Out of sight, but not out of season: Nitrifier distributions and population dynamics in a large oligotrophic lake.

Nitrification is an important control on the form and distribution of nitrogen in freshwater ecosystems. However, the seasonality of nitrogen pools and the diversity of organisms catalyzing this process have not been well documented in oligotrophic lakes. Here, we show that nitrogen pools and nitrifying organisms in Flathead Lake are temporally and vertically dynamic, with nitrifiers displaying specific preferences depending on the season. While the ammonia-oxidizing bacteria (AOB) Nitrosomonadaceae and nitrite-oxidizing bacteria (NOB) Nitrotoga dominate at depth in the summer, the ammonia-oxidizing archaea (AOA) Nitrososphaerota and NOB Nitrospirota become abundant in the winter. Given clear seasonality in ammonium, with higher concentrations during the summer, we hypothesize that the succession between these two nitrifying groups may be due to nitrogen affinity, with AOB more competitive when ammonia concentrations are higher and AOA when they are lower. Nitrifiers in Flathead Lake share more than 99% average nucleotide identity with those reported in other North American lakes but are distinct from those in Europe and Asia, indicating a role for geographic isolation as a factor controlling speciation among nitrifiers. Our study shows there are seasonal shifts in nitrogen pools and nitrifying populations, highlighting the dynamic spatial and temporal nature of nitrogen cycling in freshwater ecosystems.

Combined impact of TiO2 nanoparticles and antibiotics on the activity and bacterial community of partial nitrification system.

The effects of TiO2 nanoparticles (nano-TiO2) together with antibiotics leaking into wastewater treatment plants (WWTPs), especially the partial nitrification (PN) process remain unclear. To evaluate the combined impact and mechanisms of nano-TiO2 and antibiotics on PN systems, batch experiments were carried out with six bench-scale sequencing batch reactors. Nano-TiO2 at a low level had minimal effects on the PN system. In combination with tetracycline and erythromycin, the acute impact of antibiotics was enhanced. Both steps of nitrification were retarded due to the decrease of bacterial activity and abundance, while nitrite-oxidizing bacteria were more sensitive to the inhibition than ammonia-oxidizing bacteria. Proteobacteria at the phylum level and Nitrosospira at the genus level remained predominant under single and combined impacts. The flow cytometry analysis showed that nano-TiO2 enhanced the toxicity of antibiotics through increasing cell permeability. Our results can help clarify the risks of nano-TiO2 combined with antibiotics to PN systems and explaining the behavior of nanoparticles in WWTPs.

Changes in soil ammonia oxidizers and potential nitrification after clear-cutting of boreal forests in China.

The Korean pine and broad-leaved mixed forests are the most typical and complete ecosystem among the global boreal forests, with extremely important ecological functions. However, few studies on the changes of soil ammonia oxidizers and potential nitrification after clear-cutting of forests are reported. In this study, in contrast to primary Korean pine forests, nitrate (NO3-) was significantly higher in secondary broad-leaved forests, while ammonium (NH4+) was on the contrary. The abundance of ammonia-oxidizing bacteria (AOB) was greatly higher in secondary broad-leaved forests, while levels of ammonia-oxidizing archaea (AOA) were not significantly different between them. The significant differences of community structure of AOA and AOB were observed in different forest types and soil layers. Compared with AOA, community compositions of AOB was more sensitive to forest type. The dominant groups of AOA were Nitrososphaera and Nitrosotalea, and the dominant group of AOB was Nitrosospira, of which Nitrosospira cluster 2 and 4 were functional groups with highly activity. Soil potential nitrification rate (PNR) was higher in secondary broad-leaved forests. Furthermore, PNR and AOB abundance had a significant positive correlation, but no significant correlation with AOA abundance. These results provide insights into the soil nitrogen balance and effects on forest restoration after clear-cutting.

Physical mixing in coastal waters controls and decouples nitrification via biomass dilution.

