Extracellular Protein Concentration Research Articles

Chemotaxis gene of a bacterium impacts larval settlement and metamorphosis in the marine mussel Mytilus coruscus via c-di-GMP controlling extracellular protein production

Bacterial chemotaxis enhances bacterial adaptation to the environment and is important for biofilm formation. Biofilms play a key role in inducing larval settlement and metamorphosis in many marine invertebrates. However, the specific mechanisms by which bacterial chemotaxis influences larval settlement and metamorphosis in mussels remain unknown. The findings indicate that the absence of the chemotaxis gene cheW resulted in reduced motility of Pseudoalteromonas marina, accompanied by an increase in c-di-GMP content. The ΔcheW strain exhibited a higher capacity for biofilm formation compared to the wild-type strain. The extracellular protein content of the ΔcheW strain exhibited a significant 77% reduction, specifically in the flagellin content. The inducing activity of ΔcheW was reduced by 56% compared to the wild-type strain. This study highlights that the deficiency of the chemotaxis gene cheW inhibited larval settlement and metamorphosis in mussels through c-di-GMP regulation of extracellular protein production. It provides a novel ecological function of bacterial chemotaxis in regulating the larval settlement and metamorphosis of marine invertebrates.

Antifungal Efficacy of Ultrashort β-Peptides against Candida Species: Mechanistic Understanding and Therapeutic Implications.

Candidiasis, a condition spurred by the unchecked proliferation of Candida species, poses a formidable global health threat, particularly in immunocompromised individuals. The emergence of drug-resistant strains complicates management strategies, necessitating novel therapeutic avenues. Antimicrobial peptides (AMPs) have garnered attention for their potent antifungal properties and broad-spectrum activity against Candida species. This study assessed the antifungal effectiveness of ultrashort β-peptides against Candida strains, with a specific focus on peptide P3 (LAU-β3,3-Pip-β2,2-Ac6c-PEA). Our findings showed P3's remarkable fungistatic and fungicidal activities against Candida albicans, exhibiting an MIC of 4 μg/mL, comparable to those of standard antifungal drugs. The MIC value remained unchanged in the presence of ADC and BSA, indicating that serum albumin does not diminish the activity of P3. P3 demonstrates synergistic effects when combined with Fluconazole (FLU), Itraconazole (ITR), and Nystatin (NYS) to the extent that it becomes effective at 0.125, 0.125, and 0.03125 μg/mL, respectively. Concentration versus time-kill kinetics showed its time-dependent activity up to the first 12 h against C. albicans, and later concentration also played a role; indeed, at 24 h the whole culture was sterilized at 8× MIC. Post-antifungal effect assays confirmed prolonged suppression of pathogen growth after the removal of P3 from the media for significant durations. More importantly, P3 inhibits hyphae formation and biofilm development of Candida, outperforming Fluconazole with respect to these properties. Mechanistic insights display P3's potential to disrupt fungal cell membrane integrity and dose-dependent inhibition of ergosterol biosynthesis, essential for fungal cell wall integrity. Using the Bradford assay, it was observed that extracellular protein concentrations increased with higher doses of the compound, thereby validating the effect of P3 on membrane integrity. A comparative gene analysis using RT-PCR showed that P3 downregulates ERG3, ERG11, and HWP1, which are crucial for the survival and pathogenicity of C. albicans. The impact of P3 on ERG11 and ERG3 is more effective than that of Fluconazole. Molecular docking studies revealed strong binding of P3 to various isoforms of lanosterol 14-α-demethylase, a key enzyme in ergosterol synthesis. Furthermore, molecular dynamic simulations validated the stability of the most promising docking complex. Overall, our findings underscore P3's potential as a leading candidate for the development of innovative antifungal therapies, warranting further investigation and optimization.

