NirS-type denitrifying bacteria in aerobic water layers of two drinking water reservoirs: Insights into the abundance, community diversity and co-existence model.

The nirS-type denitrifying bacterial community is the main drivers of the nitrogen loss process in drinking water reservoir ecosystems. The temporal patterns in nirS gene abundance and nirS-type denitrifying bacterial community harbored in aerobic water layers of drinking water reservoirs have not been studied well. In this study, quantitative polymerase chain reaction (qPCR) and Illumina Miseq sequencing were employed to explore the nirS gene abundance and denitrifying bacterial community structure in two drinking water reservoirs. The overall results showed that the water quality parameters in two reservoirs had obvious differences. The qPCR results suggested that nirS gene abundance ranged from (2.61 ± 0.12) × 105 to (3.68 ± 0.16) × 105 copies/mL and (3.01 ± 0.12) × 105 to (5.36 ± 0.31) × 105 copies/mL in Jinpen and Lijiahe reservoirs, respectively. The sequencing results revealed that Paracoccus sp., Azoarcus sp., Dechloromonas sp. and Thauera sp. were the dominant genera observed. At species level, Cupriavidus necator, Dechloromonas sp. R-28400, Paracoccus denitrificans and Pseudomonas stutzeri accounted for more proportions in two reservoirs. More importantly, the co-occurrence network analysis demonstrated that Paracoccus sp. R-24615 and Staphylococcus sp. N23 were the keystone species observed in Jinpen and Lijiahe reservoirs, respectively. Redundancy analysis indicated that water quality (particularly turbidity, water temperature, pH and Chlorophyll a) and sampling time had significant influence on the nirS-type denitrifying bacterial community in both reservoirs. These results will shed new lights on exploring the dynamics of nirS-type denitrifying bacteria in aerobic water layers of drinking water reservoirs.

Geographical Patterns of nirS Gene Abundance and nirS-Type Denitrifying Bacterial Community Associated with Activated Sludge from Different Wastewater Treatment Plants.

Denitrifying bacteria is a driver of nitrogen removal process in wastewater treatment ecosystem. However, the geographical characteristics of denitrifying bacterial communities associated with activated sludge from diverse wastewater treatment plants (WWTPs) are still unclear. Here, quantitative PCR and next-generation sequencing of the nirS gene were applied to characterize the abundance and denitrifying bacterial communities from 18 geographically distributed WWTPs. The results showed that the nirS abundance ranged from 4.6 × 102 to 2.4 × 103 copies per ng DNA, while nirS-type denitrifying bacterial populations were diverse and distinct from activated sludge communities. Among WWTPs, total nitrogen removal efficiencies varied from 25.8 to 84%, which was positively correlated with diversity indices, whereas abundance-based coverage estimator index decreased with an increase in latitude. The dominant phyla across all samples were proteobacteria, accounting for 46.23% (ranging from 17.98 to 87.07%) of the sequences. Eight of the 22 genera detected were dominant: Thauera sp., Alicycliphilus sp., and Pseudomonas sp., etc. Based on network analysis, the coexistence and interaction between dominant genera may be vital for regulating the nitrogen and carbon removal behaviors. Multivariate statistical analysis revealed that both geographic location and wastewater factors concurrently govern the distribution patterns of nirS-type denitrifying bacterial community harbored in WWTPs. Taking together, these results from the present study provide novel insights into the nirS gene abundance and nirS-type denitrifying bacterial community composition in geographically distributed WWTPs. Moreover, the knowledge gained will improve the operation and management of WWTPs for nitrogen removal.

A Novel Sucrose-Regulatory MADS-Box Transcription Factor GmNMHC5 Promotes Root Development and Nodulation in Soybean (Glycine max [L.] Merr.).