Nitrification is a central process of the aquatic nitrogen cycle that controls the supply of nitrate used in other key processes, such as phytoplankton growth and denitrification. Through time series observation and modeling of a seasonally stratified, eutrophic coastal basin, we demonstrate that physical dilution of nitrifying microorganisms by water column mixing can delay and decouple nitrification. The findings are based on a 4-y, weekly time series in the subsurface water of Bedford Basin, Nova Scotia, Canada, that included measurement of functional (amoA) and phylogenetic (16S rRNA) marker genes. In years with colder winters, more intense winter mixing resulted in strong dilution of resident nitrifiers in subsurface water, delaying nitrification for weeks to months despite availability of ammonium and oxygen. Delayed regrowth of nitrifiers also led to transient accumulation of nitrite (3 to 8 µmol · kgsw-1) due to decoupling of ammonia and nitrite oxidation. Nitrite accumulation was enhanced by ammonia-oxidizing bacteria (Nitrosomonadaceae) with fast enzyme kinetics, which temporarily outcompeted the ammonia-oxidizing archaea (Nitrosopumilus) that dominated under more stable conditions. The study reveals how physical mixing can drive seasonal and interannual variations in nitrification through control of microbial biomass and diversity. Variable, mixing-induced effects on functionally specialized microbial communities are likely relevant to biogeochemical transformation rates in other seasonally stratified water columns. The detailed study reveals a complex mechanism through which weather and climate variability impacts nitrogen speciation, with implications for coastal ecosystem productivity. It also emphasizes the value of high-frequency, multiparameter time series for identifying complex controls of biogeochemical processes in aquatic systems.

Community diversity and abundance of ammonia-oxidizing archaea and bacteria in shrimp pond sediment at different culture stages.

AIMS: Ammonia oxidation is a significant process of nitrogen cycles in a lot of ecosystems sediments while there are few studies in shrimp culture pond (SCP) sediments. This paper attempted to explore the community diversity and abundance of ammonia-oxidizing archaea (AOA) and ammonia-oxidizing bacteria (AOB) in SCP sediments at different culture stages. METHODS AND RESULTS: We collected SCP sediments and analysed the community diversity and abundance of AOA and bacteria in shrimp pond sediment at different culture stages using the ammonia monooxygenase (amoA) gene with quantitative PCR (qPCR) and 16S rRNA gene sequencing. The AOB-amoA gene abundance was showed higher than AOA-amoA gene abundance in SCP sediments on Day 50 and Day 60 after shrimp larvae introducing into the pond, and the diversity of AOA in SCP sediments was higher than that of AOB. The phylogenetic tree revealed that the most of AOA were the member of Nitrosopumilus and Nitrososphaera, and the majority of AOB sequences were clustered into Nitrosospira, Nitrosomonas clusters 6a and 7. The AOA community has close relationship with total organic carbon (TOC), pH, total phosphorus (TP), nitrate reductase, urease, acid phosphatase and ß-glucosidase. The AOB community was related to TOC, C/N and nitrate reductase. CONCLUSIONS: AOA and AOB play the different ecological roles in SCP sediments at different culture stages. SIGNIFICANCE AND IMPACT OF THE STUDY: Our results suggested that the different community diversity and abundance of AOA and AOB in SCP sediments, which may improve our ecological cognition of shrimp culture stages in SCP ecosystems.

Shifts in gut microbiome and metabolome are associated with risk of recurrent atrial fibrillation.

