Spatial transcriptomic brain imaging reveals the effects of immunomodulation therapy on specific regional brain cells in a mouse dementia model.

Increasing evidence of brain-immune crosstalk raises expectations for the efficacy of novel immunotherapies in Alzheimer's disease (AD), but the lack of methods to examine brain tissues makes it difficult to evaluate therapeutics. Here, we investigated the changes in spatial transcriptomic signatures and brain cell types using the 10x Genomics Visium platform in immune-modulated AD models after various treatments. To proceed with an analysis suitable for barcode-based spatial transcriptomics, we first organized a workflow for segmentation of neuroanatomical regions, establishment of appropriate gene combinations, and comprehensive review of altered brain cell signatures. Ultimately, we investigated spatial transcriptomic changes following administration of immunomodulators, NK cell supplements and an anti-CD4 antibody, which ameliorated behavior impairment, and designated brain cells and regions showing probable associations with behavior changes. We provided the customized analytic pipeline into an application named STquantool. Thus, we anticipate that our approach can help researchers interpret the real action of drug candidates by simultaneously investigating the dynamics of all transcripts for the development of novel AD therapeutics.

CellDART: cell type inference by domain adaptation of single-cell and spatial transcriptomic data.

Deciphering the cellular composition in genome-wide spatially resolved transcriptomic data is a critical task to clarify the spatial context of cells in a tissue. In this study, we developed a method, CellDART, which estimates the spatial distribution of cells defined by single-cell level data using domain adaptation of neural networks and applied it to the spatial mapping of human lung tissue. The neural network that predicts the cell proportion in a pseudospot, a virtual mixture of cells from single-cell data, is translated to decompose the cell types in each spatial barcoded region. First, CellDART was applied to a mouse brain and a human dorsolateral prefrontal cortex tissue to identify cell types with a layer-specific spatial distribution. Overall, the proposed approach showed more stable and higher accuracy with short execution time compared to other computational methods to predict the spatial location of excitatory neurons. CellDART was capable of decomposing cellular proportion in mouse hippocampus Slide-seq data. Furthermore, CellDART elucidated the cell type predominance defined by the human lung cell atlas across the lung tissue compartments and it corresponded to the known prevalent cell types. CellDART is expected to help to elucidate the spatial heterogeneity of cells and their close interactions in various tissues.

Multichannel Microfluidic Platform for Temporal-Spatial Investigation of Niche Roles of Pseudomonas aeruginosa and Escherichia coli within a Dual-Species Biofilm.

In natural or man-made environments, microorganisms exist predominantly as biofilms forming surface-associated bacterial communities embedded in extracellular polymeric substances (EPSs). Often, biofilm reactors used for endpoint and disruptive analyses of biofilm are not suitable for periodic observation of biofilm formation and development. In this study, a microfluidic device designed with multiple channels and a gradient generator was used for high-throughput analysis and real-time monitoring of dual-species biofilm formation and development. We compared the structural parameters of monospecies and dual-species biofilms containing Pseudomonas aeruginosa (expressing mCherry) and Escherichia coli (expressing green fluorescent protein [GFP]) to understand the interactions in the biofilm. The rate of biovolume increase of each species in monospecies biofilm (2.7 × 105 µm3) was higher than those in a dual-species biofilm (9.68 × 104 µm3); however, synergism was still observed in the dual-species biofilm due to overall increases in biovolume for both species. Synergism was also observed in a dual-species biofilm, where P. aeruginosa forms a "blanket" over E. coli, providing a physical barrier against shear stress in the environment. The microfluidic chip was useful for monitoring the dual-species biofilm in the microenvironment, indicating that different species in a multispecies biofilm exhibit different niches for the survival of the biofilm community. Finally, we demonstrated that the nucleic acids can be extracted from the dual-species biofilm in situ after biofilm imaging analysis. In addition, gene expression supported that the activation and suppression of different quorum sensing genes resulted in the different phenotype seen in the biofilm. This study showed that the integration of microfluidic device with microscopy analysis and molecular techniques could be a promising tool for studying biofilm structure and gene quantification and expression simultaneously. IMPORTANCE In natural or man-made environments, microorganisms exist predominantly as biofilms forming surface-associated bacterial communities embedded in extracellular polymeric substances (EPSs). Often, biofilm reactors used for endpoint and disruptive analyses of biofilm are not suitable for periodic observation of biofilm formation and development. Here, we demonstrate that a microfluidic device with multiple channels and a gradient generator can be useful for high-throughput analysis and real-time monitoring of dual-species biofilm formation and development. Our study revealed synergism in the dual-species biofilm, where P. aeruginosa forms a "blanket" over E. coli, providing a physical barrier against shear stress in the environment. Furthermore, different species in a multispecies biofilm exhibit different niches for the survival of the biofilm community. This study showed that the integration of microfluidic device with microscopy analysis and molecular techniques could be a promising tool for studying biofilm structure and gene quantification and expression simultaneously.

