Screening and identification of functional bacterial attachment genes in aerobic granular sludge.

The screening and identification of attachment genes is important to exploring the formation mechanism of biofilms at the gene level. It is helpful to the development of key culture technologies for aerobic granular sludge (AGS). In this study, genome-wide sequencing and gene editing were employed for the first time to investigate the effects and functions of attachment genes in AGS. With the help of whole-genome analysis, ten attachment genes were screened from thirteen genes, and the efficiency of gene screening was greatly improved. Then, two attachment genes were selected as examples to further confirm the gene functions by constructing gene-knockout recombinant mutants of Stenotrophomonas maltophilia; when the two attachment genes were knocked out, the attachment potential was reduced by 50.67% and 43.93%, respectively. The results provide a new theoretical principle and efficient method for the development of AGS from the perspective of attachment genes.

Construction of a CRISPR Interference System for Gene Knockdown in Stenotrophomonas maltophilia AGS-1 from Aerobic Granular Sludge.

To identify the function of attachment genes involved in biofilm formation in Stenotrophomonas maltophilia AGS-1 isolated from aerobic granular sludge, an effective gene molecular tool is needed. We developed a two-plasmid CRISPRi system in Stenotrophomonas maltophilia AGS-1. One plasmid expressed dCas9 protein with the l-arabinose inducible promoter, and the other plasmid contained the sgRNA cassette complementary to the target gene. Under control of the araC-inducible promoter, this system exhibited little leaky basal expression and highly induced expression that silenced endogenous and exogenous genes with reversible knockdown. This system achieved up to 211-fold suppression for mCherry expression on the nontemplate strand compared to the template strand (91-fold). The utility of the developed CRISPRi platform was also characterized by suppressing the xanA and rpfF genes. The expression of these two genes was rapidly depleted and the adhesion ability decreased, which demonstrated that the modulation of either gene was an important factor for biofilm formation of the AGS-1 strain. The system also tested the ability to simultaneously silence transcriptional suppression of multiple targeted genes, an entire operon, or part of it. Lastly, the use of CRISPRi allowed us to dissect the gene intricacies involved in flagellar biosynthesis. Collectively, these results demonstrated that the CRISPRi system was a simple, feasible, and controllable manipulation system of gene expression in the AGS-1 strain.

Construction and application of a mCherry fluorescent labeling system for Stenotrophomonas AGS-1 from aerobic granular sludge.

To elucidate the specific mechanism by which high-attachment bacteria promote aerobic granular sludge (AGS) formation, a red fluorescent protein mCherry-based biomarker system was developed in the high-attachment strain Stenotrophomonas AGS-1 from AGS. The fluorescent labeling system used plasmid-mediated mCherry expression driven by a Ptac constitutive promoter. mCherry-labeled AGS-1 had normal unimpaired growth, strong fluorescent signals, and good fluorescence imaging. Also, the mCherry labeling system had no effect on the attachment ability of AGS-1. In addition, mCherry-labeled AGS-1 maintained high plasmid stability, even after more than 100 generations. Notably, after the addition of mCherry-labeled AGS-1 into the activated sludge system, the mCherry fluorescence of the sludge system can be used as a good reflection of the relative amount of AGS-1. Moreover, the spatial distribution of mCherry-labeled AGS-1 in the sludge system could be visualized and remained clear even after 5 days by fluorescence imaging. These results revealed that the mCherry-based biomarker system would provide a valuable tool for labeling AGS-1 to monitor the spatial distribution and fate of AGS-1 in AGS, which would help to better understand the mechanism of AGS formation and facilitate the development of AGS technology.

Comparative analysis of membrane fouling mechanisms induced by operation modes of membrane bioreactors with aerobic granular sludge.

This experimental work investigated fouling characteristics induced by two different configurations of membrane bioreactor (MBR), which are submerged MBR and sidestream MBR with aerobic granular sludge. Submerged membrane bioreactor with granular sludge (Sub-MGSBR) ran the longest operation time 61 days with a steady overall TMP increase rate; Sidestream membrane bioreactor with granular sludge (SS-MGSBR) performed only 39 days, which exhibited Sub-MGSBR had more efficiently retarding membrane fouling. In both membrane bioreactors with flocculent sludge (MFSBRs) as a control, membrane foulants were compact, and cake resistance was the dominant fouling factor. In MGSBRs, however, pore blocking resistance turned out the key fouling factor. Especially in Sub-MGSBR, it went beyond 75%, and there was the most conglomeration of microorganisms of foulants with the highest porosity. Extracellular polymeric substances (EPS) content of foulants proved membrane fouling was hardly just for granules accumulation into cake but microorganisms' growth in MGSBRs.

