Staphylococcal Biofilm Research Articles

Isolation and characterisation of novel lytic bacteriophages for therapeutic applications in biofilm-associated prosthetic joint infections

Abstract Prosthetic joint infections are devastating complications of joint arthroplasties. Without effective management, they can lead to limb amputation and even death. A significant proportion of these infections is caused by the primarily commensal Coagulase-negative Staphylococci pathogens, which form thick, antibiotic-resistant biofilms at the site of infection. Combinatorial therapy involving antibiotics and bacteriophages may represent a strategy to overcome resistance. Previous research indicates that as bacteria develop resistance to antibiotics, they often become more susceptible to bacteriophages. In this study, we produced a cocktail of novel bacteriophages and assessed their viability to eradicate nosocomial staphylococcal biofilms. Here, we used clinical isolates from prosthetic joint infections to isolate and identify four new bacteriophages from sewage effluent. These novel phages were characterized through electron microscopy and full genome sequencing. Subsequently, we combined them into a phage cocktail, which effectively re-sensitized biofilms to vancomycin and flucloxacillin. Notably, this phage cocktail demonstrated low cytotoxicity in vitro to human epithelial cells, even when used alongside antibiotic treatments. These findings highlight the potential of the phage cocktail as a tool to increase antibiotic treatment success in prosthetic joint infections.

Combinatorial Effects of Ursodeoxycholic Acid and Antibiotic in Combating Staphylococcus aureus Biofilm: The Roles of ROS and Virulence Factors.

Staphylococcus aureus is a biofilm-forming bacterium responsible for various human infections, one particularly challenging to treat due to its antibiotic resistance. Biofilms can form on both soft tissues and medical devices, leading to persistent and hard-to-treat infections. Combining multiple antimicrobials is a potential approach to overcoming this resistance. This study explored the effects of ursodeoxycholic acid (UDCA) combined with the antibiotic ciprofloxacin against S. aureus biofilms, aiming to evaluate any synergistic effects. Results showed that UDCA and ciprofloxacin co-treatment significantly reduced biofilm formation and disrupted pre-formed biofilms more effectively than either agent alone (p < 0.01). The combination also displayed a slight synergistic effect, with a fractional inhibitory concentration of 0.65. Additionally, the treatment reduced the production of extracellular polymeric substances, increased reactive oxygen species production, decreased metabolic activity, altered cell membrane permeability, and lowered cell surface hydrophobicity in S. aureus. Furthermore, it diminished biofilm-associated pathogenic factors, including proteolytic activity and staphyloxanthin production. Overall, the UDCA-ciprofloxacin combination shows considerable promise as a strategy to combat infections related to staphylococcal biofilms, offering a potential solution to the healthcare challenges posed by antibiotic-resistant S. aureus.

The effectiveness of antimicrobial photodynamic therapy on catheter infection model

Background/AimThis experimental study aimed to examine the effectiveness of transdermal antimicrobial photodynamic therapy (APDT) with and without antimicrobial lock therapy (ALT), on catheter biofilms. MethodsS. epidermidis and C. orthopsilosis biofilms were formed within peripheral venous catheters positioned in the marginal ear veins of New Zealand white rabbits. Biofilm formation was confirmed with scanning electron microscopy in two catheters. 24 catheters with staphylococcal biofilms and 24 with fungal biofilms were treated with APDT, ALT or “APDT plus ALT” for five days. Six catheters were separated as controls. APDT was applied with a red colored LED lamp and methylene blue as the photosensitizer. Vancomycin lock solutions were used as ALT for staphylococcal biofilms and amphotericin B for fungal biofilms. The effect of treatment procedures was evaluated by intraluminal biofilm viability testing based on spectrophotometric evaluation, and a quantitative (OD) value was obtained for each catheter. ResultsThe mean OD values obtained by 600 nm spectrophotometric reading at 24 h (biofilm viability) after “ALT”, “APDT” and “ALT plus APDT” procedures were 0.363, 0.151 and 0.128 for S. epidermidis and 0.092, 0.104 and 0.227 for C. orthopsilosis, respectively. All these OD values obtained after treatment procedures were lower than controls for both S. epidermidis (OD: 0,802) and C. orthopsilosis (OD: 0,315), although there were large fluctuations in our results. ConclusionsOur results suggest that transdermal APDT may be an effective method for treating staphylococcal and candida biofilms formed within intravenous catheters in our rabbit ear model. The combined use of APDT and ALT might be beneficial in these staphylococcal biofilms.

