Harbin University of Commerce

Abstract The amino acid antiporter system Xc − is important for the synthesis of glutathione (GSH) that functions to prevent lipid peroxidation and protect cells from nonapoptotic, iron-dependent death (i.e., ferroptosis). While the activity of system Xc − often positively correlates with the expression level of its light chain encoded by SLC7A11 , inhibition of system Xc − activity by small molecules (e.g., erastin) causes a decrease in the intracellular GSH level, leading to ferroptotic cell death. How system Xc − is regulated during ferroptosis remains largely unknown. Here we report that activating transcription factor 3 (ATF3), a common stress sensor, can promote ferroptosis induced by erastin. ATF3 suppressed system Xc − , depleted intracellular GSH, and thereby promoted lipid peroxidation induced by erastin. ATF3 achieved this activity through binding to the SLC7A11 promoter and repressing SLC7A11 expression in a p53-independent manner. These findings thus add ATF3 to a short list of proteins that can regulate system Xc − and promote ferroptosis repressed by this antiporter.

BACKGROUND: The identification of human disease-related microRNAs (disease miRNAs) is important for further investigating their involvement in the pathogenesis of diseases. More experimentally validated miRNA-disease associations have been accumulated recently. On the basis of these associations, it is essential to predict disease miRNAs for various human diseases. It is useful in providing reliable disease miRNA candidates for subsequent experimental studies. METHODOLOGY/PRINCIPAL FINDINGS: It is known that miRNAs with similar functions are often associated with similar diseases and vice versa. Therefore, the functional similarity of two miRNAs has been successfully estimated by measuring the semantic similarity of their associated diseases. To effectively predict disease miRNAs, we calculated the functional similarity by incorporating the information content of disease terms and phenotype similarity between diseases. Furthermore, the members of miRNA family or cluster are assigned higher weight since they are more probably associated with similar diseases. A new prediction method, HDMP, based on weighted k most similar neighbors is presented for predicting disease miRNAs. Experiments validated that HDMP achieved significantly higher prediction performance than existing methods. In addition, the case studies examining prostatic neoplasms, breast neoplasms, and lung neoplasms, showed that HDMP can uncover potential disease miRNA candidates. CONCLUSIONS: The superior performance of HDMP can be attributed to the accurate measurement of miRNA functional similarity, the weight assignment based on miRNA family or cluster, and the effective prediction based on weighted k most similar neighbors. The online prediction and analysis tool is freely available at http://nclab.hit.edu.cn/hdmpred.

Plant basic helix-loop-helix (bHLH) transcription factors play essential roles in abiotic stress tolerance. However, most bHLHs have not been functionally characterized. Here, we characterized the functional role of a bHLH transcription factor from Arabidopsis, AtbHLH112, in response to abiotic stress. AtbHLH112 is a nuclear-localized protein, and its nuclear localization is induced by salt, drought and abscisic acid (ABA). In addition, AtbHLH112 serves as a transcriptional activator, with the activation domain located at its N-terminus. In addition to binding to the E-box motifs of stress-responsive genes, AtbHLH112 binds to a novel motif with the sequence 'GG[GT]CC[GT][GA][TA]C' (GCG-box). Gain- and loss-of-function analyses showed that the transcript level of AtbHLH112 is positively correlated with salt and drought tolerance. AtbHLH112 mediates stress tolerance by increasing the expression of P5CS genes and reducing the expression of P5CDH and ProDH genes to increase proline levels. AtbHLH112 also increases the expression of POD and SOD genes to improve reactive oxygen species (ROS) scavenging ability. We present a model suggesting that AtbHLH112 is a transcriptional activator that regulates the expression of genes via binding to their GCG- or E-boxes to mediate physiological responses, including proline biosynthesis and ROS scavenging pathways, to enhance stress tolerance.

