Background: The periodontal pathogen Tannerella forsythia is auxotrophic for muramic acid (MurNAc), a key component of bacterial peptidoglycan, and dependent on an external supply of MurNAc to maintain pure laboratory cultures. The focus of this study was to find a source of muramic acid and peptidoglycan fragments from a Staphylococcus aureus strain. This would facilitate the isolation of T. forsythia by incorporating peptidoglycan into conventional anaerobic media. Methods: The S. aureus strain ATCC 29213 was chosen as the source. The standardization and quantification of the method included verifying concentrations via spectrophotometry and developing a linear regression model with standard curves for muramic acid and lactic acid. The resulting lysate was used to seed Fastidious Anaerobe Agar (FAA) plates, which were inoculated with strain T. forsythia (ATCC 43037) and incubated in an anaerobic chamber for seven days. Results: The resulting lysate had an optical density ranging from 0.061 to 0.083, which corresponds to a muramic acid concentration of approximately 12 µg/mL. Pure cultures of T. forsythia could then be obtained on FAA plates supplemented with muramic acid (MurNAc) (FAA-Mur). The viability of the axenic T. forsythia culture was confirmed using muramic acid/peptidoglycan fragments of microbial origin. Conclusions: The method presented improves the growth of T. forsythia. Consequently, T. forsythia is available for further investigation into the regular performance of sensitivity tests in periodontics and the routine generation of growth curves for quantitative polymerase chain reaction (qPCR) analysis.

Anaerobic Pyrolysis Chamber: An atmosphere-controlled furnace for controlled-atmosphere thermal decomposition

BackgroundVolatile anesthetics are gaining recognition for their benefits in long-term sedation of mechanically ventilated patients with bacterial pneumonia and acute respiratory distress syndrome. In addition to their sedative role, they also exhibit anti-bacterial and anti-inflammatory properties, though the mechanisms behind these effects remain only partially understood. In vitro studies examining the prolonged impact of volatile anesthetics on bacterial growth, inflammatory cytokine response, and surfactant proteins — key to maintaining lung homeostasis — are still lacking.MethodsUsing an anaerobic chamber setup, we evaluated the effects of the most commonly used volatile anesthetics, Sevoflurane and Desflurane, at clinically relevant concentrations on the growth of Pseudomonas aeruginosa, Escherichia coli, and Staphylococcus aureus. Bacterial growth was monitored over 24 h, assessing OD600, CFU/ml, and growth rate during the log phase. In the same setup, but with aerobic conditions, we investigated the immunomodulatory properties of both anesthetics on human A549 cells, either with or without bacterial lipopolysaccharide (LPS, 1 µg/ml) stimulation. Over 48 h, we analyzed pro-inflammatory chemokine release using ELISA and assessed surfactant protein expression with Western blot analysis.ResultsSevoflurane and Desflurane significantly reduced Pseudomonas aeruginosa growth as expressed consistently in OD600 and CFU/ml starting after 12 h. Both volatile anesthetics also significantly reduced Staphylococcus aureus OD600 starting after 21 h. Sevoflurane (p < 0.01) and Desflurane (p < 0.001) counteracted LPS-induced interleukin-8 release by A549 cells after 48 h and significantly ( p < 0.01 and p < 0.05) enhanced the expression of the propeptide of surfactant protein C after 24 h.ConclusionsProlonged anti-bacterial and anti-inflammatory effects of Sevoflurane and Desflurane include both the reduction of Pseudomonas aeruginosa and Staphylococcus aureus growth as well as the inhibition of LPS-induced chemokine release by A549 epithelial cells paralleled by an increase of surfactant protein expression. These effects highlight the potential of volatile anesthetics beyond sedation in supporting lung function in ventilated patients with respiratory failure.Supplementary InformationThe online version contains supplementary material available at 10.1186/s12890-025-03849-w.