Alternations of gut microbiota (GM) in atrial fibrillation (AF) with elevated diversity, perturbed composition and function have been described previously. The current work aimed to assess the association of GM composition with AF recurrence (RAF) after ablation based on metagenomic sequencing and metabolomic analyses and to construct a GM-based predictive model for RAF. Compared with non-AF controls (50 individuals), GM composition and metabolomic profile were significantly altered between patients with recurrent AF (17 individuals) and non-RAF group (23 individuals). Notably, discriminative taxa between the non-RAF and RAF groups, including the families Nitrosomonadaceae and Lentisphaeraceae, the genera Marinitoga and Rufibacter and the species Faecalibacterium spCAG:82, Bacillus gobiensis and Desulfobacterales bacterium PC51MH44, were selected to construct a taxonomic scoring system based on LASSO analysis. After incorporating the clinical factors of RAF, taxonomic score retained a significant association with RAF incidence (HR = 2.647, P = .041). An elevated AUC (0.954) and positive NRI (1.5601) for predicting RAF compared with traditional clinical scoring (AUC = 0.6918) were obtained. The GM-based taxonomic scoring system theoretically improves the model performance, and the nomogram and decision curve analysis validated the clinical value of the predicting model. These data provide novel possibility that incorporating the GM factor into future recurrent risk stratification.

Effects of thiosulfate addition on ammonia and nitrogen removal in biofilters packed with Oyaishi (pumice tuff).

Ammonia removal is achieved partly by absorption and nitrification in biofilters, resulting in the accumulation of nitrogen and the necessity of treating the effluent water. We investigated the effects of thiosulfate addition to a biofilter containing pumice tuff for ammonia and nitrogen removal in a laboratory-scale experiment. The addition of thiosulfate to the circulating water led to a decreased nitrate and nitrite along with an increase of sulfate. The inorganic nitrogen in the circulating water decreased by up to 44% with thiosulfate addition compared to without thiosulfate. Batch experiments revealed that denitrification activity decreased exponentially along with increases in dissolved oxygen; however, approximately 30% of denitrification activity was maintained at dissolved oxygen concentration of 3.3 mg/L. Metabarcoding of 16S rRNA genes indicated that the genus Thiobacillus had a relative abundance of 0.002%-0.016% of total bacteria in the biofilter packing material. The circulating water pH was decreased below 5 with sulfur oxidation, and ammonium was accumulated without pH control resulting in a decrease in the relative abundance of the family Nitrosomonadaceae. Its relative abundance increased with control of pH to near neutral, indicating that ammonia-oxidizing activity could be maintained by adjusting pH. Thiosulfate addition could stimulate nitrogen removal by sulfur-dependent denitrification in biofiltration systems.

Long-term fertilization regimes change soil nitrification potential by impacting active autotrophic ammonia oxidizers and nitrite oxidizers as assessed by DNA stable isotope probing.

Chemoautotrophic ammonia-oxidizers and nitrite-oxidizers are responsible for a significant amount of soil nitrate production. The identity and composition of these active nitrifiers in soils under different long-term fertilization regimes remain largely under-investigated. Based on that soil nitrification potential significantly decreased in soils with chemical fertilization (CF) and increased in soils with organic fertilization (OF), a microcosm experiment with DNA stable isotope probing was further conducted to clarify the active nitrifiers. Both ammonia-oxidizing archaea (AOA) and bacteria (AOB) were found to actively respond to urea addition in soils with OF and no fertilizer (CK), whereas only AOB were detected in soils with CF. Around 98% of active AOB were Nitrosospira cluster 3a.1 in all tested soils, and more than 90% of active AOA were Nitrososphaera subcluster 1.1 in unfertilized and organically fertilized soils. Nitrite oxidation was performed only by Nitrospira-like bacteria in all soils. The relative abundances of Nitrospira lineage I and VI were 32% and 61%, respectively, in unfertilized soils, and that of Nitrospira lineage II was 97% in fertilized soils, indicating long-term fertilization shifted the composition of active Nitrospira-like bacteria in response to urea. This finding indicates that different fertilizer regimes impact the composition of active nitrifiers, thus, impacting soil nitrification potential.

Nitrification inhibitors effectively target N2 O-producing Nitrosospira spp. in tropical soil.