The MADS-box protein family includes many transcription factors that have a conserved DNA-binding MADS-box domain. The proteins in this family were originally recognized to play prominent roles in floral development. Recent findings, especially with regard to the regulatory roles of the AGL17 subfamily in root development, have greatly broadened their known functions. In this study, a gene from soybean (Glycine max [L.] Merr.), GmNMHC5, was cloned from the Zigongdongdou cultivar and identified as a member of the AGL17 subfamily. Real-time fluorescence quantitative PCR analysis showed that GmNMHC5 was expressed at much higher levels in roots and nodules than in other organs. The activation of expression was first examined in leaves and roots, followed by shoot apexes. GmNMHC5 expression levels rose sharply when the plants were treated under short-day conditions (SD) and started to pod, whereas low levels were maintained in non-podding plants under long-day conditions (LD). Furthermore, overexpression of GmNMHC5 in transgenic soybean significantly promoted lateral root development and nodule building. Moreover, GmNMHC5 is upregulated by exogenous sucrose. These results indicate that GmNMHC5 can sense the sucrose signal and plays significant roles in lateral root development and nodule building.

Feeding Strategies and Trophic Niche Divergence of Three Groups of Dosidicus gigas off Peru: Based on Stable Carbon and Nitrogen Isotopes and Morphology of Feeding Apparatuses.

Dosidicus gigas (D. gigas) is a pelagic cephalopod of ecological and economic importance widely distributed in the eastern Pacific Ocean. Generally, small-, medium-, and large-sized groups of the squids have been respectively identified on the basis of the mantle length (ML) of adults. Intraspecific feeding variability maximizes the utilization of available food resources by D. gigas. However, the coexistence mechanism of three groups has not been fully understood yet. In our study, based on the analyses of beak morphology and stable carbon and nitrogen isotopes, the feeding strategies and coexistence patterns of large-, medium-, and small-sized groups of D. gigas were investigated. D. gigas had a wide range of 13C/12C (δ13C) and 15N/14N (δ15N) values in muscle tissue, variable feeding behaviors, and wide food sources. The δ13C or δ15N values showed no significant difference between the small- and medium-sized groups, which shared the same habitats and fed on preys with the similar trophic level. Compared to small- and medium-sized groups, the large-sized group had a smaller range of habitats and consumed more nearshore foods. Both isotopes and feeding apparatus morphology indicated a high degree of niche overlap between the small- and medium-sized groups, whereas the large-sized group differed significantly from other groups. In addition, the niche width of the female was larger than that of the male in all three groups. We inferred that the sex differences in body length and reproductive behavior led to the difference in niche width. The isotopic niche overlap between female and male samples was the most significant in large-sized group and the least significant in the small-sized group, indicating that different feeding strategies were adopted by the three groups. These findings proved that the three groups of D. gigas off Peruvian waters adopted a feeding strategy with inter- and intra-group regulation. This feeding strategy maximizes the use of food and habitat resources and ensures that different size groups can coexist in the same waters.

Realizing Spontaneously Regular Stacking of Pristine Graphene Oxide by a Chemical-Structure-Engineering Strategy for Mechanically Strong Macroscopic Films.

Mechanical-electrical properties of macroscopic graphene films derived from graphene oxide (GO) sheets are substantially restricted by their surface wrinkles and structural misalignment. Herein, we propose a chemical-structure-engineering strategy to realize the spontaneously regular stacking of modified GO (GO-m) with trace carboxyl. The highly aligned GO-m film delivers a fracture strength and modulus of nearly 3- and 5-fold higher than a wrinkled film with conventional Hummer's method derived GO (GO-c). The favorable assembly pattern of GO-m sheets is attributed to their decreased interfacial friction on the atomic scale, which weakens their local gelation capability for freer configuration adjustment during the assembly process. The chemical structure of GO-m can be further engineered by an epoxide-to-hydroxyl reaction, achieving a record high tensile strength of up to 631 MPa for the pristine GO film. By exploring the relationship between the surface terminations of GO and its stacking mode, this work proves the feasibility to realize high-performance macroscopic materials with optimized microstructure through the chemical modulation of nanosheet assembly.

Nitrate removal characteristics and 13C metabolic pathways of aerobic denitrifying bacterium Paracoccus denitrificans Z195.

