The name Bradyrhizobiaceae Garrity et al. 2006 contains Nitrobacter Winogradsky 1892 (Approved Lists 1980), the nomenclatural type of the family Nitrobacteraceae Buchanan 1917 (Approved Lists 1980), is illegitimate and proposals to alter the wording of Rule 54 of the International Code of Nomenclature of Prokaryotes to clarify the fact that the family name Bradyrhizobiaceae Garrity et al. 2006 is replaced by the family name Nitrobacteraceae Buchanan 1917 (Approved Lists 1980) the only correct name.

The new name at the rank of family Bradyrhizobiaceae Garrity et al. 2006 was created to include the genera Afipia Brenner et al. 1992, Agromonas Ohta and Hattori 1985, Blastobacter Zavarzin 1961 (Approved Lists 1980), Bosea Das et al. 1996, Bradyrhizobium Jordan 1982 (the nomenclatural type), Nitrobacter Winogradsky 1892 (Approved Lists 1980), Oligotropha Meyer et al. 1994, Rhodoblastus Imhoff 2001 and Rhodopseudomonas Czurda and Maresch 1937 (Approved Lists 1980). However, Nitrobacter Winogradsky 1892 (Approved Lists 1980) is the nomenclatural type of Nitrobacteraceae Buchanan 1917 (Approved Lists 1980) a name at the rank of family that was validly published prior to the valid publication of Bradyrhizobiaceae Garrity et al. 2006 and has priority. In addition Rule 51b (1) of the International Code of Nomenclature of Prokaryotes rules that under these circumstances Bradyrhizobiaceae Garrity et al. 2006 is an illegitimate name. Illegitimate names may not be used (Rule 51a) and illegitimate names are also not taken into consideration when determining priority (Rule 23a). Nitrobacteraceae Buchanan 1917 (Approved Lists 1980) is the only correct name (Rule 23a). Despite these facts the name Bradyrhizobiaceae Garrity et al. 2006 continues to be used, perhaps because the fact that it is an illegitimate name and the consequences of that status are not fully understood. A revision of Rule 54 would also appear to be appropriate in order to further emphasise the fact that the family name Bradyrhizobiaceae Garrity et al. 2006 must be replaced by the family name Nitrobacteraceae Buchanan 1917 (Approved Lists 1980), which is the oldest legitimate name and is the only correct name that may be used for the taxon that includes Bradyrhizobium Jordan 1982 and Nitrobacter Winogradsky 1892 (Approved Lists 1980).

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.

Artificial Intelligence for the Evaluation of Operational Parameters Influencing Nitrification and Nitrifiers in an Activated Sludge Process.

Nitrification at a full-scale activated sludge plant treating municipal wastewater was monitored over a period of 237 days. A combination of fluorescent in situ hybridization (FISH) and quantitative real-time polymerase chain reaction (qPCR) were used for identifying and quantifying the dominant nitrifiers in the plant. Adaptive neuro-fuzzy inference system (ANFIS), Pearson's correlation coefficient, and quadratic models were employed in evaluating the plant operational conditions that influence the nitrification performance. The ammonia-oxidizing bacteria (AOB) abundance was within the range of 1.55 × 10(8)-1.65 × 10(10) copies L(-1), while Nitrobacter spp. and Nitrospira spp. were 9.32 × 10(9)-1.40 × 10(11) copies L(-1) and 2.39 × 10(9)-3.76 × 10(10) copies L(-1), respectively. Specific nitrification rate (qN) was significantly affected by temperature (r 0.726, p 0.002), hydraulic retention time (HRT) (r -0.651, p 0.009), and ammonia loading rate (ALR) (r 0.571, p 0.026). Additionally, AOB was considerably influenced by HRT (r -0.741, p 0.002) and temperature (r 0.517, p 0.048), while HRT negatively impacted Nitrospira spp. (r -0.627, p 0.012). A quadratic combination of HRT and food-to-microorganism (F/M) ratio also impacted qN (r (2) 0.50), AOB (r (2) 0.61), and Nitrospira spp. (r (2) 0.72), while Nitrobacter spp. was considerably influenced by a polynomial function of F/M ratio and temperature (r (2) 0.49). The study demonstrated that ANFIS could be used as a tool to describe the factors influencing nitrification process at full-scale wastewater treatment plants.

Nitrite-Oxidizing Bacterium Nitrobacter winogradskyi Produces N-Acyl-Homoserine Lactone Autoinducers.