The nitrification inhibitors (NIs) 3,4-dimethylpyrazole (DMPP) and dicyandiamide (DCD) can effectively reduce N2 O emissions; however, which species are targeted and the effect of these NIs on the microbial nitrifier community is still unclear. Here, we identified the ammonia oxidizing bacteria (AOB) species linked to N2 O emissions and evaluated the effects of urea and urea with DCD and DMPP on the nitrifying community in a 258 day field experiment under sugarcane. Using an amoA AOB amplicon sequencing approach and mining a previous dataset of 16S rRNA sequences, we characterized the most likely N2 O-producing AOB as a Nitrosospira spp. and identified Nitrosospira (AOB), Nitrososphaera (archaeal ammonia oxidizer) and Nitrospira (nitrite-oxidizer) as the most abundant, present nitrifiers. The fertilizer treatments had no effect on the alpha and beta diversities of the AOB communities. Interestingly, we found three clusters of co-varying variables with nitrifier operational taxonomic units (OTUs): the N2 O-producing AOB Nitrosospira with N2 O, NO3 - , NH4 + , water-filled pore space (WFPS) and pH; AOA Nitrososphaera with NO3 - , NH4 + and pH; and AOA Nitrososphaera and NOB Nitrospira with NH4 + , which suggests different drivers. These results support the co-occurrence of non-N2 O-producing Nitrososphaera and Nitrospira in the unfertilized soils and the promotion of N2 O-producing Nitrosospira under urea fertilization. Further, we suggest that DMPP is a more effective NI than DCD in tropical soil under sugarcane.

Comparison of chemical and microbiological changes during the aerobic composting and vermicomposting of green waste.

This research was conducted to compare chemical and microbiological properties during aerobic composting (AC) and vermicomposting (VC) of green waste. Relative to AC, VC significantly decreased the pH and lignin and cellulose contents, and significantly increased the electrical conductivity and total N and available P contents. For AC, BIrii41_norank (order Myxococcales) was the major bacterial genus at 30 d and again became dominant genus from 90-150 d, with relative abundances of 2.88% and 4.77-5.19%, respectively; at 45 d and 60 d, the dominant bacterial genus was Nitrosomonadaceae_uncultured (order Nitrosomonadales) with relative abundances of 2.83-7.17%. For VC, the dominant bacterial genus was BIrii41_norank (except at 45 d), which accounted for 2.11-7.96% of the total reads. The dominant fungal class was Sordariomycetes in AC (relative abundances 39.2-80.6%) and VC (relative abundances 42.1-69.5%). The abundances of microbial taxa and therefore the bacterial and fungal community structures differed between VC and AC. The quality of the green waste compost product was higher with VC than with AC. These results will also help to achieve further composting technology breakthroughs in reducing the composting time and improving compost quality.

Nitrosospira Cluster 8a Plays a Predominant Role in the Nitrification Process of a Subtropical Ultisol under Long-Term Inorganic and Organic Fertilization.

Long-term effects of inorganic and organic fertilization on nitrification activity (NA) and the abundances and community structures of ammonia-oxidizing bacteria (AOB) and ammonia-oxidizing archaea (AOA) were investigated in an acidic Ultisol. Seven treatments applied annually for 27 years comprised no fertilization (control), inorganic NPK fertilizer (N), inorganic NPK fertilizer plus lime (CaCO3) (NL), inorganic NPK fertilizer plus peanut straw (NPS), inorganic NPK fertilizer plus rice straw (NRS), inorganic NPK fertilizer plus radish (NR), and inorganic NPK fertilizer plus pig manure (NPM). In nonfertilized soil, the abundance of AOA was 1 order of magnitude higher than that of AOB. Fertilization reduced the abundance of AOA but increased that of AOB, especially in the NL treatment. The AOA communities in the control and the N treatments were dominated by the Nitrososphaera and B1 clades but shifted to clade A in the NL and NPM treatments. Nitrosospira cluster 8a was found to be the most dominant AOB in all treatments. NA was primarily regulated by soil properties, especially soil pH, and the interaction with AOB abundance explained up to 73% of the variance in NA. When NL soils with neutral pH were excluded from the analysis, AOB abundance, especially the relative abundance of Nitrosospira cluster 8a, was positively associated with NA. In contrast, there was no association between AOA abundance and NA. Overall, our data suggest that Nitrosospira cluster 8a of AOB played an important role in the nitrification process in acidic soil following long-term inorganic and organic fertilization.IMPORTANCE The nitrification process is an important step in the nitrogen (N) cycle, affecting N availability and N losses to the wider environment. Ammonia oxidation, which is the first and rate-limiting step of nitrification, was widely accepted to be mainly regulated by AOA in acidic soils. However, in this study, nitrification activity was correlated with the abundance of AOB rather than that of AOA in acidic Ultisols. Nitrosospira cluster 8a, a phylotype of AOB which preferred warm temperatures, and low soil pH played a predominant role in the nitrification process in the test Ultisols. Our results also showed that long-term application of lime or pig manure rather than plant residues altered the community structure of AOA and AOB. Taken together, our findings contribute new knowledge to the understanding of the nitrification process and ammonia oxidizers in subtropical acidic Ultisol under long-term inorganic and organic fertilization.

