Molecular analysis of enrichment cultures of ammonia oxidizers from the Salar de Huasco, a high altitude saline wetland in northern Chile

We analyzed enrichment cultures of ammonia-oxidizing bacteria (AOB) collected from different areas of Salar de Huasco, a high altitude, saline, pH-neutral water body in the Chilean Altiplano. Samples were inoculated into mineral media with 10 mM NH4+ at five different salt concentrations (10, 200, 400, 800 and 1,400 mM NaCl). Low diversity (up to three phylotypes per enrichment) of beta-AOB was detected using 16S rDNA and amoA clone libraries. Growth of beta-AOB was only recorded in a few enrichment cultures and varied according to site or media salinity. In total, five 16S rDNA and amoA phylotypes were found which were related to Nitrosomonas europaea/Nitrosococcus mobilis, N. marina and N. communis clusters. Phylotype 1-16S was 97% similar with N. halophila, previously isolated from Mongolian soda lakes, and phylotypes from amoA sequences were similar with yet uncultured beta-AOB from different biofilms. Sequences related to N. halophila were frequently found at all salinities. Neither gamma-AOB nor ammonia-oxidizing Archaea were recorded in these enrichment cultures

Ammonia-oxidizing archaea and ammonia-oxidizing bacteria in six full-scale wastewater treatment bioreactors

In this study, dideoxy sequencing and 454 high-throughput sequencing were used to analyze diversities of the ammonia monooxygenase (amoA) genes and the 16S rRNA genes of ammonia-oxidizing archaea (AOA) and ammonia-oxidizing bacteria (AOB) in six municipal wastewater treatment plants. The results showed that AOB amoA genes were quite diverse in different wastewater treatment plants while the 16S rRNA genes were relatively conserved. Based on the observed complexity of amoA and 16S rRNA genes, most of the AOB can be assigned to the Nitrosomonas genus, with Nitrosomonas ureae, Nitrosomonas oligotropha, Nitrosomonas marina, and Nitrosomonas aestuarii being the four most dominant species. From the sequences of the AOA amoA genes, most AOA observed in this study belong to the CGI.1b group, i.e., the soil lineage. The AOB amoA and 16S rRNA genes were quantified by quantitative PCR and 454 high-throughput pyrosequencing, respectively. Although the results from the two approaches show some disconcordance, they both indicated that the abundance of AOB in activated sludge was very low

Diversity and quantity of ammonia-oxidizing Archaea and Bacteria in sediment of the Pearl River Estuary, China

The diversity and abundance of ammonia-oxidizing archaea (AOA) and ammonia-oxidizing bacteria (AOB) in the sediment of the Pearl River Estuary were investigated by cloning and quantitative real-time polymerase chain reaction (qPCR). From one sediment sample S16, 36 AOA OTUs (3% cutoff) were obtained from three clone libraries constructed using three primer sets for amoA gene. Among the 36 OTUs, six were shared by all three clone libraries, two appeared in two clone libraries, and the other 28 were only recovered in one of the libraries. For AOB, only seven OTUs (based on 16S rRNA gene) and eight OTUs (based on amoA gene) were obtained, showing lower diversity than AOA. The qPCR results revealed that AOA amoA gene copy numbers ranged from 9.6 × 106 to 5.1 × 107 copies per gram of sediment and AOB amoA gene ranged from 9.5 × 104 to 6.2 × 105 copies per gram of sediment, indicating that the dominant ammonia-oxidizing microorganisms in the sediment of the Pearl River Estuary were AOA. The terminal restriction fragment length polymorphism results showed that the relative abundance of AOB species in the sediment samples of different salinity were significantly different, indicating that salinity might be a key factor shaping the AOB community composition

Community shift of ammonia-oxidizing bacteria along an anthropogenic pollution gradient from the Pearl River Delta to the South China Sea

The phylogenetic diversity and abundance of ammonia-oxidizing beta-proteobacteria (beta-AOB) was analyzed along an anthropogenic pollution gradient from the coastal Pearl River Delta to the South China Sea using the ammonia monooxygenase subunit A (amoA) gene. Along the gradient from coastal to the open ocean, the phylogenetic diversity of the dominant genus changed from Nitrosomonas to Nitrosospira, indicating the niche specificity by these two genera as both salinity and anthropogenic influence were major factors involved. The diversity of bacterial amoA gene was also variable along the gradient, with the highest in the deep-sea sediments, followed by the marshes sediments and the lowest in the coastal areas. Within the Nitrosomonas-related clade, four distinct lineages were identified including a putative new one (A5-16) from the different sites over the large geographical area. In the Nitrosospira-related clade, the habitat-specific lineages to the deep-sea and coastal sediments were identified. This study also provides strong support that Nitrosomonas genus, especially Nitrosomonas oligotropha lineage (6a) could be a potential bio-indicator species for pollution or freshwater/wastewater input into coastal environments. A suite of statistical analyses used showed that water depth and temperature were major factors shaping the community structure of beta-AOB in this study area

Characterization and quantification of ammonia-oxidizing archaea (AOA) and bacteria (AOB) in a nitrogen-removing reactor using T-RFLP and qPCR