Different organic carbon sources affect microalgal growth and extracellular polymeric substances synthesis to trigger biomass flocculation process

The addition of organic carbon sources into the culture medium represents an effective strategy for microalgae cultivation. However, its influence on the subsequent flocculation process remains poorly understood. In this study, four representative organic carbon sources were supplemented to microalgal cultures with the aim of evaluating their potential to enhance biomass accumulation, optimize fatty acid composition and improve phototolerance. The results demonstrated that the addition of ethanol, sodium acetate, glucose and fructose to the cultures resulted in extracellular protein contents of 90.9, 125.7, 67.5 and 46.0 mg/g, respectively, compared to 36.8 mg/g observed in the control culture with pure BG-11 medium. The addition of organic carbon sources led to alterations in protein content and the composition ratio of charged groups within the extracellular polymeric substances of microalgae, which in turn influenced the flocculation process. This study provides valuable insights into the selection of appropriate organic carbon sources for promoting microalgal growth and clarifies the flocculation mechanisms under various carbon utilization strategies.

Assessment of Chondrogenic Potential of Human Dermal Fibroblasts with Various Protein Modifications

BACKGROUND: Hyaline cartilage is an avascular tissue that covers surface of large joints. This tissue, with a small number of cells and a high content of extracellular matrix proteins, has limited regenerative capacity. Recovery the articular cartilage remains a relevant and not yet fully resolved issue today. A promising approach is the use of biomedical products containing modified cells. AIM: To investigate the effects of different culture media on the chondrogenic modification of cells and to compare the results. METHODS: Human dermal fibroblasts and rabbit mesenchymal stem cells were modified using chondrogenic differentiation medium StemPro or recombinant TGF-β3 protein. Changes in the expression of genes responsible for chondrogenesis (Acan, Tgfβ3, Col2α1, Comp) were measured using the Real-Time PCR method -2ΔΔCt. RESULTS: It was shown that human dermal fibroblasts are capable of chondrogenic modification using protein media. This cell type is easily accessible for harvesting, does not require special conditions for cultivation, is easily scalable, and can be used for allogeneic transplantation. CONCLUSION: The obtained data can be used in the design of tissue engineering products for the regeneration of hyaline cartilage based on allogeneic dermal fibroblasts.

LC-MS based metabolomics identification of natural metabolites against Fusarium oxysporum.

Fusarium is a soil-borne pathogen that poses a serious threat to the quality and yield of hundreds of crops worldwide, particularly tobacco production. Using metabolomics technology, we investigated natural metabolites from disease-conducting soil (DCS) and disease-suppressing soil (DSS) of tobacco rhizosphere as fungicides to control tobacco Fusarium wilt (TFW), which is mainly caused by Fusarium oxysporum. Furthermore, the antifungal mechanisms of these natural metabolites were preliminarily elucidated through various assessments, including antifungal activity determination, chemotaxis effect tests, PI staining experiments, and measurements of extracellular conductivity and protein content. Metabolomics results showed that the DCS with three different disease grades (G1, G5 and G9 groups) had significantly higher levels of 15, 14 and 233 differential rhizosphere metabolites (DRMs) and significantly lower levels of 72, 152 and 170 DRMs compared to the DSS (G0 group). According to KEGG pathway analysis, these DRMs were found to be enriched in the caffeine metabolism, biosynthesis of phenylpropanoids, galactose metabolism and tyrosine metabolism, etc. Linustatin, scopoletin and phenylpropiolic acid were picked out from these DRMs and found to have suppressive activity against F. oxysporum through correlation analysis and antifungal experiments. The three DRMs showed strong inhibitory effects on the growth and spore germination of F. oxysporum at concentrations of 0.5 mM or higher in each test period. Furthermore, F. oxysporum showed a phobotaxis effect against these three DRMs at concentrations as low as 0.25 mM. Finally, we found that the three DRMs had an inhibitory effect on F. oxysporum by destroying the integrity of the cell membrane and increasing the membrane permeability of F. oxysporum. This study firstly reports the inhibition activity of phenylpropiolic acid and linustatin on F. oxysporum, providing a practical and environmentally friendly method for biocontrol of TFW by using natural fungicides.

