Search Results (1,020)

Cell culture methods are indispensable strategies for studies in biological sciences and for drug discovery and testing. Most cell cultures have been developed using two-dimensional (2D) culture methods, but three-dimensional (3D) culture techniques enable the establishment of in vitro models that replicate various pathogenic conditions and they provide valuable insights into the pathophysiology of various diseases as well as more precise results in tests for drug efficacy. However, one difficulty in the use of 3D cultures is selection of the appropriate 3D cell culture technique for the study purpose among the various techniques ranging from the simplest single cell type-derived spheroid culture to the more sophisticated organoid cultures. In the simplest single cell type-derived spheroid cultures, there are also various scaffold-assisted methods such as hydrogel-assisted cultures, biofilm-assisted cultures, particle-assisted cultures, and magnet particle-assisted cultures, as well as non-assisted methods, such as static suspension cultures, floating cultures, and hanging drop cultures. Since each method can be differently influenced by various factors such as gravity force, buoyant force, centrifugal force, and magnetic force, in addition to non-physiological scaffolds, each method has its own advantages and disadvantages, and the methods have different suitable applications. We have been focusing on the use of a hanging drop culture method for modeling various non-cancerous and cancerous diseases because this technique is affected only by gravity force and buoyant force and is thus the simplest method among the various single cell type-derived spheroid culture methods. We have found that the biological natures of spheroids generated even by the simplest method of hanging drop cultures are completely different from those of 2D cultured cells. In this review, we focus on the biological aspects of single cell type-derived spheroid culture and its applications in in vitro models for various diseases. Full article

The accumulation, growth, and re-mobilization of pathogens on the pipe walls in drinking water distribution systems are processes that affect the risk of exposure at the tap. We present a model that uses the Buckingham Pi theory to embody the physics of Pseudomonas aeruginosa accumulation and move within the system. We apply it to model experimental data from a biofilm annular reactor operated in conditions that are commensurate with the flow in DWDS. By calibrating the model for this benchtop system, we intend to identify the most important physical parameters for use in a simpler, more prudent model, for application in large-scale DWDS. Full article

Phage therapy has been proposed as a therapeutic alternative to antibiotics for the treatment of chronic, biofilm-related P. aeruginosa infections. To gain a deeper insight into the complex biofilm–phage interactions, we investigated in the present study the effect of three successive exposures to lytic phages of biofilms formed by the reference strains PAO1 and PA14 as well as of two sequential clinical P. aeruginosa isolates from the sputum of a patient with cystic fibrosis (CF). The Calgary device was employed as a biofilm model and the efficacy of phage treatment was evaluated by measurements of the biomass stained with crystal violet (CV) and of the cell density of the biofilm bacterial population (CFU/mL) after each of the three phage exposures. The genetic alterations of P. aeruginosa isolates from biofilms exposed to phages were investigated by whole-genome sequencing. We show here that the anti-biofilm efficacy of the phage treatment decreased rapidly with repeated applications of lytic phages on P. aeruginosa strains with different genetic backgrounds. Although we observed the maintenance of a small subpopulation of sensitive cells after repeated phage treatments, a fast recruitment of mechanisms involved in the persistence of biofilms to the phage attack occurred, mainly by mutations causing alterations of the phage receptors. However, mutations causing phage-tolerant phenotypes such as alginate-hyperproducing mutants were also observed. In conclusion, a decreased anti-biofilm effect occurred after repeated exposure to lytic phages of P. aeruginosa biofilms due to the recruitment of different resistance and tolerance mechanisms. Full article

Helicobacter pylori is one of the most common bacterial pathogens worldwide and the main etiological agent of numerous gastric diseases. The frequency of multidrug resistance of H. pylori is growing and the leading factor related to this phenomenon is its ability to form biofilm. Therefore, the establishment of a proper model to study this structure is of critical need. In response to this, the aim of this original article is to validate conditions of the optimal biofilm development of H. pylori in monoculture and co-culture with a gastric cell line in media simulating human fluids. Using a set of culture-based and microscopic techniques, we proved that simulated transcellular fluid and simulated gastric fluid, when applied in appropriate concentrations, stimulate autoaggregation and biofilm formation of H. pylori. Additionally, using a co-culture system on semi-permeable membranes in media imitating the stomach environment, we were able to obtain a monolayer of a gastric cell line with H. pylori biofilm on its surface. We believe that the current model for H. pylori biofilm formation in monoculture and co-culture with gastric cells in media containing host-mimicking fluids will constitute a platform for the intensification of research on H. pylori biofilms in in vitro conditions that simulate the human body. Full article

