Crotonylation of NAE1 Modulates Cardiac Hypertrophy via Gelsolin Neddylation.

BACKGROUND: Cardiac hypertrophy and its associated remodeling are among the leading causes of heart failure. Lysine crotonylation is a recently discovered posttranslational modification whose role in cardiac hypertrophy remains largely unknown. NAE1 (NEDD8-activating enzyme E1 regulatory subunit) is mainly involved in the neddylation modification of protein targets. However, the function of crotonylated NAE1 has not been defined. This study aims to elucidate the effects and mechanisms of NAE1 crotonylation on cardiac hypertrophy. METHODS: Crotonylation levels were detected in both human and mouse subjects with cardiac hypertrophy through immunoprecipitation and Western blot assays. TMT-labeled quantitative lysine crotonylome analysis was performed to identify the crotonylated proteins in a mouse cardiac hypertrophic model induced by transverse aortic constriction. We generated NAE1 knock-in mice carrying a crotonylation-defective lysine to arginine K238R (lysine to arginine mutation at site 238) mutation (NAE1 K238R) and NAE1 knock-in mice expressing a crotonylation-mimicking lysine to glutamine K238Q (lysine to glutamine mutation at site 238) mutation (NAE1 K238Q) to assess the functional role of crotonylation of NAE1 at K238 in pathological cardiac hypertrophy. Furthermore, we combined coimmunoprecipitation, mass spectrometry, and dot blot analysis that was followed by multiple molecular biological methodologies to identify the target GSN (gelsolin) and corresponding molecular events contributing to the function of NAE1 K238 crotonylation. RESULTS: The crotonylation level of NAE1 was increased in mice and patients with cardiac hypertrophy. Quantitative crotonylomics analysis revealed that K238 was the main crotonylation site of NAE1. Loss of K238 crotonylation in NAE1 K238R knock-in mice attenuated cardiac hypertrophy and restored the heart function, while hypercrotonylation mimic in NAE1 K238Q knock-in mice significantly enhanced transverse aortic constriction-induced pathological hypertrophic response, leading to impaired cardiac structure and function. The recombinant adenoviral vector carrying NAE1 K238R mutant attenuated, while the K238Q mutant aggravated Ang II (angiotensin II)-induced hypertrophy. Mechanistically, we identified GSN as a direct target of NAE1. K238 crotonylation of NAE1 promoted GSN neddylation and, thus, enhanced its protein stability and expression. NAE1 crotonylation-dependent increase of GSN promoted actin-severing activity, which resulted in adverse cytoskeletal remodeling and progression of pathological hypertrophy. CONCLUSIONS: Our findings provide new insights into the previously unrecognized role of crotonylation on nonhistone proteins during cardiac hypertrophy. We found that K238 crotonylation of NAE1 plays an essential role in mediating cardiac hypertrophy through GSN neddylation, which provides potential novel therapeutic targets for pathological hypertrophy and cardiac remodeling.

The prognostic predictive value of the components of the PR interval in hospitalized patients with heart failure.

OBJECTIVE: Previous reports on the epidemiology, influencing factors, and the prognostic value of the components of PR interval in hospitalized heart failure patients were limited. METHODS: This study retrospectively enrolled 1182 patients hospitalized with heart failure from 2014 to 2017. Multiple linear regression analysis was used to explore the association between the components of PR interval and the baseline parameters. The primary outcome was all-cause death or heart transplantation. Multivariable-adjusted Cox proportional hazard regression models were constructed to explore the predictive value of the components of PR interval for the primary outcome. RESULTS: In multiple linear regression analysis, higher height (for every 10 cm increase in height: regression coefficient 4.83, P < 0.001) as well as larger atrial and ventricular size were associated with larger P wave duration but not with PR segment. The primary outcome occurred in 310 patients after an average follow-up of 2.39 years. Cox regression analyses revealed that the increase in PR segment was an independent predictor of the primary outcome (every 10 ms increase: hazard ratio 1.041, 95% confidence interval [CI] 1.010-1.083, P = 0.023), whereas the P wave duration did not show significant correlation. When adding the PR segment to an initial prognostic prediction model, the likelihood ratio test and categorical net reclassification index (NRI) showed a significant improvement, but the increase in C-index was not significant. In subgroup analysis, increased PR segment was an independent predictor of the primary endpoint in patients taller than 170 cm (each 10 ms increase: hazard ratio 1.153, 95% CI 1.085-1.225, P < 0.001) but not the shorter group (P for interaction = 0.006). CONCLUSIONS: In hospitalized patients with heart failure, longer PR segment was an independent predictor of the composite endpoint of all-cause death and heart transplantation, especially in the taller group, but it had limited significance in improving the prognostic risk stratification of this population.