Nitrobacter winogradskyi is a chemolithotrophic bacterium that plays a role in the nitrogen cycle by oxidizing nitrite to nitrate. Here, we demonstrate a functional N-acyl-homoserine lactone (acyl-HSL) synthase in this bacterium. The N. winogradskyi genome contains genes encoding a putative acyl-HSL autoinducer synthase (nwi0626, nwiI) and a putative acyl-HSL autoinducer receptor (nwi0627, nwiR) with amino acid sequences 38 to 78% identical to those in Rhodopseudomonas palustris and other Rhizobiales. Expression of nwiI and nwiR correlated with acyl-HSL production during culture. N. winogradskyi produces two distinct acyl-HSLs, N-decanoyl-l-homoserine lactone (C10-HSL) and a monounsaturated acyl-HSL (C10:1-HSL), in a cell-density- and growth phase-dependent manner, during batch and chemostat culture. The acyl-HSLs were detected by bioassay and identified by ultraperformance liquid chromatography with information-dependent acquisition mass spectrometry (UPLC-IDA-MS). The C=C bond in C10:1-HSL was confirmed by conversion into bromohydrin and detection by UPLC-IDA-MS.

[Start-up Characteristics of Four-zone Integrated Reactor for Nitrogen Removal in Winter and Analysis of Nitrobacteria Community].

The treatment of decentralized sewage has gained more and more attention in China in recent years. A four-zone integrated reactor was designed by combining biofilm system and activated sludge system and start-up by gradient shorten the HRT. The removal of COD, NH4+ -N and TN were studied at 8-15°C. Fluorescence in situ hybridization(FISH) was used to detect nitrobacteria population (AOB,NOB) so as to study the relationship between the reactor effect and functional micro-bacteria. The results showed that, when the HRT was 9.2 h, the removal efficiencies of COD, ammonia and TN were 92. 11%, 99. 21% and 61. 63%, respectively. Compared to the initial stage, the numbers of AOB and NOB in the late phase were increased by 5. 82 and 6. 14 times, respectively. In addition, the proportion of nitrobacteria was increased from 6. 12% to 16. 38% , which became the dominant bacteria in biofilms. Moreover, the nitrification efficiency was increased from 78. 49% to 97. 52% , while the number of NOB was 5. 61-fold increased and the value of AOB/NOB was optimized to 1. 47. The effluent quality is guaranteed by the enrichment of AOB and NOB and suitable value of AOB/NOB.

Cultivation, growth physiology, and chemotaxonomy of nitrite-oxidizing bacteria.

Lithoautotrophic nitrite-oxidizing bacteria (NOB) are known as fastidious microorganisms, which are hard to maintain and not many groups are trained to keep them in culture. They convert nitrite stoichiometrically to nitrate and growth is slow due to the poor energy balance. NOB are comprised of five genera, which are scattered among the phylogenetic tree. Because NOB proliferate in a broad range of environmental conditions (terrestrial, marine, acidic) and have diverse lifestyles (lithoautotrophic, mixotrophic, and heterotrophic), variation in media composition is necessary to match their individual growth requirements in the laboratory. From Nitrobacter and Nitrococcus relatively high cell amounts can be achieved by consumption of high nitrite concentrations, whereas accumulation of cells belonging to Nitrospira, Nitrospina, or the new candidate genus Nitrotoga needs prolonged feeding procedures. Isolation is possible for planktonic cells by dilution series or plating techniques, but gets complicated for strains with a tendency to develop microcolonies like Nitrospira. Physiological experiments including determination of the temperature or pH-optimum can be conducted with active laboratory cultures of NOB, but the attainment of reference values like cell protein content or cell numbers might be hard to realize due to the formation of flocs and the low cell density. Monitoring of laboratory enrichments is necessary especially if several species or genera coexist within the same culture and due to population shifts over time. Chemotaxonomy is a valuable method to identify and quantify NOB in biofilms and pure cultures alike, since fatty acid profiles reflect their phylogenetic heterogeneity. This chapter focusses on methods to enrich, isolate, and characterize NOB by various cultivation-based techniques.

Seasonal variations of nitrifying community in trickling filter-solids contact (TF/SC) activated sludge systems.