Discovery of molecular features underlying the morphological landscape by integrating spatial transcriptomic data with deep features of tissue images.

Profiling molecular features associated with the morphological landscape of tissue is crucial for investigating the structural and spatial patterns that underlie the biological function of tissues. In this study, we present a new method, spatial gene expression patterns by deep learning of tissue images (SPADE), to identify important genes associated with morphological contexts by combining spatial transcriptomic data with coregistered images. SPADE incorporates deep learning-derived image patterns with spatially resolved gene expression data to extract morphological context markers. Morphological features that correspond to spatial maps of the transcriptome were extracted by image patches surrounding each spot and were subsequently represented by image latent features. The molecular profiles correlated with the image latent features were identified. The extracted genes could be further analyzed to discover functional terms and exploited to extract clusters maintaining morphological contexts. We apply our approach to spatial transcriptomic data from different tissues, platforms and types of images to demonstrate an unbiased method that is capable of obtaining image-integrated gene expression trends.

Assessment of physiological responses of bacteria to chlorine and UV disinfection using a plate count method, flow cytometry and viability PCR.

AIMS: This study aimed to investigate the physiological responses of two gram-negative bacteria (Escherichia coli and Pseudomonas aeruginosa) and two gram-positive bacteria (Enterococcus faecalis and Bacillus sphaericus) to ultraviolet (UV) and chlorine disinfection. METHODS AND RESULTS: Bacterial inactivation by UV and chlorine disinfection were evaluated with a plate count method for culturability, FCM and PMA-qPCR for membrane integrity and DyeTox13-qPCR for enzymatic activity, respectively. Both UV and chorine disinfection caused complete loss of culturability while membrane integrity remained intact after UV disinfection. Both DyeTox13-qPCR and PMA-qPCR showed high ΔCt values up to 8.9 after chlorine disinfection, indicating that both methods were able to distinguish non-treated from chlorine-treated cells. Although PMA-qPCR could not differentiate membrane integrity of cells on UV exposure, DyeTox13-qPCR showed significant differences in ΔCt values of 5.05 and 10.4 for gram-negative (E. coli) and gram-positive (Enterococcus) bacteria, respectively. However, DyeTox13-qPCR for gram-negative bacteria displayed relatively small differences in ΔCt values compared with gram-positive bacteria. CONCLUSION: UV and chlorine disinfection led to changes in physiological state of gram-negative and gram-positive bacteria. Particularly, UV disinfection could induce active but non-culturable (ABNC) for gram-negative bacteria and dormant cell for gram-positive bacteria where intact cells no longer showed the enzymatic activity. SIGNIFICANCE AND IMPACT OF THE STUDY: UV and chlorine are commonly used to disinfect water, food and fomites to inactivate pathogenic bacteria. However, a viable but non-culturable (VBNC) state of bacteria induced by disinfection may underestimate the health risks because of the potential resuscitation of VBNC cells. This study highlighted that bacteria could undergo different physiological (ABNC or dormant) states during UV and chlorine disinfection. In addition, viability PCR techniques could provide insight into the changes in physiological states during disinfection processes.

Changes in physiological states of Salmonella Typhimurium measured by qPCR with PMA and DyeTox13 Green Azide after pasteurization and UV treatment.

