Draft Genome Sequences of Xylella fastidiosa subsp. fastidiosa Strains OK3, VB11, and NOB1, Isolated from Bunch and Muscadine Grapes Grown in Southern Mississippi.

We report here high-quality draft whole-genome assemblies of Xylella fastidiosa subsp. fastidiosa strains OK3, VB11, and NOB1, which were isolated from symptomatic bunch and muscadine grape plants grown in southern Mississippi.

ANNOUNCEMENT

Xylella fastidiosa is an economically important pathogen that occurs worldwide and affects numerous species of wild and cultivated plants (1, 2). The pathogen is endemic to the southeastern United States, where it causes Pierce’s disease (PD) and represents a major limiting factor for the expansion of the grape and wine industry (3, 4). The Gulf South region has particularly high PD pressure due to the abundance of native glassy-winged sharpshooters (Homalodisca vitripennis [formerly Homalodisca coagulata]), which act as an efficient vector of X. fastidiosa (5). Although the first reports of PD in Mississippi date back to the 1950s (6), many local aspects of this economically significant disease remain poorly understood. To address this gap, we sequenced and analyzed the genomes of strains OK3, VB11, and NOB1, which were isolated from symptomatic hybrid and muscadine grapes and represent the complex of X. fastidiosa strains that circulate in the state of Mississippi.

The strains OK3 and VB11 were isolated from interspecific bunch hybrid grape vines, OK392 and Villard Blanc, grown in Beaumont, Mississippi (31°14′5.5ʺN, 88°55′8.0ʺW), whereas NOB1 was isolated from muscadine cv. Noble collected at the USDA-ARS Stone County Research Farm (30°47′25.09ʺN, 88°59′22.25ʺW). The strains were isolated by shaking the surface-sterilized petioles and midribs of symptomatic plants for 2 h in PD2 broth (7), followed by plating onto PD2 agar amended with cycloheximide (100 μg ml−1). The taxonomic identity of isolates was confirmed by sequencing 16S amplicons generated with primers 8F and 1492R (8). BLASTn analysis of the 16S rRNA sequences from strains OK3, VB11, and NOB1 revealed 99.9% identity to X. fastidiosa strains ATCC 35879T and AB4 (GenBank accession numbers CP044352.1 and JN990279.1, respectively).

Samples of bacterial DNA were prepared from 1-week-old PD2 cultures using the cetyltrimethylammonium bromide protocol (9), quantified with a Qubit double-stranded DNA (dsDNA) high-sensitivity (HS) assay kit (Thermo Fisher Scientific, Waltham, MA), and sheared to a size of 350 bp. The libraries were prepared with a NEBNext DNA library preparation kit (New England Biolabs, Ipswich, MA) and sequenced in the 150-bp paired-end mode on a HiSeq 4000 platform (Illumina, Sacramento, CA). The sequencing of OK3, VB11, and NOB1 produced 11.8 million, 15.2 million, and 18.4 million reads, respectively, which were quality checked with FastQC (https://www.bioinformatics.babraham.ac.uk/projects/fastqc) and trimmed with Trim Galore v.0.6.1 (10). De novo genome assemblies were prepared with SPAdes v.3.13.0 (11), assessed with QUAST v.5.0.2 (12), and annotated with the NCBI Prokaryotic Genome Annotation Pipeline (PGAP) (13) and RASTtk (14) in PATRIC (15). All analyses were conducted using default parameters. The genome coverage, genome size, number of contigs, G+C content, and results of the PGAP annotation are summarized in Table 1. Multilocus sequence typing based on the housekeeping genes cysG, gltT, holC, leuA, malF, nuoL, and petC (16) revealed that the studied strains share an identical sequence type (ST) profile (ST2) and belong to the X. fastidiosa subsp. fastidiosa group.

TABLE 1

Characteristics and accession numbers of genomes of Xylella fastidiosa subsp. fastidiosa OK3, VB11, and NOB1 strains

Strain	Fold coverage	Genome size (bp)	No. of contigs	N50 (bp)	G+C content (%)	Total no. of genes	No. of RNA genes	Total no. of CDSsa	No. of PECsb	No. of hypothetical proteins	GenBank accession no.	SRA accession no.
OK3	137	2,415,390	54	116,099	51.50	2,207	54	2,153	416	532	JABCJH000000000	SRR11592759
VB11	135	2,444,001	75	103,260	51.45	2,268	55	2,213	416	559	JABCJI000000000	SRR11592758
NOB1	136	2,418,556	53	104,057	51.48	2,211	54	2,157	416	534	JABCJG000000000	SRR11592760

CDSs, coding sequences for proteins.

PECs, proteins with Enzyme Commission numbers.

Data availability.

The annotated genomes were deposited in GenBank under the accession numbers described in Table 1. Table 1 also lists the accession numbers for raw reads deposited in the Sequence Read Archive (SRA). All genome annotation and SRA data were included in BioProject number PRJNA627531.