Strain Z195 was isolated and identified as Paracoccus denitrificans. Z195 exhibited efficient aerobic denitrification and carbon removal abilities, and removed 93.74% of total nitrogen (TN) and 97.81% of total organic carbon.71.88% of nitrogen was lost as gaseous products.13C-metabolic flux analysis revealed that 95% and 132% of the carbon fluxes entered the Entner-Doudoroff (ED) pathway and tricarboxylic acid (TCA) cycle, respectively. Electrons produced by carbon metabolism markedly promoted the processes of nitrogen metabolism process and aerobic respiration. A response surface methodology model demonstrated that the optimal conditions for the maximum TN removal were a C/N ratio of 7.47, shaking speed of 108 rpm, temperature of 31 °C and initial pH of 8.02. Additionally, the average TN and chemical oxygen demand removal efficiencies of raw wastewater were 89% and 91%, respectively. The results give new insight for understanding metabolic flux analysis of aerobic denitrifying bacteria.

Water Bacterial and Fungal Community Compositions Associated with Urban Lakes, Xi'an, China.

Urban lakes play a vital role in the sustainable development of urbanized areas. In this freshwater ecosystem, massive microbial communities can drive the recycling of nutrients and regulate the water quality. However, water bacterial and fungal communities in the urban lakes are not well understood. In the present work, scanning electron microscopy (SEM) was combined with community level physiological profiles (CLPPs) and Illumina Miseq sequence techniques to determine the diversity and composition of the water bacterial and fungal community in three urban lakes, namely Xingqing lake (LX), Geming lake (LG) and Lianhu lake (LL), located in Xi'an City (Shaanxi Province, China). The results showed that these three lakes were eutrophic water bodies. The highest total nitrogen (TN) was observed in LL, with a value of 12.1 mg/L, which is 2 times higher than that of LG. The permanganate index (CODMn) concentrations were 21.6 mg/L, 35.4 mg/L and 28.8 mg/L in LG, LL and LX, respectively (p < 0.01). Based on the CLPPs test, the results demonstrated that water bacterial communities in the LL and LX urban lakes had higher carbon source utilization ability. A total of 62,742 and 55,346 high quality reads were grouped into 894 and 305 operational taxonomic units (OTUs) for bacterial and fungal communities, respectively. Water bacterial and fungal community was distributed across 14 and 6 phyla. The most common phyla were Proteobacteriaand Cyanobacteria. Cryptomycota was particularly dominant in LL, while Chytridiomycota and Entomophthormycota were the most abundant fungal phyla, accounting for 95% of the population in the LL and 56% in the LG. Heat map and redundancy analysis (RDA) highlighted the dramatic differences of water bacterial communities among three urban lakes. Meanwhile, the profiles of fungal communities were significantly correlated with the water quality parameters (e.g., CODMn and total nitrogen, TN). Several microbes (Legionella sp. and Streptococcus sp.) related to human diseases, such as infectious diseases, were also found. The results from this study provides useful information related to the water quality and microbial community compositions harbored in the aquatic ecosystems of urban lakes.

Disentangling the Drivers of Diversity and Distribution of Fungal Community Composition in Wastewater Treatment Plants Across Spatial Scales.

Activated sludge microbial community composition is a key bio-indicator of the sustainability of wastewater treatment systems. Therefore, a thorough understanding of the activated sludge microbial community dynamics is critical for environmental engineers to effectively manage the wastewater treatment plants (WWTPs). However, fungal communities associated with activated sludge have been poorly elucidated. Here, the activated sludge fungal community in 18 geographically distributed WWTPs was determined by using Illumina sequencing. The results showed that differences in activated sludge fungal community composition were observed among all WWTPs and also between oxidation ditch and anaerobic-anoxic-aerobic (A/A/O) systems. Ascomycota was the largest phyla, followed by Basidiomycota in all samples. Sporidiobolales and Pezizales were the most abundant order in oxidation ditch and A/A/O systems, respectively. The network analysis indicated cooperative and co-occurrence interactions between fungal taxa in order to accomplish the wastewater treatment process. Hygrocybe sp., Sporobolomyces sp., Rhodotorula sp., Stemphylium sp., Parascedosporium sp., and Cylindrocarpon sp., were found to have statistically significant interactions. Redundancy analysis revealed that temperature, total phosphorus, pH, and ammonia nitrogen were significantly affected the fungal community. This study sheds light on providing the ecological characteristics of activated sludge fungal communities and useful guidance for improving wastewater treatment performance efficiency.