Ferrous ion as a reducing agent in the generation of antibiofilm nitric oxide from a copper-based catalytic system.

The work found that the electron-donating properties of ferrous ions (Fe2+) can be used for the conversion of nitrite (NO2-) into the biofilm-dispersing signal nitric oxide (NO) by a copper(II) complex (CuDTTCT) catalyst, a potentially applicable biofilm control technology for the water industries. The availability of Fe2+ varied depending on the characteristics of the aqueous systems (phosphate- and carbonate-containing nitrifying bacteria growth medium, NBGM and phosphate buffered saline, PBS at pH 6 to 8, to simulate conditions typically present in the water industries) and was found to affect the production of NO from nitrite by CuDTTCT (casted into PVC). Greater amounts of NO were generated from the CuDTTCT-nitrite-Fe2+ systems in PBS compared to those in NBGM, which was associated with the reduced extent of Fe2+-to-Fe3+ autoxidation by the iron-precipitating moieties phosphates and carbonate in the former system. Further, acidic conditions at pH 6.0 were found to favor NO production from the catalytic system in both PBS and NBGM compared to neutral or basic pH (pH 7.0 or 8.0). Lower pH was shown to stabilize Fe2+ and reduce its autoxidation to Fe3+. These findings will be beneficial for the potential implementation of the NO-generating catalytic technology and indeed, a 'non-killing' biofilm dispersal activity of CuDTTCT-nitrite-Fe2+ was observed on nitrifying bacteria biofilms in PBS at pH 6.

Mixed nitrifying bacteria culture under different temperature dropping strategies: Nitrification performance, activity, and community.

In this study, the nitrification performance, metabolic activity, antioxidant enzyme activity as well as bacterial community of mixed nitrifying bacteria culture under different temperature dropping strategies [(#1) growth temperature kept at 20 °C; (#2) sharp1 decreased from 20 °C to 10 °C; (#3) growth at 20 °C for 6 days followed by sharp decrease to 10 °C; and (#4) gradual decreased from 20 °C to 10 °C] were evaluated. It was shown that acclimation at 20 °C for 6 days allowed to maintain better nitrification activity at 10 °C. The nitrite oxidation capacity of nitrifiers was significantly correlated with the relative light unit (RLU) (p < .05) and the fluctuation of superoxide dismutase (SOD) enzyme activity (p < .01). With serial #3 showed the highest RLU levels and the least SOD enzyme fluctuation as compared to serials #2 and #4. Throughout the experimental period, Nitrosospira and Nitrosomonas as well as Nitrospira were identified as the predominant ammonia-oxidizing bacteria (AOB) and nitrate-oxidizing bacteria (NOB). The dynamic change of AOB/NOB ratios and nitrification activity in serials #2-#4 demonstrated that AOB recovered better than NOB with long-term 10 °C exposure, and the nitrification performance was mainly limited by the nitrite oxidation capacity of NOB. Applying 6 days acclimation at 20 °C was beneficial for the mixed nitrifying bacteria culture to cope with low temperature (10 °C) stress, possibly due to the maintenance of metabolic activity, antioxidant enzyme activity stability as well as appropriate AOB/NOB ratio.