Extracellular glypican-1 affects tumor progression and prognosis in esophageal cancer.

Cells are covered with a glycan surface layer that is referred to as the glycocalyx (GCX). It has been reported that the formation of the GCX is promoted on cancer cells and is associated with tumor growth and metastasis. Heparan sulfate proteoglycan glypican-1 (GPC1) is a core protein of the GCX that is overexpressed in esophageal squamous cell carcinoma (ESCC) and is involved in the development and progression of cancer cells. The purpose of the present study is to analyze the utility of GPC1 as a new biomarker ralated to glycocalyx that reflects therapeutic effect and prognosis of ESCC. We measured the concentration of GPC1 protein in preoperative plasma from advanced esophageal cancer patients and examined its relationships with clinicopathological factors and therapeutic efficacy, and the effects of extracellular GPC1 were investigated. The following clinical factors were significantly correlated with the preoperative high GPC1 concentration: male, tumor size ≥30 mm, venous invasion, pT factor ≥2, pStage ≥3, residual tumor, and distant metastatic recurrence. Both overall and recurrence-free survival were significantly worse in the high GPC1 group. Extracellular GPC1 protein concentration reflected intracellular GPC1 expression. Furthermore, we examined the effects of extracellular recombinant human (rh)GPC1 on ESCC cells, and found that extracellular rhGPC1 affects cell motility, including migration and invasion. These results demonstrated the utility of extracellular GPC1 as a biomarker, which can be assayed from a less invasive blood sample-based liquid biopsy. Extracellular GPC1 protein plays a role in both tumor cell motility and cancer progression. Thus, plasma GPC1 is a useful biomarker for esophageal cancer progression and may be a potential candidate of therapeutic target.

Revealing detrimental effects of various DC electrical energy conditions on different multidrug resistant bacteria: a comprehensive study

The arbitrary discharge of contaminated wastes, especially that encompass multidrug resistant microbes (MDR), would broaden the circle of epidemic diseases such as COVID-19, which in turn deteriorate definitely the whole socioeconomics. Therefore, the employment of electrical stimulation techniques such as direct current (DC) with low energy considers being effective tool to impede spontaneous changes in microbial genetic makeup, which increases the prevalence of MDR phenomenon. Herein, the influence of different electric energies generated by DC electric field, volts and time on MDR-bacteria that are categorized among the highly ranked nosocomial pathogens, was scrutinized. Wherein, Pseudomonas aeruginosa, Escherichia coli, Staphylococcus aureus and Enterococcus faecalis were examined as paradigms of Gram-negative and Gram-positive pathogens. The results declared the significant superior antagonizing potency of electric energy in a dose-dependent modality rather than the applied volts or exposure time. Notably, the exposure of bacterial cultures to140 J inhibited the bacterial count by > 78% and the range of 47–73% for Gram-negative and Gram-positive, respectively. While the suppression in their metabolic activity assessed by > 75% and 41–68%, respectively; reflecting the capability of electrical energy to induce viable but non-culturable (VBNC) state. Similarly, the results of total protein, extracellular protein content and lactate dehydrogenase activity emphasized the cell wall deterioration and losing of cell membrane integrity. Additionally, the elevating in ROS upon DC-exposure participated in DNA fragmentation and plasmid decomposability by the range of 33–60%. Further, SEM micrographs depicted drastic morphological deformations after electrical treatment. Strikingly, DC-treatment impaired antibiotic resistance of the examined strains against several antibiotics by > 64.2%. Generally, our comparative detailed study revealed deleterious potentiality of different DC-protocols in defeating microbial pollution, which could be invested as efficient disinfectant alternative in various sectors such as milk sterilization and wastewater purification.

Enhanced biodegradation of benzo[a]pyrene with Trametes versicolor stimulated by citric acid.