MOTIVATION: Identifying microRNAs associated with diseases (disease miRNAs) is helpful for exploring the pathogenesis of diseases. Because miRNAs fulfill function via the regulation of their target genes and because the current number of experimentally validated targets is insufficient, some existing methods have inferred potential disease miRNAs based on the predicted targets. It is difficult for these methods to achieve excellent performance due to the high false-positive and false-negative rates for the target prediction results. Alternatively, several methods have constructed a network composed of miRNAs based on their associated diseases and have exploited the information within the network to predict the disease miRNAs. However, these methods have failed to take into account the prior information regarding the network nodes and the respective local topological structures of the different categories of nodes. Therefore, it is essential to develop a method that exploits the more useful information to predict reliable disease miRNA candidates. RESULTS: miRNAs with similar functions are normally associated with similar diseases and vice versa. Therefore, the functional similarity between a pair of miRNAs is calculated based on their associated diseases to construct a miRNA network. We present a new prediction method based on random walk on the network. For the diseases with some known related miRNAs, the network nodes are divided into labeled nodes and unlabeled nodes, and the transition matrices are established for the two categories of nodes. Furthermore, different categories of nodes have different transition weights. In this way, the prior information of nodes can be completely exploited. Simultaneously, the various ranges of topologies around the different categories of nodes are integrated. In addition, how far the walker can go away from the labeled nodes is controlled by restarting the walking. This is helpful for relieving the negative effect of noisy data. For the diseases without any known related miRNAs, we extend the walking on a miRNA-disease bilayer network. During the prediction process, the similarity between diseases, the similarity between miRNAs, the known miRNA-disease associations and the topology information of the bilayer network are exploited. Moreover, the importance of information from different layers of network is considered. Our method achieves superior performance for 18 human diseases with AUC values ranging from 0.786 to 0.945. Moreover, case studies on breast neoplasms, lung neoplasms, prostatic neoplasms and 32 diseases further confirm the ability of our method to discover potential disease miRNAs. AVAILABILITY AND IMPLEMENTATION: A web service for the prediction and analysis of disease miRNAs is available at http://bioinfolab.stx.hk/midp/.

Nickel oxide (NiO) is a promising candidate for future electrochromic supercapacitors due to its pronounced electrical properties and low cost. Unfortunately, the weak interaction between NiO films and conductive substrates results in poor cycling stability. In addition, the long color-switching time and low capacitance by the small lattice spacing in dense NiO impede its practical applications seriously. Herein, a hierarchical porous NiO film/ITO glass bifunctional electrode has been prepared via the solvothermal and subsequent calcination process of growing MOF-74 in situ on ITO, which shows outstanding cycle reversibility, excellent capacitance, high coloration efficiency and short color-switching time. Because of the strong binding force between the NiO film and substrate, and large surface areas with a hierarchical porous structure which are beneficial to the ion transport, the NiO film demonstrates perfect capacitive and electrochromic properties. As a bifunctional electrode, the NiO film shows a specific capacitance of 2.08 F cm-2 at 1 mA cm-2, large optical modulation of 41.08% and about 86% of optical modulation retention after 10 000 cycles. Furthermore, we assembled a bifunctional device whose energy condition can be roughly estimated according to the color state of the device. This finding can provide us with a new application of MOFs in the dual device of electrochromic supercapacitors.

As an emerging e-commerce model that combines the convenience of traditional e-commerce with the real-time and interactive nature of live streaming, live-streaming (LS) e-commerce is loved and recognized by consumers. At the same time, LS e-commerce also faces many difficulties such as homogenization of marketing content and consumers’ low willingness to repeat purchase. Therefore, how to better stimulate consumers’ continuous purchase willingness in LS has become a hot topic of current research. Based on the stimulus–organism–response (SOR) model, this paper constructs a model of consumers’ purchase intention mediated by consumers’ trust in LS e-commerce, measuring consumers’ perceived value in LS e-commerce in three dimensions: utilitarian value, hedonic value and social value, and using consumers’ trust in the streamer and trust in the product as mediators to investigate the influence of perceived value on consumers’ continuous purchase intention. Data of Chinese users were collected by questionnaire survey to prove the assumptions of this paper. The results show that consumers’ perceived utilitarian value, hedonic value and social value significantly and positively influence their trust in the streamer; consumers’ perceived utilitarian value and social value significantly and positively influence their trust in product; trust in streamer significantly and positively influences their trust in the product; and consumer trust partially mediates the relationship between perceived value and consumers’ willingness to continue to purchase.