Fear and anxiety during dental visits are common issues that can lead to avoidance of appointments and deterioration of oral health. Effectively managing patients’ emotions during dental treatments is crucial to improving their experiences, increasing adherence to regular visits, and achieving better treatment outcomes. This study aimed to assess the levels of fear and anxiety in patients undergoing hygiene treatments utilizing Guided Biofilm Therapy (GBT) and identify factors that could reduce these negative emotions. A total of 247 patients were evaluated using standardized questionnaires (MDAS, STAI X1, STAI X2, and Gatchel), custom questions, and heart rate monitoring as a physiological stress indicator. Clinical factors, including dental status confirmed by AI-based radiographic analysis (Diagnocat system), as well as sociodemographic influences, were analyzed. Results indicated significant reductions in fear and anxiety after the procedure, as shown by both heart rate and questionnaire scores. Factors such as pain, the presence of caries, and implants were linked to higher anxiety, while strategies like avoiding visible needles and postprocedure interaction with staff were associated with lower stress levels. The findings underscore the importance of personalized care and emotional support to enhance patient experiences. Full article

Biofilm-associated infections present a significant challenge in modern medicine, primarily due to their resilience and resistance to conventional treatments. These infections occur when bacteria form biofilms, protective layers formed by bacterial communities, which are notoriously resistant to traditional antibiotics on surfaces such as medical implants and biological surfaces, making eradication with standard antibiotics difficult. This resilience leads to persistent infections, imposing a substantial economic burden on healthcare systems. The urgency to find alternative treatments is critical as current methods are insufficient and costly. Innovative approaches, such as nanotechnology-based therapies, offer promising alternatives by targeting biofilms more effectively and reducing the need for invasive procedures. Nanocarriers hold significant promise in the fight against biofilm-associated infections. Nanocarriers can penetrate biofilms more effectively than conventional treatments, delivering higher concentrations of antibiotics or other antimicrobial agents precisely where they are needed. This targeted approach not only enhances the efficacy of treatments but also minimizes potential side effects. The development of nanocarrier-based therapies is crucial for overcoming the limitations of current treatments and ultimately improving patient outcomes and reducing the economic burden of biofilm-associated infections on healthcare systems. In this review, nanotechnology-based systems, their characteristics, limitations, and potential benefits are explored to address biofilms-related infections. Additionally, biofilm evaluation models and the tests necessary for the preclinical validation of these nanosystems to facilitate their clinical application are addressed. Full article

Predicting Legionella concentrations reaching users through building water systems requires a comprehensive water quality modeling approach. We integrate various frameworks and data to test the effect of nutrient availability, temperature, chlorine, and biofilm interactions in modeling Legionella. We show that neglecting biofilm detachment underestimates concentrations up to 5.5 logs, while including it increases estimates by 4.2 logs. This study identifies critical factors and uncertainty sources for characterizing the Legionella fate and transport phenomena within buildings. Full article

We present the results of water quality measurements in a pilot-scale, continuously circulated test rig consisting of HDPE pipe segments, where pH, conductivity, turbidity, salinity, temperature, and dissolved oxygen were measured daily. Microbiological measurements (CFU) on the pipe wall and in the bulk water were measured at least once every week. The measurement campaign lasted for 18 weeks. In the first part of the paper, we provide an overview of the results and our experiences. In particular, the time histories of the measured quantities are presented and assessed. Additionally, the flow velocity was increased in six steps from 0.4 to 1.1 m/s to study biofilm detachment once every week. In the second part of the paper, we attempt to use these measurement results for the parameter identification of standard biofilm models. In particular, we search for indirect connections between our measurement results and model parameters (e.g., yield and growth-limiting parameters) via optimising, where the objective is to recover the measured CFU concentration results as closely as possible. Finally, we present preliminary results on the critical wall shear stress resulting in biofilm detachment. Full article