Polycyclic aromatic hydrocarbons (PAHs) are a class of persistent organic pollutants with carcinogenic, mutagenic and teratogenic effects. The white-rot fungi in the fungal group have significant degradation ability for high molecular weight organic pollutants. However, exogenous fungi are easily antagonized by indigenous microorganisms. Low molecular weight organic acids, a small molecular organic matter secreted by plants, can provide carbon sources for soil microorganisms. Combining organic acids with white rot fungi may improve the nutritional environment of fungi. In this study, immobilized Trametes versicolor was used to degrade benzo[a]pyrene in soil, and its effect on removing benzo[a]pyrene in soil mediated by different low molecular weight organic acids was investigated. The results showed that when the degradation was 35days, the removal effect of the experimental group with citric acid was the best, reaching 43.7%. The degradation effect of Trametes versicolor on benzo[a]pyrene was further investigated in the liquid medium when citric acid was added, and the effects of citric acid on the biomass, extracellular protein concentration and laccase activity of Trametes versicolor were investigated by controlling different concentrations of citric acid. In general, citric acid can act as a carbon source for Trametes versicolor and promote its extracellular protein secretion and laccase activity, thereby accelerating the mineralization of benzo[a]pyrene by Trametes versicolor. Therefore, citric acid can be used as a biostimulant in the remediation of PAHs contaminated soil with Trametes versicolor.

Adaptation of Ochrobactrum rhizosphaerae IMV B-7956 Bacteria to the Influence of Copper (II) Chloride

In technologically altered habitats, an increased content of organic compounds, nitrogen, phosphorus, sulfur compounds, antibiotic substances, etc. is found. Therefore, microorganisms that are systematically exposed to various stressors have developed adaptation mechanisms. The strain Ochrobactrum rhizosphaerae IMV B-7956, isolated from the infiltrate lakes of the Lviv solid waste landfill, is resistant to copper, chromium, manganese, and iron in concentrations exceeding the maximum permissible concentrations. The work aimed to study the response of O. rhizosphaerae IMV B-7956 cells to CuCl2 exposure by detecting changes in the content of lipid peroxidation products, products of the oxidative modification of proteins, activity of antioxidant defense system enzymes, and synthesis of extracellular polymers. Methods. To study the effect of CuCl2 on prooxidant indicators and the activity of enzymes of the antioxidant defense system, bacteria were pre-incubated in Tris-HCl buffer containing 2–10 mM CuCl2. After one hour of incubation, the bacterial cells were washed and cultured for 1, 12, 24, and 48 h in metal-free media. The copper content in the bacterial cells was determined by atomic absorption spectrometry. The content of lipid peroxidation indicators, carbonyl groups in proteins, total low-molecular-weight thiols, enzymatic activity, and the content of exopolysaccharides and extracellular proteins were determined photometrically. Results. Within an hour, O. rhizosphaerae IMV B-7956 bacteria accumulated 2.3–7.8 mg Cu/g of biomass. Under these conditions, an increased content of lipid peroxidation products was detected. During the first hour of growth in bacterial cells, enzymes with catalase and peroxidase activity were activated. During further cultivation, an increase in the activity of other antioxidant defense enzymes was detected. Carbonyl groups in proteins are probably formed due to an increase in the content of lipid peroxidation products, as they are formed later. Within 12–48 h of growth, the copper content in the bacterial cells decreases. This leads to the restoration of growth. Conclusions. The main damaging effect of CuCl2 on bacterial cells was detected during the first 24 h of growth. Activation of the enzymes of the antioxidant system and synthesis of exopolysaccharides are among the adaptations that ensure the survival of bacteria under these conditions.