Stroke is a leading cause of death and disability worldwide, and approximately 87% of cases are attributed to ischemia. The main factors that cause ischemic stroke include excitotoxicity, energy metabolism disorder, Ca + overload, oxidative damage, apoptosis, autophagy, and inflammation. However, no effective drug is currently available for the comprehensive treatment of ischemic stroke in clinical applications; thus, there is an urgent need to find and develop comprehensive and effective drugs to treat postischemic stroke. Traditional Chinese medicine (TCM) has unique advantages in treating ischemic stroke, with overall regulatory effects at multiple levels and on multiple targets. Many researchers have studied the effective components of TCMs and have achieved undeniable results. This paper reviews studies on the anticerebral ischemia effects of TCM monomers such as tetramethylpyrazine (TMP), dl‐3‐n‐butylphthalide (NBP), ginsenoside Rg1 (Rg1), tanshinone IIA (TSA), gastrodin (Gas), and baicalin (BA) as well as effective extracts such as Ginkgo biloba extract (EGB). Research on the anticerebral ischemia effects of TCMs has focused mostly on their antioxidative stress, antiapoptotic, anti‐inflammatory, proangiogenic, and proneurogenic effects. However, the research on the use of TCM to treat ischemic stroke remains incompletely characterized. Thus, we summarized and considered this topic from the perspective of pharmacokinetics, pharmacological effects, and mechanistic research, and we have provided a reference basis for future research and development on anticerebral ischemia TCM drugs.

BACKGROUND: Agarwood is an expensive resinous heartwood derived from Aquilaria plants that is widely used in traditional medicines, incense and perfume. Only wounded trees can produce agarwood, and the huge demand for the agarwood products has led all Aquilaria spp. being endangered and listed in the Appendix II of the CITES (http://www.cites.org). The major components of agarwood are sesquiterpenes and phenylethyl chromones. Owing to a lack of genomic information, the molecular basis of wound-induced sesquiterpenes biosynthesis and agarwood formation remains unknown. RESULTS: To identify the primary genes that maybe related to agarwood formation, we sequenced 2 cDNA libraries generated from healthy and wounded A. sinensis (Lour.) Gilg. A total of 89,137 unigenes with an average length of 678.65 bp were obtained, and they were annotated in detail at bioinformatics levels. Of those associated with agarwood formation, 30 putatively encoded enzymes in the sesquiterpene biosynthesis pathway, and a handful of transcription factors and protein kinases were related to wound signal transduction. Three full-length cDNAs of sesquiterpene synthases (ASS1-3) were cloned and expressed in Escherichia coli, and enzyme assays revealed that they are active enzymes, with the major products being δ-guaiene. A methyl jasmonate (MJ) induction experiment revealed that the expression of ASS was significantly induced by MJ, and the production of sesquiterpenes was elevated accordingly. The expression of some transcription factors and protein kinases, especially MYB4, WRKY4, MPKK2 and MAPK2, was also induced by MJ and coordinated with ASS expression, suggesting they maybe positive regulators of ASS. CONCLUSIONS: This study provides extensive transcriptome information for Aquilaria spp. and valuable clues for elucidating the mechanism of wound-induced agarwood sesquiterpenes biosynthesis and their regulation.

Drug-drug interactions play a vital role in drug research. However, they may also cause adverse reactions in patients, with serious consequences. Manual detection of drug-drug interactions is time-consuming and expensive, so it is urgent to use computer methods to solve the problem. There are two ways for computers to identify drug interactions: one is to identify known drug interactions, and the other is to predict unknown drug interactions. In this paper, we review the research progress of machine learning in predicting unknown drug interactions. Among these methods, the literature-based method is special because it combines the extraction method of DDI and the prediction method of DDI. We first introduce the common databases, then briefly describe each method, and summarize the advantages and disadvantages of some prediction models. Finally, we discuss the challenges and prospects of machine learning methods in predicting drug interactions. This review aims to provide useful guidance for interested researchers to further promote bioinformatics algorithms to predict DDI.

In this study, an emulsion stabilized by soy protein isolate (SPI)-pectin (PC) complexes was prepared to investigate the effects of high-intensity ultrasound (HIU) treatment (150–600 W) on the physicochemical properties, microstructure, and stability of emulsions. The results found that the emulsion treated at 450 W showed the best emulsion stability index (ESI) (25.18 ± 1.24 min), the lowest particle size (559.82 ± 3.17 nm), the largest ζ-potential absolute value (16.39 ± 0.18 mV), and the highest adsorbed protein content (27.31%). Confocal laser scanning microscopy (CLSM) and atomic force microscopy (AFM) revealed that the emulsion aggregation was significantly improved by ultrasound treatment, and the average roughness value (Rq) was the smallest (10.3 nm) at 450 W. Additionally, HIU treatment reduced the interfacial tension and apparent viscosity of the emulsion. Thermal stability was best when the emulsion was treated at 450 W, D43 was minimal (907.95 ± 31.72 nm), and emulsion separation also improved. Consequently, the creaming index (CI) was significantly decreased compared to the untreated sample, indicating that the storage stability of the emulsion was enhanced.