Antimicrobial dosing can be a complex challenge. Although a solid rationale exists for a link between antibiotic exposure and outcome, conflicting data suggest a poor correlation between pharmacokinetic/pharmacodynamic targets and infection control. Different reasons may lead to this discrepancy: poor tissue penetration by β-lactams due to inflammation and inadequate tissue perfusion; different bacterial response to antibiotics and biofilms; heterogeneity of the host’s immune response and drug metabolism; bacterial tolerance and acquisition of resistance during therapy. Consequently, either a fixed dose of antibiotics or a fixed target concentration may be doomed to fail. The role of biomarkers in understanding and monitoring host response to infection is also incompletely defined. Nowadays, with the ever-growing stream of data collected in hospitals, utilizing the most efficient analytical tools may lead to better personalization of therapy. The rise of artificial intelligence and machine learning has allowed large amounts of data to be rapidly accessed and analyzed. These unsupervised learning models can apprehend the data structure and identify homogeneous subgroups, facilitating the individualization of medical interventions. This review aims to discuss the challenges of β-lactam dosing, focusing on its pharmacodynamics and the new challenges and opportunities arising from integrating machine learning algorithms to personalize patient treatment. Full article

Intravascular catheter-associated infections pose a significant threat to the health of patients because of biofilm formation. Hence, it is imperative to exploit cost-effective approaches to improve patient compliance. With this aim, our present study reported the potential of an antimicrobial polymeric gel coating of chitosan (CS) and chlorhexidine (CHX) on the marketed urinary catheters to minimize the risk of biofilm formation. The study involved the implementation of the Quality by Design (QbD) approach by identifying the critical parameters that can affect the coating of the catheter’s surface in any possible way. Later, design of experiments (DoE) analysis affirmed the lack of linearity in the model for the studied responses in a holistic manner. Moreover, in vitro studies were conducted for the evaluation of various parameters followed by the antibiofilm study. The coating exhibited promising release of CHX in the artificial urinary media together with retention of the coating on the catheter’s surface. Therefore, this study aims to emphasize the importance of a systematic and quality-focused approach by contributing to the development of a safe, effective, and reliable catheter coating to enhance intravascular catheter safety. Full article

Infections are one of the main complications in arthroplasties. These infections are difficult to treat because the bacteria responsible for them settle in the prosthesis and form a biofilm that does not allow antimicrobials to reach the infected area. This study is part of a research project aimed at developing 3D-printed spacers (temporary prostheses) capable of incorporating antibacterials for the personalized treatment of arthroplasty infections. The main objective of this research was to analyze the impact of the layer thickness of 3D-printed constructs based on polylactic acid (PLA) for improved treatment of infections in arthroplasty. The focus is on the following parameters: resistance, morphology, drug release, and the effect of antibacterials incorporated in the printed temporary prostheses. The resistance studies revealed that the design and layer thickness of a printed spacer have an influence on its resistance properties. The thickness of the layer used in printing affects the amount of methylene blue (used as a model drug) that is released. Increasing layer thickness leads to a greater release of the drug from the spacer, probably as a result of higher porosity. To evaluate antibacterial release, cloxacillin and vancomycin were incorporated into the constructs. When incorporated into the 3D construct, both antibacterials were released, as evidenced by the growth inhibition of Staphylococcus aureus. In conclusion, preliminary results indicate that the layer thickness during the three-dimensional (3D) printing process of the spacer plays a significant role in drug release. Full article

Biofilm-associated infections account for a large proportion of chronic diseases and pose a major health challenge. Metal nanoparticles offer a new way to address this problem, by impairing microbial growth and biofilm formation and by causing degradation of existing biofilms. This review of metal nanoparticles with antimicrobial actions included an analysis of 20 years of journal papers and patent applications, highlighting the progress over that time. A network analysis of relevant publications showed a major focus on the eradication of single-species biofilms formed under laboratory conditions, while a bibliometric analysis showed growing interest in combining different types of metal nanoparticles with one another or with antibiotics. The analysis of patent applications showed considerable growth over time, but with relatively few patents progressing to be granted. Overall, this profile shows that intense interest in metal nanoparticles as anti-biofilm agents is progressing beyond the confines of simple laboratory biofilm models and coming closer to clinical application. Looking to the future, metal nanoparticles may provide a sustainable approach to combatting biofilms of drug-resistant bacteria. Full article

