Unusual case of Niemeier type II gallbladder perforation: case report and literature review.

Niemeier type II gallbladder perforation (GBP) is caused by inflammation and necrosis of the gallbladder wall followed by bile spilling into the abdominal cavity after perforation. The gallbladder then becomes adhered to the surrounding inflammatory tissue to form a purulent envelope, which communicates with the gallbladder. At present, the clinical characteristics and treatment of type II GBP are not well understood and management of GBP remains controversial. Type II GBP with gastric outlet obstruction is rare and prone to misdiagnosis and delayed treatment. Recent systematic reviews report that percutaneous drainage does not influence outcomes. In this current case, due to the high risk of bleeding and accidental injury, as well as a lack of access to safely visualize the Calot's triangle, the patient could not undergo laparoscopic cholecystectomy, which would have been the ideal option. This current case report presents the use of percutaneous laparoscopic drainage combined with percutaneous transhepatic gallbladder drainage in a patient with type II GBP associated with gastric outlet obstruction. A review of the relevant literature has been provided in addition to a summary of the clinical manifestations and treatments for type II GBP.

Glucose and HODEs regulate Aspergillus ochraceus quorum sensing through the GprC-AcyA pathway.

Aspergillus ochraceus is the traditional ochratoxin A (OTA)-producing fungus with density-dependent behaviors, which is known as quorum sensing (QS) that is mediated by signaling molecules. Individual cells trend to adapt environmental changes in a "whole" flora through communications, allowing fungus to occupy an important ecological niche. Signals perception, transmission, and feedback are all rely on a signal network that constituted by membrane receptors and intracellular effectors. However, the interference of density information in signal transduction, which regulates most life activities of Aspergillus, have yet to be elucidated. Here we show that the G protein-coupled receptor (GPCR) to cAMP pathway is responsible for transmitting density information, and regulates the key point in life cycle of A. ochraceus. Firstly, the quorum sensing phenomenon of A. ochraceus is confirmed, and identified the density threshold is 103 spores/mL, which represents the low density that produces the most OTA in a series quorum density. Moreover, the GprC that classified as sugar sensor, and intracellular adenylate cyclase (AcyA)-cAMP-PKA pathway that in response to ligands glucose and HODEs are verified. Furthermore, GprC and AcyA regulate the primary metabolism as well as secondary metabolism, and further affects the growth of A. ochraceus during the entire life cycle. These studies highlight a crucial G protein signaling pathway for cell communication that is mediated by carbohydrate and oxylipins, and clarified a comprehensive effect of fungal development, which include the direct gene regulation and indirect substrate or energy supply. Our work revealed more signal molecules that mediated density information and connected effects on important adaptive behaviors of Aspergillus ochraceus, hoping to achieve comprehensive prevention and control of mycotoxin pollution from interrupting cell communication.

A medical big data access control model based on smart contracts and risk in the blockchain environment.

The rapid development of the Hospital Information System has significantly enhanced the convenience of medical research and the management of medical information. However, the internal misuse and privacy leakage of medical big data are critical issues that need to be addressed in the process of medical research and information management. Access control serves as a method to prevent data misuse and privacy leakage. Nevertheless, traditional access control methods, limited by their single usage scenario and susceptibility to single point failures, fail to adapt to the polymorphic, real-time, and sensitive characteristics of medical big data scenarios. This paper proposes a smart contracts and risk-based access control model (SCR-BAC). This model integrates smart contracts with traditional risk-based access control and deploys risk-based access control policies in the form of smart contracts into the blockchain, thereby ensuring the protection of medical data. The model categorizes risk into historical and current risk, quantifies the historical risk based on the time decay factor and the doctor's historical behavior, and updates the doctor's composite risk value in real time. The access control policy, based on the comprehensive risk, is deployed into the blockchain in the form of a smart contract. The distributed nature of the blockchain is utilized to automatically enforce access control, thereby resolving the issue of single point failures. Simulation experiments demonstrate that the access control model proposed in this paper effectively curbs the access behavior of malicious doctors to a certain extent and imposes a limiting effect on the internal abuse and privacy leakage of medical big data.

Heat-Killed Saccharomyces boulardii Alleviates Dextran Sulfate Sodium-Induced Ulcerative Colitis by Restoring the Intestinal Barrier, Reducing Inflammation, and Modulating the Gut Microbiota.