Effect of sodium-glucose cotransporter 2 inhibitors on cardiac structure and function in type 2 diabetes mellitus patients with or without chronic heart failure: a meta-analysis.

BACKGROUND: Although the benefits of sodium-glucose cotransporter 2 inhibitors (SGLT2i) on cardiovascular events have been reported in patients with type 2 diabetes mellitus (T2DM) with or without heart failure (HF), the impact of SGLT2i on cardiac remodelling remains to be established. METHODS: We searched the PubMed, Embase, Cochrane Library and Web of Science databases up to November 16th, 2020, for randomized controlled trials reporting the effects of SGLT2i on parameters of cardiac structure, cardiac function, plasma N-terminal pro-brain natriuretic peptide (NT-proBNP) level or the Kansas City Cardiomyopathy Questionnaire (KCCQ) score in T2DM patients with or without chronic HF. The effect size was expressed as the mean difference (MD) or standardized mean difference (SMD) and its 95% confidence interval (CI). Subgroup analyses were performed based on the stage A-B or stage C HF population and HF types. RESULTS: Compared to placebo or other antidiabetic drugs, SGLT2i showed no significant effects on left ventricular mass index, left ventricular end diastolic volume index, left ventricular end systolic volume index, or left atrial volume index. SGLT2i improved left ventricular ejection fraction only in the subgroup of HF patients with reduced ejection fraction (MD 3.16%, 95% CI 0.11 to 6.22, p = 0.04; I2 = 0%), and did not affect the global longitudinal strain in the overall analysis including stage A-B HF patients. SGLT2i showed benefits in the E/e' ratio (MD - 0.45, 95% CI - 0.88 to - 0.03, p = 0.04; I2 = 0%), plasma NT-proBNP level (SMD - 0.09, 95% CI - 0.16 to - 0.03, p = 0.004; I2 = 0%), and the KCCQ score (SMD 3.12, 95% CI 0.76 to 5.47, p  = 0.01; I2 = 0%) in the overall population. CONCLUSION: The use of SGLT2i was associated with significant improvements in cardiac diastolic function, plasma NT-proBNP level, and the KCCQ score in T2DM patients with or without chronic HF, but did not significantly affect cardiac structural parameters indexed by body surface area. The LVEF level was improved only in HF patients with reduced ejection fraction.

A simple method for in vitro preparation of natural killer cells from cord blood.

BACKGROUND: Cord Blood (CB) has been considered a promising source of natural killer (NK) cells for cellular immunotherapy. However, it is difficult to expand the large numbers of highly pure NK cells from CB without cell sorting and feeder cells/multiple cytokines. In this study, we try to develop a simple, safe and economical method for ex vivo expansion and purification of NK cells from CB without cell sorting and feeder cells/multiple cytokines. RESULTS: The large numbers (mean: 1.59 × 1010) of highly pure (≥90%) NK cells from CB could be obtained through interleukin-2, group A streptococcus and zoledronate stimulation of mononuclear cells using the 21-day culture approach. When compared to resting NK cells, expanded NK cells were a higher expression of activating receptors CD16, NKG2D, NKp30, NKp44, NKp46 and activating markers CD62L and CD69, while the inhibitory receptors, CD158a and CD158b remained largely unchanged. In addition, these cells showed a higher concentration of IFN-γ, TNF-α and GM-CSF secretion and cytotoxicity to K562 cells and acute myeloid leukemia targets than resting NK cells. CONCLUSION: We develop a simple, safe and economical method to obtain high yield, purity, and functionality NK cells from CB without cell sorting and feeder cells/multiple cytokines.

