Complete Genome Sequence of Paenibacillus mucilaginosus 3016, a Bacterium Functional as Microbial Fertilizer

Abstract

Paenibacillus mucilaginosus is a ubiquitous functional bacterium in microbial fertilizer. Here we report the complete sequence of P. mucilaginosus 3016. Multiple sets of functional genes have been found in the genome. To the best of our knowledge, this is the first announcement about the complete genome sequence of a P. mucilaginosus strain.

GENOME ANNOUNCEMENT

Paenibacillus mucilaginosus (formerly Bacillus mucilaginosus) (6), the type species of Paenibacillus, is consequently widely used as a microbial fertilizer in agricultural applications due to its growth-promoting properties (7, 12). P. mucilaginosus 3016 was isolated from rhizosphere soil and has been widely used in microbial fertilizer by most manufacturers in China (20).

The P. mucilaginosus genome was determined by Roche 454 pyrosequencing (15), and Illumina sequencing by synthesis. The Illumina adapters were ligated onto fragmented P. mucilaginosus genomic DNA, and then gel electrophoresis was used to select DNA fragments about 3 kbp in size. Libraries were PCR amplified using Phusion polymerase. Sequencing libraries were denatured with sodium hydroxide and diluted in hybridization buffer for loading onto a single lane of an Illumina genome analyzer (GA) flow cell. Cluster formation, primer hybridization, and sequencing were performed using proprietary reagents according to manufacturer-recommended protocols. The paired-end DNA library (8-kbp span) for the 454 platform was prepared according to the manual from Roche. All the low-quality sequences were trimmed before assembly. The mate-paired reads (718 Mbp; 82.53× coverage) were generated by a Solexa sequencer and assembled by the SOAPdenovo method (11). Then, the 454 reads (190 Mbp; 21.84× coverage) and the split fragments of contigs generated by SOAPdenovo were used for a hybrid assembly with the Newbler sequence assembler (version 2.6). To finish the genome, conventional Sanger sequencing technologies were used to fill the gaps. Coding sequences were predicted by Glimmer3 (4). Functional assignment and classification were determined by performing sequence similarity searches with BLAST (1) (E-value cutoff, 1e−5) against the eggNOG database (16), the KEGG reference database (8), and the nonredundant GenBank CDS database. The tRNA and the rRNA genes were predicted by tRNASCAN-SE (13) and RNAmmer (9), respectively. A set of in-house Perl scripts and EMBOSS (17) were used for sequence manipulation.

The complete genome sequence of strain 3016 is a circular 8,739,048-bp chromosome with mean GC contents of 58.3%. There are 7,285 coding genes, 42 rRNA operons, and 170 tRNAs in the chromosome. Many essential genes were detected in the chromosome, such as the genes involving the metabolism of nitrogen, phosphorus, and potassium, as well as nitrogen-fixing NifU domain-containing protein, potassium channel protein, and potassium-transporting ATPase subunit A, indicating its importance to the strain's life.

In comparison to Paenibacillus polymyxa SC2, a strain of plant growth-promoting rhizobacteria in the same genus, there are 1,641 shared genes and 5,416 unique genes in the chromosome of strain 3016. The unique genes, such as the ywlF gene (NCBI M3016-39), the ppdK gene (M3016-1649), the ytiB gene (M3016-3117), the yfkO gene (M3016-4045), the yvdA gene (M3016-4842), and the yurN gene (M3016-5050), are most involved in carbohydrate transport and metabolism (10.73% of the unique genes), transcription (9.31% of the unique genes), signal transduction mechanisms (6.96% of the unique genes), and amino acid transport and metabolism (5.89% of the unique genes), based on their similarities to previously published genes (2, 3, 5, 10, 14, 18, 19, 21).

Nucleotide sequence accession number.

The Paenibacillus mucilaginosus 3016 chromosome sequence has been deposited in GenBank under accession number CP003235.