Complete Genome Sequence of Bacillus thuringiensis subsp. jinghongiensis Reference Strain YGd22-03

ABSTRACT

Bacillus thuringiensis is widely used in producing ecofriendly microbial agents for the purpose of controlling insect pests. In this study, we determined the complete genome sequence of B. thuringiensis subsp. jinghongiensis reference strain YGd22-03, which contains three cry genes and one cerecidin biosynthetic gene cluster.

GENOME ANNOUNCEMENT

Bacillus thuringiensis is an ecofriendly biopesticide for controlling a wide variety of insect pests of several orders, such as Lepidoptera, Diptera, Coleoptera, Hemiptera, Hymenoptera, and Mallophaga (1). Until now, B. thuringiensis strains have been classified by their flagellar immunological reactions into at least 71 H serotypes containing 84 subspecies (2, 3). YGd22-03 is a novel B. thuringiensis strain isolated from China and was identified as the reference strain of B. thuringiensis subsp. jinghongiensis (serotype H42) (4). The complete genome sequence of YGd22-03 was determined in this study.

Genomic DNA of YGd22-03 was extracted and a 500-bp paired-end insert library was sequenced on an Illumina HiSeq 2500 platform with read lengths of 150 bp. The reads were quality filtered by Quake (5), resulting in a 2.169-Gbp total length that comprised 17,358,564 high-quality reads and represented 400-fold coverage of the genome. The clean reads were then de novo assembled with a SPAdes 3.5.0 assembler (6), and 79 contigs longer than 500 bp were generated. The contigs were assembled by comparing them to multiple complete genome sequences of B. thuringiensis to obtain a draft genome with gaps. The gaps were further filled by high-fidelity PCR and Sanger sequencing to construct the complete genome. The genome was annotated using the NCBI Prokaryotic Genome Annotation Pipeline (https://www.ncbi.nlm.nih.gov/genome/annotation_prok/) with the best-placed reference protein set and GeneMarkS+ methods.

The complete genome of YGd22-03 contains one circular chromosome and five circular plasmids. The length of the chromosome is 5,420,545 bp, and it harbors 5,721 genes, including 36 rRNA genes and 99 tRNA genes. The G+C content of the chromosome is 35.2%. The five circular plasmids containing 315 protein-coding genes are named pYGD98 (98,137 bp), pYGD83 (83,894 bp), pYGD36 (36,974 bp), pYGD30 (30,343 bp), and pYGD5 (5,650 bp), and the G+C contents of the plasmids range from 32.0% to 34.5%.

Three parasporal crystal protein-coding genes (BVH75-30100, BVH75-30365, and BVH75-30370) and one vegetative insecticidal protein (VIP)-like protein-coding gene (BVH75-29980) are found in plasmid pYGD98. The gene products of BVH75-30365 and BVH75-30370 showed 100% amino acid sequence similarity to the proteins PS5-1 (Cry64Ba1) and PS5-2 (Cry64Ca1), respectively. The gene product of gene BVH75-30100 shows 99% similarity to Cry11-like protein (GenBank accession number AKS28920). According to a previous report, the gene products from one or all three of these cry genes might exhibit cytocidal activity against cancer cells (7). In addition, a cerecidin biosynthetic gene cluster, whose product was reported to exhibit bactericidal activity against a broad spectrum of Gram-positive pathogens (8), was found in the chromosome by analysis of the genome with antiSMASH (9). Therefore, YGd22-03 might be a candidate for potential medical usage for harboring anticancer proteins and producing the antimicrobial substance cerecidin.

Accession number(s).

The genome sequence of the B. thuringiensis strain YGd22-03 has been deposited in GenBank under the accession numbers CP019230 to CP019235. The strain is available in the Bacillus Genetic Stock Center (http://www.bgsc.org/) under the identification number BGSCID 4AR1.