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. 2016 Mar 31:6:23747.
doi: 10.1038/srep23747.

A novel ammonia-oxidizing archaeon from wastewater treatment plant: Its enrichment, physiological and genomic characteristics

Yuyang Li  1 , Kun Ding  1 , Xianghua Wen  1 , Bing Zhang  1 , Bo Shen  1 , Yunfeng Yang  1
Affiliations

A novel ammonia-oxidizing archaeon from wastewater treatment plant: Its enrichment, physiological and genomic characteristics

Yuyang Li et al. Sci Rep. .

Abstract

Ammonia-oxidizing archaea (AOA) are recently found to participate in the ammonia removal processes in wastewater treatment plants (WWTPs), similar to their bacterial counterparts. However, due to lack of cultivated AOA strains from WWTPs, their functions and contributions in these systems remain unclear. Here we report a novel AOA strain SAT1 enriched from activated sludge, with its physiological and genomic characteristics investigated. The maximal 16S rRNA gene similarity between SAT1 and other reported AOA strain is 96% (with "Ca. Nitrosotenuis chungbukensis"), and it is affiliated with Wastewater Cluster B (WWC-B) based on amoA gene phylogeny, a cluster within group I.1a and specific for activated sludge. Our strain is autotrophic, mesophilic (25 °C-33 °C) and neutrophilic (pH 5.0-7.0). Its genome size is 1.62 Mb, with a large fragment inversion (accounted for 68% genomic size) inside. The strain could not utilize urea due to truncation of the urea transporter gene. The lack of the pathways to synthesize usual compatible solutes makes it intolerant to high salinity (>0.03%), but could adapt to low salinity (0.005%) environments. This adaptation, together with possibly enhanced cell-biofilm attachment ability, makes it suitable for WWTPs environment. We propose the name "Candidatus Nitrosotenuis cloacae" for the strain SAT1.

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Figures

Figure 1
Figure 1. Phylogenetic tree showing the relationships of amoA gene sequence of strain SAT1 to reference sequences from the GenBank database.
The tree was constructed with the neighbor-joining method. Bootstrap values shown at nodes where the value was greater than 50, are based on 1000 trials. For sequences inside wastewater cluster B, those from wastewater treatment plant were marked with circles (●), those from freshwater rivers/lakes were marked triangles (▲), and those from drinking water treatment plant were marked block (■).
Figure 2
Figure 2
Photomicrographs of the SAT1 enrichment culture using SEM (a) and TEM (b).
Figure 3
Figure 3. Autotrophic growth and ammonia oxidation by strain SAT1.
(a) Cell growth is represented by archaeal amoA and 16S rRNA gene abundance. Error bars represent the standard deviations from triplicate experiments. (b) Distribution of the relative abundance of archaeal amoA gene in CsCl gradient for 13C-NaHCO3, or 12C-NaHCO3 treatment. The number in parentheses (1 or 2) means the cycles of incubation. Vertical and horizontal error bars represent standard deviations of the relative abundance and buoyant density of fractions from duplicate cultivation samples respectively.
Figure 4
Figure 4
Influence of (a) salinity, (b) ammonia, (c) nitrite, (d) temperature, (e) pH and (f) Allylthiourea on the growth activity of strain SAT1. Values represent percentage of specific growth rates of cultures grown under different environmental values relative to those at optimal ones. Error bars represent the standard deviations of triplicate cultures.
Figure 5
Figure 5
(a) Organization of urease utilizing genes of SAT1 genome and (b) the experimental test of its urea utilizing ability. The organization of genes was in comparison with “Ca. Nitrosopumilus sediminis” AR2 (above) and “Ca. Nitrososphaera evergladensis” (below). Urease core protein genes were shown in red, urease accessory protein genes were shown in blue, urea transporter (UT) genes were shown in green, and histidine kinase (HK) genes were shown in orange. For comparison, forward matches colored in red and reverse matches colored in green. The numbers between two genes indicated the number of interval ORFs. For the test of the urea utilizing ability of SAT1, the nitrite production and archaeal 16S rRNA abundance during one cycle of incubation, using 0.5 mM NH4+ (purple line and blue column) and 0.25 mM Urea (light green line and red column) as substrate respectively.
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