Ulcerative colitis (UC) is a global intestinal disease, and conventional therapeutic drugs often fail to meet the needs of patients. There is an urgent need to find efficient and affordable novel biological therapies. Saccharomyces boulardii has been widely used in food and pharmaceutical research due to its anti-inflammatory properties and gut health benefits. However, there is still a relatively limited comparison and evaluation of different forms of S. boulardii treatment for UC. This study aimed to compare the therapeutic effects of S. boulardii, heat-killed S. boulardii, and S. boulardii ß-glucan on UC, to explore the potential of heat-killed S. boulardii as a new biological therapy. The results demonstrate that all three treatments were able to restore body weight, reduce the disease activity index (DAI), inhibit splenomegaly, shorten colon length, and alleviate histopathological damage to colonic epithelial tissues in DSS-induced colitis mice. The oral administration of S. boulardii, heat-killed S. boulardii, and S. boulardii ß-glucan also increased the levels of tight junction proteins (Occludin and ZO-1), decreased the levels of pro-inflammatory cytokines (TNF-α, IL-1ß, and IL-6) in the serum, and suppressed the expressions of TNF-α, IL-1ß, and IL-6 mRNA in the colon. In particular, in terms of gut microbiota, S. boulardii, heat-killed S. boulardii, and S. boulardii ß-glucan exhibited varying degrees of modulation on DSS-induced dysbiosis. Among them, heat-killed S. boulardii maximally restored the composition, structure, and functionality of the intestinal microbiota to normal levels. In conclusion, heat-killed S. boulardii showed greater advantages over S. boulardii and S. boulardii ß-glucan in the treatment of intestinal diseases, and it holds promise as an effective novel biological therapy for UC. This study is of great importance in improving the quality of life for UC patients and reducing the burden of the disease.

Construction Strategy of Functionalized Liposomes and Multidimensional Application.

Liposomes are widely used in the biological field due to their good biocompatibility and surface modification properties. With the development of biochemistry and material science, many liposome structures and their surface functional components have been modified and optimized one by one, pushing the liposome platform from traditional to functionalized and intelligent, which will better satisfy and expand the needs of scientific research. However, a main limiting factor effecting the efficiency of liposomes is the complicated environmental conditions in the living body. Currently, in order to overcome the above problem, functionalized liposomes have become a very promising strategy. In this paper, binding strategies of liposomes with four main functional elements, namely nucleic acids, antibodies, peptides, and stimuli-responsive motif have been summarized for the first time. In addition, based on the construction characteristics of functionalized liposomes, such as drug-carrying, targeting, long-circulating, and stimulus-responsive properties, a comprehensive overview of their features and respective research progress are presented. Finally, the paper critically presents the limitations of these functionalized liposomes in the current applications and also prospectively suggests the future development directions, aiming to accelerate realization of their industrialization.

Acid-resistant enzymes: the acquisition strategies and applications.

Enzymes have promising applications in chemicals, food, pharmaceuticals, and other variety products because of their high efficiency, specificity, and environmentally friendly properties. However, due to the complexity of raw materials, pH, temperature, solvents, etc., the application range of enzymes is greatly limited in the industry. Protein engineering and enzyme immobilization are classical strategies to overcome the limitations of industrial applications. Although the pH tendency of enzymes has been extensively researched, the mechanism underlying enzyme acid resistance is unclear, and a less practical strategy for altering the pH propensity of enzymes has been suggested. This review proposes that the optimum pH of enzyme is determined by the pKa values of active center ionizable amino acid residues. Three levels of acquiring acid-resistant enzymes are summarized: mining from extreme environments and enzyme databases, modification with protein engineering and enzyme microenvironment engineering, and de novo synthesis. The industrial applications of acid-resistant enzymes in chemicals, food, and pharmaceuticals are also summarized. KEY POINTS: • The mechanism of enzyme acid resistance is fundamentally determined. • The three aspects of the method for acquiring acid-resistant enzymes are summarized. • Computer-aided strategies and artificial intelligence are used to obtain acid-resistant enzymes.

Enhancing the thermostability of carboxypeptidase A by a multiple computer-aided rational design based on amino acids preferences at ß-turns.