The human microbiome comprises diverse and dynamic microbial communities that play essential roles in host health. Understanding these communities and their responses to environmental factors is critical for advancing microbiome-based therapeutics. Traditional in vitro models for cultivating human-derived microbiota often lack scalability and require extensive technical expertise, limiting their accessibility and throughput. To address these limitations, we developed the Minibioreactor Array (MBRA) system -- a modular, single-stage, continuous-flow platform for high-throughput cultivation of microbial communities. This system enables parallel cultivation of up to 48 distinct microbial communities, supporting experimental flexibility while maintaining the stable growth of complex ecosystems. This protocol provides detailed guidance on MBRA fabrication, assembly, sterilization, and operation. The system's modular design allows for easy integration into anaerobic chambers and supports customization for a wide range of experimental applications. It has been used to study microbial responses to antibiotics, dietary compounds, and pathogen invasion, and to screen for pathogen-resistant communities. With its accessibility, scalability, and reproducibility, the MBRA represents a powerful model system for investigating microbial interactions and advancing microbiome research.

Using anaerobic cultivation chambers to decipher geomicrobiological processes

Aim of the study: Traditional Chinese herbs have a unique therapeutic effect on stroke and numerous successful clinical cases. However, these clinical cases are highly dispersed, creating challenges for translational research. This study employs a new paradigm to identify treatment patterns and the active compound interactions contained within these clinical cases, with experimental validation after target screening. Methods and Materials: Stroke-related targets were identified through GEO, DisGeNET, and Genecards. Active ingredients were extracted from BATMAN-TCM 2.0. All herbs and diseases were confirmed by the Pharmacopoeia of the People's Republic of China (2020 edition) and Medical Subject Heading (MeSH). All networks in this study were constructed by Cytoscape, and data analysis was done by Python. All formulations and herbs were retrieved from the literature review. For the molecular docking process, Autodock was applied as the docking platform, and all the protein structures were downloaded from PDB. For experimental validation after target screening, HT22 cells were incubated with glucose-free DMEM and placed in an anaerobic chamber for 2 h. Subsequently, HT22 cells were reoxygenated for 24 h. Estrogen Receptor 1 (ESR1) protein levels were measured in vitro. Results: seven materials, including Angelicae Sinensis Radix, Pheretima, Chuanxiong Rhizoma, Persicae Semen, Astragali Radix, Carthami Flos, and Radix Paeoniae Rubra, were identified as the core herbs for the treatment of stroke. The targets of the stroke mechanism were screened, and the herbs-compound-target network was constructed. Among them, paeoniflorin (PF) was identified as the core active compound, and its interaction with ESR1 was verified by molecular docking as the key interaction for the treatment of stroke. In vitro experiments showed that PF inhibited cell apoptosis under hypoxia by increasing the expression of ESR1 compared with the oxygen-glucose deprivation-reperfusion (OGD/R) model group. Western showed that PF (100 μM, 200 μM) can significantly increase the decreased ESR1 protein level caused by the OGD/R model. Conclusions: seven key herbs were screened. Further bioinformatics and network pharmacology studies suggested that PF is expected to become a new active compound for the treatment of stroke. In vitro validation further demonstrated that PF enhanced neuronal survival and ESR1 expression under ischemic conditions, supporting its therapeutic candidacy.

Fecal microbiota transplantation (FMT) has been used as a treatment option for horses (Equus caballus) with gastrointestinal diseases. Several preparation and conservation protocols to improve bacterial survival have been studied in other species. This study aimed to evaluate the impact of oxygen exposure and different protectant solutions on bacterial viability before and after freezing using horse feces. Fecal samples from 10 healthy horses were aliquoted and diluted in cryoprotectant solutions containing antioxidants (n = 40) or 10% glycerol (n = 40). Half of the aliquots from each dilution condition were prepared inside an anaerobic chamber, while the other half were prepared under ambient air conditions. Each sample was also analyzed fresh and after freezing at -20 °C for 90 days. Bacterial viability was assessed using flow cytometry. A mixed linear model and the Friedman and Wilcoxon tests were used depending on data distribution. Freeze-thawing decreased bacterial viability by 47% (mean ± SD: 51 ± 27% before, 27 ± 8% after; p < 0.001). Glycerol was superior to the cryoprotectant after freezing (32 ± 8% glycerol, 24 ± 8% cryoprotectant; p < 0.001). Oxygen exposure did not affect viability (p = 0.13). There was no statistical difference between protectant solutions in fresh samples (p = 0.16). Fresh FMT solutions may be better for treating horses with dysbiosis, but if freezing cannot be avoided, glycerol should be used to dilute feces.