Construction and application of a new CRISPR/Cas12a system in Stenotrophomonas AGS-1 from aerobic granular sludge.

Aerobic granular sludge (AGS) is a microbial aggregate with a biofilm structure. Thus, investigating AGS in the aspect of biofilm and microbial attachment at the genetic level would help to reveal the mechanism of granule biofilm formation. In this work, a two-plasmid clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated proteins (Cas)12a genome editing system was constructed to identify attachment genes for the first time in Stenotrophomonas AGS-1 from AGS. One plasmid contained a Cas12a cassette driven by an arabinose-inducible promoter, and another contained the specific crRNA and homologous arms (HAs). Acidaminococcus sp. Cas12a (AsCas12a) was adopted and proven to have mild toxicity (compared to Cas9) and strong cleavage activity for AGS-1. CRISPR/Cas12a-mediated rmlA knockout decreased attachment ability by 38.26%. Overexpression of rmlA in AGS-1 resulted in an increase of 30.33% in attachment ability. These results showed that the modulation of rmlA was an important factor for the biofilm formation of AGS-1. Moreover, two other genes (xanB and rpfF) were knocked out by CRISPR/Cas12a and identified as attachment-related genes in AGS-1. Also, this system could achieve point mutations. These data indicated that the CRISPR/Cas12a system could be an effective molecular platform for attachment gene function identification, which would be useful for the development of AGS in wastewater treatment.

Reference-frame-independent quantum key distribution with advantage distillation.

Advantage distillation (AD) provides a means of separating highly correlated raw key bits from weakly correlated information in quantum key distribution (QKD). In this Letter, we apply the AD method to improve the performance of reference-frame-independent QKD (RFI-QKD). Simulation results show that, compared with RFI-QKD without AD, RFI-QKD with AD can tolerate higher system errors and obtain better performance on the secret key rate and transmission distance. Furthermore, we extend the AD method to RFI measurement-device-independent QKD (RFI-MDI-QKD) and demonstrate that the AD method can improve the performance of RFI-MDI-QKD more significantly.

Improved reference-frame-independent quantum key distribution: erratum.

We present an erratum to our Letter [Opt. Lett.47, 4219 (2022)10.1364/OL.470558]. This erratum corrects the error results of Figs. 2 and 4 due to the error in the simulation code. The corrections have no influence on the conclusions of the original Letter.

The novel HLA-A*24:520 allele was identified in a Chinese individual.

HLA-A*24:520 differs from HLA-A*24:02:01:01 by one nucleotide substitution at position 668 C→T in exon 4.

Development and Application of a New Arabinose-Inducible Vector in High-Attachment Strain Stenotrophomonas AGS-1 from Aerobic Granular Sludge.

To explore the molecular structure of attachment genes, we constructed and characterized a new arabinose-inducible vector for the high-attachment strain Stenotrophomonas AGS-1 isolated from aerobic granular sludge (AGS). mCherry was used as a simple observation biomarker, and the araC-PBAD-inducible promoter was chosen to artificially regulate the expression of target genes. The system achieved little leaky basal expression and high maximal induced expression. The araC-PBAD-based inducible expression was modulated over a wide range of 0.0005 to 0.2% l-arabinose. Notably, a "lag expression" phenomenon was observed in which mCherry was expressed after bacterial growth in LB medium. Using the system and the strategy of fusion expression of target genes (rmlA and AsCas12a) plus mCherry, the recombinant AGS-1 strain achieved the effective induction of rmlA and AsCas12a-mCherry gene expression in the range of 0.0005 to 0.1% l-arabinose. These results demonstrate that the new arabinose-inducible vector could be used as an important molecular tool in the gene function and genome-editing research of strain AGS-1.

Repetitive accumulation of interstitial cells generates the branched structure of Cladonema medusa tentacles.