Diarrheal diseases caused by Salmonella pose a major threat to public health, and assessment of bacterial viability is critical in determining the safety of food and drinking water after disinfection. Viability PCR could overcome the limitations of traditional culture-dependent methods for a more accurate assessment of the viability of a microbial sample. In this study, the physiological changes in Salmonella Typhimurium induced by pasteurization and UV treatment were evaluated using a culture-based method, RT-qPCR, and viability PCR. The plate count results showed no culturable S. Typhimurium after the pasteurization and UV treatments, while viability PCR with propidium monoazide (PMA) and DyeTox13-qPCR indicated that the membrane integrity of S. Typhimurium remained intact with no metabolic activity. The RT-qPCR results demonstrated that invasion protein (invA) was detectable in UV-treated cells even though the log2-fold change ranged from - 2.13 to - 5.53 for PMA treatment. However, the catalytic activity gene purE was under the detection limit after UV treatment, indicating that most Salmonella entered metabolically inactive status after UV disinfection. Also, viability PCRs were tested with artificially contaminated eggs to determine physiological status on actual food matrices. DyeTox13-qPCR methods showed that most Salmonella lost their metabolic activity but retained membrane integrity after UV disinfection. RT-qPCR may not determine the physiological status of Salmonella after UV disinfection because mRNA could be detectable in UV-treated cells depending on the choice of target gene. Viability PCR demonstrated potential for rapid and specific detection of pathogens with physiological states such as membrane integrity and metabolic activity.Key Points• Membrane integrity of Salmonella remained intact with no metabolic activity after UV.• mRNA could be detectable in UV-treated cells depending on the choice of target gene.• Viability PCR could rapidly detect specific pathogens with their physiological states.

Acute exposure to microplastics induces metabolic disturbances and gut dysbiosis in adult zebrafish (Danio rerio).

There is limited knowledge of the ecotoxicological impacts of MPs at the environmentally relevant concentration on freshwater animals, even though numerous studies have demonstrated the toxic effects of MPs on living organisms. In this study, zebrafish (Danio rerio) was used as a model organism to investigate the ecotoxicological effects of acute exposure of virgin MPs on changes in metabolome and gut microbiota. High-throughput untargeted metabolomics using liquid chromatography with tandem mass spectrometry (LC-MS/MS) provided comprehensive insights into the metabolic responses of zebrafish exposed to PE (polyethylene) and PES (polyester) MPs. Statistical analysis of metabolomics data indicated that 39 and 27 metabolites, such as lysophosphatidylcholine, phosphocholine, phosphatidylserine, triglyceride, glycosphingolipid, psychosine, 8-amino-7-oxononanoate, cholesterol fatty acid ester, phosphatidylinositol, n-Triacontanol, were significantly altered in PE- and PES-exposed zebrafish, respectively. Furthermore, the enrichment pathway analysis unveiled the synthesis of the structural and functional lipids, signaling molecules, fatty alcohol metabolism, and amino acid metabolism, which was considerably perturbated in MPs-exposed zebrafish. In addition, high-throughput DNA sequencing was conducted to examine changes in gut microbiota in the MPs-treated zebrafish. The MPs exposure increased in the relative abundance of Fusobacteria and Proteobacteria, while the relative abundance of Firmicutes declined in MPs-treated zebrafish. Also, microbial diversity and linear discriminant analyses indicated microbiota dysbiosis, metabolomic dysregulation, and oxidative stress. Taken together, the acute exposure of MPs at environmentally relevant concentrations could disrupt the metabolic interaction via the microbiota-gut-liver-brain relationship, implying gastrointestinal and neurological/immune disorders in zebrafish.

Acute exposure effects of tetracycline, ampicillin, sulfamethoxazole, and their mixture on nutrient removal and microbial communities in the activated sludge of air-scouring and reciprocation membrane bioreactors.