Stepwise Assembly of MII7 Clusters Revealed by Mass Spectrometry, EXAFS, and Crystallography.

The square-planar monomer NiL2 (Ni1 ), L=2-ethoxy-6-(N-methyl-iminomethyl)phenolate, reacts with M(H2 O)6 (ClO4 )2 , M=Ni or Co, to form heptanuclear disks [Cox Ni7-x (OH)6 (L)6 ](ClO4 )2 ⋅2 CH3 CN (Cox Ni7-x , x=0-7) and the co-crystal [Cox Ni7-x (OH)6 L6 ][NiL2 ](ClO4 )2 ⋅2 CH3 CN (Cox Ni7-x -Ni1 ) under ambient conditions. It has proved possible to explore the bottom-up assembly process of Cox Ni7-x and Cox Ni7-x -Ni1 in real time. The final products have been characterized by thermogravimetric analysis, IR, elemental analysis, ICP-MS, and single-crystal X-ray diffraction. Time-dependent mass spectrometry (MS) revealed the following reaction steps: Ni1 →[M2 L3 ]+ →[M4 (OH)2 L4 ]2+ →[M7 (OH)6 L6 ]2+ . In contrast, the reaction of Ni1 with Zn2+ only reaches halfway, and crystallographic evidence indicates a butterfly structure for [Zn2 Ni2 (OH)2 Cl2 ] (Zn2 Ni2 ), an intermediate that is difficult to isolate in the above Ni-Co series. A summation method has been used to analyze the MS of bimetallic clusters with very similar atomic masses, as is the case for Co and Ni. The results provide ample information on the distribution of Co and Ni within each cluster and their statistical distribution within selected crystals.

Carbohydrate antigens as potential biomarkers for the malignancy in patients with idiopathic deep venous thrombosis: a retrospective cohort study.

A variety of biomarkers have been identified in recent prospective and retrospective reports as being potentially predictive of venous thromboembolis (VTE), particularly idiopathic deep venous thrombosis (IDVT). This study identified a serum tumor biomarker for early screening of IDVT. A total of 128 IDVT patients (54 females and 74 males; average age: 50.9±17.4 years) were included. Carcinoembryonic antigen (CEA), ferritin, ß2-microglobulin, cancer antigen (CA) 125, CA 15-3, CA 19-9, squamous cell carcinoma antigen (SCC), alpha-fetoprotein (AFP), prostate specific antigen (PSA), free PSA (f-PSA), and beta-human chorionic gonadotropin (ß-HCG) in patients with IDVT were detected. Malignancies were histo- or cytopathologically confirmed. Of the 128 IDVT patients, 16 (12.5%) were found to have malignancies. Serum CEA, CA 125, CA 15-3, and CA 19-9 were found to be helpful for detecting malignancies in IDVT patients. Our study revealed a positive association between these markers and tumors in IDVT patients. On the other hand, SCC and AFP were not sensitive enough to be markers for detecting tumors in patients with IDVT. No significant differences were found in positive rates of ferritin and ß2-microglobulin between tumor and non-tumor groups, and no significant difference exists in serum levels of ferritin and ß2-microglobulin between the two groups. Carbohydrate antigens, CA 15-3 in particular, may be useful for differential diagnosis and prediction of malignancies in patients with IDVT.

Recent advances of PIWI-interacting RNA in cardiovascular diseases.