Combining metabolomics and transcriptomics to analyze key response metabolites and molecular mechanisms of Aspergillus fumigatus under cadmium stress

The co-cultivation of fungi with microalgae facilitates microalgae harvesting and enhances heavy metal adsorption. However, the mechanisms of fungal tolerance to cadmium (Cd) have not yet been studied in detail. In this study, functional groups of fungi were analyzed under Cd stress using Fourier transform infrared spectrometer (FTIR), X-ray photoelectron spectroscopy (XPS), scanning electron microscope (SEM), and transmission electron microscope (TEM) to explore their morphology. Confocal laser scanning microscope (CLSM) was used to characterize the changes in the content of extracellular polysaccharides and proteins, and a decrease in the ratio of glutathione (GSH) to oxidized glutathione (GSSG) was monitored. The GSH and GSSG contents in mycelium were 7.4 and 7.9 times higher than that in the control, respectively. After 72 h of Cd treatment, the fungal extracellular polysaccharide and extracellular protein contents increased by 16 and 11.4 mg/g, respectively, compared to the control. This provided several functional groups for the complexation of Cd ions to enhance fungal Cd tolerance. The metabolomic and transcriptomic results revealed a total of 358 differential metabolites after 20, 48, and 72 h in the positive and negative ion modes, and the number of differential metabolites specific to each group was 104, 14, and 89, respectively. There were 927, 1167, and 1287 up-regulated genes, and 1301, 1480, and 1683 down-regulated genes at 20, 48, and 72 h, respectively. Energy metabolism, amino acid metabolism, and the ABC transport system are the key metabolic pathways for tolerance enhancement and heavy metal detoxification in fungi. The expression of S-cysteinosuccinic acid was significantly up-regulated after Cd stress and associated with enhanced fungal tolerance and resistance to Cd.

Performance evaluation of typical flocculants for efficient harvesting of Chlorella sorokiniana under different carbon application modes

In this study, the growth characteristics of microalgae cultured with different carbon sources were analyzed, and the flocculation characteristics under the influence of carbon sources were evaluated using three typical flocculants. The results showed that the organic carbon sources could significantly increase the content of extracellular proteins in microalgae. Specifically, the extracellular protein concentrations of microalgae cultured with pure BG-11, ethanol, sodium acetate and glucose were 18.2 29.2, 97.3, and 34.7 mg/g, respectively. During the flocculation process, microalgae cultured with sodium acetate exhibited a weak response to the flocculant because of excessive extracellular proteins inhibited flocculation. In addition, the flocculation efficiency was also less than 50.0% cultured with sodium acetate in all pH test ranges when alum and chitosan were used as flocculants. It could be inferred that the flocculant initially happened to charge neutralization with the negatively charged proteins in the solution and then bridged the charges with the microalgae. These findings provide insights into the effects of different carbon sources on microalgal flocculation, promising organic integration of microalgae wastewater treatment and harvesting.

2-Heptanol inhibits Botrytis cinerea by accelerating amino acid metabolism and retarding membrane transport.

During the postharvest storage of tomatoes, they are susceptible to infection by Botrytis cinerea, leading to significant economic losses. This study evaluated the antifungal potential of 2-heptanol (2-HE), a volatile biogenic compound, against B. cinerea and explored the underlying antifungal mechanism. The results indicated that 2-HE effectively suppressed the growth of B. cinerea mycelia both in vivo and in vitro and stimulated the activities of antioxidative enzymes, including superoxide dismutase (SOD), peroxidase (POD), and catalase (CAT) in tomatoes. Furthermore, 2-HE reduced spore viability, compromised membrane integrity, and resulted in increased levels of extracellular nucleic acids, protein content, and membrane lipid peroxidation. Transcriptomic analysis revealed that 2-HE disrupted the membrane transport system and enhanced amino acid metabolism, which led to intracellular nutrient depletion and subsequent B. cinerea cell death. Additionally, the 2-HE treatment did not negatively impact the appearance or quality of the tomatoes. In conclusion, the findings of this study offer insights into the use of 2-HE as a biocontrol agent in food and agricultural applications.

Sodium acetate increases the productivity of HEK293 cells expressing the ECD-Her1 protein in batch cultures: experimental results and metabolic flux analysis.

