Complete Genome Sequence of the Nonylphenol-Degrading Bacterium Sphingobium cloacae JCM 10874^T

GENOME ANNOUNCEMENT

Environmentally persistent alkylphenols, such as 4-nonylphenol (NP), are known to be estrogenic in animals (1). The chemicals are breakdown products of microbial degradation of man-made nonionic surfactants, namely, nonylphenol polyethoxylates (2). Tanghe et al. published the first report regarding the isolation of a bacterium able to degrade NP in 1999 (3). Following that study, two NP-degrading bacteria were isolated from sewage treatment plant wastewater and river sediment, which were later described as new species, Sphingomonas (Sphingobium) cloacae and Sphingobium amiense (4, 5). Further analysis of biodegradation activity revealed that S. cloacae S-3^T (JCM 10874^T) displays a catabolic activity specific to branched alkylphenols, while S. amiense YT^T is a versatile organism capable of utilizing a wide range of phenolic compounds (6). Here, we investigated the genome of S. cloacae JCM 10874^T to obtain important information regarding the catabolic genes of NP-degrading Sphingomonas (Sphingobium) species.

Genomic DNA was isolated from S. cloacae JCM 10874^T and sequenced using PacBio RSII instruments. Reads were assembled using FALCON version 0.2.1 to produce seven contigs. The genome sequence was annotated using the Microbial Genome Annotation Pipeline (http://www.migap.org) and Rapid Annotations using Subsystems Technology version 2.0 (7), followed by manual annotation with the NCBI-nr databases using the BLASTP program (8). tRNAs were predicted using tRNAscan-SE (9).

The genome of S. cloacae JCM 10874^T consists of 3,767,292 bp in one circular chromosome (SCLO1), with a coverage of 322-fold and 64.6% G+C content, 3,293 coding sequences (CDSs), six rRNA operons, and 49 tRNA genes. There are also five circular plasmids. The plasmids are pSCLO2 (sequence coverage, 301-fold; total nucleotide sequences, 375,493 bp; G+C content, 64.9%; total number of CDSs, 334), pSCLO3 (sequence coverage, 340-fold; total nucleotide sequences, 151,712 bp; G+C content, 62.8%; total number of CDSs, 137), pSCLO4 (sequence coverage, 225-fold; total nucleotide sequences, 108,910 bp; G+C content, 63.7%; total number of CDSs, 92), pSCLO5 (sequence coverage, 85-fold; total nucleotide sequences, 57,701 bp; G+C content, 63.5%; total number of CDSs, 53), pSCLO7 (sequence coverage, 32-fold; total nucleotide sequences, 33,768 bp; G+C content, 62.9%; total number of CDSs, 33), and linear pSCLO6 (sequence coverage, 99-fold; total nucleotide sequences, 52,690 bp; G+C content, 62.4%; total number of CDSs, 51).

Two genes responsible for alkylphenol oxygenation at the initial step of degradation were detected: one copy of the octylphenol 4-monooxygenase gene (opdA) was present in pSCLO3, and one copy of the nonylphenol monooxygenase gene (nmoA) was present in pSCLO4. opdA shows an amino acid sequence identity of 99.1% with OpdA of Sphingomonas sp. TTNP3 (3, 10, 11), and nmoA shows 100.0% identity with NmoA of Sphingomonas sp. NP5 (12). Monooxygenase and dioxygenase genes related to cyclic hydrocarbon (cyclopentanone and hydroxyquinol) degradation were also found in the circular plasmids. A gene cluster (7,939 bp) that encodes a hydroquinone catabolic pathway, hqdRAB-orf1-orf2-hqdCDEF, was found in the linear plasmid. Amino acid sequence identities were 99.4% (HqdA) and 100% (HqdR, HqdB, HqdC, HqdD, HqdE, and HqdF) between S. cloacae JCM 10874^T and the well-characterized Sphingomonas sp. strain TTNP3 (3, 13, 14). These results suggest that the degradation of alkylphenols is conducted by the concerted action of different plasmid systems in the bacterium.