BACKGROUND: The relationship between noncoding RNAs (ncRNAs) and human diseases has been a hot topic of research, but the study of ncRNAs in cardiovascular diseases (CVDs) is still in its infancy. PIWI-interacting RNA (piRNA), a small ncRNA that binds to the PIWI protein to maintain genome stability by silencing transposons, was widely studied in germ lines and stem cells. In recent years, piRNA has been shown to be involved in key events of multiple CVDs through various epigenetic modifications, revealing the potential value of piRNA as a new biomarker or therapeutic target. CONCLUSION: This review explores origin, degradation, function, mechanism and important role of piRNA in CVDs, and the promising therapeutic targets of piRNA were summarized. This review provide a new strategy for the treatment of CVDs and lay a theoretical foundation for future research. KEY POINTS: piRNA can be used as a potential therapeutic target and biomaker in CVDs. piRNA influences apoptosis, inflammation and angiogenesis by regulating epigenetic modificaions. Critical knowledge gaps remain in the unifying piRNA nomenclature and PIWI-independent function.

[Analysis of intraoperative molecular assessment of sentinel lymph nodes in breast carcinoma].

OBJECTIVE: To evaluate the reliability and application of GeneSearch(TM) breast lymph node assay (Genesearch), a real-time fluorescence quatitative PCR method, in intraoperative assay of metastasis in sentinel lymph nodes (SLNs) from breast cancer patients. METHODS: Totally 140 SLNs from 80 patients with breast carcinoma were prospectively studied from May 2010 to August 2010. The 80 patients included 78 women and 2 men who ranged in age from 29 to 85 years, and the median age is 49 years. The expression of CK19 and mammaglobulin in all 140 SLNs were detected by Genesearch, and the results were compared with that of histological evaluation of both frozen and paraffin-embedded sections. RESULTS: Among SLNs, by histological analyses, there were 121 without metastasis, 17 with macrometastasis, 2 with micrometastasis, and none of isolated tumor cell. By Genesearch, there were 119 without metastasis and 21 with metastasis. Genesearch showed sensitivity of 89.4%, positive predictive value of 81.0%, negative predictive value of 98.3% and specificity of 96.7% by comparing to histological analyses. The concordance between Genesearch and histological analysis was 95.7%. The sensitivity of Genesearch was 15/17 for macrometastasis and 2/2 for micrometastasis. CONCLUSIONS: Genesearch detection presents high sensitivity and specificity in evaluating metastasis of sentinel lymph nodes in breast cancer, but strict performance technically is necessary to avoid false positive and false negative results. Inability of further subtyping for the positive cases might be the key limitations for wide application of this method.

TMEM11 regulates cardiomyocyte proliferation and cardiac repair via METTL1-mediated m7G methylation of ATF5 mRNA.

The mitochondrial transmembrane (TMEM) protein family has several essential physiological functions. However, its roles in cardiomyocyte proliferation and cardiac regeneration remain unclear. Here, we detected that TMEM11 inhibits cardiomyocyte proliferation and cardiac regeneration in vitro. TMEM11 deletion enhanced cardiomyocyte proliferation and restored heart function after myocardial injury. In contrast, TMEM11-overexpression inhibited neonatal cardiomyocyte proliferation and regeneration in mouse hearts. TMEM11 directly interacted with METTL1 and enhanced m7G methylation of Atf5 mRNA, thereby increasing ATF5 expression. A TMEM11-dependent increase in ATF5 promoted the transcription of Inca1, an inhibitor of cyclin-dependent kinase interacting with cyclin A1, which suppressed cardiomyocyte proliferation. Hence, our findings revealed that TMEM11-mediated m7G methylation is involved in the regulation of cardiomyocyte proliferation, and targeting the TMEM11-METTL1-ATF5-INCA1 axis may serve as a novel therapeutic strategy for promoting cardiac repair and regeneration.

Increased risks of antibiotic resistant genes (ARGs) induced by chlorine disinfection in the reverse osmosis system for potable reuse of reclaimed water.