Biological nitrogen removal and metabolic characteristics of a novel aerobic denitrifying fungus Hanseniaspora uvarum strain KPL108.

A novel aerobic denitrifying fungal strain KPL108 was isolated from the sediment of Jinpen drinking water reservoir and identified as Hanseniaspora uvarum. Strain KPL108 removed 99% of nitrate without nitrite accumulation under aerobic conditions, while the total organic carbon removal efficiency was 93%. KPL108 expressed fungal specific denitrifying gene p450nor. Nitrogen balance exhibited that approximately 92% of the initial nitrate was removed as gaseous products. Based on 13C-isotope labeling tracer, pentose phosphate pathway and tricarboxylic acid cycle were highly active in intracellular central carbon metabolism of strain KPL108. Response surface methodology revealed that the maximum total nitrogen removal efficiency occurred with the optimized parameters: C/N ratio of 6.4, pH of 8.2, 28.5 °C and 109.7 rpm. Collectively, the results from the present study indicate that strain KPL108 has aerobic denitrification ability, which has a great potential application for nitrogenous wastewater treatment.

Geographical patterns of denitrifying bacterial communities associated with different urban lakes.

The geographical variation of denitrifying bacterial communities and water quality parameters in urban lakes distributed across nine provinces in China were determined. The Illumina sequencing data of the denitrifying encoding gene nirS was examined in the samples collected from nine localities (pairwise geographical distance: 200-2600 km). The results showed that fundamental differences in water quality were observed among different urban lakes. The highest nitrate (2.02 mg L-1) and total nitrogen (3.82 mg L-1) concentrations were observed in Pingzhuang (P < 0.01). The algal cell concentration ranged from 1.29 × 108 to 3.0 × 109 cell per L. The sequencing data generated a total of 421058 high quality nirS gene reads that resulted in 6369 OTUs (97% cutoff), with Proteobacteria and Firmicutes being the dominant taxa. A co-occurrence network analysis indicated that the top five genera identified as keystone taxa were Dechlorospirillum sp., Alicycliphilus sp., Dechloromonas sp., Pseudogulbenkiania sp., and Paracoccus sp. A redundancy analysis (RDA) further revealed that distinct denitrifying bacterial communities inhabited the different urban lakes, and influenced by urban lake water ammonia nitrogen, manganese and algal cell concentrations. A variance partitioning analysis (VPA) also showed that geographic location was more important than water quality factors in structuring the denitrifying bacterial communities. Together, these results provide new insight into understanding of denitrifying bacterial communities associated with geographically distributed urban lakes on a larger scale, and these results also expand our exploration of aquatic microbial ecology in freshwater bodies.

Dynamics of Bacterial and Fungal Communities during the Outbreak and Decline of an Algal Bloom in a Drinking Water Reservoir.