The herbicide glyphosate inhibits the shikimate pathway of the synthesis of amino acids such as phenylalanine, tyrosine, and tryptophan. However, much uncertainty remains concerning precisely how glyphosate kills plants or affects cellular redox homeostasis and related processes in glyphosate-sensitive and glyphosate-resistant crop plants. To address this issue, we performed an integrated study of photosynthesis, leaf proteomes, amino acid profiles, and redox profiles in the glyphosate-sensitive soybean (Glycine max) genotype PAN809 and glyphosate-resistant Roundup Ready Soybean (RRS). RRS leaves accumulated much more glyphosate than the sensitive line but showed relatively few changes in amino acid metabolism. Photosynthesis was unaffected by glyphosate in RRS leaves, but decreased abundance of photosynthesis/photorespiratory pathway proteins was observed together with oxidation of major redox pools. While treatment of a sensitive genotype with glyphosate rapidly inhibited photosynthesis and triggered the appearance of a nitrogen-rich amino acid profile, there was no evidence of oxidation of the redox pools. There was, however, an increase in starvation-associated and defense proteins. We conclude that glyphosate-dependent inhibition of soybean leaf metabolism leads to the induction of defense proteins without sustained oxidation. Conversely, the accumulation of high levels of glyphosate in RRS enhances cellular oxidation, possibly through mechanisms involving stimulation of the photorespiratory pathway.

The relationship between environmental protection and economic development has long been a controversial issue. This paper presents a literature survey on Environmental Kuznets Curve (EKC), which includes genesis, explanations and empirical evidence of EKC. The conclusion of this paper are, on the one hand, some factors are key reasons to EKC, including income elasticity of environmental quality demand, scale, technological and composition effects, international trade, FDI and history accidents, etc., on the other hand, from the empirical literature of EKC, the environmental quality indicators improved with public health. Moreover, Different data type will lead to different empirical result, so it is important to choose suitable indicators and data.

In this study, we present an effective strategy to enhance energy-storage density by the Mn2+ substitution of Ti4+ into 0.7(Na0.5Bi0.5)TiO3-0.3SrTiO3 (0.7NBT-0.3ST) relaxor ferroelectric thin films. The influence of Mn doping on the microstructures, ferroelectric properties, and energy-storage performances of the as-prepared films was investigated in detail. The results show that the values of electric break-down field strength and the difference values between maximum polarization and remnant polarization of the thin films are markedly improved by appropriate Mn doping. Owing to the high break-down field strength of 1894 kV/cm and the huge difference value between the maximum polarization and the remnant polarization of 56 μC/cm2, a giant recoverable energy-storage density of 27 J/cm3 was obtained for the 1 mol. % Mn-doped 0.7NBT-0.3ST thin film. These results indicate that the appropriately Mn-doped 0.7NBT-0.3ST thin films are promising for the application of advanced capacitors with high-energy storage density.

In this study, a soy protein isolate (SPI)-pectin (PC) complex was prepared, and the effects of different high intensity ultrasound (HIU) powers on the structure and solubility of the complex were studied. Fourier transform infrared (FTIR) spectroscopy analysis exhibited that with increasing HIU power, the α-helix content of the SPI in the complex was significantly reduced, and the random coil content increased; however, an opposite trend appeared after higher power treatments. Fluorescence spectra showed that HIU treatment increased the fluorescence intensity of the complex, and the surface hydrophobicity was increased. The trend of the protein structure studied by Raman spectroscopy was similar to that of FTIR and fluorescence spectroscopy. When the HIU treatment was performed for 15 min and at 450 W power, the particle size of the complex was 451.85 ± 2.17 nm, and the solubility was 89.04 ± 0.19 %, indicating that the HIU treatment caused the spatial conformation of the protein to loosen and improved the functional properties of the complex. Confocal laser scanning microscopy (CLSM) revealed that the complex after HIU treatment exhibited improved dispersibility in water and smaller particle size. Gel electrophoresis results indicated that HIU treatment did not affect the protein subunits of the complex. Therefore, the selection of a suitable HIU treatment power can effectively improve the structural properties and solubility of SPI in the complex, and promote the application of the SPI-PC complex in food processing and industries.