Two full-scale trickling filter/solids contact (TF/SC) basin plants, each successfully performing nitrification, were sampled throughout various seasons over a period of one year. Concentrations of ammonia, nitrate and nitrite were measured at various sampling locations along the treatment train. DNA was also extracted from mixed liquor in the solids contact basins. These DNA samples were subjected to terminal restriction fragment length polymorphism (TRFLP) in order to profile the ammonia oxidizing bacteria and nitrite oxidizing bacteria communities. In both plants, there was a prevalence of Nitrosomonas europaea among the ammonia oxidizing bacteria (AOBs). However, during the summer months, there was increased diversity of Nitrosomonas species. Likewise, Nitrospira spp. was the dominant nitrite oxidizing bacteria (NOBs) in both plants regardless of season. Yet there was an increased presence of Nitrobacter among the NOBs in the summer months. These results add an important understanding of the ecology and dynamics in nitrifying population in full-scale TF/SC wastewater treatment plants.

[Analysis of microbial diversity of nitrifying bacteria by terminal restriction fragment length polymorphism].

We analyzed the microbial diversity and quantity of nitrifying bacteria in the enrichment reactor by Terminal Restriction Fragment Length Polymorphism (T_RFLP), a cultured-independent molecular technique. The result indicated that nitrobacteria enriched the best, and the diversity index decreased 62.80% compared with the initial data. Nitrobacteria were predominant in the reactor. Meanwhile, we studied the microbial diversity before and after adding Nitrobacteria into shrimp ponds, and analyzed several major bacterial species that existed stably in the pond. According to the analysis by T_RFLP program, species including Brevibacillus brevis, Microbacterium lactium, Azoarcus indigens and Bordetella holmesii were the dominant bacteria in the ponds.

[Characteristics of N forms and N-transforming bacteria in paddy soil under different tillage patterns].

Paddy soil samples were collected in layers (0-5, 5-12, and 12-20 cm) during rice growth period to investigate the characteristics of the N forms and N-transforming bacteria in the soil profile under different tillage patterns (no-tillage with straw returning, NTS; conventional tillage with straw returning, CTS; no-tillage, NT; and conventional tillage, CT). In the whole rice growth period, ammonifying bacteria in 0-5 cm soil layer had the highest number under NTS, and nitrosobacteria in 0-5 cm and 5-12 cm soil layers were more abundant but in 12-20 cm soil layer were lesser under CT than under NT. Nitrosobacteria and denitrobacteria in 0-20 cm soil layer were lesser under NTS than under CTS. At elongating and ripening stages, anaerobic N-fixing bacteria in 0-5 cm soil layer were more abundant under NT than under CT. In the whole rice growth period, the alkali-hydrolyzable N and total N contents in 0-5 cm soil layer were significantly higher but in 5-12 cm and 12-20 cm soil layers were lower under NT than under CT, and the NH4(+)-N and NO3(-)-N contents in 0-20 cm soil layer were higher under NTS but in 12-20 cm soil layer had no significant differences between NT and CT. Correlation analysis and multiple polynomial regression analysis further revealed that there were significant relationships between soil NH4(+)-N and soil ammonifying bacteria, nitrosobacteria and denitrobacteria, and between soil alkali-hydrolyzable N and soil anaerobic N-fixing bacteria. Among the test tillage patterns, NTS could be the more desirable one for the N supply and fertility maintenance of paddy soil.

Nitrifying community structures and nitrification performance of full-scale municipal and swine wastewater treatment plants.

This study evaluated nitrification performance and microbial ecology of nitrifying sludge in two full-scale wastewater treatment plants (WWTPs) including a municipal WWTP treating 20mgNL(-1) of ammonium and a swine WWTP treating 220mgNL(-1) of ammonium. These two plants differed in both wastewater characteristics and operating parameters, such as influent COD, TKN, ammonium, hydraulic retention time, and solids retention time, even though both plants achieve >85% nitrification efficiency. By employing molecular techniques, including terminal restriction fragment length polymorphism, cloning-sequencing and phylogenetic analyses targeting the 16S ribosomal RNA and group specific ammonia-monooxygenase functional gene (amoA), microbial community structures of nitrifying sludge and their significance to nitrification performance were evaluated. The results reveal that for the municipal WWTP Nitrosomonas marina-like AOB (ammonia-oxidizing bacteria) and Nitrospira-like NOB (nitrite-oxidizing bacteria) were the ubiquitously dominant nitrifiers, while Nitrosomonas europaea-, Nitrosomonas oligotropha-, and Nitrosospira-like AOB and Nitrobacter- and Nitrospira-like NOB were the major nitrifying populations found in the swine WWTP. The observed dissimilar nitrifying populations prevailing in these two plants may be related to niche differentiation concerning ammonium concentrations, system operation, and salinity. Moreover, our results suggest that the swine nitrifying sludge, involving relatively diverse AOB and NOB populations that perform the same task but with distinct growth and survival characters, may allow communities to maintain nitrifying capabilities when conditions change such as sudden increases in ammonium concentrations as examined with nitrification kinetic batch tests.

