Effect of food-to-microorganisms ratio on aerobic granular sludge settleability: Microbial community, potential roles and sequential responses of extracellular proteins and polysaccharides.

The food-to-microorganism ratio (F/M) is an important parameter in wastewater biotreatment that significantly affects the granulation and settleability of aerobic granular sludge (AGS). Hence, understanding the long-term effects and internal mechanisms of F/M on AGS settling performance is essential. This study investigated the relationship between F/M and the sludge volume index (SVI) within a range of 0.23-2.50 kgCOD/(kgMLVSS·d). Thiothrix and Candidatus_Competibacter were identified as two dominant bacterial genera influencing AGS settling performance. With F/M increased from 0.27 kgCOD/(kgMLVSS·d) to 1.53 kgCOD/(kgMLVSS·d), the abundance of Thiothrix significantly increased from 0.20% to 27.02%, and the hydrophobicity of extracellular proteins (PN) decreased, which collectively reduced AGS settling performance. However, under high-F/M conditions, the gel-like polysaccharides (PS) effectively retained the granular biomass by binding to the highly abundant Thiothrix (53.65%). The progressive increment in biomass led to a concomitant reduction in F/M, resulting in the recovery of AGS settleability. In addition, two-dimensional correlation infrared spectroscopy analysis revealed the preferential responses of PN and PS to the increase and decrease of F/M, and the content and characteristics of PN and PS played important roles in granular settling. The study provides insight into the microbial composition and the potential role of extracellular polymer substances in the AGS sedimentation behavior, offering valuable theoretical support for stable AGS operation.

Cells-in-Touch: 3D Printing in Reconstruction and Modelling of Microscopic Biological Geometries for Education and Future Research Applications.

Additive manufacturing (3D printing) and computer-aided design (CAD) still have limited uptake in biomedical and bioengineering research and education, despite the significant potential of these technologies. The utility of organ-scale 3D-printed models of living structures is widely appreciated, while the workflows for microscopy data translation into tactile accessible replicas are not well developed yet. Here, we demonstrate an accessible and reproducible CAD-based methodology for generating 3D-printed scalable models of human cells cultured in vitro and imaged using conventional scanning confocal microscopy with fused deposition modeling (FDM) 3D printing. We termed this technology CiTo-3DP (Cells-in-Touch for 3D Printing). As a proof-of-concept, we created dismountable CiTo-3DP models of human epithelial, mesenchymal, and neural cells by using selectively stained nuclei and cytoskeletal components. We also provide educational and research context for the presented cellular models. In the future, the CiTo-3DP approach can be adapted to different imaging and 3D printing modalities and comprehensively present various cell types, subcellular structures, and extracellular matrices. The resulting CAD and 3D printed models could be used for a broad spectrum of education and research applications.

Non-filamentous sludge bulking induced by exopolysaccharide variation in structure and properties during aerobic granulation.

The forming mechanism of non-filamentous sludge bulking during aerobic granulation were investigated basing on three feeding strategies (R1 direct aeration after fast feeding, R2 anaerobic stirring after fast feeding and R3 anaerobic plug-flow slow feeding). Results showed that strong selection stress (shortening settling time) led to a sharp flocs washout and the subsequent increase of food to microorganisms (F/M) in R1 and R3 reactors, but not found in R2 due to the different strategies of feeding modes. With the increase of F/M, zeta potential and hydrophobicity of sludge surfaces significantly decreased and thus leading to an enhanced repulsive force and energy barriers for sludge aggregation. Particularly, when F/M exceeded 1.2 kgCOD/(kgMLSS·d), non-filamentous sludge bulking was ultimately triggered in R1 and R3. Further analysis showed that massive extracellular exopolysaccharide (PS) accumulated on the surfaces of non-filamentous bulking sludge due to the increased abundance of the microorganisms related to PS secretion during sludge bulking. In addition, significantly increased intracellular second messenger (c-di-GMP), a key substance regulating PS biosynthesis, was confirmed via its concentration determination as well as microbial function prediction analysis, which played a critical role in sludge bulking. Combing with the systematic detection from surface plasmon resonance system, rheometer and size-exclusion chromatography-multiangle laser light detection-refractive index system, higher molecular weight, compact conformation, higher viscosity and higher hydrophilicity was determined in sludge bulking PS relative to PS extracted from non-filamentous bulking sludge. Clearly, the changes of PS (content, structures and properties) driven by c-di-GMP are the dominant mechanism for the formation of non-filamentous sludge bulking during aerobic granulation. This work could provide theoretical support for successful start-up and application of aerobic granular sludge technology.