The reverse osmosis (RO) process has been applied extensively in wastewater reclamation for industrial and potable reuse. To prevent biofouling, chlorine disinfection was usually used in pretreatment. However, this study found that chlorine disinfection could significantly increase risks of antibiotic resistant genes (ARGs) in the RO system. With the increase of chlorine concentration from 0 to 5 mg/L, the accumulative relative abundance of 14 common ARGs in the membrane foulants increased by 49.6%. Among these ARGs, tolC, acrA and acrB (resistance to multiple drugs) showed the highest increament after chlorine disinfection. Especially, the relative abundance of tolC in the group with 5 mg/L chlorine increased by 113.3% compared with the control group. These ARGs tended to be enriched in a few bacterial genus, including Candidatus, Thiomonas, Silanimonas, Xanthomonas and Pseudomonas. These results indicated that the foulants on RO membranes might become a potential sink of ARGs. Considering the possibility of membrane breach, the ARGs may contaminate the permeate and bring great biological risks.

Emerging roles of ferroptosis in cardiovascular diseases.

The mechanism of cardiovascular diseases (CVDs) is complex and threatens human health. Cardiomyocyte death is an important participant in the pathophysiological basis of CVDs. Ferroptosis is a new type of iron-dependent programmed cell death caused by excessive accumulation of iron-dependent lipid peroxides and reactive oxygen species (ROS) and abnormal iron metabolism. Ferroptosis differs from other known cell death pathways, such as apoptosis, necrosis, necroptosis, autophagy and pyroptosis. Several compounds have been shown to induce or inhibit ferroptosis by regulating related key factors or signalling pathways. Recent studies have confirmed that ferroptosis is associated with the development of diverse CVDs and may be a potential therapeutic drug target for CVDs. In this review, we summarize the characteristics and related mechanisms of ferroptosis and focus on its role in CVDs, with the goal of inspiring novel treatment strategies.

The circRNA CNEACR regulates necroptosis of cardiomyocytes through Foxa2 suppression.

Circular RNAs (circRNAs) are differentially expressed in various cardiovascular disease including myocardial ischemia-reperfusion (I/R) injury. However, their functional impact on cardiomyocyte cell death, in particular, in necrotic forms of death remains elusive. In this study, we found that the level of mmu_circ_000338, a cardiac- necroptosis-associated circRNA (CNEACR), was reduced in hypoxia-reoxygenation (H/R) exposed cardiomyocytes and I/R-injured mice hearts. The enforced expression of CNEACR attenuated the necrotic form of cardiomyocyte death caused by H/R and suppressed of myocardial necrosis in I/R injured mouse heart, which was accompanied by a marked reduction of myocardial infarction size and improved cardiac function. Mechanistically, CNEACR directly binds to histone deacetylase (HDAC7) in the cytoplasm and interferes its nuclear entry. This leads to attenuation of HDAC7-dependent suppression of forkhead box protein A2 (Foxa2) transcription, which can repress receptor-interacting protein kinase 3 (Ripk3) gene by binding to its promoter region. In addition, CNEACR-mediated upregulation of FOXA2 inhibited RIPK3-dependent necrotic/necroptotic death of cardiomyocytes. Our study reveals that circRNAs such as CNEACR can regulate the cardiomyocyte necroptosis associated activity of HDACs, promotes cell survival and improves cardiac function in I/R-injured heart. Hence, the CNEACR/HDAC7/Foxa2/ RIPK3 axis could be an efficient target for alleviating myocardial damage caused by necroptotic death in ischemia heart diseases.

Microwave versus traditional solvothermal synthesis of Ni7(II) discs: effect of ligand on exchange reaction in solution studied by electrospray ionization-mass spectroscopy and magnetic properties.

