to colonize the surface and form biofilms.

The Role of Nanoscale Distribution of Fibronectin in the Adhesion of Studied by Protein Patterning and DNA-PAINT.

is a widespread and highly virulent pathogen that can cause superficial and invasive infections. Interactions between surface receptors and the extracellular matrix protein fibronectin mediate the bacterial invasion of host cells and is implicated in the colonization of medical implant surfaces. In this study, we investigate the role of distribution of both fibronectin and cellular receptors on the adhesion of to interfaces as a model for primary adhesion at tissue interfaces or biomaterials.

The giant staphylococcal protein Embp facilitates colonization of surfaces through Velcro-like attachment to fibrillated fibronectin.

causes some of the most hard-to-treat clinical infections by forming biofilms: Multicellular communities of bacteria encased in a protective matrix, supporting immune evasion and tolerance against antibiotics. Biofilms occur most commonly on medical implants, and a key event in implant colonization is the robust adherence to the surface, facilitated by interactions between bacterial surface proteins and host matrix components. is equipped with a giant adhesive protein, extracellular matrix-binding protein (Embp), which facilitates bacterial interactions with surface-deposited, but not soluble fibronectin.

Aptamer-Targeted Drug Delivery for Biofilm.

Treatment of biofilm infections using conventional antibiotic therapy is challenging as only doses that are sublethal to the biofilm can be administered safely to patients. A potential solution to this challenge is targeted drug delivery. In this study, we tailored an aptamer-targeted liposomal drug delivery system for accumulation and delivery of antibiotics locally in biofilm.

The Bacterial Life Cycle in Textiles is Governed by Fiber Hydrophobicity.

Colonization of textiles and subsequent metabolic degradation of sweat and sebum components by axillary skin bacteria cause the characteristic sweat malodor and discoloring of dirty clothes. Once inside the textile, the bacteria can form biofilms that are hard to remove by conventional washing. When the biofilm persists after washing, the textiles retain the sweat odor.