The fate of antibiotics, their effects on non-target species, and the spread of antibiotic resistance in wastewater treatment systems have been of concern in recent years. Despite its importance, the effects of these antibiotics on biological nutrient removal in WWTPs have not been completely elucidated. To evaluate the effects of antimicrobial compounds on nutrient removal performance and microbiome, batch experiments were performed using activated sludge samples taken from two distinct membrane bioreactor systems (reciprocation MBR vs. air-scouring MBR). We exposed the activated sludge to 0 mg/L, 0.1 mg/L, and 1.0 mg/L of tetracycline (TET), ampicillin (AMP), sulfamethoxazole (SUL), and their mixture. The mixture of antibiotics significantly decreased ammonia removal efficiency in the reciprocation MBR (rMBR) and air-scouring MBR (AS MBR) by 5% and 12%, respectively. A significant reduction (p < 0.05) in the amoA-AOB gene was observed in AS MBR, while this gene remained unaffected in the rMBR. Interestingly, the gene abundance of amoA from comammox Nitrospira increased from 2.8 × 108 gene copies per gram sludge (0 mg/L) to 5.0 × 108 gene copies per gram sludge (1.0 mg/L) in the setup with antibiotics in the mixture. Correlation analysis of the relative abundance of prevalent taxa and antibiotic concentrations showed that the microbial communities of the AS MBR were more susceptible to TET and MXD antibiotics than the rMBR microbiome.

Effects of monochloramine on culturability, viability and persistence of Pseudomonas putida and tap water mixed bacterial community.

Bacterial biofilms are able to persist in drinking water distribution systems (DWDS) even if disinfectants such as monochloramine are used to inhibit bacterial colonization and biofilm formation. While studies have determined the monochloramine concentrations required to inhibit bacterial biofilms, not much is known about how bacterial biofilms develop resistance towards monochloramine. This study covers the development of resistance to monochloramine in both single species and mixed bacterial biofilms. Through culturability tests and flow cytometry, exposing bacterial biofilms to monochloramine disinfection using a sub-lethal concentration (1.5 mg/L Cl2, experimentally determined) was sufficient to cause an increase of the monochloramine's inhibitory concentrations by as much as two times than what is initially required to inhibit biofilm growth. Through persister cultures and 16S rRNA next generation sequencing (NGS) studies, mixed bacterial biofilms experienced to monochloramine exposure resulted in more bacterial genera becoming persistent and resistant towards monochloramine. Through this study, bacterial genera that were persistent towards monochloramine were suggested to share common traits including the ability (1) to readily enter a persister or viable but non-culturable (VBNC) state and (2) to form biofilms primarily comprising proteinaceous extra-polymeric substances (EPS). Both of these traits also suggested that selected bacterial genera tended to be more persistent to monochloramine and produce EPS. This study advances our understanding of bacterial biofilm resistance towards monochloramine and showed the importance of maintaining monochloramine concentrations in DWDS to prevent the development of bacterial resistance towards monochloramine. KEY POINTS: • Monochloramine-resistant biofilm was developed after sub-lethal disinfection. • Mixed-species culture experienced monochloramine showed more persistence to monochloramine. • Ability to enter a persister/VBNC state is a common trait of persistent bacteria genera.

Limited implication of initial bone scintigraphy on long-term condylar bone change in temporomandibular disorders-Comparison with cone beam computed tomography at 1 year.

BACKGROUND: The current diagnostic criteria for temporomandibular disorders (TMD) do not require imaging for the diagnosis of degenerative joint disease (DJD) of the temporomandibular joint (TMJ) condyle, and there is a lack of data investigating the effectiveness of imaging modalities in predicting long-term TMJ DJD prognosis. OBJECTIVES: To verify the association between initial bone scintigraphy results and long-term DJD bone changes occurring in the TMJ condyle on cone beam computed tomography (CBCT). METHODS: Initial bone scintigraphy, panoramic radiography and CBCT results were analysed in relation to long-term (12 months) TMJ DJD bone change on CBCTs in 55 TMD patients (110 joints). Clinical and radiographic indices were statistically analysed among three groups (improved, no change, and worsened) based on long-term TMJ DJD prognosis calculated by destructive change index (DCI). RESULTS: Neither the uptake ratio nor visual assessment results from initial bone scintigraphy showed a significant difference according to long-term condylar bone change groups. The cut-off value of bone scintigraphy uptake ratio was 2.53 for long-term worsening of TMJ DJD. Worsening of TMJ DJD was significantly associated with the diagnosis based on panoramic radiography (p = .011) and CBCT (p < .001). Initial DCI (ß = -.291, p = .046) had a significant association with long-term worsening of TMJ DJD. CONCLUSION: Initial bone scintigraphy results did not show sufficiently close associations with long-term TMJ DJD prognosis. This should be considered in the selection process of imaging modalities for TMJ DJD patients. Future studies are needed to develop prognostic indices that comprise both clinical and imaging contents for improved predictive ability.