Carboxypeptidase A (CPA) with efficient hydrolysis ability has shown vital potential in food and biological fields. In addition, it is also the earliest discovered enzyme with Ochratoxin A (OTA) degradation activity. Thermostability plays an imperative role to catalyze the reactions at high temperatures in industry, but the poor thermostability of CPA restricts its industrial application. In order to improve the thermostability of CPA, flexible loops were predicted through molecular dynamics (MD) simulation. Based on the amino acid preferences at ß-turns, three ΔΔG-based computational programs (Rosetta, FoldX and PoPMuSiC) were employed to screen three variants from plentiful candidates and MD simulations were then used to verify two potential variants with enhanced thermostability (R124K and S134P). Results showed that compared to the wild-type CPA, the variants S134P and R124K exhibited rise of 4.2 min and 7.4 min in half-life (t1/2) at 45 °C, 3 °C and 4.1 °C in the half inactivation temperature (T5010), in addition to increase by 1.9 °C and 1.2 °C in the melting temperature (Tm), respectively. The mechanism responsible for the enhanced thermostability was elucidated through the comprehensive analysis of molecular structure. This study shows that the thermostability of CPA can be improved by the multiple computer-aided rational design based on amino acid preferences at ß-turns, broadening its industrial applicability of OTA degradation and providing a valuable strategy for the protein engineering of mycotoxin degrading enzymes.

Quorum Sensing-Mediated Lipid Oxidation Further Regulating the Environmental Adaptability of Aspergillus ochraceus.

Quorum sensing (QS) is a cellular strategy of communication between intra- and inter-specific microorganisms, characterized by the release of quorum sensing molecules (QSMs) that achieve coordination to adaptation to the environment. In Aspergillus, lipids carry population density-mediated stresses, and their oxidative metabolite oxylipins act as signaling to transmit information inside cells to regulate fungal development in a synchronized way. In this study, we investigated the regulation of density-dependent lipid metabolism in the toxigenic fungi Aspergillus ochraceus by the oxidative lipid metabolomics in conjunction with transcriptomics. In addition to proven hydroxyoctadecadienoic acids (HODEs), prostaglandins (PGs) also appear to have the properties of QSM. As a class of signaling molecule, oxylipins regulate the fungal morphology, secondary metabolism, and host infection through the G protein signaling pathway. The results of combined omics lay a foundation for further verification of oxylipin function, which is expected to elucidate the complex adaptability mechanism in Aspergillus and realize fungal utilization and damage control.

Establishment, optimization, and application of genetic technology in Aspergillus spp.

Aspergillus is widely distributed in nature and occupies a crucial ecological niche, which has complex and diverse metabolic pathways and can produce a variety of metabolites. With the deepening of genomics exploration, more Aspergillus genomic informations have been elucidated, which not only help us understand the basic mechanism of various life activities, but also further realize the ideal functional transformation. Available genetic engineering tools include homologous recombinant systems, specific nuclease based systems, and RNA techniques, combined with transformation methods, and screening based on selective labeling. Precise editing of target genes can not only prevent and control the production of mycotoxin pollutants, but also realize the construction of economical and efficient fungal cell factories. This paper reviewed the establishment and optimization process of genome technologies, hoping to provide the theoretical basis of experiments, and summarized the recent progress and application in genetic technology, analyzes the challenges and the possibility of future development with regard to Aspergillus.

Purification and characterization of the enzymes from Brevundimonas naejangsanensis that degrade ochratoxin A and B.

Ochratoxin A (OTA) and Ochratoxin B (OTB) co-contaminate many types of agricultural products. Screening enzymes that degrade both OTA and OTB has significance in food safety. In this study, four novel OTA and OTB degrading enzymes, namely BnOTase1, BnOTase2, BnOTase3, and BnOTase4, were purified from the metabolites of the Brevundimonas naejangsanensis ML17 strain. These four enzymes hydrolyzed OTA into OTα and hydrolyzed OTB into OTß. BnOTase1, BnOTase2, BnOTase3, and BnOTase4 have the apparent Km values for hydrolyzing OTA of 19.38, 0.92, 12.11, 1.09 µmol/L and for hydrolyzing OTB of 0.76, 2.43, 0.60, 0.64 µmol/L respectively. OTα and OTß showed no significant cytotoxicity to HEK293 cells, suggesting that these enzymes mitigate the toxicity of OTA and OTB. The discovery of the novel OTA and OTB degrading enzymes enriches the research on ochratoxin control and provides objects for protein rational design.