The shaping of tissues and organs in many animals relies on interactions between the epithelial cell layer and its underlying mesoderm-derived tissues. Inductive signals, such as receptor tyrosine kinase (RTK) signaling emanating from mesoderm, act on cells of the epithelium to initiate three-dimensional changes. However, how tissues are shaped in a diploblastic animal with no mesoderm remains largely unknown. In this study, the jellyfish Cladonema pacificum was used to investigate branch formation. The tentacles on its medusa stage undergo branching, which increases the epithelial surface area available for carrying nematocytes, thereby maximizing prey capture. Pharmacological and cellular analyses of the branching process suggest a two-step model for tentacle branch formation, in which mitogen-activated protein kinase kinase signaling accumulates interstitial cells in the future branch-forming region, and fibroblast growth factor signaling regulates branch elongation. This study highlights an essential role for these pluripotent stem cells in the tissue-shaping morphogenesis of a diploblastic animal. In addition, it identifies a mechanism involving RTK signaling and cell proliferative activity at the branch tip for branching morphogenesis that is apparently conserved across the animal kingdom.

Isolation and characterization of a strain with high microbial attachment in aerobic granular sludge.

Aerobic granule is a special microbial aggregate associated with biofilm structure. The formation of aerobic granular sludge is primarily depending on its bacterial community and relevant microbiological properties. In this experiment, a strain with high microbial attachment was isolated from aerobic granular sludge, and the detailed characteristics were examined. Its high attachment ability could reach 2.34 (OD600nm), while other low attachment values were only around 0.06-0.32, which indicated a big variation among the different bacteria. The strain exhibited a very special morphology with many fibric fingers under SEM observation. A distinctive behaviour was to form a spherical particle by themselves, which would be very beneficial for the formation and development of granular sludge. The EPS measurement showed that its PN content was higher than low attachment bacteria, and 3D-EEM confirmed that there were some different components. Based on the 16S rRNA analysis, it was identified to mostly belong to Stenotrophomonas. Its augmentation to particle sludge cultivation demonstrated that the strain could significantly promote the formation of aerobic granule. Conclusively, it was strongly suggested that it might be used as a good and potential model strain or chassis organism for the aerobic granular sludge formation and development.

Extracorporeal membrane oxygenation as a bridge vs. non-bridging for lung transplantation: A systematic review and meta-analysis.

Whether extracorporeal membrane oxygenation (ECMO) as a bridge to lung transplantation (BTT) can achieve a similar survival to non-BTT remains controversial. We conducted this meta-analysis to compare the outcomes between ECMO BTT and non-BTT to facilitate better clinical decision-making. Seven databases were searched for eligible studies comparing ECMO BTT and non-BTT. The primary endpoints included survival, intraoperative indicators, postoperative hospitalization indicators, and postoperative complications. Nineteen studies (involving 7061 participants) were included in the final analysis. The outcomes of overall survival, overall survival rate, graft survival rate, in-hospital mortality, postoperative hospital days, postoperative intensive care unit days, postoperative ventilation time, blood transfusion volume, and postoperative complications were all better in the non-BTT group. The total mortality in ECMO bridging was 23.03%, in which the top five causes of death were right heart failure (8.03%), multiple organ failure (7.03%), bleeding (not cranial) (4.67%), cranial bleeding (3.15%), and sepsis (2.90%). In summary, Non-BTT is associated with better survival and fewer complications compared to BTT. When ECMO may be the only option, the patient and medical team need to realize the increased risk of ECMO by complications and survival.

[Measurements of Bacterial Community and Biodiversity from Activated Sludge for a Wastewater Treatment Containing Starch].

This study analyzes the microbial community and diversity composition of activated sludge in anoxic/oxic (A/O) treatment systems at different operation stages using Illumina MiSeq high-throughput sequencing to investigate the microbial community structure and diversity in activated sludge for starch wastewater treatment. The experimental results showed that the microbial community structure of activated sludge for starch production wastewater treatment in A/O systems was quite stable under the same wastewater condition, and that the dominant bacteria of the activated sludge were Proteobacteria, Bacteroidetes, Chloroflexi, Firmicutes, and Actinobacteria. The most important dominant bacterial group was Proteobacteria (45.66%-66.30%), of which γ-subclass bacteria were the main member and occupied 36.38%-66.65%. The proportion of Sphingobacteria, the main member of the Bacteroidetes, decreased when the sludge settling performance was better, but the proportion of Anaerolineae, the main member of Chloroflexi, increased significantly when the sludge sedimentation performance was better. These changes may have been closely related to the behavior of sludge settleability. There were a large number of functional bacteria in the activated sludge, which played an important role in the degradation of pollutants and in nitrogen and/or phosphorus removal.