Biofilms in premise plumbing systems as a double-edged sword: microbial community composition and functional profiling of biofilms in a tropical region.

To understand distributions of opportunistic premise plumbing pathogens (OPPPs) and microbial community structures governed by sample location, pipe materials, water temperature, age of property and type of house, 29 biofilm samples obtained from faucets, pipes, and shower heads in different households in Singapore were examined using next-generation sequencing technology. Predictive functional profiling of the biofilm communities was also performed to understand the potential of uncultivated microorganisms in premise plumbing systems and their involvement in various metabolic pathways. Microbial community analysis showed Proteobacteria, Bacteroidetes, Acidobacteria, Nitrospira, and Actinobacteria to be the most abundant phyla across the samples which was found to be significantly different when grouped by age of the properties, location, and the type of house. Meanwhile, opportunistic premise plumbing pathogens such as Mycobacterium, Citrobacter, Pseudomonas, Stenotrophomonas, and Methylobacterium were observed from the samples at 0.5% of the total reads. Functional prediction using 16S gene markers revealed the involvement of the biofilm communities in different metabolic pathways like nitrogen metabolism, biodegradation of xenobiotics, and bacterial secretion implying diverse functionalities that are yet to be studied in this environment. This study serves as a preliminary survey on the microbial communities harboring premise plumbing systems in a tropical region like Singapore.

The toxic effect of triclosan and methyl-triclosan on biological pathways revealed by metabolomics and gene expression in zebrafish embryos.

The omnipresence of antimicrobial triclosan (TCS) and by-products in aquatic environments is a threat to aquatic organisms. Traditionally, the adverse effects of TCS and its by-products have been evaluated by examining the phenotypic output relevant to predicting acute toxicity rather than studying the perturbation of biological pathways. Identifying alterations in the key pathways and molecular mechanisms caused by toxic chemicals helps researchers assess the ecological risks of TCS and its by-products to aquatic environments. In this study, we used metabolomics and reverse transcription qPCR to investigate the adverse effects of a wide range of concentrations of triclosan and its derivative methyl-triclosan (MTCS), ranging from relatively low environmentally relevant levels (ng/L) to high-dose concentrations (sublethal concentration), on zebrafish (Danio rerio) embryos. The metabolism and transcriptome analysis revealed changes in the metabolite and transcripts expression of zebrafish embryos after 96 h exposure at 30 µg/L and 300 µg/L of TCS, 400 µg/L of MTCS and the TCS/MTCS mixture (30 µg/L TCS + 3 µg/L MTCS and 300 µg/L TCS + 30 µg/L MTCS). Significant dysregulations in the expression of the urea transporter (UT), glucose-6-phosphate dehydrogenase (G6PD), alanine transaminase (ALT), glutamate dehydrogenase (GDH), phosphoglucomutase (PGM), and fatty acid synthase (FASN), together with changes in alanine, urea, glucose, 6-phosphogluconalactone, and palmitic acid were observed in the TCS, MTCS, and TCS/MTCS treatments. Particularly, the MTCS treatment group showed fold changes in the mRNA expression of nitrogen metabolism, energy metabolism, and fatty acid synthesis, indicating a disruption of the zebrafish embryos' biological pathways. The changes in the metabolites and gene expressions induced by the TCS, MTCS and the TCS/MTCS mixture treatment demonstrate the pathway changes in starch and sucrose metabolism, nitrogen metabolism, fatty acid synthesis, and phenylalanine, tyrosine and tryptophan biosynthesis. Therefore, our study provides better insights into the risks of the parental compound (TCS) and its by-product (MTCS), as well as the perturbation in biological pathways induced by these two compounds in aquatic environments.

Fabrication and Evaluation of Aluminum Laminated Film for a Pouch-Type Secondary Battery.