Background: Microplastics (MPs) are small plastic fragments with diameters less than 5 mm in size and are prevalent in everyday essentials and consumables. Large global plastic production has now led to a flooding of MPs in our natural environment. Due to their detrimental impacts on the planet's ecosystems and potentially our health, MPs have emerged as a significant public health concern. In this pilot study, we hypothesize that MPs exposure will negatively affect gut microbiota composition and function, in which metabolic reprogramming plays an important role. Methods: Using in vitro experiments, three bacterial strains (Escherichia coli MG1655, Nissle 1917, and Lactobacillus rhamnosus) were selected to investigate the impacts of MPs exposure. The bacterial strains were individually cultured in an anaerobic chamber and exposed to 1 µm polystyrene MPs at various concentrations (0, 10, 20, 50, 100, and 500 µg/mL) in the culture medium. Results: MPs exposure reduced the growth of all three bacterial strains in a dose-dependent manner. Liquid chromatography mass spectrometry (LC-MS)-based untargeted metabolomics revealed significant differences in multiple metabolic pathways, such as sulfur metabolism and amino sugar and nucleotide sugar metabolism. In addition, we extracted gut microbiota from C57BL/6 mice, and 16S rRNA sequencing results showed a significant upregulation of Lactobacillales and a significant reduction in Erysipelotrichales due to MPs exposure. Furthermore, targeted and untargeted metabolomics corroborated the in vitro results and revealed alterations in microbial tryptophan metabolism and energy producing pathways, such as glycolysis/gluconeogenesis and the pentose phosphate pathway. Conclusions: These findings provide evidence that MPs exposure causes comprehensive changes to healthy gut microbiota, which may also provide insights into the mechanistic effects of MPs exposure in humans.

Introduction:Platelets are discoidal and anuclear structures which are formed in bone marrow from megakaryocytes. They play a major role in the release of growth factors at the site of injury to initiate wound healing. Platelet rich fibrin which belongs to second generation of platelet concentrates has an important role in hard and soft tissue regeneration.Aim:This research aims to determine advanced PRF (A-PRF) as a medium for prolonged release of antibiotics locally and its effects on Porphyromonas gingivalis.Materials and Methods:This in vitro study consisted of four groups, where for each group 10 ml of blood was collected from healthy volunteers after obtaining informed consent. Before centrifugation, to each of the tubes, one among the three antibiotics, such as 0.5 ml amoxicillin, 0.5 ml of metronidazole, and 0.5 ml of azithromycin was added. The fourth tube had 0.5 ml of saline solution, which served as the control. Tubes were centrifuged at 1500 rpm for 14 min at room temperature. A-PRF membrane containing different antibiotics and control was placed on the blood agar plates containing four wells. These plates were then transferred to the anaerobic chamber at 37°C for 7 days. Antimicrobial activity of the A-PRF was assessed at 24 h, 48 h, 72 h, and 96 h.Results:This study compared the antibacterial action of A-PRF against P. gingivalis at the 24 h, 48 h, 72 h, and 96 h. A-PRF with amoxicillin at 24 h showed less anti-P. gingivalis activity, whereas maximum activity was noted at 72 h and 96 h.Conclusion:Results showed that among antibiotic solutions with A-PRF, metronidazole had more antimicrobial efficacy compared to amoxicillin and azithromycin against P. gingivalis.