Both Saccharomyces boulardii and Its Postbiotics Alleviate Dextran Sulfate Sodium-Induced Colitis in Mice, Association with Modulating Inflammation and Intestinal Microbiota.

OBJECTIVE: To investigate the effect of Saccharomyces boulardii and its freeze-dried and spray-dried postbiotics on the intervention and potential mechanism of dextran sulfate sodium (DSS)-induced ulcerative colitis in mice. [Methods] After the acclimation period of C67BL/6J mice, a colitis model was constructed by applying 2% DSS for 7 d, followed by 7 d of intervention. Subsequently, the disease activity index (DAI), organ index, colon length, colon HE staining of pathological sections, ELISA for blood inflammatory factors (Interleukin (IL)-1ß, IL-6, IL-10, Tumor necrosis factor (TNF)-α), Real time quantitative polymerase chain reaction (RT-qPCR) to determine the levels of colonic inflammatory factors (IL-1ß, IL-6, IL-10, TNF-α), Occludin gene expression, and intestinal flora were assessed to evaluate the protective effects of S. boulardii and its postbiotics on colitis in mice. RESULTS: Compared with the DSS group, S. boulardii and the postbiotics interventions effectively improved colonic shortening and tissue damage, increased the expression of intestinal tight junction protein, reduced the secretion of pro-inflammatory factors, increased the secretion of anti-inflammatory factors, and maintained the homeostasis of intestinal microorganisms. Postbiotics intervention is better than probiotics. CONCLUSIONS: S. boulardii and its postbiotics can effectively alleviate DSS-induced colitis in mice through modulating host immunity and maintaining intestinal homeostasis. Postbiotics are promising next-generation biotherapeutics for ulcerative colitis treatment.

Small Peptides in the Detection of Mycotoxins and Their Potential Applications in Mycotoxin Removal.

Mycotoxins pose significant risks to humans and livestock. In addition, contaminated food- and feedstuffs can only be discarded, leading to increased economic losses and potential ecological pollution. Mycotoxin removal and real-time toxin level monitoring are effective approaches to solve this problem. As a hot research hotspot, small peptides derived from phage display peptide libraries, combinatorial peptide libraries, and rational design approaches can act as coating antigens, competitive antigens, and anti-immune complexes in immunoassays for the detection of mycotoxins. Furthermore, as a potential approach to mycotoxin degradation, small peptides can mimic the natural enzyme catalytic site to construct artificial enzymes containing oxidoreductases, hydrolase, and lyase activities. In summary, with the advantages of mature synthesis protocols, diverse structures, and excellent biocompatibility, also sharing their chemical structure with natural proteins, small peptides are widely used for mycotoxin detection and artificial enzyme construction, which have promising applications in mycotoxin degradation. This paper mainly reviews the advances of small peptides in the detection of mycotoxins, the construction of peptide-based artificial enzymes, and their potential applications in mycotoxin control.

Co-Occurrence of Staphylococcus aureus and Ochratoxin A in Pasteurized Milk.

Pathogens and mycotoxins are serious public health risks for humans and food safety in milk. This study concentrated on detecting Staphylococcus aureus and Ochratoxin A (OTA) in 210 pasteurized milk from ten urban Beijing districts to suggest the co-occurrence of S. aureus with toxin-producing genes and OTA in milk and the possible risk. S. aureus was identified by physiological and biochemical experiments and molecular biology experiments, and enterotoxin genes were identified by PCR. OTA was detected by LC-MS/MS. The study found 29 isolates of S. aureus, of which 17.24% had the sea gene encoding enterotoxin A. OTA was detected in 31 out of 120 samples and the maximum amount of detection was 18.8 µg/kg. The results of this study indicate that when failing to guarantee the cold chain, the presence of S. aureus with enterotoxin genes in milk will present a risk to food safety. Furthermore, the high detection rates and levels of OTA in milk suggest that OTA is a hidden risk. The co-occurrence of S. aureus and OTA in milk is a food safety concern and there is a need to control the occurrence of these two biohazards in milk.

Genome mining reveals the genes of carboxypeptidase for OTA-detoxification in Bacillus subtilis CW14.