In this study, we intend to find the laminate condition for improvement of the chemical resistance of the aluminum pouch film widely used as the coating material of the secondary battery. Here, we investigated the properties including the initial adhesive strength and electrolyte resistance between the metal film layer aluminum and the sealant layer cast polypropylene (CPP) film. As for the laminating condition, we changed the temperature and line speed and maintained the identical pressure conditions. A roll-to-roll dry laminate coating system was used in surface treatment agent coating, adhesive coating, and film laminate. As for the laminate condition of the surface treated aluminum and CPP film, the initial adhesive strength of the laminated pouch film manufactured with 110 °C temperature and 30 m/min line speed was 1300 gf/15 mm. The adhesive strength of the 85 °C electrolyte resistance measured after being immersed for 7 days was found to be 800 gf/15 mm. The initial adhesive strength of CPP film laminated to aluminum without surface treatment showed 900 gf/15 mm and decreased sharply to 150 gf/15 mm in the electrolyte at 85 °C after an hour. The initial adhesive strength and electrolyte resistance of the aluminum and CPP were measured by using the universal testing machine.

Discriminating activated sludge flocs from biofilm microbial communities in a novel pilot-scale reciprocation MBR using high-throughput 16S rRNA gene sequencing.

Membrane bioreactors (MBRs) are a well-established filtration technology that has become a popular solution for treating wastewater. One of the drawbacks of MBRs, however, is the formation of biofilm on the surface of membrane modules. The occurrence of biofilms leads to biofouling, which eventually compromises water quality and damages the membranes. To prevent this, it is vital to understand the mechanism of biofilm formation on membrane surfaces. In this pilot-scale study, a novel reciprocation membrane bioreactor was operated for a period of 8 months and fed with domestic wastewater from an aerobic tank of a local WWTP. Water quality parameters were monitored and the microbial composition of the attached biofilm and suspended aggregates was evaluated in this reciprocating MBR configuration. The abundance of nitrifiers and composition of microbial communities from biofilm and suspended solids samples were investigated using qPCR and high throughput 16S amplicon sequencing. Removal efficiencies of 29%, 16%, and 15% of chemical oxygen demand, total phosphorus and total nitrogen from the influent were observed after the MBR process with average effluent concentrations of 16 mg/L, 4.6 mg/L, and 5.8 mg/L respectively. This suggests that the energy-efficient MBR, apart from reducing the total energy consumption, was able to maintain effluent concentrations that are within regulatory standards for discharge. Molecular analysis showed the presence of amoA Bacteria and 16S Nitrospira genes with the occurrence of nitrification. Candidatus Accumulibacter, a genus with organisms that can accumulate phosphorus, was found to be present in both groups which explains why phosphorus removal was observed in the system. High-throughput 16S rRNA amplicon sequencing revealed the genus Saprospira to be the most abundant species from the total OTUs of both the membrane tank and biofilm samples.

Effect of Sensors on the Reliability and Control Performance of Power Circuits in the Web of Things (WoT).

In order to realize a true WoT environment, a reliable power circuit is required to ensure interconnections among a range of WoT devices. This paper presents research on sensors and their effects on the reliability and response characteristics of power circuits in WoT devices. The presented research can be used in various power circuit applications, such as energy harvesting interfaces, photovoltaic systems, and battery management systems for the WoT devices. As power circuits rely on the feedback from voltage/current sensors, the system performance is likely to be affected by the sensor failure rates, sensor dynamic characteristics, and their interface circuits. This study investigated how the operational availability of the power circuits is affected by the sensor failure rates by performing a quantitative reliability analysis. In the analysis process, this paper also includes the effects of various reconstruction and estimation techniques used in power processing circuits (e.g., energy harvesting circuits and photovoltaic systems). This paper also reports how the transient control performance of power circuits is affected by sensor interface circuits. With the frequency domain stability analysis and circuit simulation, it was verified that the interface circuit dynamics may affect the transient response characteristics of power circuits. The verification results in this paper showed that the reliability and control performance of the power circuits can be affected by the sensor types, fault tolerant approaches against sensor failures, and the response characteristics of the sensor interfaces. The analysis results were also verified by experiments using a power circuit prototype.

Effects of chloride and ionic strength on physical morphology, dissolution, and bacterial toxicity of silver nanoparticles.