Bioemulsifier from sponge-associated bacteria reduces staphylococcal biofilm

Biofilm formation is a major health concern and studies have been pursued to find compounds able to prevent biofilm establishment and remove pre-existing biofilms. While biosurfactants (BS) have been well-known for possessing antibiofilm activities, bioemulsifiers (BE) are still scarcely explored for this purpose. The present study aimed to evaluate the bioemulsifying properties of cell-free supernatants produced by Bacillaceae and Vibrio strains isolated from marine sponges and investigate their antiadhesive and antibiofilm activities against different pathogenic Gram-positive and Gram-negative bacteria. The BE production by the marine strains was confirmed by the emulsion test, drop-collapsing, oil-displacement, cell hydrophobicity and hemolysis assays. Notably, Bacillus cereus 64BHI1101 displayed remarkable emulsifying activity and the ultrastructure analysis of its BE extract (BE64-1) revealed the presence of structures typically observed in macromolecules composed of polysaccharides and proteins. BE64-1 showed notable antiadhesive and antibiofilm activities against Staphylococcus aureus, with a reduction of adherence of up to 100 % and a dispersion of biofilm of 80 %, without affecting its growth. BE64-1 also showed inhibition of Staphylococcus epidermidis and Escherichia coli biofilm formation and adhesion. Thus, this study provides a starting point for exploring the antiadhesive and antibiofilm activities of BE from sponge-associated bacteria, which could serve as a valuable tool for future research to combat S. aureus biofilms.

Emergence of rifampicin-resistant staphylococci on the skin and nose of rifampicin-treated patients with an orthopaedic-device-related infection.

Rifampicin is a key antibiotic in the treatment of staphylococcal biofilm infections. In this pilot study, we found that patients who received rifampicin for treatment of an orthopaedic-device-related infection (ODRI) were colonized with rifampicin-resistant staphylococci during treatment and this persisted for up to 2months after cessation of treatment.

Investigating the translational value of Periprosthetic Joint Infection (PJI) models to determine the risk and severity of Staphylococcal biofilms.

With the advent of antibiotic-eluting polymeric materials for targeting recalcitrant infections, using preclinical models to study biofilm is crucial for improving the treatment efficacy in periprosthetic joint infections. The stratification of risk and severity of infections is needed to develop an effective clinical dosing framework with better outcomes. Here, using in-vivo and in-vitro implant-associated infection models, we demonstrate that methicillin-sensitive and resistant Staphylococcus aureus (MSSA and MRSA) have model-dependent distinct implant and peri-implant tissue colonization patterns. The maturity of biofilms and the location (implant vs tissue) were found to influence the antibiotic susceptibility evolution profiles of MSSA and MRSA and the models could capture the differing host-microbe interactions in vivo. Gene expression studies revealed the molecular heterogeneity of colonizing bacterial populations. The comparison and stratification of the risk and severity of infection across different preclinical models provided in this study can guide clinical dosing to effectively prevent or treat PJI.

Cuminaldehyde and Tobramycin Forestall the Biofilm Threats of Staphylococcus aureus: A Combinatorial Strategy to Evade the Biofilm Challenges.