The microbial communities associated with algal blooms play a pivotal role in organic carbon, nitrogen and phosphorus cycling in freshwater ecosystems. However, there have been few studies focused on unveiling the dynamics of bacterial and fungal communities during the outbreak and decline of algal blooms in drinking water reservoirs. To address this issue, the compositions of bacterial and fungal communities were assessed in the Zhoucun drinking water reservoir using 16S rRNA and internal transcribed spacer (ITS) gene Illumina MiSeq sequencing techniques. The results showed the algal bloom was dominated by Synechococcus, Microcystis, and Prochlorothrix. The bloom was characterized by a steady decrease of total phosphorus (TP) from the outbreak to the decline period (p < 0.05) while Fe concentration increased sharply during the decline period (p < 0.05). The highest algal biomass and cell concentrations observed during the bloom were 51.7 mg/L and 1.9×108 cell/L, respectively. The cell concentration was positively correlated with CODMn (r = 0.89, p = 0.02). Illumina Miseq sequencing showed that algal bloom altered the water bacterial and fungal community structure. During the bloom, the dominant bacterial genus were Acinetobacter sp., Limnobacter sp., Synechococcus sp., and Roseomonas sp. The relative size of the fungal community also changed with algal bloom and its composition mainly contained Ascomycota, Basidiomycota and Chytridiomycota. Heat map profiling indicated that algal bloom had a more consistent effect upon fungal communities at genus level. Redundancy analysis (RDA) also demonstrated that the structure of water bacterial communities was significantly correlated to conductivity and ammonia nitrogen. Meanwhile, water temperature, Fe and ammonia nitrogen drive the dynamics of water fungal communities. The results from this work suggested that water bacterial and fungal communities changed significantly during the outbreak and decline of algal bloom in Zhoucun drinking water reservoir. Our study highlights the potential role of microbial diversity as a driving force for the algal bloom and biogeochemical cycling of reservoir ecology.

[Vertical Distribution of Fungal Community Composition and Water Quality During the Deep Reservoir Thermal Stratification].

Freshwater fungal community composition plays pivotal roles in sustaining the function and health of drinking water reservoir ecosystems. To investigate the vertical evaluative characteristics of an aquatic fungal community under conditions of stable thermal stratification, water samples were collected at 0.5, 10, 25, 40, 60 and 70 m depths of Jinpen Reservoir during thermal stratification. Fungal community composition was determined using a high-throughput DNA sequencing technique and combined with water quality parameters. The results showed that the epilimnion, metalimnion, and hypolimnion water layers were formed in Jinpen Reservoir. The temperature decreased steadily from the surface at 22.33℃ to the bottom with 7.21℃ (P<0.05). The dissolved oxygen (DO) concentration decreased significantly from the surface to the bottom (P<0.05). The conductivity and the concentration total phosphorus and iron increased significantly from the surface to the bottom (P<0.05). The high-throughput DNA sequencing revealed a total of 1247 operational taxonomic units (OTUs), which were affiliated with four phylum, 14 classes, and 39 genera, including Zygomycota, Basidiomycota, Ascomycota, and Chytridiomycota. The highest Shannon diversity and Chao 1 were 3.45 and 360 at 60 m, respectively. The Shannon diversity and Chao 1 were significantly higher in the hypolimnion than in the metalimnion (P<0.05). Rhodotorula (27.23%), Alternaria (24.28%),Cladosporium (22.98%), Alternaria (32.00%), Didymella (17.47%), and Cladosporium (28.17%) were the dominant species at 0.5 m, 10 m, 25 m, 40 m, 60 m, and 70 m, respectively. There are a number of unclassified fungi at different water depths. The heat map profile indicated significant differences in the vertical distribution of the fungal community composition in Jinpen Reservoir. A principle component analysis (PCA) indicated that water temperature, DO, TP, and conductivity had dramatic influences on the vertical distribution of the fungal community composition. The results provide new insights on the relationship between water quality and fungal community composition during reservoir thermal stratified periods.

Paracoccus versutus KS293 adaptation to aerobic and anaerobic denitrification: Insights from nitrogen removal, functional gene abundance, and proteomic profiling analysis.