Bacillus subtilis CW14, isolated from fresh elk droppings in Beijing Zoo, is a Gram-positive, conferred Generally Recognized as Safe (GRAS) bacterium with the capacity of ochratoxin A (OTA) detoxification. The genome sequence of the CW14 strain showed a size of 4,287,522 bp with 44.06% GC content. It was predicted many putative enzymes involved in degrading mycotoxin by analyzing the signal peptides and the transmembrane regions. Nine extracellular enzymes were predicted relating to OTA detoxification, including four D-Ala-D-Ala carboxypeptidases, two hydrolases, two amidases, and one lactamase. Indeed, two of the carboxypeptidase genes dacA and dacB, expressed in Escherichia coli, were verified contributing to OTA detoxification. DacA and OTA were mixed incubated for 24 h, and the degradation rate reached 71.3%. After purification, the concentration of recombinant DacA protein was 0.5 mg/mL. Bacillus subtilis CW14 and its carboxypeptidases may be used as OTA detoxification agents in food and feed industry production.

Perfluoroalkyl substances (PFASs) in leachate, fly ash, and bottom ash from waste incineration plants: Implications for the environmental release of PFAS.

Perfluoroalkyl substances (PFASs) are a family of chemicals widely distributed in daily use consumer products. Most of these products become municipal solid wastes (MSWs) after they have been used. In the present study, we examined different types of PFASs in leachate, fly ash and bottom ash produced from three MSW incineration plants in southern China. High PFAS levels were found in leachate (mean concentration 215 ng/mL, range 21.4-682 ng/mL) from the incineration plants, which indicated large amounts of PFASs in the wastes leached out. The average quantities of PFASs annually discharged from the leachates of the three plants were estimated to be approximately 384 kg (Plant A), 47.3 kg (Plant B), and 2.82 kg (Plant C). Relatively lower levels of PFASs in fly ash (mean 16.4 ng/g, range 1.46-87.6 ng/g) and bottom ash (mean 14.6 ng/g, range 3.11-77.4 ng/g) indicated that high-temperature incineration destroyed most of the PFASs. The wide array of PFASs concentrations in all three matrices illustrated that some PFASs-containing industrial wastes were still entered into local MSW. In general, short chain PFASs, including perfluorobutyric acid (PFBA) and perfluorobutane sulfonate (PFBS), were the primary PFASs in leachate samples. In addition, PFOS was the predominant PFASs in fly ash samples. The results showed that leachate, fly ash, and bottom ash from MSW incineration plants are important vectors of PFASs.

The degradation mechanisms of perfluorooctanoic acid (PFOA) and perfluorooctane sulfonic acid (PFOS) by different chemical methods: A critical review.

Per- and polyfluoroalkyl substances (PFASs) are a class of artificial compounds comprised of a perfluoroalkyl main chain and a terminal functional group. With them being applied in a wide range of applications, PFASs have drawn increasing regulatory attention and research interests on their reductions and treatments due to their harmful effects on environment and human beings. Among numerous studies, chemical treatments (e.g., photochemical, electrochemical, and thermal technologies) have been proved to be important methods to degradation PFASs. However, the pathways and mechanisms for the degradation of PFASs through these chemical methods still have not been well documented. This article therefore provides a comprehensive review on the degradation mechanisms of two important PFASs (perfluorooctanoic acid (PFOA) and perfluorooctane sulfonic acid (PFOS)) with photochemical, electrochemical and thermal methods. Different decomposition mechanisms of PFOA and PFOS are reviewed and discussed. Overall, the degradation pathways of PFASs are associated closely with their head groups and chain lengths, and H/F exchange and chain shortening were found to be predominant degradation mechanisms. The clear study on the degradation mechanisms of PFOA and PFOS should be very useful for the complete degradation or mineralization of PFASs in the future.

Occurrence and Ecological Risk Assessment of Perfluoroalkyl Substances (PFASs) in Water and Sediment from an Urban River in South China.