Staphylococcus aureus, an opportunistic Gram-positive pathogen, is known for causing various infections in humans, primarily by forming biofilms. The biofilm-induced antibiotic resistance has been considered a significant medical threat. Combinatorial therapy has been considered a reliable approach to combat antibiotic resistance by using multiple antimicrobial agents simultaneously, targeting bacteria through different mechanisms of action. To this end, we examined the effects of two molecules, cuminaldehyde (a natural compound) and tobramycin (an antibiotic), individually and in combination, against staphylococcal biofilm. Our experimental observations demonstrated that cuminaldehyde (20μg/mL) in combination with tobramycin (0.05μg/mL) exhibited efficient reduction in biofilm formation compared to their individual treatments (p < 0.01). Additionally, the combination showed an additive interaction (fractional inhibitory concentration value 0.66) against S. aureus. Further analysis revealed that the effective combination accelerated the buildup of reactive oxygen species (ROS) and increased the membrane permeability of the bacteria. Our findings also specified that the cuminaldehyde in combination with tobramycin efficiently reduced biofilm-associated pathogenicity factors of S. aureus, including fibrinogen clumping ability, hemolysis property, and staphyloxanthin production. The selected concentrations of tobramycin and cuminaldehyde demonstrated promising activity against the biofilm development of S. aureus on catheter models without exerting antimicrobial effects. In conclusion, the combination of tobramycin and cuminaldehyde presented a successful strategy for combating staphylococcal biofilm-related healthcare threats. This combinatorial approach holds the potential for controlling biofilm-associated infections caused by S. aureus.

1,25 dihydroxyvitamin D3-mediated effects on bovine innate immunity and on biofilm-forming Staphylococcus spp. isolated from cattle with mastitis

Mastitis is one the most widespread and serious diseases in dairy cattle. Recurrent and chronic infections are often attributable to certain pathogenicity mechanisms in mastitis-causing pathogens such as Staphylococcus spp. These include growing in biofilm and invading cells, both of which make it possible to resist or evade antimicrobial therapies and the host’s immune system. This study tested the effects of active vitamin D3 (i.e., calcitriol or 1,25-dihydroxyvitamin D3) on the internalization and phagocytosis of biofilm-forming Staphylococcus spp. isolated from animals with mastitis. Two established bovine cell lines were used: MAC-T (mammary epithelial cells) and BoMac (macrophages). Calcitriol (0–200 nM) did not affect the viability of MAC-T cells nor that of BoMac cells after 24 and 72 h. Concentrations of 0–100 mM for 24 h upregulated the expression of 24-hydroxylase in MAC-T cells, but did not alter that of VDR. Pre-treatment of the cells with calcitriol for 24 h decreased the internalization of S. aureus V329 into MAC-T cells (0–100 nM), and stimulated the phagocytosis of the same strain and of S. xylosus 4913 (0–10 nM). Calcitriol and two conditioned media, obtained by treating the cells with 25–200 nM of the metabolite for 24 h, were also assessed in terms of their antimicrobial and antibiofilm activity. Neither calcitriol by itself nor the conditioned media affected staphylococcal growth or biofilm formation (0–200 nM for 12 and 24 h, respectively). In contrast, the conditioned media (0–100 nM for 24 h) decreased the biomass of preformed non-aureus staphylococcal biofilms and killed the bacteria within them, without affecting metabolic activity. These effects may be mediated by reactive oxygen species and proteins with antimicrobial and/or antibiofilm activity. In short, calcitriol could make pathogens more accessible to antimicrobial therapies and enhance bacterial clearance by professional phagocytes. Moreover, it may modulate the host’s endogenous defenses in the bovine udder and help combat preformed non-aureus staphylococcal biofilms (S. chromogenes 40, S. xylosus 4913, and/or S. haemolyticus 6). The findings confirm calcitriol’s potential as an adjuvant to prevent and/or treat intramammary infections caused by Staphylococcus spp., which would in turn contribute to reducing antibiotic use on dairy farms.

Surface Topography, Microbial Adhesion, and Immune Responses in Silicone Mammary Implant-Associated Capsular Fibrosis.