In this study, we comprehensively evaluate chloride- and ionic-strength-mediated changes in the physical morphology, dissolution, and bacterial toxicity of silver nanoparticles (AgNPs), which are one of the most-used nanomaterials. The findings isolate the impact of ionic strength from that of chloride concentration. As ionic strength increases, AgNP aggregation likewise increases (such that the hydrodynamic radius [HR] increases), fractal dimension (Df) strongly decreases (providing increased available surface relative to suspensions with higher Df), and the release of Ag(aq) increases. With increased Ag(+) in solution, Escherichia coli demonstrates reduced tolerance to AgNP exposure (i.e., toxicity increases) under higher ionic strength conditions. As chloride concentration increases, aggregates are formed (HR increases) but are dominated by AgCl(0)(s) bridging of AgNPs; relatedly, Df increases. Furthermore, AgNP dissolution strongly increases under increased chloride conditions, but the dominant, theoretical, equilibrium aqueous silver species shift to negatively charged AgClx((x-1)-) species, which appear to be less toxic to E. coli. Thus, E. coli demonstrates increased tolerance to AgNP exposure under higher chloride conditions (i.e., toxicity decreases). Expression measurements of katE, a gene involved in catalase production to alleviate oxidative stress, support oxidative stress in E. coli as a result of Ag(+) exposure. Overall, our work indicates that the environmental impacts of AgNPs must be evaluated under relevant water chemistry conditions.

Quantitative detection and survival analysis of VBNC Salmonella Typhimurium in flour using droplet digital PCR and DNA-intercalating dyes.

The difficulty in detecting viable but non-culturable (VBNC) Salmonella by culture-dependent methods poses a risk to food safety. In our study, we applied a viability test to Salmonella following a lethal treatment and to flour samples inoculated with Salmonella to evaluate the effectiveness of viability polymerase chain reaction (PCR). Our findings revealed that the combination of both ddPCR and qPCR with those DNA-intercalating dyes could quantify viable cells at low concentrations when the plate counting method failed to detect them post-inactivation. Prolonged UV exposure did not induce cell membrane disruption, as confirmed with PMA-ddPCR, with insignificant differences in gene copies. However, samples exposed to DyeTox13 and DyeTox13 + EMA showed lower gene copy numbers, implying that enzymatic activity was decreased by UV exposure duration. In addition, temperature-dependent survival in flour revealed uniform decay rates and D values (time required for a 1 log reduction) of DNA in untreated samples across various temperatures. By contrast, different decay rates were observed with DNA-intercalating dyes (DyeTox13 and DyeTox13 + EMA), showing faster metabolic activity loss at higher temperatures in flour. The decay rates and D values, determined through plate counting and those DNA-intercalating dyes, indicated the potential presence of VBNC Salmonella. A strong correlation between DyeTox13 dyes and the plate counting method suggested DyeTox13 as a rapid alternative for detecting Salmonella in flour. The ddPCR with DNA-intercalating dyes could effectively evaluate Salmonella viability, facilitating more precise monitoring of VBNC in food. IMPORTANCE: Salmonella, a major foodborne pathogen, poses significant risks, particularly to vulnerable groups like infants, older people, and the immunocompromised. Accurate detection is vital for public health and food safety, given its potential to cause severe and life-threatening symptoms. Our study demonstrated digital polymerase chain reaction (ddPCR) with DNA-intercalating dyes for identifying the different physiological statuses of Salmonella. Also, the application of ddPCR with DNA-intercalating dyes offers quantification of viable cells post-disinfection as an alternative method in food. Utilizing ddPCR and DNA-intercalating dyes, we enhanced the detection of VBNC Salmonella, a form often undetectable by conventional methods. This innovative approach could significantly improve the precision and efficiency of detection for viable Salmonella. By providing deeper insights into its transmission potential, our method is a critical tool in preventing outbreaks and ensuring the safety of food products. This research contributes substantially to global efforts in controlling foodborne illnesses and safeguarding public health.

Lumen air pressure regulated multifunctional microbiotas in membrane-aerated biofilm reactors for simultaneous nitrogen removal and antibiotic elimination from aquaculture wastewater.