Activated packed bed bioreactor for rapid nitrification in brackish water hatchery systems.

A packed bed bioreactor (PBBR) was developed for rapid establishment of nitrification in brackish water hatchery systems in the tropics. The reactors were activated by immobilizing ammonia-oxidizing (AMONPCU-1) and nitrite-oxidizing (NIONPCU-1) bacterial consortia on polystyrene and low-density polyethylene beads, respectively. Fluorescence in situ hybridization demonstrated the presence of autotrophic nitrifiers belong to Nitrosococcus mobilis, lineage of beta ammonia oxidizers and nitrite oxidizer Nitrobacter sp. in the consortia. The activated reactors upon integration to the hatchery system resulted in significant ammonia removal (P < 0.01) culminating to its undetectable levels. Consequently, a significantly higher percent survival of larvae was observed in the larval production systems. With spent water the reactors could establish nitrification with high percentage removal of ammonia (78%), nitrite (79%) and BOD (56%) within 7 days of initiation of the process. PBBR is configured in such a way to minimize the energy requirements for continuous operation by limiting the energy inputs to a single stage pumping of water and aeration to the aeration cells. The PBBR shall enable hatchery systems to operate under closed recirculating mode and pave the way for better water management in the aquaculture industry.

First exploration of Nitrobacter diversity in soils by a PCR cloning-sequencing approach targeting functional gene nxrA.

Nitrite oxidoreductase (NXR) is the key enzyme responsible for the oxidation of NO(2)(-) to NO(3)(-) in nitrite-oxidizing bacteria. For the first time a molecular approach for targeting the nxrA gene was developed, encoding the catalytic subunit of the NXR, to study diversity of Nitrobacter-like organisms based on the phylogeny of nxrA gene sequences in soils. NxrA sequences of the Nitrobacter strains analysed (Nitrobacter hamburgensis, Nitrobacter vulgaris, Nitrobacter winogradskyi, Nitrobacter alkalicus) by PCR, cloning and sequencing revealed the occurrence of multiple copies of nxrA genes in these strains. The copy number and similarity varied among strains. The diversity of Nitrobacter-like nxrA sequences was explored in three soils (a French permanent pasture soil, a French fallow soil, and an African savannah soil) using a cloning and sequencing approach. Most nxrA sequences found in these soils (84%) differed from nxrA sequences obtained from Nitrobacter strains. Moreover, the phylogenetic distribution and richness of nxrA-like sequences was extremely variable depending on soil type. This nxrA tool extends the panel of functional genes available for studying bacteria involved in the nitrogen cycle.

The phylogeny of the genus Nitrobacter based on comparative rep-PCR, 16S rRNA and nitrite oxidoreductase gene sequence analysis.

Strains of Nitrobacter mediate the second step in the nitrification process by oxidizing nitrite to nitrate. The phylogenetic diversity of the genus is currently not well investigated. In this study, a rep-PCR profile and the nearly complete 16S rRNA gene sequence of 30 strains, comprising a wide physiological as well as ecological diversity and encompassing representatives of the four species, were determined. The sequence diversity of the 16S rRNA gene between different species was low, indicating the need for additional phylogenetic markers. Therefore, primers were developed for amplifying the complete nxrX gene and a 380bp fragment of the nxrB1 gene, which are both genes involved in the nitrite oxidation process. These genes confirmed the division into phylogenetic groups revealed by the 16S rRNA gene but showed a better discriminatory power. They can be a valuable additional tool for phylogenetic analysis within the genus Nitrobacter and can assist in the identification of new Nitrobacter isolates.

Development and calibration of a nitrification PDE model based on experimental data issued from biofilter treating drinking water.