Breast cancer is the most common cancer in women globally, often necessitating mastectomy and subsequent breast reconstruction. Silicone mammary implants (SMIs) play a pivotal role in breast reconstruction, yet their interaction with the host immune system and microbiome remains poorly understood. This study investigates the impact of SMI surface topography on host antimicrobial responses, wound proteome dynamics, and microbial colonization. Biological samples were collected from ten human patients undergoing breast reconstruction with SMIs. Mass spectrometry profiles were analyzed for acute and chronic wound proteomes, revealing a nuanced interplay between topography and antimicrobial response proteins. 16S rRNA sequencing assessed microbiome dynamics, unveiling topography-specific variations in microbial composition. Surface topography alterations influenced wound proteome composition. Microbiome analysis revealed heightened diversity around rougher SMIs, emphasizing topography-dependent microbial invasion. In vitro experiments confirmed staphylococcal adhesion, growth, and biofilm formation on SMI surfaces, with increased texture correlating positively with bacterial colonization. This comprehensive investigation highlights the intricate interplay between SMI topography, wound proteome dynamics, and microbial transmission. The findings contribute to understanding host-microbe interactions on SMI surfaces, essential for optimizing clinical applications and minimizing complications in breast reconstruction.

Poly(d-amino acid) Nanoparticles Target Staphylococcal Growth and Biofilm Disassembly by Interfering with Peptidoglycan Synthesis.

d-Amino acids are signals for biofilm disassembly. However, unexpected metabolic pathways severely attenuate the utilization of d-amino acids in biofilm disassembly, resulting in unsatisfactory efficiency. Herein, three-dimensional poly(d-amino acid) nanoparticles (NPs), which possess the ability to block intracellular metabolism, are constructed with the aim of disassembling the biofilms. The obtained poly(α-N-acryloyl-d-phenylalanine)-block-poly(β-N-acryloyl-d-aminoalanine NPs (denoted as FA NPs) present α-amino groups and α-carboxyl groups of d-aminoalanine on their surface, which guarantees that FA NPs can effectively insert into bacterial peptidoglycan (PG) via the mediation of PG binding protein 4 (PBP4). Subsequently, the FA NPs trigger the detachment of amyloid-like fibers that connect to the PG and reduce the number of polysaccharides and proteins in extracellular polymeric substances (EPS). Finally, FA NPs damage the structural stability of EPS and lead to the disassembly of the biofilm. Based on this feature, FA NPs significantly enhance the killing efficacy of encapsulated sitafloxacin sesquihydrate (Sita) by facilitating the penetration of Sita within the biofilm, achieving complete elimination of Staphylococcal biofilm in mice. Therefore, this study strongly demonstrates that FA NPs can effectively improve biofilm disassembly efficacy and provide great potential for bacterial biofilm infection treatment.

Effect of Potassium Permanganate on Staphylococcal Isolates Derived from the Skin of Patients with Atopic Dermatitis.

In atopic dermatitis (AD), Staphylococcus aureus frequently colonizes lesions, leading to superinfections that can then lead to exacerbations. The presence of biofilm-producing isolates has been associated with worsening of the disease. Potassium permanganate is used as a topical treatment of infected eczema, blistering conditions, and wounds. Little is known of its effects against microbes in AD skin. The aim of this study was to explore antibacterial and antibiofilm properties of potassium permanganate against staphylococcal isolates derived from AD skin. Viable count and radial diffusion assays were used to investigate antibacterial effects of potassium permanganate against planktonic staphylococcal isolates. The antibiofilm effects were assessed using biofilm assays and scanning electron microscopy. The Staphylococcus aureus isolates were completely killed when exposed to 0.05% of potassium permanganate. In concentrations of 0.01%, potassium permanganate inhibited bacterial biofilm formation. Eradication of established staphylococcal biofilm was observed in concentrations of 1%. Electron microscopy revealed dense formations of coccoidal structures in growth control and looser formations of deformed bacteria when exposed to potassium permanganate. This suggests antibacterial and antibiofilm effects of potassium permanganate against staphylococcal isolates derived from AD skin, when tested in vitro, and a potential role in the treatment of superinfected AD skin.