In this study, two membrane-aerated biofilm reactors (MABRs) were constructed: one solely utilizing biofilm and another hybrid MABR (HMABR) incorporating both suspended-sludge and biofilm to treat low C/N aquaculture wastewater under varying lumen air pressure (LAP). Both HMABR and MABR demonstrated superior nitrogen removal than conventional aeration reactors. Reducing LAP from 10 kPa to 2 kPa could enhance denitrification processes without severely compromising nitrification, resulting in an increase in total inorganic nitrogen (TIN) removal from 50.2±3.1 % to 71.6±1.0 %. The HMABR exhibited better denitrification efficacy than MABR, underscoring its potential for advanced nitrogen removal applications. A decline in LAP led to decreased extracellular polymeric substance (EPS) production, which could potentially augment reactor performance by minimizing mass transfer resistance while maintaining microbial matrix stability and function. Gene-centric metagenomics analysis revealed decreasing LAP impacted nitrogen metabolic potentials and electron flow pathways. The enrichment of napAB at higher LAP and the presence of complete ammonia oxidation (Comammox) Nitrospira at lower LAP indicated aerobic denitrification and Comammox processes in nitrogen removal. Multifunctional microbial communities developed under LAP regulation, diversifying the mechanisms for simultaneous nitrification-denitrification. Increased denitrifying gene pool (narGHI, nirK, norB) and enzymatic activity at a low LAP can amplify denitrification by promoting denitrifying genes and electron flow towards denitrifying enzymes. Sulfamethoxazole (SMX) was simultaneously removed with efficiency up to 80.2 ± 3.7 %, mainly via biodegradation, while antibiotic resistome and mobilome were propagated. Collectively, these findings could improve our understanding of nitrogen and antibiotic removal mechanisms under LAP regulation, offering valuable insights for the effective design and operation of MABR systems in aquaculture wastewater treatment.

Psychometric properties and measurement invariance of the short form of grit scale in Korean adolescents.

This study aimed to identify the factor structure of the Korean version of the Short Grit Scale (Grit-S) and examine its cross-sectional and longitudinal measurement invariance (MI). Data from the Korean Children and Youth Panel Survey 2018 were analyzed, which included two cohorts, comprising 2,327 and 2,325 fourth-year elementary and first-year middle school students, respectively. It was found that the two-factor model fit the data well for the elementary and middle school samples. The results of the cross-sectional MI tests across genders indicated that the full threshold and loading invariance were also supported for the elementary school sample, and the partial threshold and loading invariance were supported for the middle school sample. The analyses of the longitudinal MI revealed that the partial threshold and loading invariance were supported for both samples. The reliability analysis revealed satisfactory McDonald's Omega values for both samples at each time point and moderate stability coefficients over time. Based on these findings, it was concluded that the Korean version of the Grit-S demonstrated satisfactory psychometric properties and exhibited MI across gender and time in Korean adolescents.

Unveiling the impact of short-term polyethylene microplastics exposure on metabolomics and gut microbiota in earthworms (Eudrilus euganiae).

Microplastics (MPs) pose a significant environmental concern, particularly for terrestrial fauna. In this study, earthworms were used as a model organism to investigate the ecotoxicological effects of short-term exposure to virgin MPs on changes in metabolome and gut microbiota. High-throughput untargeted metabolomics showed significant internal reactions in the earthworms' metabolic processes due to MPs exposure, even when no visible stress signs, such as changes in growth or mortality rates, were present. Earthworms exposed to different concentrations of polyethylene (PE) MP exhibited significant disruption in 39 and 199 molecular features related to energy and lipid metabolism, anti-inflammatory, cell signaling, and membrane integrity. The activities of enzymes and transport proteins in earthworms were dysregulated when exposed to PE. Changes in the gut microbiota's community structure and complexity were observed in response to PE MPs exposure. Despite the relative stability in alpha-diversity and relative abundance, shifts in beta-diversity and network analysis in the PE-exposed group were indicative of an adaptive response to MPs. Earthworms exhibited resilience or adaptation in response to MPs exposure, potentially maintaining their functionality. This study provides preliminary insights into the impact of MPs on soil invertebrates like earthworms and highlights the need for further exploration of long-term effects and underlying molecular mechanisms.