Digital Quantification Method for Sensitive Point-of-Care Detection of Salivary Uric Acid Using Smartphone-Assisted µPADs.

Uric acid (UA) is an important biomarker for many diseases. A sensitive point-of-care (POC) testing platform is designed for the digital quantification of salivary UA based on a colorimetric reaction on an easy-to-build smartphone-assisted microfluidic paper-based analytical device (SµPAD). UA levels are quantified according to the color intensity of Prussian blue on the SµPAD with the aid of a MATLAB code or a smartphone APP. A color correction method is specifically applied to exclude the light effect. Together with the engineering design of SµPADs, the background calibration function with the APP increases the UA sensitivity by 100-fold to reach 0.1 ppm with a linear range of 0.1-200 ppm. The assay time is less than 10 min. SµPADs demonstrate a correlation of 0.97 with a commercial UA kit for the detection of salivary UA in clinical samples. SµPADs provide a sensitive, fast, affordable, and reliable tool for the noninvasive POC quantification of salivary UA for early diagnosis of abnormal UA level-associated health conditions.

Deciphering the concurrence of comammox, partial denitrification and anammox in a single low-oxygen mainstream nitrogen removal reactor.

One-stage anammox-based autotrophic nitrogen removal technology has attracted increasing interest to sustainable biological nitrogen removal for future wastewater treatment. However, its application in mainstream municipal wastewater treatment is still challenging due to low nitrogen and high organics of raw wastewater. Herein, a novel Simultaneous Carbon Oxidation, partial Comammox, Denitratation and Anammox (SCOCDA) was firstly developed in a single sequencing batch biofilm reactor operated at a dissolved oxygen concentration of ∼0.5 mg/L for treating synthetic municipal wastewater (50 mg/L NH4+-N and 100-250 mg/L COD). The long-term operation showed that almost complete COD and nitrogen removal performance could be achieved at a carbon/nitrogen ratio (COD/NH4+-N) of 3-5 with the corresponding effluent total nitrogen (TN)＜5 mg/L. Microbial community and amoA-targeting amplicon sequencing analysis further verified that comammox Nitrospira spp., denitrifier Thauera and other aerobic/facultative heterotrophs could work synergistically with anammox bacteria, Candidatus Kuenenia. Moreover, nitrogen metabolic and inorganic carbon fixation pathways through the interaction between comammox and anammox were also revealed with the aid of Kyoto Encyclopedia of Genes and Genomes (KEGG). Lastly, potential application of proposed SCOCDA process was illustrated. This research sheds new light on advanced nitrogen removal towards limit of technology via the synergy of comammox and anammox.

Selective myelinated nerve fiber stimulation via temporal interfering electric fields.

We have investigated selective electrical stimulation of myelinated nerve fibers using a computational model of temporal interfering (TI) fields. The model consists of two groups of electrodes placed on the outer bundle surface, each group stimulated at a different frequency. We manipulated the stimulus waveform, magnitude and frequency of short-duration stimuli (70ms), and investigated fiber-specific stimulus-elicited compound action potentials. Results show that under 100Hz & 200Hz TI stimulation with 0.6mA total current shared by the electrodes, continuous action potentials were generated in deeper nerve fibers, and that the firing region was steerable by changing individual electrode currents. This study provides a promising platform for non-invasive nerve bundle stimulation by TI fields.

Study on Factors Affecting the Performance of a CRISPR/Cas-Assisted New Immunoassay: Detection of Salivary Insulin as an Example.

The clustered regularly interspaced short palindromic repeat (CRISPR)/Cas is now playing a significant role in biosensing applications, especially when the trans-cleavage activity of several Cas effectors is discovered. Taking advantages of both CRISPR/Cas and the enzyme-linked immunosorbent assay (ELISA) in analytical and clinical investigations, CRISPR/Cas-powered ELISA has been successfully designed to detect a spectrum of analytes beyond nucleic acid. Herein, we developed a CRISPR/Cas12a-assisted new immunoassay (CANi) for detection of salivary insulin as an example. Specifically, factors (antibody selection, temperature, and assay time) affecting the CRISPR/Cas-based ELISA system's performance were investigated. It was observed that the concentration of blocking solution, selection of the capture antibody pairs, and the sequences of triggering ssDNA and guiding RNA affected this immunoassay sensitivity. In contrast, the preincubation of CRISPR/Cas12a working solution and pre-mixture of detection antibody with anti-IgG-ssDNA did not show influence on the performance of CANi for the detection of insulin. Under optimized conditions, the sensitivity for detection of salivary insulin was 10 fg/ml with a linear range from 10 fg/ml to 1 ng/ml.