Effectiveness of Ya-Samarn-Phlae in diabetic wound healing: Evidence from in vitro studies and a multicenter randomized controlled clinical trial

Ethnopharmacological relevanceYa-Samarn-Phlae (YaSP) has traditionally been widely used in southern Thailand for treating chronic and infected wounds, including diabetic foot ulcers. However, there are only a limited number of clinical studies supporting the use of this polyherbal formulation. Therefore, the present work aims to provide clinical evidence to support the application of YaSP, prepared according to a standardized traditional procedure (T-YaSP). Additionally, its potential chemical markers and wound healing-related biological activities were examined. Materials and methodsThe in vitro wound healing-related biological activities of YaSP ethanol extract and T-YaSP, including antibacterial activity against Staphylococcus epidermidis, inhibition and eradication of staphylococcal biofilm, anti-inflammatory effects, and enhancement of human dermal fibroblast migration in scratch wounds, were examined using well-established protocols. The chemical profiles of the ethanol extract of YaSP and T-YaSP were compared, and with promising chemical markers, arecoline, alpha-mangostin, and curcumin were selected and quantified using the HPLC method. A prospective, multicenter, randomized, controlled, parallel-group study was conducted over 12 weeks to evaluate the efficacy of the YaSP solution as an adjunct therapy, combined with standard wound care, for diabetic ulcers compared to standard treatment. ResultsThe YaSP extract reduces NO production and can scavenge NO radicals in LPS-induced RAW 264.7 macrophage cells. Additionally, in a scratch assay, this extract and one of its herbal components, Curcuma longa, enhance the migration of human dermal fibroblasts. T-YaSP, containing 2.412 ± 0.002 mg/g of arecoline, 2.399 ± 0.005 mg/g of curcumin, and 0.017 ± 0.000 mg/g of α-mangostin, has shown the ability to inhibit the development and eradicate the mature biofilm of S. epidermidis. The use of T-YaSP as an adjunct therapy led to a significantly higher proportion of patients achieving healing within six weeks compared to the standard treatment group (36%/9 patients vs. 4%/1 patient; p = 0.013). After 12 weeks, 19 out of 25 patients in the T-YaSP group experienced complete healing, whereas only four patients in the standard treatment group achieved complete wound healing (76% in the T-YaSP group vs. 16% in the control group; p < 0.001). ConclusionThe results presented here represent the first randomized controlled trial to demonstrate the effectiveness of the traditional polyherbal solution, T-YaSP, which exhibits a wide range of wound healing-related activities. Utilizing T-YaSP as an adjunctive treatment resulted in a significant improvement in the number of type 2 diabetic patients achieving complete healing. However, to explore and utilize YaSP further, conducting a double-blind, randomized controlled trial with a larger population is necessary.