To remove ammonia for production of drinking water, nitrification can be performed in a bio-filter. At least 1 month is necessary to capture from the groundwater and then grow a sufficient amount of nitrifying bacteria to reach the desired removal efficiency. Improving start-up of bio-filters at low substrate concentration is therefore a major challenge. In this connection, it is important to develop appropriate models for designing, monitoring or analysing biofilm systems during start-up or following disinfection events. This study discusses the development and calibration of a nitrification PDE model which reflects the compromise between the complexity associated with the description of the full physical and biochemical mechanisms and the search for a simplified model with identifiable parameters. This model takes only the relevant phenomena (considering the full operating range) into account. The validity of the calibrated model has been evaluated through experiments under very different operational conditions, at the laboratory and under real industrial conditions, involving the full upstream chain of water treatment (iron oxidation and sand filter).

Genome sequence of the chemolithoautotrophic nitrite-oxidizing bacterium Nitrobacter winogradskyi Nb-255.

The alphaproteobacterium Nitrobacter winogradskyi (ATCC 25391) is a gram-negative facultative chemolithoautotroph capable of extracting energy from the oxidation of nitrite to nitrate. Sequencing and analysis of its genome revealed a single circular chromosome of 3,402,093 bp encoding 3,143 predicted proteins. There were extensive similarities to genes in two alphaproteobacteria, Bradyrhizobium japonicum USDA110 (1,300 genes) and Rhodopseudomonas palustris CGA009 CG (815 genes). Genes encoding pathways for known modes of chemolithotrophic and chemoorganotrophic growth were identified. Genes encoding multiple enzymes involved in anapleurotic reactions centered on C2 to C4 metabolism, including a glyoxylate bypass, were annotated. The inability of N. winogradskyi to grow on C6 molecules is consistent with the genome sequence, which lacks genes for complete Embden-Meyerhof and Entner-Doudoroff pathways, and active uptake of sugars. Two gene copies of the nitrite oxidoreductase, type I ribulose-1,5-bisphosphate carboxylase/oxygenase, cytochrome c oxidase, and gene homologs encoding an aerobic-type carbon monoxide dehydrogenase were present. Similarity of nitrite oxidoreductases to respiratory nitrate reductases was confirmed. Approximately 10% of the N. winogradskyi genome codes for genes involved in transport and secretion, including the presence of transporters for various organic-nitrogen molecules. The N. winogradskyi genome provides new insight into the phylogenetic identity and physiological capabilities of nitrite-oxidizing bacteria. The genome will serve as a model to study the cellular and molecular processes that control nitrite oxidation and its interaction with other nitrogen-cycling processes.

Influence of inorganic nitrogen management regime on the diversity of nitrite-oxidizing bacteria in agricultural grassland soils.

To assess links between the diversity of nitrite-oxidizing bacteria (NOB) in agricultural grassland soils and inorganic N fertilizer management, NOB communities in fertilized and unfertilized soils were characterized by analysis of clone libraries and denaturing gradient gel electrophoresis (DGGE) of 16S rRNA gene fragments. Previously uncharacterized Nitrospira-like sequences were isolated from both long-term-fertilized and unfertilized soils, but DGGE migration patterns indicated the presence of additional sequence types in the fertilized soils. Detailed phylogenetic analysis of Nitrospira-like sequences suggests the existence of one newly described evolutionary group and of subclusters within previously described sublineages, potentially representing different ecotypes; the new group may represent a lineage of noncharacterized Nitrospira species. Clone libraries of Nitrobacter-like sequences generated from soils under different long-term N management regimes were dominated by sequences with high similarity to the rhizoplane isolate Nitrobacter sp. strain PJN1. However, the diversity of Nitrobacter communities did not differ significantly between the two soil types. This is the first cultivation-independent study of nitrite-oxidizing bacteria in soil demonstrating that nitrogen management practices influence the diversity of this bacterial functional group.

Nitrification and nitrifying bacteria in the lower Seine River and estuary (France).