Experimental quantum key distribution with uncharacterized sources and projective measurements.

In theory, quantum key distribution (QKD) can offer information-theoretic secure communication based on the laws of quantum mechanics. However, the vast majority of practical QKD implementations assume the perfect state preparation to ensure security, which is a demanding requirement with current technology. Here, by incorporating the mismatched-basis data, we report an experimental decoy-state QKD demonstration with uncharacterized encoding sources, which only requires that the encoding states are two-dimensional. Furthermore, the measurement operation of the receiver is loosened to be projective measurements. With a rigorous statistical fluctuation analysis, we can distribute secret keys when the transmission distances of the standard fiber link are 101 and 202 km. Our experimental demonstration represents a significant step toward realizing long-distance quantum communication, even with uncharacterized sources and projective measurements.

The viscosity behaviors of bacterial suspensions or extracellular polymeric substances and their effects on aerobic granular sludge.

Although the viscosity behavior of bacteria and extracellular polymeric substances (EPS) in flocculent activated sludge (FAS) and aerobic granular sludge (AGS) has been investigated, no studies have explored the role of viscosity in microbial attachment in pure culture. This study investigated the viscosity behavior of bacteria and EPS. The results showed that bacteria and their EPS exhibited non-Newtonian fluid and shear-thinning behavior. The viscosity of bacteria and EPS was 1.55-3.80 cP and 1.10-2.40 cP, respectively, while the attachment of bacteria (optical density at 600 nm) was 0.1426-3.1015. Bacteria with high attachment secreted EPS with a higher viscosity (2.40 cP), whereas those with weak attachment expressed EPS with a lower viscosity (1.10 cP). Viscosity and microbial attachment or extracellular polysaccharide (PS) content were significantly positively correlated. PS content was the source of bacterial viscosity, and ß-polysaccharide played a more important role in viscosity and microbial attachment than α-polysaccharide. Thus, viscosity plays a critical role in microbial attachment, and high viscosity and PS content result in high microbial attachment, which is beneficial to the granulation process of AGS.

Bilateral Lung Transplantation Provides Better Long-term Survival and Pulmonary Function Than Single Lung Transplantation: A Systematic Review and Meta-analysis.

BACKGROUND: Both bilateral lung transplantation (BLT) and single lung transplantation (SLT) are commonly used, but which method is better remains controversial. This meta-analysis was conducted to compare the 2 surgical procedures to identify a better clinical choice. METHODS: Cohort studies comparing SLT and BLT were identified by conducting searches of databases and screening references of retrieved articles. Survival, pulmonary function, surgical indicators, and complications were compared between the 2 groups. RESULTS: Thirty studies (1980 recipients in the SLT group and 2112 recipients in the BLT group) were pooled in the meta-analysis. The long-term overall survival rate (OSR) (OSR-4y and OSR-5y), bronchiolitis obliterans syndrome (BOS)-free survival, BOS-free survival rate (BFSR) (2-5 y), 6-minute walking distance, forced expiratory volume in 1 second (%), forced vital capacity (%), oxygenation index, pulmonary arterial pressure, Arterial partial pressure of oxygen (Pao2), diffusing capacity of the lung for carbon monoxide (Dlco), and BOS were better in the BLT group than in the SLT group. The advantages shown in the BLT group compared with the SLT group in regard to these variables increased with the prolongation of survival time. However, surgical time, ischemic time, postoperative intensive care unit days, and postoperative hospital days were shorter in the SLT group than in the BLT group. Overall survival, short-term OSR (1-3 y), BSFR-1y, in-hospital mortality, postoperative ventilator days, and postoperative complications (except BOS) were similar between the 2 groups. Bacterial pneumonia, graft failure, fungal infection, cardiac arrhythmia, and hemorrhage were the top 5 causes of in-hospital mortality. CONCLUSIONS: BLT appears to be associated with better long-term survival, better postoperative lung function, and less BOS compared with SLT. In-hospital mortality and postoperative complications (except BOS) were similar between the 2 groups.