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. However, little is known about the form DNA takes in the extracellular space, and how different non-canonical DNA structures such as Z-DNA or G-quadruplexes might contribute to its function in the biofilm. The aim of this study was to determine if non-canonical DNA structures form in eDNA-rich staphylococcal biofilms, and if these structures protect the biofilm from degradation by nucleases. We grew Staphylococcus epidermidis biofilms in laboratory media supplemented with hemin and NaCl to stabilize secondary DNA structures and visualized their location by immunolabelling and fluorescence microscopy. We furthermore visualized the macroscopic biofilm structure by optical coherence tomography. We developed assays to quantify degradation of Z-DNA and G-quadruplex DNA oligos by different nucleases, and subsequently investigated how these enzymes affected eDNA in the biofilms. Z-DNA and G-quadruplex DNA were abundant in the biofilm matrix, and were often present in a web-like structures. In vitro, the structures did not form in the absence of NaCl or mechanical shaking during biofilm growth, or in bacterial strains deficient in eDNA or exopolysaccharide production. We thus infer that eDNA and polysaccharides interact, leading to non-canonical DNA structures under mechanical stress when stabilized by salt. We also confirmed that G-quadruplex DNA and Z-DNA was present in biofilms from infected implants in a murine implant-associated osteomyelitis model. Mammalian DNase I lacked activity against Z-DNA and G-quadruplex DNA, while Micrococcal nuclease could degrade G-quadruplex DNA and S1 Aspergillus nuclease could degrade Z-DNA. Micrococcal nuclease, which originates from Staphylococcus aureus, may thus be key for dispersal of biofilm in staphylococci. In addition to its structural role, we show for the first time that the eDNA in biofilms forms a DNAzyme with peroxidase-like activity in the presence of hemin. While peroxidases are part of host defenses against pathogens, we now show that biofilms can possess intrinsic peroxidase activity in the extracellular matrix.

Synergistic use of anti-inflammatory ketorolac and gentamicin to target staphylococcal biofilms

BackgroundWhile antibiotics remain our primary tools against microbial infection, increasing antibiotic resistance (inherent and acquired) is a major detriment to their efficacy. A practical approach to maintaining or reversing the efficacy of antibiotics is the use of other commonly used therapeutics, which show synergistic antibacterial action with antibiotics. Here, we investigated the extent of antibacterial synergy between the antibiotic gentamicin and the anti-inflammatory ketorolac regarding the dynamics of biofilm growth, the rate of acquired resistance, and the possible mechanism of synergy.MethodsControl (ATCC 12600, ATCC 35984) and clinical strains (L1101, L1116) of Staphylococcus aureus and Staphylococcus epidermidis with varying antibiotic susceptibility profiles were used in this study to simulate implant-material associated low-risk and high-risk biofilms in vitro. The synergistic action of gentamicin sulfate (GS) and ketorolac tromethamine (KT), against planktonic staphylococcal strains were determined using the fractional inhibitory concentration measurement assay. Nascent (6 h) and established (24 h) biofilms were grown on 316L stainless steel plates and the synergistic biofilm eradication activity was determined and characterized using adherent bacteria count, minimum biofilm eradication concentration (MBEC) measurement for GS, visualization by live/dead imaging, scanning electron microscopy, gene expression of biofilm-associated genes, and bacterial membrane fluidity assessment.ResultsGentamicin-ketorolac (GS-KT) combination demonstrated synergistic antibacterial action against planktonic Staphylococci. Control and clinical strains showed distinct biofilm growth dynamics and an increase in biofilm maturity was shown to confer further resistance to gentamicin for both ‘low-risk’ and ‘high-risk’ biofilms. The addition of ketorolac enhanced the antibiofilm activity of gentamicin against acquired resistance in staphylococcal biofilms. Mechanistic studies revealed that the synergistic action of gentamicin–ketorolac interferes with biofilm morphology and subverts bacterial stress response altering bacterial physiology, membrane dynamics, and biofilm properties.ConclusionThe results of this study have a significant impact on the local administration of antibiotics and other therapeutic agents commonly used in the prevention and treatment of orthopaedic infections. Further, these results warrant the study of synergy for the concurrent or sequential administration of non-antibiotic drugs for antimicrobial effect.