Advances in Biosensors for Continuous Glucose Monitoring Towards Wearables.

Continuous glucose monitors (CGMs) for the non-invasive monitoring of diabetes are constantly being developed and improved. Although there are multiple biosensing platforms for monitoring glucose available on the market, there is still a strong need to enhance their precision, repeatability, wearability, and accessibility to end-users. Biosensing technologies are being increasingly explored that use different bodily fluids such as sweat and tear fluid, etc., that can be calibrated to and therefore used to measure blood glucose concentrations accurately. To improve the wearability of these devices, exploring different fluids as testing mediums is essential and opens the door to various implants and wearables that in turn have the potential to be less inhibiting to the wearer. Recent developments have surfaced in the form of contact lenses or mouthguards for instance. Challenges still present themselves in the form of sensitivity, especially at very high or low glucose concentrations, which is critical for a diabetic person to monitor. This review summarises advances in wearable glucose biosensors over the past 5 years, comparing the different types as well as the fluid they use to detect glucose, including the CGMs currently available on the market. Perspectives on the development of wearables for glucose biosensing are discussed.

Nanozyme Applications: A Glimpse of Insight in Food Safety.

Nanozymes own striking merits, including high enzyme-mimicking activity, good stability, and low cost. Due to the powerful and distinguished functions, nanozymes exhibit widespread applications in the field of biosensing and immunoassay, attracting researchers in various fields to design and engineer nanozymes. Recently, nanozymes have been innovatively used to bridge nanotechnology with analytical techniques to achieve the high sensitivity, specificity, and reproducibility. However, the applications of nanozymes in food applications are seldom reviewed. In this review, we summarize several typical nanozymes and provide a comprehensive description of the history, principles, designs, and applications of nanozyme-based analytical techniques in food contaminants detection. Based on engineering and modification of nanozymes, the food contaminants are classified and then discussed in detail via discriminating the roles of nanozymes in various analytical methods, including fluorescence, colorimetric and electrochemical assay, surface-enhanced Raman scattering, magnetic relaxing sensing, and electrochemiluminescence. Further, representative examples of nanozymes-based methods are highlighted for contaminants analysis and inhibition. Finally, the current challenges and prospects of nanozymes are discussed.

[Relationship and risk factors of osteonecrosis of femoral head after internal fixation of femoral neck fracture with serum bone metabolism and vasoactive factors].

OBJECTIVE: To study the relationship between osteonecrosis of femoral head after internal fixation of femoral neck fracture and serum bone metabolism, vascular active factors, and analyze the risk factors. METHODS: Total 150 patients with femoral neck fracture who underwent reduction and internal fixation from April 2016 to April 2019 were selected, including 83 males and 67 females. According to whether there was necrosis of femoral head after operation, they were divided into necrosis group(32 cases) and non necrosis group (118 cases). Before operation and 1, 3, 5 days after operation, the serum levels of beta-C terminal cross-linked telopeptides of typeⅠ collagen(ß-CTX), N-telopeptide of typeⅠ procollagen(PINP), nitric oxide (NO), Endothelin-1 (ET-1) were measured. The clinical characteristics of the two groups were compared. The risk factors of postoperative femoral head necrosis were analyzed by logistic regression model. The value of serum indexes in predicting postoperative femoral head necrosis was analyzed by ROC curve. RESULTS: There was no significant difference in the levels of serum PINP and ß-CTX between necrotic group and non necrotic group before operation and 1, 3 and 5 days after operation(P>0.05). There was no significant difference in the levels of serum NO and ET-1 before operation and 5 days after operation(P>0.05). There were significant differences in the levels of serum NO and ET-1(P<0.05), fracture type, preoperative traction ratio and reduction quality (P<0.05). Logistic regression analysis showed that the decrease of serum NO content and the increase of ET-1 content at 1 day after operation, Ⅲ-Ⅳ fracture and Ⅲ-Ⅳ reduction were the risk factors of femoral head necrosis;ROC curve analysis showed that serum NO and ET-1 content at 1 day after operation had predictive value for femoral head necrosis, and the best cut-off points were 26.55 µmol / L and 7.785 µg / L, respectively. CONCLUSION: The content of serum NO and ET-1 on the first day after operation can predict the necrosis of femoral head.