Human Embryonic Kidney cells (HEK293) are a popular host for recombinant protein expression and production in the biotechnological industry. This has driven within both, the scientific and the engineering communities, the search for strategies to increase their protein productivity. The present work is inserted into this search exploring the impact of adding sodium acetate (NaAc) into a batch culture of HEK293 cells. We monitored, as a function of time, the cell density, many external metabolites, and the supernatant concentration of the heterologous extra-cellular domain ECD-Her1 protein, a protein used to produce a candidate prostate cancer vaccine. We observed that by adding different concentrations of NaAc (0, 4, 6 and 8mM), the production of ECD-Her1 protein increases consistently with increasing concentration, whereas the carrying capacity of the medium decreases. To understand these results we exploited a combination of experimental and computational techniques. Metabolic Flux Analysis (MFA) was used to infer intracellular metabolic fluxes from the concentration of external metabolites. Moreover, we measured independently the extracellular acidification rate and oxygen consumption rate of the cells. Both approaches support the idea that the addition of NaAc to the culture has a significant impact on the metabolism of the HEK293 cells and that, if properly tuned, enhances the productivity of the heterologous ECD-Her1 protein.

Enhanced bioremediation of organically combined contaminated soil by white rot fungal agent: physiological characteristics and contaminants degradation

ABSTRACT Microbial remediation of organically combined contaminated sites is currently facing technical challenges. White rot fungi possess broad-spectrum degradation capabilities, but most of the studies are conducted on polluted water bodies, and few research focus on the degradation of combined organically contaminated soils. This study aimed to investigate the physiological changes in Trametes versicolor to enhance its simultaneous degradation ability towards benzo(a)pyrene (BaP) and TPH. The results demonstrated that Trametes versicolor, when subjected to liquid fermentation, achieved an 88.08% degradation of individual BaP within 7 days. However, under the combined contamination conditions of BaP and TPH, the BaP degradation rate decreased to 69.25%, while the TPH degradation rate was only 16.95%. Furthermore, the degradation rate of BaP exhibited a significant correlation with the extracellular protein concentration and laccase activities. Conversely, the TPH degradation rate exhibited a significant and positive correlation with the intracellular protein concentration. Solid-state fermentation utilizing fungal agents proved to be the most effective method for removing BaP and TPH, yielding degradation rates of 56.16% and 15.73% respectively within 60 days. Overall, Trametes versicolor demonstrated a commendable capability for degrading combined PAHs-TPH pollutants, thereby providing theoretical insights and technical support for the remediation of organically combined contaminated sites.

Extracellular Vesicle Protein Expression in Doped Bioactive Glasses: Further Insights Applying Anomaly Detection.

Proteomic analysis of extracellular vesicles presents several challenges due to the unique nature of these small membrane-bound structures. Alternative analyses could reveal outcomes hidden from standard statistics to explore and develop potential new biological hypotheses that may have been overlooked during the initial evaluation of the data. An analysis sequence focusing on deviating protein expressions from donors' primary cells was performed, leveraging machine-learning techniques to analyze small datasets, and it has been applied to evaluate extracellular vesicles' protein content gathered from mesenchymal stem cells cultured on bioactive glass discs doped or not with metal ions. The goal was to provide additional opportunities for detecting details between experimental conditions that are not entirely revealed with classic statistical inference, offering further insights regarding the experimental design and assisting the researchers in interpreting the outcomes. The methodology extracted a set of EV-related proteins whose differences between conditions could be partially explainable with statistics, suggesting the presence of other factors involved in the bioactive glasses' interactions with tissues. Outlier identification of extracellular vesicles' protein expression levels related to biomaterial preparation was instrumental in improving the interpretation of the experimental outcomes.

