ABSTRACT
This report describes the whole-genome sequence of a microcystin-degrading bacterium, Novosphingobium sp. strain MD-1, isolated from a lake in Japan. The Novosphingobium sp. strain MD-1 genome had a total length of 4,617,766 bp. Moreover, strain MD-1 showed a conserved microcystin-degrading gene cluster (mlrA to mlrF), similar to Sphingopyxis sp. strain C-1.
ANNOUNCEMENT
Harmful algal blooming cyanobacteria such as Microcystis spp. produce microcystin, which is a potent hepatotoxin. Microcystin is very stable (1, 2) but can be degraded by specific enzymes (MlrA, MlrB, MlrC, and MlrD) in microcystin-degrading bacteria (3–6). Lakes and reservoirs that have harmful algal blooms generally have highly alkaline pH (7), and alkaline conditions provide the optimal pH for most microcystin-degrading bacteria (8). However, the microcystin-degrading bacterium Novosphingobium sp. strain MD-1 (formerly classified as Sphingomonas sp. strain MD-1) prefers a neutral pH (9, 10). Therefore, the whole gene structure may differ greatly from that of other microcystin-degrading bacteria. Here, we report the whole-genome sequence of Novosphingobium sp. strain MD-1.
Novosphingobium sp. strain MD-1 was isolated from Lake Kasumigaura in 1999 (9). Novosphingobium sp. strain MD-1 has been maintained in a glycerol stock at −80°C. Culturing was conducted using 1:5 peptone-yeast extract (PY) at 28°C, and then total DNA was extracted in the late logarithmic growth phase. Genomic DNA was prepared as a paired-end library (350 bp) with the TruSeq Nano DNA low-throughput (LT) sample preparation kit (Illumina, San Diego, CA, USA) and as a mated-pair library (8 to 10 kb) with a Nextera mate pair sample preparation kit (Illumina). Whole-genome sequencing was carried out using an Illumina HiSeq 2500 system with a paired-end library and a mated-pair library. A total of 22,852,894 reads, averaging 99 bp long, were obtained, for a total of 2,262,436,506 bases of the sequence. The final coverage (depth) of the filtered genome sequence was 146×. All reads were assembled de novo using Velvet v1.2.08 (11). Gaps between the resultant 292 contigs were closed using Platanus v1.2.1 (12). The whole genome was annotated using the RAST server (http://rast.nmpdr.org/rast.cgi), which predicted protein-coding sequences (CDSs). Default parameters were used for all software.
The Novosphingobium sp. strain MD-1 genome consisted of 34 contigs, with an N50 value of 406,083 bp, and had a total length of 4,617,766 bp. The draft genome had a G+C content of 65.80%, with 4,178 CDSs and 60 RNA-coding genes (i.e., 3 sets of rRNA genes and 51 tRNA genes). The annotation revealed that 3,060 CDSs exhibited homology to genes with known functions; the remaining 1,118 CDSs encoded hypothetical proteins with unknown functions.
Okano et al. reported that microcystin is degraded by six enzymes in Sphingopyxis sp. strain C-1, namely, MlrA, MlrB, MlrC, MlrD, MlrE, and MlrF (13). The mlrF, mlrE, mlrB, mlrD, mlrA, and mlrC genes of Novosphingobium sp. strain MD-1 were designated with locus tags NMD1_03009 to NMD1_03013, respectively (positions 426855 to 435216), in the genome (GenBank accession number BBXA01000030). MlrA, MlrB, MlrC, MlrD, MlrE, and MlrF had 93.3, 95.7, 93.7, 95.3, 96.8, and 90.7% identities, respectively, to those of Sphingopyxis sp. strain C-1 on an amino acid basis, using the Align Sequences Protein BLAST. Therefore, it was suggested that the six genes participated in microcystin degradation independent of the suitable pH for growth.
Data availability.This whole-genome shotgun project was deposited in DDBJ/EMBL/GenBank under accession number BBXA00000000. Raw HiSeq data were deposited in the DDBJ Sequence Read Archive under accession number DRA009321.
ACKNOWLEDGMENTS
This work was supported by the Kakenhi Challenge Project of Akita Prefectural University and a Grant-in-Aid for Scientific Research (B) from JSPS KAKENHI (grant 18H03396).
FOOTNOTES
- Received 5 December 2019.
- Accepted 28 February 2020.
- Published 19 March 2020.
- Copyright © 2020 Okano et al.
This is an open-access article distributed under the terms of the Creative Commons Attribution 4.0 International license.