Development of aerobic methane oxidation, denitrification coupled to methanogenesis (AMODM) in a microaerophilic expanded granular sludge blanket biofilm reactor.

The issue of enhancing nitrogen removal and managing dissolved methane emission in anaerobic treatment systems is a major bottleneck in its wider application to treat high-strength organic wastewater with nitrate. Herein, a novel aerobic methane oxidation, denitrification coupled to methanogenesis (AMODM) process was developed in a glucose-fed microaerobic expanded granular sludge blanket biofilm reactor (EGSBBR) through in-situ utilization of produced methane for nitrogen removal. The 162-day operation demonstrated that long-term treatment performance under the decreased COD/NO3--N (C/N) ratio from 66.7 to 10 and the optimal C/N ratio for completing AMODM was found to be 16.7. Microbial community analysis further evidenced that Methanothrix as key methanogen predominated in the sludge bed, while Methlogaea as aerobic methane oxidizer was mainly detected in the packing bed of the hybrid system. Meanwhile, some facultative heterotrophic and dissimilated nitrate-reduction (DNRA) genera also co-existed. The profiling of key functional genes further proved concurrent occurrence of methanogenesis, aerobic methane oxidation and denitrification. Furthermore, possible microbial mechanism on AMODM process was elucidated from the prospective of targeted species interaction within the reactor. This research provides a robust and environment-friendly alternative process treating nitrate-containing organic wastewater towards efficient nitrogen removal, low resource consumption, bioenergy recovery and greenhouse gas reduction.

Unravelling nitrogen removal and nitrous oxide emission from mainstream integrated nitrification-partial denitrification-anammox for low carbon/nitrogen domestic wastewater.

Stable supply of nitrite is often a major obstacle for achieving mainstream anammox due to washout failure of nitrite oxidizers (NOB) at low influent ammonia of municipal wastewater. In this study, an integrated nitrification, partial denitrification and anammox (INPDA) as a one-stage mainstream nitrogen removal alternative was established in a low-oxygen sequencing batch biofilm reactor treating synthetic sewage. The overall nitrogen removal and nitrous oxide (N2O) emission were mainly investigated at 50 mg/L NH4+-N influent with a low carbon/nitrogen (C/N) of 2.5. Continuous operation demonstrated that as high as 98.8% NH4+-N and 94.1% TN were removed in SBBR system. Cyclic experiment verified sequential completion of nitrification, partial denitrification and anammox were responsible for high-rate TN removal. During one typical cycle, the trend of N2O emission was characterized by firstly rapid rise, then fluctuant decrease followed by rapid decrease and finally slow disappearance. The maximum N2O emission rate reached up to 6.7 µg/(L·min) occurred at 75 min. High-throughput sequencing revealed the co-existence of nitrifying, denitrifying and anammox species and large detection of key functional genes (Hzs, Hdh, Hao, Nor) in an oxygen-limited SBBR, thereby highly correlating nitrogen removal and N2O emission characteristics. Nitrogen metabolic pathways analysis further suggest denitratation(NO3--N to NO2--N)-based anammox is a main route for mainstream nitrogen removal. Moreover, N2O might be generated by both hydroxylamine oxidation step in nitrification and also heterotrophic denitrification pathway. The research findings provide more deep understandings of enhanced nitrogen removal and mitigated N2O footprint from a single mainstream anammox-based system.

Performance and microbial community in a single-stage simultaneous carbon oxidation, partial nitritation, denitritation and anammox system treating synthetic coking wastewater under the stress of phenol.

As a highly toxic pollutant, phenol is typically present in some high-strength nitrogenous wastewater. In this study, a synthetic coking wastewater with 400 mg L-1 ammonia-nitrogen and 50-250 mg L-1 phenol was treated. Results showed that simultaneous carbon oxidation, partial nitritation, denitritation and anammox (SCONDA) was successfully achieved by step-wise phenol addition. At 200 mg L-1 phenol, 99.8% phenol, 97.5% COD and 89.8% nitrogen could be together removed. However, further increase in phenol concentration caused significant deterioration of the short-terms nitrogen removal efficiency. High-throughput sequencing revealed remarkable evolution in microbial biodiversity, community composition, especially functional species at different phenol concentrations. When the phenol addition was increased from 200 to 250 mg L-1, the relative abundance of Candidatus Kuenenia as predominant anammox species decreased by 87.1%, while phenol-degrading bacteria was increasingly abundant. Furthermore, the removal mechanism of phenol and nitrogen was elucidated by the collaboration among different key functional microbial consortia.