BACKGROUND: Liver cancer is the sixth most commonly diagnosed cancer and the fourth most common cause of cancer death. The purpose of this work is to find new diagnostic biomarkers or prognostic biomarkers and explore the biological functions related to the prognosis of liver cancer. METHODS: GSE25097 datasets were firstly obtained and compared with TCGA LICA datasets and an analysis of the overlapping differentially expressed genes (DEGs) was conducted. Cytoscape was used to screen out the Hub Genes among the DEGs. ROC curve analysis was used to screen the Hub Genes to determine the genes that could be used as diagnostic biomarkers. Kaplan-Meier analysis and Cox proportional hazards model screened genes associated with prognosis biomarkers, and further Gene Set Enrichment Analysis was performed on the prognosis genes to explore the mechanism affecting the survival and prognosis of liver cancer patients. RESULTS: 790 DEGs and 2162 DEGs were obtained respectively from the GSE25097 and TCGA LIHC data sets, and 102 Common DEGs were identified by overlapping the two DEGs. Further screening identified 22 Hub Genes from 102 Common DEGs. ROC and survival curves were used to analyze these 22 Hub Genes and it was found that there were 16 genes with a value of AUC > 90%. Among these, the expression levels of ESR1,SPP1 and FOSB genes were closely related to the survival time of liver cancer patients. Three common pathways of ESR1, FOBS and SPP1 genes were identified along with seven common pathways of ESR1 and SPP1 genes and four common pathways of ESR1 and FOSB genes. CONCLUSIONS: SPP1, AURKA, NUSAP1, TOP2A, UBE2C, AFP, GMNN, PTTG1, RRM2, SPARCL1, CXCL12, FOS, DCN, SOCS3, FOSB and PCK1 can be used as diagnostic biomarkers for liver cancer, among which FOBS and SPP1 genes can also be used as prognostic biomarkers. Activation of the cell cycle-related pathway, pancreas beta cells pathway, and the estrogen signaling pathway, while on the other hand inhibition of the hallmark heme metabolism pathway, hallmark coagulation pathway, and the fat metabolism pathway may promote prognosis in liver cancer patients.

In this study, rice bran protein–chlorogenic acid (RBP–CA) emulsion was subjected to an ultrasonic-assisted treatment technique. The encapsulation efficiency and loading capacity of chlorogenic acid (CA), and the morphology, particle size, zeta (ζ)-potential, atomic force microscopy image, viscosity, turbidity, and interfacial protein content of the emulsion under different ultrasonic power were investigated. The results revealed that the emulsion exhibited an encapsulation efficiency and loading capacity of 86.26 ± 0.11% and 17.25 ± 0.06 g/100 g, respectively, at an ultrasonic power of 400 W. In addition, the size of the emulsion droplets decreased and became more evenly distributed. Furthermore, the viscosity of the emulsion decreased significantly, and it exhibited a turbidity and interfacial protein content of 24,758 and9.34 mg/m2, respectively. Next, the storage, oxidation, thermal, and salt ion stabilities of the emulsion were evaluated. The results revealed that the ultrasonic-assisted treatment considerably improved the stability of the emulsion.

This research aimed to determine the prevalence of workplace violence (WPV) against healthcare workers, explore the frequency distribution of violence in different occupational groups, and determine which healthcare occupation suffers from WPV most frequently. Furthermore, the current study aimed to compare risk factors affecting different types of WPV in Chinese hospitals. A cross-sectional design was utilized. A total of 1899 healthcare workers from Heilongjiang, a province in Northeastern China, completed the questionnaire. Of the respondents, 83.3% reported exposure to workplace violence, and 68.9% reported non-physical violence. Gender, education, shift work, anxiety level, and occupation were significantly correlated with physical violence (p < 0.05 for all correlations). Additionally, age, professional title, and occupation were correlated with non-physical violence, which critically affected doctors. Thus, gender, age, profession, anxiety, and shift work were predictive of workplace violence toward healthcare workers. Doctors appeared to experience non-physical workplace violence with particularly higher frequency when compared to nurses and other workers in hospitals. For healthcare workers, interventions aimed at WPV reduction should be enacted according to the types of violence, profession, and other factors underlying the various types of WPV in hospitals.