A comparative evaluation of the solvothermal method of synthesis of magnetic Ni(7) discs with four different ligands using the same concentrations of reagents and temperature found microwave heating is more effective than the traditional oven one. Where the former only needs minutes, the latter needs days with an equivalence of 10 min microwave to 1 day of traditional. The size of crystals has a narrow distribution and increases with time for the microwave but is a rather wide distribution for the traditional one. Furthermore the shape of the crystals is more regular for the microwave. The four Ni(7)(II) discs of formulas [Ni(7)L(6)(µ(3)-OMe)(6)](ClO(4))(2) (1-3) and {[Ni(7)(L(4))(6)(µ(3)-OMe)(6)][Ni(L(4))(2)]}(ClO(4))(2) (4) were synthesized as green hexagonal rods from Ni(ClO(4))(2)·6H(2)O in mixed MeOH/MeCN solution and salicylalde Schiff base ligands (L(1) = 2-methoxy-6-(iminomethyl)phenol, L(2) = 2-ethoxy-6-(iminomethyl)phenol, L(3) = 2-methoxy-6-((methylimino)methyl)phenol, L(4) = 2-ethoxy-6-((methylimino)methyl)phenol). X-ray structural analyses show six symmetrically positioned Ni(2) around a central Ni(1) bridged by the µ(3)-methoxide and surrounded by the ligand L which also isolates the discs from one another. The perchlorate sits in the interstices, and the planar [Ni(L(4))(2)] of 4 also inserts itself between the discs. The structures of 1-3 can be regarded as ordered discotic liquid crystals. Electrospray ionization mass spectrometry of solutions showed an exchange of methoxide for hydroxide and a different distribution of [Ni(7)] phase with mixed (MeO/OH) core bridges, confirming a probable "step by step" substitution of MeO(-) by OH(-). Magnetic studies indicate ferromagnetic interaction between Ni(1) and Ni(2) and possible antiferromagnetic between Ni(2) and Ni(2), resulting in a noncollinear system which only reaches half of the moment in 50 kOe at 2 K.

[Clinicopathologic and immunohistochemical study of 187 cases of intraductal papillary neoplasm of breast].

OBJECTIVE: To evaluate the diagnostic approach and criteria for intraductal papillary neoplasms of breast. METHODS: According to the criteria of 2003 WHO classification, 187 cases of intraductal papillary neoplasm of breast were identified and enrolled into the study. The clinical and histologic features were reviewed and immunohistochemical study for CD10, p63, CK14, CK5/6, CK7, MGB1 and p53 were carried out on 53 cases. RESULTS: Amongst the 187 cases studied, there were 128 cases of intraductal papilloma, 16 cases of atypical intraductal papilloma and 43 cases of intraductal papillary carcinoma. They showed a spectrum of morphologic features including epithelial and stromal hyperplasia and secondary changes. The expression of myoepithelial markers, including CD10 and p63, significantly decreased in ascending order from intraductal papillomas, atypical intraductal papillomas and intraductal papillary carcinomas (P < 0.001). The expression of basal cell markers, including CK5/6 and CK14, showed a mosaic pattern in benign lesions and significantly decreased or was absent in atypical and carcinomatous lesions (P < 0.001). In contrast, the luminal cell marker CK7 expressed in the three groups with no statistically significant difference (P = 0.06). On the other hand, the expression of MGB1 in intraductal papillary carcinomas was much lower than that in the other two groups (P = 0.002 and P = 0.007). The staining for p53 was negative in all of the three groups. CONCLUSIONS: Intraductal papillary neoplasms of breast represent a heterogeneous group of lesions with various morphologic appearances. Correlation with immunostaining results for myoepithelial markers, basal-type cytokeratins and luminal epithelial markers are helpful in arriving at a definitive diagnosis.

When old information is intermixed with new elements: An event-related potential study.

Previous research has focused on the event-related potential of the old/new effect, but the mechanism for cognitive conflict and subsequent integration processing both induced by intermixed (old and new) stimuli have not been widely researched. This paper describes the effect of familiar stimuli mixed with new information and presented as intermixed stimuli. Three conditions were set within a study-test paradigm incorporating old, new, and intermixed conditions. The intermixed condition resulted in the lowest accuracy, thereby implying the greatest old/new effect. Moreover, compared with the old condition, the intermixed condition elicited a greater N270 as well as a stronger N400, while the new condition only induced a stronger N270. These results elucidate that when new information is intermixed with old information, the old/new effect is more pronounced, indicating that intermixed stimuli are possibly more difficult to integrate than old and new stimuli.