A trinuclear Zn (II) schiff base dicyanamide complex attenuates bacterial biofilm formation by ROS generation and membrane damage and exhibits anticancer activity

A trinuclear Zn (II) complex, [(ZnL{N(CN)2})2Zn], termed complex 1 has been synthesized by the reaction of an aqueous solution of sodium dicyanamide to the methanolic solution of Zn (CH3COO)2, 2H2O and corresponding Schiff base (H2L) which is derived from 1:2 condensation of 1, 4 butane diamine with 3-ethoxy salicylaldehyde. Complex 1 is characterized by elemental analysis, IR, UV and Single X-ray diffraction study. Drug resistance is a growing global public health concern that has prompted researchers to look into advanced alternative treatment modalities. In this context, complex 1 has shown promising antibacterial and antibiofilm efficacy against gram-positive Staphylococcus aureus and Methicillin-resistant Staphylococcus aureus strains. Complex 1 attenuated Staphylococcal biofilm formation by reducing several virulence factors including the formation of extracellular polysaccharide matrix, slime, haemolysin, staphyloxanthin, auto-aggregation, cell surface hydrophobicity, and motility. Notably, complex 1 mechanistically potentiated Reactive Oxygen Species (ROS) generation within the bacterial cells, leading to the damage of bacterial cell membrane followed by DNA leakage and thereby impeding the growth of Staphylococcus aureus. Furthermore, complex 1 significantly exhibited anticancer activity by reducing the growth of prostate adenocarcinoma cells. It obstructed the migration of cancer cells by potentiating apoptosis and arresting the cell cycle at the G2/M phase. In summary, complex 1 could act as a potent candidate for the generation of novel antibacterial, antibiofilm as well as anticancer treatment regimens for the management of drug-resistant biofilm-mediated Staphylococcus aureus infection and lethal prostate malignancy.

Non-biofilm-forming Staphylococcus epidermidis planktonic cell supernatant induces alterations in osteoblast biological function

Staphylococcal biofilms significantly contribute to prosthetic joint infection (PJI). However, 40% of S. epidermidis PJI isolates do not produce biofilms, which does not explain the role of biofilms in these cases. We studied whether the supernatant from planktonic S. epidermidis alters osteoblast function. Non-biofilm-forming S. epidermidis supernatants (PJI− clinical isolate, healthy skin isolate (HS), and ATCC12228 reference strain) and biofilm-forming supernatants (PJI+ clinical isolate, ATCC35984 reference strain, and Staphylococcus aureus USA300 reference strain) were included. Osteoblasts stimulated with supernatants from non-biofilm-forming isolates for 3, 7, and 14 days showed significantly reduced cellular DNA content compared with unstimulated osteoblasts, and apoptosis was induced in these osteoblasts. Similar results were obtained for biofilm-forming isolates, but with a greater reduction in DNA content and higher apoptosis. Alkaline phosphatase activity and mineralization were significantly reduced in osteoblasts treated with supernatants from non-biofilm-forming isolates compared to the control at the same time points. However, the supernatants from biofilm-forming isolates had a greater effect than those from non-biofilm-forming isolates. A significant decrease in the expression of ATF4, RUNX2, ALP, SPARC, and BGLAP, and a significant increase in RANK-L expression were observed in osteoblasts treated with both supernatants. These results demonstrate that the supernatants of the S. epidermidis isolate from the PJI− and HS (commensal) with a non-biofilm-forming phenotype alter the function of osteoblasts (apoptosis induction, failure of cell differentiation, activation of osteoblasts, and induction of bone resorption), similar to biofilm-forming isolates (PJI+, ATCC35984, and S. aureus USA300), suggesting that biofilm status contributes to impaired osteoblast function and that the planktonic state can do so independently of biofilm production.

Unravelling staphylococcal small-colony variants in cardiac implantable electronic device infections: clinical characteristics, management, and genomic insights.