A patchwork pathway for oxygenase-independent degradation of side chain containing steroids.

The denitrifying betaproteobacterium Sterolibacterium denitrificans serves as model organism for studying the oxygen-independent degradation of cholesterol. Here, we demonstrate its capability of degrading various globally abundant side chain containing zoo-, phyto- and mycosterols. We provide the complete genome that empowered an integrated genomics/proteomics/metabolomics approach, accompanied by the characterization of a characteristic enzyme of steroid side chain degradation. The results indicate that individual molybdopterin-containing steroid dehydrogenases are involved in C25-hydroxylations of steroids with different isoprenoid side chains, followed by the unusual conversion to C26-oic acids. Side chain degradation to androsta-1,4-diene-3,17-dione (ADD) via aldolytic C-C bond cleavages involves acyl-CoA synthetases/dehydrogenases specific for the respective 26-, 24- and 22-oic acids/-oyl-CoAs and promiscuous MaoC-like enoyl-CoA hydratases, aldolases and aldehyde dehydrogenases. Degradation of rings A and B depends on gene products uniquely found in anaerobic steroid degraders, which after hydrolytic cleavage of ring A, again involves CoA-ester intermediates. The degradation of the remaining CD rings via hydrolytic cleavage appears to be highly similar in aerobic and anaerobic bacteria. Anaerobic cholesterol degradation employs a composite repertoire of more than 40 genes partially known from aerobic degradation in gammaproteobacteria/actinobacteria, supplemented by unique genes that are required to circumvent oxygenase-dependent reactions.

Acyl-Homoserine Lactone Production in Nitrifying Bacteria of the Genera Nitrosospira, Nitrobacter, and Nitrospira Identified via a Survey of Putative Quorum-Sensing Genes.

The genomes of many bacteria that participate in nitrogen cycling through the process of nitrification contain putative genes associated with acyl-homoserine lactone (AHL) quorum sensing (QS). AHL QS or bacterial cell-cell signaling is a method of bacterial communication and gene regulation and may be involved in nitrogen oxide fluxes or other important phenotypes in nitrifying bacteria. Here, we carried out a broad survey of AHL production in nitrifying bacteria in three steps. First, we analyzed the evolutionary history of AHL synthase and AHL receptor homologs in sequenced genomes and metagenomes of nitrifying bacteria to identify AHL synthase homologs in ammonia-oxidizing bacteria (AOB) of the genus Nitrosospira and nitrite-oxidizing bacteria (NOB) of the genera Nitrococcus, Nitrobacter, and Nitrospira Next, we screened cultures of both AOB and NOB with uncharacterized AHL synthase genes and AHL synthase-negative nitrifiers by a bioassay. Our results suggest that an AHL synthase gene is required for, but does not guarantee, cell density-dependent AHL production under the conditions tested. Finally, we utilized mass spectrometry to identify the AHLs produced by the AOB Nitrosospira multiformis and Nitrosospira briensis and the NOB Nitrobacter vulgaris and Nitrospira moscoviensis as N-decanoyl-l-homoserine lactone (C10-HSL), N-3-hydroxy-tetradecanoyl-l-homoserine lactone (3-OH-C14-HSL), a monounsaturated AHL (C10:1-HSL), and N-octanoyl-l-homoserine lactone (C8-HSL), respectively. Our survey expands the list of AHL-producing nitrifiers to include a representative of Nitrospira lineage II and suggests that AHL production is widespread in nitrifying bacteria.IMPORTANCE Nitrification, the aerobic oxidation of ammonia to nitrate via nitrite by nitrifying microorganisms, plays an important role in environmental nitrogen cycling from agricultural fertilization to wastewater treatment. The genomes of many nitrifying bacteria contain genes associated with bacterial cell-cell signaling or quorum sensing (QS). QS is a method of bacterial communication and gene regulation that is well studied in bacterial pathogens, but less is known about QS in environmental systems. Our previous work suggested that QS might be involved in the regulation of nitrogen oxide gas production during nitrite metabolism. This study characterized putative QS signals produced by different genera and species of nitrifiers. Our work lays the foundation for future experiments investigating communication between nitrifying bacteria, the purpose of QS in these microorganisms, and the manipulation of QS during nitrification.