Improved statistical fluctuation analysis for measurement-device-independent quantum key distribution with four-intensity decoy-state method.

Recently Zhang et al [ Phys. Rev. A95, 012333 (2017)] developed a new approach to estimate the failure probability for the decoy-state BB84 QKD system when taking finite-size key effect into account, which offers security comparable to Chernoff bound, while results in an improved key rate and transmission distance. Based on Zhang et al's work, now we extend this approach to the case of the measurement-device-independent quantum key distribution (MDI-QKD), and for the first time implement it onto the four-intensity decoy-state MDI-QKD system. Moreover, through utilizing joint constraints and collective error-estimation techniques, we can obviously increase the performance of practical MDI-QKD systems compared with either three- or four-intensity decoy-state MDI-QKD using Chernoff bound analysis, and achieve much higher level security compared with those applying Gaussian approximation analysis.

A comprehensive sediment dynamics study of a major mud belt system on the inner shelf along an energetic coast.

Globally mud areas on continental shelves are conduits for the dispersal of fluvial-sourced sediment. We address fundamental issues in sediment dynamics focusing on how mud is retained on the seabed on shallow inner shelves and what are the sources of mud. Through a process-based comprehensive study that integrates dynamics, provenance, and sedimentology, here we show that the key mechanism to keep mud on the seabed is the water-column stratification that forms a dynamic barrier in the vertical that restricts the upward mixing of suspended sediment. We studied the 1000 km-long mud belt that extends from the mouth of the Changjiang (Yangtze) River along the coast of Zhejiang and Fujian Provinces of China and ends on the west coast of Taiwan. This mud belt system is dynamically attached to the fluvial sources, of which the Changjiang River is the primary source. Winter is the constructive phase when active deposition takes place of fine-grained sediment carried mainly by the Changjiang plume driven by Zhe-Min Coastal Currents southwestward along the coast.

Anhydrates and hemihydrate of tasimelteon: Synthesis, structure, and pharmacokinetic study.

Two new crystal forms of tasimelteon TSM-I and TSM-II were reported here. Crystallization of crude in methanol or mixture solvent results in anhydrate crystal form (TSM-I) and hemihydrate crystal form (TSM-II) respectively. To the best of our knowledge, it is the first report about crystalline form of tasimelteon. The two crystal forms were exhaustively characterized by Scanning Electron Microscopy, Fourier Transform Infrared Spectroscopy, Raman Spectra, Differential Scanning Calorimetry, Thermogravimetric Analysis, Solid State NMR Spectroscopy and Powder X-ray diffraction and Single Crystal X-ray Diffraction. Furthermore, the pharmacokinetic behavior of TSM-I and TSM-II in rats were measured. We found that though TSM-II is considerably more soluble than TSM-I under water (pH = 7.0) and pH 1.2 buffer conditions, the bioavailability of TSM-Ivia oral administration was better compared to that of TSM-II.

Comparison of the crystal structures and thermochemistry of a novel soluble guanylate cyclase stimulator riociguat and its solvates.

Riociguat (Rio) is the first oral soluble guanylate cyclase stimulator to be approved for pulmonary arterial hypertension. In this study, form (II) of riociguat and three solvates with acetonitrile [form (III)], N,N-dimethylformamide [form (IV)] and ethyl acetate [form (V)] were crystallized. They were identified and characterized by differential scanning calorimetry, thermogravimetric analysis, X-ray powder diffraction, and their crystal structures were determined by single-crystal X-ray diffraction. No crystal structure has previously been reported for the known form (II) of riociguat. Crystal structure determination of Rio and its new solvates revealed that the dimeric R22(14) motif is common in both structures. The crystal packing of solvates adopts channel-like patterns, whereas form (II) of riociguat adopts sheet-like patterns. Strong π-π interactions exist in the above four forms. The conformation of the riociguat in one molecule of 0.5-DMF solvate was found to be significantly different from the conformations found in the other solvates. Desolvation of the three solvates was studied by thermogravimetric analysis and X-ray diffraction, and was shown to transform them into form (I) of riociguat.