The propagation of antibiotic resistance in environments, particularly aquatic environments that serve as primary pathways for antibiotic resistance genes (ARGs), poses significant health risks. The impact of nutrients, as key determinants of bacterial growth and metabolism, on the propagation of ARGs, particularly extracellular ARGs (eARGs), remains poorly understood. In this study, we collected microorganisms from the Yangtze River and established a series of microcosms to investigate how variations in nutrient levels and delivery frequency affect the relative abundance of intracellular ARGs (iARGs) and eARGs in bacterial communities. Our results show that the relative abundance of 7 out of 11 representative eARGs in water exceeds that of iARGs, while 8 iARGs dominate in biofilms. Notably, iARGs and eARGs consistently exhibited opposite responses to nutrient variation. When nutrient levels increased, iARGs in the water also increased, with the polluted group (COD = 333.3 mg/L, COD:N:P = 100:3:0.6, m/m) and the eutrophic group (COD = 100 mg/L, COD:N:P = 100:25:5, m/m) showing 1.2 and 3.2 times higher levels than the normal group (COD = 100 mg/L, COD:N:P = 100:10:2, m/m), respectively. In contrast, eARGs decreased by 6.7% and 8.4% in these groups. On the other hand, in biofilms, higher nutrient levels led to an increase in eARGs by 1.5 and 1.7 times, while iARGs decreased by 17.5% and 50.1% in the polluted and eutrophic groups compared to the normal group. Moreover, while increasing the frequency of nutrient delivery (from 1 time/10 d to 20 times/10 d) generally did not favor iARGs in either water or biofilm, it selectively enhanced eARGs in both. To further understand these dynamics, we developed an ARGs-nutrient model by integrating the Lotka–Volterra and Monod equations. The results highlight the complex interplay of bacterial growth, nutrient availability, and mechanisms such as horizontal gene transfer and secretion influencing ARGs’ propagation, driving the opposite trend between these two forms of ARGs. This contrasting response between iARGs and eARGs contributes to a dynamic balance that stabilizes bacterial resistance levels amid nutrient fluctuations. This study offers helpful implications regarding the persistence of bacterial resistance in the environment. Full article

This review describes the discovery, structure, activity, engineered constructs, and applications of KR-12, the smallest antibacterial peptide of human cathelicidin LL-37, the production of which can be induced under sunlight or by vitamin D. It is a moonlighting peptide that shows both antimicrobial and immune-regulatory effects. Compared to LL-37, KR-12 is extremely appealing due to its small size, lack of toxicity, and narrow-spectrum antimicrobial activity. Consequently, various KR-12 peptides have been engineered to tune peptide activity and stability via amino acid substitution, end capping, hybridization, conjugation, sidechain stapling, and backbone macrocyclization. We also mention recently discovered peptides KR-8 and RIK-10 that are shorter than KR-12. Nano-formulation provides an avenue to targeted delivery, controlled release, and increased bioavailability. In addition, KR-12 has been covalently immobilized on biomaterials/medical implants to prevent biofilm formation. These constructs with enhanced potency and stability are demonstrated to eradicate drug-resistant pathogens, disrupt preformed biofilms, neutralize endotoxins, and regulate host immune responses. Also highlighted are the safety and efficacy of these peptides in various topical and systemic animal models. Finaly, we summarize the achievements and discuss future developments of KR-12 peptides as cosmetic preservatives, novel antibiotics, anti-inflammatory peptides, and microbiota-restoring agents. Full article

Borrelia burgdorferi, the bacterium responsible for Lyme disease, has been shown to form antimicrobial-tolerant biofilms, which protect it from unfavorable conditions. Bacterial biofilms are known to significantly contribute to severe inflammation, such as carditis, a common manifestation of Lyme disease. However, the role of B. burgdorferi biofilms in the development of Lyme carditis has not been thoroughly investigated due to the absence of an appropriate model system. In this study, we examined heart tissues from mice infected with B. burgdorferi for the presence of biofilms and inflammatory markers using immunohistochemistry (IHC), combined fluorescence in situ hybridization FISH/IHC, 3D microscopy, and atomic force microscopy techniques. Our results reveal that B. burgdorferi spirochetes form aggregates with a known biofilm marker (alginate) in mouse heart tissues. Furthermore, these biofilms induce inflammation, as indicated by elevated levels of murine C-reactive protein near the biofilms. This research provides evidence that B. burgdorferi can form biofilms in mouse heart tissue and trigger inflammatory processes, suggesting that the mouse model is a valuable tool for future studies on B. burgdorferi biofilms. Full article