Optimization of overexpression of a chaperone protein of steroid C25 dehydrogenase for biochemical and biophysical characterization.

Molybdenum is an essential nutrient for metabolism in plant, bacteria, and animals. Molybdoenzymes are involved in nitrogen assimilation and oxidoreductive detoxification, and bioconversion reactions of environmental, industrial, and pharmaceutical interest. Molybdoenzymes contain a molybdenum cofactor (Moco), which is a pyranopterin heterocyclic compound that binds a molybdenum atom via a dithiolene group. Because Moco is a large and complex compound deeply buried within the protein, molybdoenzymes are accompanied by private chaperone proteins responsible for the cofactor's insertion into the enzyme and the enzyme's maturation. An efficient recombinant expression and purification of both Moco-free and Moco-containing molybdoenzymes and their chaperones is of paramount importance for fundamental and applied research related to molybdoenzymes. In this work, we focused on a D1 protein annotated as a chaperone of steroid C25 dehydrogenase (S25DH) from Sterolibacterium denitrificans Chol-1S. The D1 protein is presumably involved in the maturation of S25DH engaged in oxygen-independent oxidation of sterols. As this chaperone is thought to be a crucial element that ensures the insertion of Moco into the enzyme and consequently, proper folding of S25DH optimization of the chaperon's expression is the first step toward the development of recombinant expression and purification methods for S25DH. We have identified common E. coli strains and conditions for both expression and purification that allow us to selectively produce Moco-containing and Moco-free chaperones. We have also characterized the Moco-containing chaperone by EXAFS and HPLC analysis and identified conditions that stabilize both forms of the protein. The protocols presented here are efficient and result in protein quantities sufficient for biochemical studies.

Treatment of petrochemical wastewater by microaerobic hydrolysis and anoxic/oxic processes and analysis of bacterial diversity.

Microaerobic hydrolysis-acidification (MHA)-anoxic-oxic (A/O) processes were developed to treat actual petrochemical wastewater. The results showed that the overall COD removal efficiency was 72-79% at HRT=20h, and MHA accounted for 33-42% of COD removal, exhibiting good efficiency of acidogenic fermentation. Ammonium removal was more than 94%. The main pollutants in the influent were identified to be benzene, ketone, alcohols, amine, nitrile and phenols by GC-MS, and the majority of pollutants could be removed by MHA-A/O treatment. Proteobacteria was the most dominant bacteria in the system, accounting for more than 55% of the reads. The predominant genera in MHA, anoxic and oxic reactors were Anaerolineaceae and Sulfuritalea, Lactococcus and Blastocatella, and Saprospiraceae uncultured and Nitrosomonadaceae, respectively. This treatment system exhibited good performance in degrading the complex compounds in the petrochemical wastewater.

Microbial community composition of a household sand filter used for arsenic, iron, and manganese removal from groundwater in Vietnam.