In vitro Preparation and Evaluation of Sustained-Release Microcapsules of Salvianolic Acid.

OBJECTIVE: This study aims to investigate the preparation of sustained-release microcapsules of salvianolic acid. METHODS: The stability of salvianolic acid microcapsules was improved, and the time of action was prolonged in the present study. This was prepared using the spray-drying method, with chitosan as the carrier. In the preparation process, the prescription and process were optimized by L9 (34) using an orthogonal design, with yield and drug loading as indexes, in order to obtain optimum conditions. RESULTS: The optimal process and prescription for the preparation of salvianolic acid microcapsules were found to be as follows: mass concentration of chitosan, 1.5%; mass ratio of salvianolic acid to chitosan, 1:3; inlet air temperature, 190°C; and peristaltic pump speed, 300 mL·h-1. The surface of the microcapsules was round, the drug loading was 25.99% ± 2.14%, the yield was 51.88% ± 2.84%, the entrapment efficiency was 86.21% ± 2.89%, and the average particle size was 105.6 ± 2.56 nm. The microcapsules in vitro had certain sustained release characteristics. The internally fitted first-order release model equation was ln(1-Q) = -0.236 t + 4.591 7, r = 0.920. In addition, the results of differential scanning calorimetry show that the properties of salvianolic acid were not changed by the microcapsules. CONCLUSION: Sustained-release microcapsules of salvianolic acid can be successfully prepared by adopting marine polysaccharide as a carrier.

Chlorine-resistant bacteria (CRB) in the reverse osmosis system for wastewater reclamation: Isolation, identification and membrane fouling mechanisms.

Chlorine disinfection is often used as a pretreatment technology to control biofouling of reverse osmosis (RO) membranes. However, previous studies showed that biofouling of the RO system was aggravated after chlorine disinfection. Chlorine-resistant bacteria (CRB) were presumed to be closely related to the aggravation of fouling caused by chlorine disinfection. In order to analyze the membrane fouling mechanisms of CRB, 5 CRB strains were isolated from the surface of fouled RO membranes for wastewater reclamation, and 3 reference bacterial strains, Sphingopyxis soli BM1-1, Pseudomonas aeruginosa PAO1 and Escherichia coli CGMCC1.3373, were selected for comparative study. The chlorine resistance, membrane fouling potential, secretion and adhesion characteristics of these strains were evaluated. Among these isolated strains, 3 strains showed much higher chlorine resistance than PAO1 under the condition of 0.5, 2, 5 mg/L-Cl2, especially Bacillus CR19 and Bacillus CR2. Furthermore, a significant positive correlation was found between membrane fouling potential and chlorine resistance of all the strains in this study. The membrane fouling potential of the above 8 strains increased monotonically with the increase of chlorine resistance (under the condition of 0.5 mg/L-Cl2). Serious fouling caused by extracellular substances was observed in biofouling layers of the strains with high chlorine resistance, which lead to more severe flux decline. Extracellular polymeric substances (EPS) amount per cell was found to be the main factor related to the chlorine resistance as well as the fouling potential. Computational fluid dynamics (CFD) simulation was used to demonstrate the filtration resistance induced by the secretion of EPS. However, CRB with higher EPS amount may not show higher membrane adhesion potential, and thus may not be the dominant strain on the RO membranes before chlorine disinfection. These CRB with high fouling potential but low membrane adhesion potential, such as Bacillus CR19 and Bacillus CR2, may become the dominant bacteria on the membrane surface after chlorine disinfection and thus aggravate membrane fouling significantly.

Effects of microbial inactivation approaches on quantity and properties of extracellular polymeric substances in the process of wastewater treatment and reclamation: A review.

