Draft Genome Sequences of Nine Bacillus Strains and Two Weizmannia Strains Isolated from Live Dietary Supplements and a Cultured Food Product.

We present the genome sequences of nine Bacillus isolates and two Weizmannia isolates from 10 different dietary supplements and one cultured food product. Strains of these species have been associated with health benefits when ingested by humans, due to their ability to survive the stomach's acidic environment and colonize the intestinal tract.

ANNOUNCEMENT

Bacillus species are Gram-positive, rod-shaped bacteria that thrive either aerobically or anaerobically. Many Bacillus species have been used in fermented food production, such as Bacillus subtilis in fermented soybean products and kimchi (1, 2). Weizmannia coagulans (previously known as Bacillus coagulans) and B. subtilis are common ingredients in commercially available dietary supplements due to their beneficial properties (3).

Concerns over Bacillus species’ ability to produce toxins and potentially transfer antimicrobial resistance genes create the need to explore strain variances (4, 5). With the increased interest in the use of species belonging to these two genera as beneficial ingredients in foods and dietary supplements, there is a need for high-quality Bacillus and Weizmannia genomic sequences to use as tools for safety assessments and strain comparisons of these species. Here, we present the draft genome sequences of nine Bacillus strains and two Weizmannia strains isolated directly from 10 different dietary supplements and one cultured food product for classification.

A serving from each one of the products was suspended in 5 mL of saline. One hundred microliters from serial dilutions was plated on MRS agar plates and incubated under different temperatures and atmospheric conditions. Pure colony isolates were grown for 48 h in MRS broth at 37°C. DNA extractions were performed with the Qiagen QIAcube using the DNeasy blood and tissue/Gram-positive bacteria or yeast protocol (Qiagen, Hilden, Germany). A DNA Clean and Concentrator kit (Zymo Research, Irvine, CA) was used to increase the quality of the DNA by removing potential inhibitors that could affect the sequencing process, as well as concentrating the sample from 150 μL to a final volume of 6 μL. DNA quality and concentration were determined using a Qubit 3.0 fluorometer (Life Technologies, Burlington, CA). Sequencing libraries were prepared with 1 ng of DNA using the Nextera XT DNA library preparation kit and the Nextera XT index kit v2 set A (Illumina, San Diego, CA) and sequenced on either the Illumina MiSeq system using the MiSeq reagent kit v2 (500 cycle) or the NextSeq platform using the NextSeq 500/550 mid output kit v2.5 (300 cycle). The resulting paired-end reads were quality controlled using FastQC v0.11.9 (Q scores of >30) (6), and then the reads were trimmed using Trimmomatic v0.38.1 (7) and de novo assembled using SPAdes v3.8.2 (8). The completeness of assemblies was assessed using BUSCO v5.0.0 and the bacillales_odb10 data set, with genome assemblies being 98.7% to 100% complete (9). Species-level taxonomic identification and abundances were inferred for all samples using our in-house kmer database (k = 30) (10). Default parameters were used for all analytical tools.

The depth of coverage for the draft genomes ranged from 29× to 99×, with genome sizes ranging from 3,352,943 to 5,678,307 bp (Table 1). The number of contigs ranged from 24 to 175, while the N50 values ranged from 46,339 to 446,425 bp. The GC contents varied from 37.1 to 47.1%. The sequences of these nine Bacillus isolates and two Weizmannia isolates were submitted to GenBank and included three Bacillus inaquosorum isolates, one Bacillus spizizenii isolate, four B. subtilis isolates, one Bacillus thuringiensis isolate, and two W. coagulans isolates.

TABLE 1

Bacillus and Weizmannia isolates sequenced in this study

Species	Strain	Source	Sequencing platform	SRA accession no.	BioSample accession no.	GenBank accession no.	No. of raw reads	Genome coverage (×)	No. of contigs	Size of largest contig (bp)	Total length (bp)	N50 (bp)	GC content (%)
Bacillus inaquosorum	DS36_4	Dietary supplement	NextSeq 500/550	SRR13955761	SAMN15078705	JAHHEC000000000	4,021,596	54	33	536,702	4,113,678	285,246	45.3
Bacillus inaquosorum	DS39_4	Dietary supplement	MiSeq	SRR11910190	SAMN15078713	JAHFZR000000000	1,664,686	58	34	578,311	4,173,056	334,226	45.3
Bacillus inaquosorum	DS43_4	Dietary supplement	NextSeq 500/550	SRR11910149	SAMN15078727	JAHHEB000000000	3,873,230	56	32	424,759	4,341,060	295,205	44.4
Bacillus spizizenii	DS19_4	Dietary supplement	MiSeq	SRR5310859	SAMN06464102	JAHHED000000000	2,269,810	60	46	315,861	3,978,163	226,004	44.8
Bacillus subtilis	DS1_4	Dietary supplement	MiSeq	SRR5310874	SAMN06464101	JAHFZX000000000	1,978,148	55	29	1,015,049	4,160,817	378,075	44.2
Bacillus subtilis	DS3_4	Dietary supplement	MiSeq	SRR5310850	SAMN06464103	JAHFZW000000000	736,374	29	65	358,756	4,000,557	144,448	44.9
Bacillus subtilis	DS6_4	Dietary supplement	MiSeq	SRR5310849	SAMN06464104	JAHFZV000000000	806,384	29	142	156,474	4,134,799	76,421	45.2
Bacillus subtilis	DS42_4	Dietary supplement	NextSeq 500/550	SRR13955744	SAMN15078726	JAHFZQ000000000	7,746,092	99	24	672,589	4,298,702	446,425	45.4
Bacillus thuringiensis	CF4_25	Cultured food	NextSeq 500/550	SRR11910240	SAMN15078641	JAHFZT000000000	8,096,266	77	76	843,051	5,678,307	209,556	37.1
Weizmannia coagulans	DS16_15	Dietary supplement	MiSeq	SRR5310867	SAMN06464119	JAHFZU000000000	2,561,696	85	171	184,398	3,361,815	46,339	46.6
Weizmannia coagulans	DS39_15	Dietary supplement	MiSeq	SRR11910189	SAMN15078709	JAHFZS000000000	1,797,890	70	175	154,030	3,352,943	47,238	47.1

Data availability.

This whole-genome shotgun project has been deposited in DDBJ/ENA/GenBank under BioProject number PRJNA336518 and accession numbers JAHFZQ000000000 to JAHHED000000000. The assemblies described in this paper are the first versions. All annotations were done by NCBI Prokaryotic Genome Annotation Pipeline (PGAP) v5.2 (11).