A novel strain KS293 exhibiting excellent aerobic and anaerobic denitrification performance was isolated and identified as Paracoccus versutus KS293. Nitrate nitrogen and total organic carbon could be effectively removed by P. versutus KS293 without nitrite accumulation, whilst 82% and 85% of total nitrogen was converted into gaseous products under aerobic and anaerobic conditions (P > .05), respectively. Based on the ratio of anaerobic to aerobic, relative abundance values were increased 1.41, 1.45, and 2.31 folds for nirS, nosZ, and narG, respectively. A comparison of the two-dimensional gel electrophoresis and principal component analysis showed significant differences in proteomic profiles between aerobic and anaerobic conditions. In total, 78 proteins that displayed fluctuations in relative expression were observed. 10 proteins including nitrate reductase, maintenance of cell membrane (TolA), and RNA polymerase-binding transcription factor (DksA) were differentially expressed. These findings demonstrated that P. versutus KS293 was effective for nitrogen removal under aerobic or anaerobic conditions.

[Denitrification Characteristics and Functional Genes of Denitrifying Bacteria Under Aerobic or Anaerobic Conditions].

An efficient aerobic denitrifying bacterium was isolated from the sediments of the Jin Pen Reservoir in Xi'an. The strain was identified by morphological features and 16S rDNA sequences as Pseudomonas stutzeri and named it KK99. The denitrification characteristics of the strain and the expression level of the functional genes narG, nirS, and nosZ in aerobic/anaerobic conditions were investigated. The results showed that the strain can carry out both anaerobic and aerobic denitrification and has a high efficiency of denitrification, which occurs under both aerobic and anaerobic conditions; after 24 h, the removal rates of total nitrogen (TN) were 85.08% and 89.05%, respectively. Under both the conditions, the expression levels of the functional genes nosZ and nirS are high. nosZ plays a vital role in denitrification in the aerobic pathway, nirS plays a vital role in denitrification in the anaerobic pathway, and narG expression is low under both the conditions. At the same time, simultaneous nitrification and denitrification (SND) capacity of the strain was observed when nitrate and ammonium salts were the nitrogen sources, with the total nitrogen removal rate being 76% within 24 h in aerobic conditions. The P. stutzeri KK99 strain can be used for controlling eutrophication of micro-polluted water, and the application of total nitrogen reduction engineering.

Early detection of rheumatoid arthritis in rats and humans with 99mTc-3PRGD2 scintigraphy: imaging synovial neoangiogenesis.

OBJECTIVES: To validate 99mTc-labeled arginylglycylaspartic acid (99mTc-3PRGD2) scintigraphy as a means to image synovial neoangiogenesis in joints afflicted by rheumatoid arthritis and to investigate its potential in the early detection and management of rheumatoid arthritis. METHODS: Rheumatoid arthritis and osteoarthritis were generated in Sprague Dawley rats by type II collagen immunization and papain injection, respectively. Rats were imaged with 99mTc-3PRGD2 and 99mTc- methyl diphosphonate (99mTc MDP). X-ray images were also obtained and assessed by a radiologist. Immunohistochemistry of αvß3 and CD31confirmed the onset of synovial neoangiogenesis. The effect of bevacizumab on rheumatoid arthritis was followed with 99mTc-3PRGD2 scintigraphy. A patient with rheumatoid arthritis and a healthy volunteer were scanned with 99mTc-3PRGD2. RESULTS: Two weeks after immunization, a significant increase in 99mTc-3PRGD2 was observed in the joints of the rheumatoid arthritis model though uptake in osteoarthritis model and untreated controls was low. 99mTc-MDP whole body scans failed to distinguish early rheumatoid arthritis joints from healthy controls. The expression of αvß3 and CD31was significantly higher in the joints of rheumatoid arthritis rats compared to normal controls. In serial 99mTc-3PRGD2 scintigraphy studies, 99mTc-3PRGD2 uptake increased in parallel with disease progression. Bevacizumab anti-angiogenetic therapy both improved the symptoms of the rheumatoid arthritis rats and significantly decreased 99mTc-3PRGD2 uptake. Significantly higher 99mTc-3PRGD2 accumulation was also observed in rheumatoid arthritis joints in the patient. CONCLUSIONS: Our findings indicate that 99mTc-3PRGD2 scintigraphy could detect early rheumatoid arthritis by imaging the associated synovial neoangiogenesis, and may be useful in disease management.