Staphylococcal small-colony variants (SCVs) are common in cardiac implantable electronic device (CIED) infections. This is the first retrospective and multi-case study on CIED infections due to staphylococcal SCVs, aiming to provide a theoretical basis for the clinical management of CIED and device-related infections caused by staphylococcal SCVs. Ninety patients with culture positive CIED infections were enrolled between 2021 and 2022. We compared the demographic and clinical characteristics of patients with and without SCVs and performed genomic studies on SCVs isolates. Compared to patients without SCVs, those with SCVs had a longer primary pacemaker implantation time and were more likely to have a history of device replacement and infection. They showed upregulated inflammatory indicators, especially higher NEUT% (52.6 vs. 26.8%, P = 0.032) and they had longer hospital stays (median 13 vs. 12 days, P = 0.012). Comparative genomics analysis was performed on Staphylococcus epidermidis wild-type and SCVs. Some genes were identified, including aap, genes encoding adhesin, CHAP domain-containing protein, LPXTG cell wall anchor domain-containing protein, and YSIRK-type signal peptide-containing protein. Staphylococcal SCVs affect the clinical characteristics of CIED infections. The process of staphylococcal SCVs adherence, biofilm formation, and interaction with neutrophils play a vital role.

Comparison of Staphylococcal Virulence Factors, Biofilm Formation and Antibiotic Susceptibility Test Among Staphylococcus aureus Strains Isolated from Various Clinical Specimens

Comparison of Staphylococcal Virulence Factors, Biofilm Formation and Antibiotic Susceptibility Test Among Staphylococcus aureus Strains Isolated from Various Clinical Specimens

Enzyme-responsive nanoparticles: enhancing the ability of endolysins to eradicate Staphylococcus aureus biofilm.

Stimuli-responsive nanomaterials show promise in eradicating Staphylococcus aureus biofilm from implants. Peptidoglycan hydrolases (PGHs) are cationic antimicrobials that can be bioengineered to improve the targeting of persisters and drug-resistant bacteria. However, these molecules can be degraded before reaching the target and/or present limited efficacy against biofilm. Therefore, there is an urgent need to improve their potency. Herein, PGH-polyphosphate nanoparticles (PGH-PP NPs) are formed by ionotropic gelation between cationic PGHs and anionic polyphosphate, with the aim of protecting PHGs and delivering them at the target site triggered by alkaline phosphatase (AP) from S. aureus biofilm. Optimized conditions for obtaining M23-PP NPs and GH15-PP NPs are presented. Size, zeta potential, and transmission electron microscopy imaging confirm the nanoscale size. The system demonstrates outstanding performance, as evidenced by a dramatic reduction in PGHs' minimum inhibitory concentration and minimum bactericidal concentration, together with protection against proteolytic effects, storage stability, and cytotoxicity towards the Caco-2 and HeLa cell lines. Time-kill experiments show the great potential of these negatively charged delivery systems in overcoming the staphylococcal biofilm barrier. Efficacy under conditions inhibiting AP proves the enzyme-triggered delivery of PGHs. The enzyme-responsive PGH-PP NPs significantly enhance the effectiveness of PGHs against bacteria residing in biofilm, offering a promising strategy for eradicating S. aureus biofilm.

Staphylococcal superantigen-like protein 10 enhances the amyloidogenic biofilm formation in Staphylococcus aureus

Staphylococcus aureus is a highly infectious pathogen that represents a significant burden on the current healthcare system. Bacterial attachment to medical implants and host tissue, and the establishment of a mature biofilm, play an important role in chronic diseases such as endocarditis, osteomyelitis and wound infections. These biofilms decrease bacterial susceptibility to antibiotics and immune defences, making the infections challenging to treatment. S. aureus produces numerous exotoxins that contribute to the pathogenesis of the bacteria. In this study, we have identified a novel function of staphylococcal superantigen-like protein 10 (SSL10) in enhancing the formation of staphylococcal biofilms. Biofilm biomass is significantly increased when SSL10 is added exogenously to bacterial cultures, whereas SSL2 and SSL12 are found to be less active. Exogenously added SSL10 mask the surface charge of the bacterial cells and lowers their zeta potential, leading to the aggregation of the cells. Moreover, the biofilm formation by SSL10 is governed by amyloid aggregation, as evident from spectroscopic and microscopic studies. These findings thereby give the first overview of the SSL-mediated amyloid-based biofilm formation and further drive the future research in identifying potential molecules for developing new antibacterial therapies against Staphylococcus aureus.