The chemical substances in urban rivers influence municipal water systems and reflect the recent use of these chemicals by humans or industries around the urban center. In this study, seven perfluoroalkyl substances (PFASs)-perfluorohexanoic acid (PFHxA), perfluoroheptanoic acid (PFHpA), perfluorooctanoic acid (PFOA), perfluorononanoic acid (PFNA), 2-perfluorohexyl ethanol (6:2 FTOH), 2-perfluorooctyl ethanol (8:2 FTOH), and 6:2 chlorinated polyfluoroalkyl ether sulfonic acids (F-53B)-could be detected and quantified in river water and sediment samples collected from one tributary of the Liuxi River, which is part of Pearl River near Guangzhou in Guangdong province, South China. The fluxes of target PFASs into Liuxi River and their related ecological risks were further estimated. The total concentrations of PFASs (ΣPFASs) ranged from 506 to 3.16 × 103 ng/L in water samples and 9.13 to 850 ng/L in sediment samples. The two dominant PFAS compounds were 6:2 FTOH and PFHpA, which accounted for more than 90.0% of ΣPFASs in river water and sediment. Correlation analysis showed that there was significant positive correlation (p < 0.01) between two selected PFASs (e.g., between 6:2 FTOH and PFHpA). Correlation analysis of PFASs in river water and sediment indicated most PFASs in sediment were partitioned from river water. The ecological risk assessment indicated that the detected PFASs have a low risk (HQ < 0.1) in river water and sediment to Daphnia magna in the Liuxi River.

Multidimensional analysis of the epigenetic alterations in toxicities induced by mycotoxins.

Mycotoxins contaminate all types of food and feed, threatening human and animal health through food chain accumulation, producing various toxic effects. Increasing attention is being focused on the molecular mechanism of mycotoxin-induced toxicity in all kinds of in vivo and in vitro models. Epigenetic alterations, including DNA methylation, non-coding RNAs (ncRNAs), and protein post-translational modifications (PTMs), were identified as being involved in various types of mycotoxin-induced toxicity. In this review, the emphasis was on summarizing the epigenetic alterations induced by mycotoxin, including aflatoxin B1 (AFB1), ochratoxin A (OTA), zearalenone (ZEA), fumonisin B1 (FB1), and deoxynivalenol (DON). This review summarized and analyzed the roles of DNA methylation, ncRNAs, and protein PTMs after mycotoxin exposure based on recently published papers. Moreover, the main research methods and their deficiencies were determined, while some remedial suggestions are proposed. In summary, this review helps to understand better the epigenetic alterations induced by the non-genotoxic effects of mycotoxin.

Fungal G-Protein-Coupled Receptors: A Promising Mediator of the Impact of Extracellular Signals on Biosynthesis of Ochratoxin A.

G-protein-coupled receptors (GPCRs) are transmembrane receptors involved in transducing signals from the external environment inside the cell, which enables fungi to coordinate cell transport, metabolism, and growth to promote their survival, reproduction, and virulence. There are 14 classes of GPCRs in fungi involved in sensing various ligands. In this paper, the synthesis of mycotoxins that are GPCR-mediated is discussed with respect to ligands, environmental stimuli, and intra-/interspecific communication. Despite their apparent importance in fungal biology, very little is known about the role of ochratoxin A (OTA) biosynthesis by Aspergillus ochraceus and the ligands that are involved. Fortunately, increasing evidence shows that the GPCR that involves the AF/ST (sterigmatocystin) pathway in fungi belongs to the same genus. Therefore, we speculate that GPCRs play an important role in a variety of environmental signals and downstream pathways in OTA biosynthesis. The verification of this inference will result in a more controllable GPCR target for control of fungal contamination in the future.

Water flushing irremovable biofilms on support material in dynamic membrane bioreactor: Formation, composition, and microbial community.

Dynamic membrane bioreactors mainly rely on the in-situ formed biofilms on support materials to reject fine particles in water. The development of irremovable biofilms on support materials can decrease the cleaning efficiency when removing the unwanted biofilms with low permeability by water flushing. In the present study, the initial formed biofilms on support materials at 5-day solids retention time (SRT) were removable by water flushing. After repeated cleaning with off-line water flushing during operation, however, irremovable biofilms were developed gradually inside the mesh pores and thus, rapid rising in transmembrane pressure occurred in every one to three days. At 20-day SRT, the biofilms formed on support materials with the same operation time were still removable. Therefore, both low SRT and repeated water flushing promoted the formation of irremovable biofilms on support materials. Further study found that the composition and microbial community between the irremovable and removable biofilms were significantly different, which differentiated the biofilm adhesion and removability. The irremovable biofilms had a greater faction of proteins (49.0%) and ß-d-glucopyranose polysaccharides (17.8%) in extracellular polymeric substance (EPS), while the removable biofilms had a greater fraction of α-d-glucopyranose polysaccharides. After repeated cleaning with off-line water flushing during operation, Nitrospiraceae was selectively enriched in the irremovable biofilms at a relative abundance of 39.1%, which could have resulted in the particular EPS matrix that strengthened the biofilm adhesion.