Background: Reduction of Faecalibacterium prausnitzii abundance is related to inflammatory bowel diseases (IBDs), and supplement of it exists protective effects. Aim: This study aimed to establish a F. prausnitzii-colonized mouse model and investigate that the presence of F. prausnitzii in the gut can ameliorate the severity of dextran sulfate sodium (DSS)-induced colitis. Methods: A F. prausnitzii (ATCC 27768) strain was maintained on the PS-BHI agar plates and manipulated in a strictly anaerobic chamber. A F. prausnitzii-colonized C57BL/6 mice model was tested by a rectal enema with 1 × 109 bacteria/day for 3 days. The 5% DSS was added to drinking water for 3 days to induce colitis and diarrhea in experimental mice. The clinical, cytological, and histological severities were compared between groups. Results: The F. prausnitzii-colonized mice model was successfully established via rectal enema with the property of transfer to offspring. DSS treatment altered gut microbiota and significantly attenuated the abundance of F. prausnitzii in colonized mice. Mice with F. prausnitzii colonization had significantly improved weight loss, anal bleeding, stool consistency, cecum weight, colon length, and serum amyloid A (SAA) level than those without after DSS treatment. Furthermore, the F. prausnitzii-colonized mice significantly reduced the transcription levels of TNF-α, INF-γ and IL-18, and epithelial damage and PMN infiltration in the lamina propria and had better preservation of goblet cells than the control group. Conclusion: We have successfully established a mouse model colonized with F. prausnitzii via rectal enema administration and showed colonization of F. prausnitzii in the gut has a protective effect against DSS-induced colitis.

Understanding abiotic carbon fixation provides insights into early Earth’s carbon cycles and life’s emergence in terrestrial hot springs, where iron sulfide (FeS), similar to cofactors in metabolic enzymes, may catalyze prebiotic synthesis. However, the role of FeS-mediated carbon fixation in such conditions remains underexplored. Here, we investigate the catalytic behaviors of FeS (pure and doped with Ti, Ni, Mn, and Co), which are capable of H2-driven CO2 reduction to methanol under simulated hot spring vapor-zone conditions, using an anaerobic flow chamber connected to a gas chromatograph. Specifically, Mn-doped FeS increases methanol production five-fold at 120 °C, with UV−visible light (300–720 nm) and UV-enhanced light (200–600 nm) further increasing this activity. Operando and theoretical investigations indicate the mechanism involves a reverse water-gas shift with CO as an intermediate. These findings highlight the potential of FeS-catalyzed carbon fixation in early Earth’s terrestrial hot springs, effective with or without UV light.

Fate of arsenic in contaminated coastal soil induced by rising temperature and seawater intrusion

This study investigates the response of biofilm characteristics to variations in fluid depth and their influence on the corrosion behavior of carbon steel (C1020) under low-flow fluid conditions, utilizing Desulfovibrio vulgaris. The experiments were conducted in an anaerobic chamber at 30 °C, utilizing modified Baar's medium as the testing medium. The findings reveal that fluid depth significantly impacts biofilm-corrosion product composite formation, with deeper depths promoting thicker and more heterogeneous biofilm-corrosion product layer compared to shallower depths, where a thinner and more uniform biofilm-corrosion product layer is observed. Moreover, the characteristics of initially attached biofilms was verified as the primary factor affecting subsequent corrosion behavior during prolonged exposure. Corrosion analysis reveals that greater fluid depth leads to increased weight loss (91 ± 13.2 mg/cm2) and deeper pit depths (540 ± 69 μm), surpassing those observed in shallower test media (21 ± 2.3 mg/cm2 and 105 ± 17 μm) after 28 days of exposure. The corrosion products within the biofilm were predominantly FeS and Fe3(PO4)2·8H2O. A direct relationship was observed between the thickness of this biofilm-corrosion product layer and the progression of pit depth, suggesting a strong correlation between carbon steel corrosion and biofilm development in limited fluid depths (e.g., 5–15 mm). Furthermore, a significant association between the deepest pits (average) and the number of sessile cells within the biofilm underscores the pivotal role of sessile cell numbers in carbon steel corrosion.