Metabolomics-based analysis of the interaction effects and stress response mechanisms in the co-culture system of fungi and microalgae under cadmium stress

Research on the co-culture of fungi and microalgae to harvest microalgae and enhance the adsorption capacity for heavy metals to reduce contamination is well demonstrated, but the study of the correlation mechanism among critical metabolites, system stability, and stress response in the system under cadmium stress based on metabolomics analyses has not been reported in detail. In this study, the co-culture system of Aspergillus allahabadii and Synechocystis sp. PCC6803 was taken as the object of investigation, and isotope labelling (13C–Na2CO3 and 15N-(NH4)2SO4) and nanoscale secondary ion mass spectrometry were used to confirm the existence of carbon and nitrogen nutrient exchange. The adsorption behavior of the system after Cd treatment was investigated by Fourier Transform Infrared Spectroscopy, X-ray Photoelectron Spectroscopy, and Scanning Electron Microscope/Transmission Electron Microscope. The content of extracellular polysaccharides in the system was increased from 40.8 mg/g to 80.12 mg/g, and the content of extracellular proteins was increased from 22.3 mg/g to 55.68 mg/g after 96 h of Cd treatment. Metabolomics results revealed 235 differential metabolites co-expressed after Cd treatment, with 95, 15, and 77 unique differential metabolites. In particular, the synthesis of extracellular polymeric substances was a factor affecting the stability of the symbiotic system, the up-regulation of the expression of the transporter system was one of the reasons for the improvement of the system's tolerance and resistance to heavy metals; and the nutrient metabolism between the fungus and the microalgae was the main defense mechanism to reduce the extent of the impairment of the system's carbon sequestration capacity by Cd. This provides a theoretical basis for the green and economically friendly application of the symbiotic system in microalgae harvesting, efficient carbon dioxide fixation and reduction of heavy metal pollution.

Antibacterial Activity and Mechanism of Self-Assembly Spermidine-Capped Carbon Dots against Staphylococcus aureus.

This paper investigated the antibacterial mechanism of spermidine-capped carbon dots (S-PCDs) against Staphylococcus aureus. The results showed that there were a large number of amino groups on the surface of S-PCDs and they had a high positive charge (+47.06 mV), which could be adsorbed on the negatively charged bacterial surface through electrostatic interaction and changed the permeability of the bacterial cell membrane. The extracellular protein and nucleic acid contents of S. aureus treated with S-PCDs were 5.4 and 1.2 times higher than those of the control group, respectively. The surface folds and defects of the bacterial cell membrane, and the leakage of cell contents were observed using SEM and TEM. The expression of metabolic oxidation regulatory genes dmpI, narJ and narK was upregulated and the intracellular ROS generation was induced, causing bacterial oxidative stress and eventually bacterial death. S-PCDs can effectively inhibit biofilm formation and had low cytotoxicity. The S-PCD treatment successfully inhibited microbial reproduction when pasteurized milk was stored at 25 °C and 4 °C. These results provide important insights into the antimicrobial mechanism of S-PCDs and lay the foundation for their application in the food field as a potentially novel bacteriostatic nanomaterial.

Regulation of σB-Dependent Biofilm Formation in Staphylococcus aureus through Strain-Specific Signaling Induced by Diosgenin.

Staphylococcus aureus is a commensal skin bacterium and a causative agent of infectious diseases. Biofilm formation in S. aureus is a mechanism that facilitates the emergence of resistant strains. This study proposes a mechanism for the regulation of biofilm formation in S. aureus through strain-specific physiological changes induced by the plant steroid diosgenin. A comparison of diosgenin-induced changes in the expression of regulatory genes associated with physiological changes revealed the intracellular regulatory mechanisms involved in biofilm formation. Diosgenin reduced biofilm formation in S. aureus ATCC 6538 and methicillin-resistant S. aureus (MRSA) CCARM 3090 by 39% and 61%, respectively. Conversely, it increased biofilm formation in S. aureus ATCC 29213 and MRSA CCARM 3820 by 186% and 582%, respectively. Cell surface hydrophobicity and extracellular protein and carbohydrate contents changed in a strain-specific manner in response to biofilm formation. An assessment of the changes in gene expression associated with biofilm formation revealed that diosgenin treatment decreased the expression of icaA and spa and increased the expression of RNAIII, agrA, sarA, and sigB in S. aureus ATCC 6538 and MRSA CCARM 3090; however, contrasting gene expression changes were noted in S. aureus ATCC 29213 and MRSA CCARM 3820. These results suggest that a regulatory mechanism of biofilm formation is that activated sigB expression sequentially increases the expression of sarA, agrA, and RNAIII. This increased RNAIII expression decreases the expression of spa, a surface-associated adhesion factor. An additional regulatory mechanism of biofilm formation is that activated sigB expression decreases the expression of an unknown regulator that increases the expression of icaA. This in turn decreases the expression of icaA, which decreases the synthesis of polysaccharide intercellular adhesins and ultimately inhibits biofilm formation. By assessing strain-specific contrasting regulatory signals induced by diosgenin in S. aureus without gene mutation, this study elucidated the signal transduction mechanisms that regulate biofilm formation based on physiological and gene expression changes.

