Complete genome sequence of the ammonia-oxidizing bacterium and obligate chemolithoautotroph Nitrosomonas europaea

Abstract

Nitrosomonas europaea (ATCC 19718) is a gram-negative obligate chemolithoautotroph that can derive all its energy and reductant for growth from the oxidation of ammonia to nitrite. Nitrosomonas europaea participates in the biogeochemical N cycle in the process of nitrification. Its genome consists of a single circular chromosome of 2,812,094 bp. The GC skew analysis indicates that the genome is divided into two unequal replichores. Genes are distributed evenly around the genome, with approximately 47% transcribed from one strand and approximately 53% transcribed from the complementary strand. A total of 2,460 protein-encoding genes emerged from the modeling effort, averaging 1,011 bp in length, with intergenic regions averaging 117 bp. Genes necessary for the catabolism of ammonia, energy and reductant generation, biosynthesis, and CO(2) and NH(3) assimilation were identified. In contrast, genes for catabolism of organic compounds are limited. Genes encoding transporters for inorganic ions were plentiful, whereas genes encoding transporters for organic molecules were scant. Complex repetitive elements constitute ca. 5% of the genome. Among these are 85 predicted insertion sequence elements in eight different families. The strategy of N. europaea to accumulate Fe from the environment involves several classes of Fe receptors with more than 20 genes devoted to these receptors. However, genes for the synthesis of only one siderophore, citrate, were identified in the genome. This genome has provided new insights into the growth and metabolism of ammonia-oxidizing bacteria.

FIG. 1.

Circular representation of the 2,812,094-bp genome of N. europaea ATCC 19718. The outer two circles represent protein-encoding and structural-RNA genes, plus and minus strand (green, energy metabolism; red, DNA replication; magenta, transcription; yellow, translation; orange, amino acid metabolism; dark blue, carbohydrate metabolism; pale red, nucleotide metabolism; black, coenzyme metabolism; cyan, lipid metabolism; light blue, cellular processes; brown, general function; gray, hypothetical and conserved hypothetical genes; pale green, structural RNAs). The third circle indicates the major IS element families as follows: black, ISne1 family; orange, ISne2; red, ISne3; green, ISne4; blue, ISne5; cyan, ISne6; magenta, ISne7; yellow, ISne8. The fourth circle indicates siderophore-receptor genes (blue) and pseudogenes (red). The fifth circle indicates amo (red), hao (green), the duplicated genes for EF-Tu (blue), the 7.5-kb tandem duplication (magenta) and the 339- and 318-bp repeats (black). RNA structural genes are indicated in the sixth circle. The inner two circles are the GC bias and GC skew.

FIG. 2.

Arrangement of hao and amo in N. europaea and Nitrosomonas sp. strain ENI-11. Arrows indicate the orientation of the genes. The nomenclature of the genes in N. europaea is with respect to their KpnI fragment sizes for hao (38) and EcoRI fragment sizes for amo (37). The nomenclature for Nitrosomonas sp. strain ENI-11 follows that described by Hirota et al. (36). Distances between regions are indicated in kilobases and are not drawn to proportion. The gray areas indicate the regions similarly arranged.

FIG. 3.

Diagram of N. europaea cell. Processes primarily associated with the generation of a proton gradient are indicated on the bottom, and processes associated with utilization of the gradient are on the other sides. ATP and NADH production are indicated on the left. Transporters are indicated on the top and right. Not all transporters are indicated; the numbers in parentheses indicate the approximate numbers of each type. Major metabolic pathways and biosynthetic pathways are indicated in the center of the cell. The roman numbers refer to the enzyme complex I (NADH-ubiquinone reductase), complex III (ubiquinol-cytochrome c reductase), and complex IV (cytochrome c oxidase) in the respiratory chain.

FIG.4.

N. europaea gene clusters. Several gene clusters described in the text are diagrammed. Each arrow represents a gene in the cluster. The N. europaea gene numbers are above each arrow and the gene names (or other identifiers) are below each arrow. In panel E, gene clusters from R. metallidurans CH34 (Rm CH34) and P. fluorescens 13525 (Pf 13525) are included for comparison.

FIG. 5.

Phylogeny of FecI and FecR homologous proteins and associated iron siderophore receptors. Distance-neighbor-joining trees were derived from CLUSTALW alignments of FecI- and FecR-homologous protein sequences that were deduced from identified ORFs in the genome of N. europaea. Sequences of identified functional FecI (FiuI) and FecR (FiuR) proteins from E. coli K-12 (U14003) and P. aeruginosa PAO1 (AF051691) were obtained from GenBank and used as a reference for the alignments. Designations of N. europaea proteins with high similarity to the FecIR proteins from E. coli and P. aeruginosa are given in boldface. Designations of FecI proteins without a respective FecR are given in italics. Putative iron siderophore sensing receptors that preceded or succeed a fecIR gene tandem in the genome are indicated with superscript letters as follows: a, ferrichrome-iron receptor 3 PCC6803 (D90899); b, ferrichrome-iron receptor B. bronchioseptica (U77671); c, hydroxamate ferrisiderophore receptor PAO1 (AF051691); d, unidentified ferric siderophore receptor PAO1 (U33150); e, ferripyoverdine receptor PAO1 (L10210); and f, TonB-dependent outer membrane receptor PAO1 (AE004674). Letters in parentheses indicate that the ORF is probably truncated. An additional gene encoding a ferrienterobactin receptor similar to AF083948 was found not associated with FecIR (data not indicated).