Removal of cadmium and zinc by calcium polysulfide in acidic groundwater: Injection ratio and precipitation mechanism

Enhanced removal of PPCPs and antibiotic resistance genes in saline wastewater using a bioelectrochemical-constructed wetland system

Biological methods do not effectively remove pharmaceutical products (PPs) and antibiotic resistance genes (ARGs) from wastewater at low temperatures, leading to environmental pollution. Therefore, anaerobic-aerobic-coupled upflow bioelectrochemical reactors (AO-UBERs) were designed to improve the removal of PPs at low temperatures (10 ± 2 °C). The result shows that diclofenac (DIC) and ibuprofen (IBU) removals in the system with aerobic anodic and anaerobic cathodic chambers were 91.7% and 94.7%, higher than that in the control system (12.2 ± 1.5%, 36.5 ± 5.9%), and aerobic zone favors DIC and IBU removal; fluoroquinolone antibiotics (FQs) removals in the system with aerobic cathodic and anaerobic anodic chambers were 17.5-22.4% higher than that in the control system (9.1-22.4%), and anaerobic zone favors FQs removal. Analysis of microbial community structure and ARGs showed that different electrotrophic microbes (Flavobacterium, Acinetobacter, and Delftia) with cold-resistant ability to degrade PPs were enriched in different electrode combinations, and the aerobic cathodic chambers could remove certain ARGs. These results showed that AO-UBERs under intermittent electrical stimulation mode are an alternative method for the effective removal of PPs and ARGs at low temperatures.

Iron-retrofitted anaerobic baffled reactor system for rural wastewater treatment: Stable performance of nutrients removal with phosphorus recovery and minimal sludge production

The current study was conducted to understand the sole role of graphite as a substrate material in a dual-chambered baffled electroactive wetland (EW) in the treatment of Methyl red dye-containing wastewater. The results obtained were compared with conventional gravel-based unplanted dual-chambered constructed wetlands (CW) at a lab scale. The highest dye decolorisation and COD removal efficiency achieved was 92.88 ± 1.6% and 95.78 ± 4.1%, respectively, in the electro-active wetland. Dissolved oxygen (DO) and pH conditions were appropriately maintained in both the microcosms because of separated aerobic and anaerobic chambers. UV-vis and gas chromatography-mass spectroscopy analysis revealed the production of by-products like 4-amino benzoic and N- N dimethyl phenyl-diamine of MR in microcosms and revealed further mineralisation of by-products in the aerobic zone of electroactive-wetland. Higher root growth of Cicer aerietinum and Vigna radiata was observed in the presence of effluents of baffled electroactive wetlands compared to constructed wetland, indicating a decrease in phytotoxicity. Metagenomic analysis revealed the abundance of potential microbes for MR and organic matter removal from phylum Proteobacteria, Firmicutes, Bacteroidetes, and Euryarchaeota. A batch adsorption study revealed a higher adsorption capability of graphite material in comparison to gravel. Hence, this study demonstrated that graphite is an appropriate substrate in electroactive wetland in facilitating microbial attachments and enhancing dye degradation, in addition to exhibiting superior adsorption quality.

Intermittent electrical stimulation removes mixed antibiotics and associated antibiotic resistance genes at low temperatures

Photosynthetic microbial fuel cells (pMFCs) are hybrid systems that enable simultaneous wastewater treatment under anaerobic conditions and the generation of electricity by utilizing the potential difference in the anaerobic anode chamber and the oxygenated cathode chamber. Dairy wastewater with a concentration of 2000 mg COD/L was treated in the anode of a batch pMFC. In the cathode chamber, Chlorella vulgaris or Arthrospira platensis was cultivated in synthetic medium, and next in diluted effluent from the anode chamber. The highest power density of 91 mW/m2 was generated by the pMFC with the cultivation of Arthrospira platensis. Higher values of dissolved oxygen remained during the dark phase in the cathodic medium with Arthrospira platensis cultivation than with Chlorella vulgaris. This depletion of oxygen significantly decreased voltage generation, which during the light phase increased again to the maximum values. The COD removal achieved in the anodic chamber was 87%. The efficiency of nitrogen removal in the cathode chamber during the cultivation of Arthrospira platensis and Chlorella vulgaris was about 78% and 69%, respectively. The efficiency of phosphorus removal in the cathode chamber with the cultivation of Arthrospira plantensis and Chlorella vulgaris was 58% and 43%, respectively. This study has shown that the introduction of Arthrospira platensis into the cathode chamber is more effective than that of Chlorella vulgaris.