Biofilm formation and development during the pyrite bioleaching of moderately thermophilic microorganisms

Bioleaching is a process that uses microorganisms to recover metals from intractable metal sulfide ores. Metal sulfide bioleaching is an interfacial process, and biofilm growth is crucial during the early phases. To study the interactions between microbes and minerals during the bioleaching of moderately thermophilic microorganisms, the formation and development of biofilms on pyrite coupons incubated in shake flasks for 40 days were evaluated. The biofilm biomass progressively increased as bioleaching proceeded. The most extensive biofilm was formed on day 30, when bacteria were attached to the mineral surface or submerged in extracellular polymeric substances. Confocal laser scanning microscopy indicated that the biofilm biomass of pyrite on day 40 was 6.55, 5.28, 3.17, and 1.70 times higher than that on days 5, 10, 20, and 30, respectively. Extracellular protein content exhibited an increasing trend throughout the process, whereas extracellular polysaccharides played a significant role during the early stages. Live/dead staining showed the highest mortality rate of 94% when cells were in contact with pyrite in the early stage, and the percentage of live bacteria increased in the later period. In the bioleaching environment, mature biofilms appeared to form relatively quickly after day 20. Real-time quantitative fluorescent quantitative analysis indicated that biofilm formation played an important role in defense against hostile external environments. In this study, biofilm formation and the associated RNA transcript numbers of moderately thermophilic microorganisms during pyrite bioleaching were investigated, contributing to the understanding of cell-mineral interactions.

Effects of flow velocity on biofilm composition and microbial molecular ecological network in reclaimed water distribution systems

The existence of biofilm on the reclaimed water pipeline seriously affects the safety of water distribution. And the flow regimes in the pipeline play a crucial role in the growth of biofilms. In this study, the biofilm composition, surface topography and bacterial community were detected under eight levels of flow velocity in the range of 0.10–1.40 m s−1. The results showed that the dry weight, the concentration of extracellular protein and extracellular polysaccharide in the biofilm reached a dynamic stable period after 640 h. The biofilm composition and surface topography of biofilm were significantly different under the different flow regimes (laminar flow belongs to [0.10, 0.19] m s−1, and turbulent flow belongs to [0.29, 1.40] m s−1). As the flow velocity range increases, the concentration of each component in the biofilm and the parameters of biofilm surface topography increased and then decreased. The flow velocity could be a strong environmental stimulus resulting in the succession of bacterial community in biofilm. As the flow velocity increased from 0.10 m s−1 to 1.40 m s−1, at the phylum level, the average relative abundance of Firmicutes mainly showed a trend of first increasing and then decreasing with the highest abundance value of 71.57% at 0.49 m s−1. The flow velocity increased from 0.10 m s−1 to 0.49 m s−1, a significant increase in microbial diversity could be detected. The increase in flow velocity promoted the proliferation of microorganisms, and the interaction between different microbial components was enhanced. At 0.49 m s−1, the function of the biofilm is complex, and the ability to resist environmental stress is the strongest. This study can effectively improve the cognition depth of biofilms under the influence of flow velocity in the reclaimed water distribution systems, and provide an important theoretical support for the safe distribution of reclaimed water.