AbstractMethanotrophs can oxidize methane to carbon dioxide through sequential reactions catalyzed by a series of enzymes including methane monooxygenase, methanol dehydrogenase, formaldehyde dehydrogenase, and formate dehydrogenase. When suspensions of methanotrophic bacteria of Methylosinus trichosporium IMV 3011 were incubated at 32°C with methane and oxygen, there was an extracellular accumulation of methanol from methane oxidation in response to carbon dioxide addition. Maximal accumulation of methanol was achieved with 40% carbon dioxide in the mixed reaction gases. A continuous experiment was performed in a continuous ultrafiltration reactor. The optimum gas mixture containing 20% (v v−1) methane, 20% oxygen, 20% nitrogen and 40% carbon dioxide was used to provide substrates and to maintain the transmembrane pressure. The product (methanol) was removed in the eluate buffer. The initial methanol concentration in the eluate buffer was 8.22 μmol L−1. The bioreactor was operated continuously for 198 h without obvious loss of productivity.KeywordsCarbon dioxidemethane oxidationmethanol biosynthesisMethanotrophMethylosinus trichosporium IMV 3011continuous reaction

Diabetic cardiomyopathy (DCM) has been increasingly considered as a main cause of heart failure and death in diabetic patients. At present, no effective treatment exists to prevent its development. In the present study, we describe the potential protective effects and mechanisms of myricitrin (Myr) on the cardiac function of streptozotosin-induced diabetic mice and on advanced glycation end products (AGEs)-induced H9c2 cardiomyocytes. In vitro experiments revealed that pretreatment with Myr significantly decreased AGEs-induced inflammatory cytokine expression, limited an increase in ROS levels, and reduced cell apoptosis, fibrosis, and hypertrophy in H9c2 cells. These effects are correlated with Nrf2 activation and NF-κB inhibition. In vivo investigation demonstrated that oral administration of Myr at 300 mg/kg/day for 8 weeks remarkably decreased the expression of enzymes associated with cardiomyopathy, as well as the expression of inflammatory cytokines and apoptotic proteins. Finally, Myr improved diastolic dysfunction and attenuated histological abnormalities. Mechanistically, Myr attenuated diabetes-induced Nrf2 inhibition via the regulation of Akt and ERK phosphorylation in the diabetic heart. Collectively, these results strongly indicate that Myr exerts cardioprotective effects against DCM through the blockage of inflammation, oxidative stress, and apoptosis. This suggests that Myr might be a potential therapeutic agent for the treatment of DCM.

Aerobic granular sludge (AGS) is promising in wastewater treatment. However, the formation and existence of AGS under low organic loading rate (OLR) is still not fully understood due to a knowledge gap in the variations and correlations of N-acyl-homoserine lactones (AHLs), the microbial community, extracellular polymeric substances (EPS) and other physiochemical granule properties. This study comprehensively investigated the AHL-mediated quorum sensing (QS) and microbial community characters in the AGS fed with ammonium-rich wastewater under a low OLR of 0.15 kg COD (m3 d)−1. The results showed that the AGS appeared within 90 days, and the size of mature granules was over 700 μm with strong settleability and ammonium removal performance. More tightly-bound extracellular polysaccharide and tightly-bound extracelluar protein were produced in the larger AGS. C10-HSL and C12-HSL gradually became dominant in sludge, and short-chain AHLs dominated in water. EPS producers and autotrophic nitrifiers were successfully retained in the AGS under low OLR. AHL-mediated QS utilized C10-HSL, C12-HSL and 3OC6-HSL as the critical AHLs to regulate the TB-EPS in aerobic granulation, and autotrophic nitrifiers may perform interspecific communication with C10-HSL. The correlations of bacterial genera with AGS properties and AHLs were complex due to the dynamic fluctuations of microbial composition and other variable factors in the mixed-culture system. These findings confirmed the participation of AHL-mediated QS in the regulation of microbial community characters and AGS properties under low OLR, which may provide guidance for the operation of AGS systems under low OLR from a microbiological viewpoint.