Microbial extracellular polymeric substances (EPS) have a profound role in various wastewater treatment and reclamation processes, in which a variety of technologies are used for disinfection and microbial growth inhibition. These treatment processes can induce significant changes in the quantity and properties of EPS, and altered EPS could further adversely affect the wastewater treatment and reclamation system, including membrane filtration, disinfection, and water distribution. To clarify the effects of microbial inactivation approaches on EPS, these effects were classified into four categories: (1) chemical reactions, (2) cell lysis, (3) changing EPS-producing metabolic processes, and (4) altering microbial community. Across these different effects, treatments with free chlorine, methylisothiazolone, TiO2, and UV irradiation typically enhance EPS production. Among the residual microorganisms in EPS matrices after various microbial inactivation treatments, one of the most prominent is Mycobacterium. With respect to EPS properties, proteins and humic acids in EPS are usually more susceptible to treatment processes than polysaccharides. The affected EPS properties include changes in molecular weight, hydrophobicity, and adhesion ability. All of these changes can undermine wastewater treatment and reclamation processes. Therefore, effects on EPS quantity and properties should be considered during the application of microbial inactivation and growth inhibition techniques.

Metagenomics analysis of the key functional genes related to biofouling aggravation of reverse osmosis membranes after chlorine disinfection.

Chlorine disinfection is a common technology to control biofouling in the pretreatment of the reverse osmosis (RO) system for wastewater reclamation. However, chlorine disinfection could even aggravate the RO membrane biofouling because of the changes of microbial community structure. In this study, the mechanism of biofilm formation and EPS secretion after chlorine disinfection was investigated by analyzing the genes coding quorum sensing, exopolysaccharide biosynthesis, and amino acid biosynthesis. After 1, 5, and 15 mg-Cl2/L chlorine disinfection, the relative abundances of the functional genes all increased significantly. Compared with the control group, chlorine-resistant bacteria (Acidovorax, Arenimonas, and Pseudomonas) also harbored higher relative abundances of these functional genes. The high relative abundances of these genes might provide the bacterial community after chlorine disinfection with high potential of biofilm formation and EPS secretion and then cause severe RO membrane biofouling. In the sample with 5 mg-Cl2/L chlorine disinfection, the correlation coefficients (r) between each two of the three kinds of functional genes were more than 0.9 and much stronger than that in the control group. These results indicated that the bacterial community selected by chlorine disinfection could build more stable biofilm to resist chlorine but also could cause more severe RO membrane biofouling.

Evaluating method and potential risks of chlorine-resistant bacteria (CRB): A review.

Chlorine-resistant bacteria (CRB) are commonly defined as bacteria with high resistance to chlorine disinfection or bacteria which can survive or even regrow in the residual chlorine. Chlorine disinfection cannot completely control the risks of CRB, such as risks of pathogenicity, antibiotic resistance and microbial growth. Currently, researchers pay more attention to CRB with pathogenicity or antibiotic resistance. The microbial growth risks of non-pathogenic CRB in water treatment and reclamation systems have been neglected to some extent. In this review, these three kinds of risks are all analyzed, and the last one is also highlighted. In order to study CRB, various methods are used to evaluate chlorine resistance. This review summarizes the evaluating methods for chlorine resistance reported in the literatures, and collects the important information about the typical isolated CRB strains including their genera, sources and levels of chlorine resistance. To our knowledge, few review papers have provided such systematic information about CRB. Among 44 typical CRB strains from 17 genera isolated by researchers, Mycobacterium, Bacillus, Legionella, Pseudomonas and Sphingomonas were the five genera with the highest frequency of occurrence in literatures. They are all pathogenic or opportunistic pathogenic bacteria. In addition, although there are many studies on CRB, information about chlorine resistance level is still limited to specie level or strain level. The difference in chlorine resistance level among different bacterial genera is less well understood. An inconvenient truth is that there is still no widely-accepted method to evaluate chlorine resistance and to identify CRB. Due to the lack of a unified method, it is difficult to compare the results about chlorine resistance level of bacterial strains in different literatures. A recommended evaluating method using logarithmic removal rate as an index and E. coli as a reference strain is proposed in this review based on the summary of the current evaluating methods. This method can provide common range of chlorine resistance of each genus and it is conducive to analyzing the distribution and abundance of CRB in the environment.