The Achères wastewater treatment plant, located just downstream of Paris, discharges its effluents into the lower Seine River. The effluents contain large numbers of heterotrophic bacteria, organic matter, and ammonium and are a source of nitrifying bacteria. As a result, degradation of organic matter by heterotrophic bacteria and subsequent oxygen depletion occur immediately downstream of the effluent outlet, whereas nitrifying bacteria apparently need to build up a significant biomass before ammonium oxidation significantly depletes the oxygen. We quantified the potential total nitrifying activity and the potential activities of the ammonia- and nitrite-oxidizing communities along the Seine River. In the summer, the maximum nitrifying activity occurs in the upper freshwater estuary, approximately 200 km downstream of Achères. The quantities of nitrifying bacteria, based on amoA gene copy numbers, and of Nitrobacter organisms, based on 16S rRNA gene copy numbers, were correlated with the potential nitrifying activities. The species composition of ammonia-oxidizing bacteria was investigated at two sites: the Triel station just downstream from Achères (km 84) and the Seine freshwater estuary at the Duclair station (km 278). By means of PCR primers targeting the amoA gene, a gene library was created. Phylogenetic analysis revealed that the majority of the analyzed clones at both sites were affiliated with the genus NITROSOMONAS: The Nitrosomonas oligotropha- and Nitrosomonas urea-related clones represented nearly 81% of the community of ammonia-oxidizing bacteria at Triel and 60% at Duclair. Two other ammonia-oxidizing clusters of the beta subclass of the Proteobacteria, i.e., Nitrosomonas europaea- and Nitrosospira-like bacteria, were found in smaller numbers. The major change in the ammonia-oxidizing community between the two stations along the Seine River-upper estuary continuum was the replacement of the N. oligotropha- and N. urea-related bacteria by the Nitrosospira-affiliated bacteria. Although the diversities of the ammonia oxidizers appear to be similar for the two sites, only half of the restriction patterns are common to both sites, which could be explained by the differences in ammonium concentrations, which are much lower in the upper estuary than in the river at the effluent outlet. These results imply a significant immigration and/or selection of the ammonia-oxidizing bacterial population along the continuum of the Seine River from Paris to the estuary.

In situ characterization of nitrifiers in an activated sludge plant: detection of Nitrobacter Spp.

AIMS: The purpose of this work was to investigate microbial ecology of nitrifiers at the genus level in a typical full-scale activated sludge plant. METHODS AND RESULTS: Grab samples of mixed liquor were collected from a plug-flow reactor receiving domestic wastewater. Fluorescent in situ hybridization technique (FISH) was used to characterize both ammonia oxidizing bacteria (AOB) and nitrite oxidizing bacteria (NOB) in combination with Confocal Scanning Laser Microscope (CSLM). Fluorescently labelled, 16S rRNA-targeted oligonucleotide probes were used in this study. Both Nitrosomonas and Nitrosospira genera as AOB and Nitrobacter and Nitrospira genera as NOB were sought with genus specific probes Nsm156, Nsv443 and NIT3 and NSR1156, respectively. CONCLUSIONS: It was shown that Nitrosospira genus was dominant in the activated sludge system studied, although Nitrosomonas is usually assumed to be the dominant genus. At the same time, Nitrobacter genus was detected in activated sludge samples. SIGNIFICANCE AND IMPACT OF THE STUDY: Previous studies based on laboratory scale pilot plants employing synthetic wastewater suggested that only Nitrospira are found in wastewater treatment plants. We have shown that Nitrobacter genus might also be present. We think that these kinds of studies may not give a valid indication of the microbial diversity of the real full-scale plants fed with domestic wastewater.

Fatty acid profiles of nitrite-oxidizing bacteria reflect their phylogenetic heterogeneity.

The fatty acid profiles of all described species of the nitrite-oxidizing genera Nitrobacter, Nitrococcus, Nitrospina and Nitrospira were analyzed. The four genera had distinct profiles, which can be used for the differentiation and allocation of new isolates to these genera. The genus Nitrobacter is characterized by vaccenic acid as the main compound with up to 92% of the fatty acids and the absence of hydroxy fatty acids. The genus Nitrococcus showed cis-9-hexadecenoic acid, hexadecanoic acid and vaccenic acid as main parts. Small amounts of 3-hydroxy-dodecanoic acid were detected. The genus Nitrospina possessed tetradecanoic acid and cis-9-hcxadecenoic acid as main compounds, also 3-hydroxy-hexadecanoic acid was detected for this genus. The genus Nitrospira showed a pattern with more variations among the two described species. These organisms are characterized by the cis-7 and cis-11-isomers of hexadecenoic acid. For Nitrospira moscoviensis a specific new fatty acid was found, which represented the major constituent in the fatty acid profiles of autotrophically grown cultures. It was identified as 11-methyl-hexadecanoic acid. Since this compound is not known for other bacterial taxa, it represents a potential lipid marker for the detection of Nitrospira moscoviensis relatives in enrichment cultures and environmental samples. A cluster analysis of the fatty acid profiles is in accordance with 16S rRNA sequence-based phylogeny of the nitrite-oxidizing bacteria.