Using ammonia monooxygenase α-subunit (amoA) gene and 16S rRNA gene, the community structure and abundance of ammonia-oxidizing archaea (AOA) and ammonia-oxidizing bacteria (AOB) in a nitrogen-removing reactor, which was operated for five phases, were characterized and quantified by cloning, terminal restriction fragment length polymorphism (T-RFLP), and quantitative polymerase chain reaction (qPCR). The results suggested that the dominant AOB in the reactor fell to the genus Nitrosomonas, while the dominant AOA belonged to Crenarchaeotal Group I.1a in phylum Crenarchaeota. Real-time PCR results demonstrated that the levels of AOB amoA varied from 2.9 × 103 to 2.3 × 105 copies per nanogram DNA, greatly (about 60 times) higher than those of AOA, which ranged from 1.7 × 102 to 3.8 × 103 copies per nanogram DNA. This indicated the possible leading role of AOB in the nitrification process in this study. T-RFLP results showed that the AOB community structure significantly shifted in different phases while AOA only showed one major peak for all the phases. The analyses also suggested that the AOB community was more sensitive than that of AOA to operational conditions, such as ammonia loading and dissolved oxygen

Spatial distribution and abundances of ammonia-oxidizing archaea (AOA) and ammonia-oxidizing bacteria (AOB) in mangrove sediments

We investigated the diversity, spatial distribution, and abundances of ammonia-oxidizing archaea (AOA) and ammonia-oxidizing bacteria (AOB) in sediment samples of different depths collected from a transect with different distances to mangrove forest in the territories of Hong Kong. Both the archaeal and bacterial amoA genes (encoding ammonia monooxygenase subunit A) from all samples supported distinct phylogenetic groups, indicating the presences of niche-specific AOA and AOB in mangrove sediments. The higher AOB abundances than AOA in mangrove sediments, especially in the vicinity of the mangrove trees, might indicate the more important role of AOB on nitrification. The spatial distribution showed that AOA had higher diversity and abundance in the surface layer sediments near the mangrove trees (0 and 10 m) but lower away from the mangrove trees (1,000 m), and communities of AOA could be clustered into surface and bottom sediment layer groups. In contrast, AOB showed a reverse distributed pattern, and its communities were grouped by the distances between sites and mangrove trees, indicating mangrove trees might have different influences on AOA and AOB community structures. Furthermore, the strong correlations among archaeal and bacterial amoA gene abundances and their ratio with NH4+, salinity, and pH of sediments indicated that these environmental factors have strong influences on AOA and AOB distributions in mangrove sediments. In addition, AOA diversity and abundances were significantly correlated with hzo gene abundances, which encodes the key enzyme for transformation of hydrazine into N2 in anaerobic ammonium-oxidizing (anammox) bacteria, indicating AOA and anammox bacteria may interact with each other or they are influenced by the same controlling factors, such as NH4+. The results provide a better understanding on using mangrove wetlands as biological treatment systems for removal of nutrients

Phylogenetic and functional marker genes to study ammonia-oxidizing microorganisms (AOM) in the environment

The oxidation of ammonia plays a significant role in the transformation of fixed nitrogen in the global nitrogen cycle. Autotrophic ammonia oxidation is known in three groups of microorganisms. Aerobic ammonia-oxidizing bacteria and archaea convert ammonia into nitrite during nitrification. Anaerobic ammonia-oxidizing bacteria (anammox) oxidize ammonia using nitrite as electron acceptor and producing atmospheric dinitrogen. The isolation and cultivation of all three groups in the laboratory are quite problematic due to their slow growth rates, poor growth yields, unpredictable lag phases, and sensitivity to certain organic compounds. Culture-independent approaches have contributed importantly to our understanding of the diversity and distribution of these microorganisms in the environment. In this review, we present an overview of approaches that have been used for the molecular study of ammonia oxidizers and discuss their application in different environments

Enrichment and characterization of ammonia-oxidizing archaea from the open ocean : phylogeny, physiology and stable isotope fractionation

Author Posting. © The Author(s), 2011. This is the author's version of the work. It is posted here by permission of Nature Publishing Group for personal use, not for redistribution. The definitive version was published in The ISME Journal 5 (2011): 1796–1808, doi:10.1038/ismej.2011.58.Archaeal genes for ammonia oxidation are widespread in the marine environment, but direct physiological evidence for ammonia oxidation by marine archaea is limited. We report the enrichment and characterization of three strains of pelagic ammonia-oxidizing archaea (AOA) from the north Pacific Ocean that have been maintained in laboratory culture for over three years. Phylogenetic analyses indicate the three strains belong to a previously identified clade of water column-associated AOA and possess 16S rRNA genes and ammonia monooxygenase subunit a (amoA) genes highly similar (98-99% identity) to those recovered in DNA and cDNA clone libraries from the open ocean. The strains grow in natural seawater-based liquid medium while stoichiometrically converting ammonium (NH4 +) to nitrite (NO2 -). Ammonia oxidation by the enrichments is only partially inhibited by allylthiourea at concentrations known to inhibit cultivated ammonia-oxidizing bacteria. The three strains were used to determine the nitrogen stable isotope effect (15εNH3) during archaeal ammonia oxidation, an important parameter for interpreting stable isotope ratios in the environment. Archaeal 15εNH3 ranged from 13- 41‰, within the range of that previously reported for ammonia-oxidizing bacteria. Despite low amino acid identity between the archaeal and bacterial Amo proteins, their functional diversity as captured by 15εNH3 is similar.This work was supported by a Woods Hole Oceanographic Institution (WHOI) Postdoctoral Scholar fellowship to AES and the WHOI Ocean Life Institute

Erratum to “Nitrifying and heterotrophic population dynamics in biofilm reactors: effects of hydraulic retention time and the presence of organic carbon”

Erratum to ‘‘Nitrifying and heterotrophic population dynamicsin biofilm reactors: effects of hydraulic retention time and thepresence of organic carbon’’[Water Research 36 (2002) 469–481]. Available at: http://hdl.handle.net/1822/1602When this paper was first published the affiliations were incorrect.(undefined