Household sand filters are used in rural areas of Vietnam to remove As, Fe, and Mn from groundwater for drinking water purposes. Currently, it is unknown what role microbial processes play in mineral oxide formation and As removal during water filtration. We performed most probable number counts to quantify the abundance of physiological groups of microorganisms capable of catalyzing Fe- and Mn-redox transformation processes in a household sand filter. We found up to 10(4) cells g(-1) dry sand of nitrate-reducing Fe(II)-oxidizing bacteria and Fe(III)-reducing bacteria, and no microaerophilic Fe(II)-oxidizing bacteria, but up to 10(6) cells g(-1) dry sand Mn-oxidizing bacteria. 16S rRNA gene amplicon sequencing confirmed MPN counts insofar as only low abundances of known taxa capable of performing Fe- and Mn-redox transformations were detected. Instead the microbial community on the sand filter was dominated by nitrifying microorganisms, e.g. Nitrospira, Nitrosomonadales, and an archaeal OTU affiliated to Candidatus Nitrososphaera. Quantitative PCR for Nitrospira and ammonia monooxygenase genes agreed with DNA sequencing results underlining the numerical importance of nitrifiers in the sand filter. Based on our analysis of the microbial community composition and previous studies on the solid phase chemistry of sand filters we conclude that abiotic Fe(II) oxidation processes prevail over biotic Fe(II) oxidation on the filter. Yet, Mn-oxidizing bacteria play an important role for Mn(II) oxidation and Mn(III/IV) oxide precipitation in a distinct layer of the sand filter. The formation of Mn(III/IV) oxides contributes to abiotic As(III) oxidation and immobilization of As(V) by sorption to Fe(III) (oxyhydr)oxides.

Nitrosospira lacus sp. nov., a psychrotolerant, ammonia-oxidizing bacterium from sandy lake sediment.

A Gram-negative, spiral-shaped, chemolithotrophic, ammonia-oxidizing bacterium, designated APG3(T), was isolated into pure culture from sandy lake sediment collected from Green Lake, Seattle, WA, USA. Phylogenetic analyses based on the 16S rRNA gene sequence showed that strain APG3(T) belongs to cluster 0 of the genus Nitrosospira, which is presently not represented by described species, with Nitrosospira multiformis (cluster 3) as the closest species with a validly published name (identity of 98.6 % to the type strain). Strain APG3(T) grew at 4 °C but could not grow at 35 °C, indicating that this bacterium is psychrotolerant. Remarkably, the strain was able to grow over a wide range of pH (pH 5-9), which was greater than the pH range of any studied ammonia-oxidizing bacteria in pure culture. The DNA G+C content of the APG3(T) genome is 53.5 %, which is similar to that of Nitrosospira multiformis ATCC 25196(T) (53.9 %) but higher than that of Nitrosomonas europaea ATCC 19718 (50.7 %) and Nitrosomonas eutropha C71 (48.5 %). The average nucleotide identity (ANI) calculated for the genomes of strain APG3(T) and Nitrosospira multiformis ATCC 25196(T) was 75.45 %, significantly lower than the value of 95 % ANI that corresponds to the 70 % species-level cut-off based on DNA-DNA hybridization. Overall polyphasic taxonomy study indicated that strain APG3(T) represents a novel species in the genus Nitrosospira, for which the name Nitrosospira lacus sp. nov. is proposed (type strain APG3(T) = NCIMB 14869(T) = LMG 27536(T) = ATCC BAA-2542(T)).

Removal of bisphenol A (BPA) in a nitrifying system with immobilized biomass.

The potential for bisphenol A (BPA) removal by mixed consortia of immobilized microorganisms with high nitrification activity was investigated with BPA concentrations in the influent from 2.5 to 10.0 mg/L. The presence of BPA limited ammonium oxidation; nitrification efficiency decreased from 91.2±1.3% in the control series to 47.4±9.4% when BPA concentration in wastewater was the highest. The efficiency of BPA removal rose from 87.1±5.5% to 92.9±2.9% with increased BPA concentration in the influent. Measurement of oxygen uptake rates by biomass exposed to BPA showed that BPA was mainly removed by heterotrophic bacteria. A strong negative correlation between the BPA removal efficiency and nitrification efficiency indicated the limited contribution of ammonia-oxidizing bacteria (AOB) to BPA biodegradation. Exposure of biomass to BPA changed the quantity and diversity of AOB in the biomass as shown by real-time PCR and denaturing gradient gel electrophoresis.