Draft Genome Sequence of Francisella tularensis Strain 410108 from Tibet, China

Abstract

Francisella tularensis is the etiological agent of the potentially fatal disease tularemia. Here, we report the draft genome sequence of a virulent human isolate from Tibet, China in 1962, F. tularensis strain 410108, an intermediate-genotype strain of F. tularensis subsp. holarctica between biovar japonica and non-japonica strains in the world.

GENOME ANNOUNCEMENT

Francisella tularensis is a Gram-negative bacterium and the causative agent of tularemia, a public health issue across the northern hemisphere (1). There are four subspecies of F. tularensis: subsp. tularensis, holarctica, novicida, and mediaasiatica (2, 3). These four subspecies are closely related genetically and share most biochemical characteristics, but only the first two subspecies are clinically important (4, 5). In China, tularemia or F. tularensis has been found in Inner Mongolia, Tibet, Xinjiang, Jilin, Shandong, Heilongjiang, and Qinghai provinces or municipalities (6). Current reports of F. tularensis in China belong to subsp. holarctica.

Here, we report the draft genome of F. tularensis subsp. holarctica strain 410108, isolated from a tularemia patient in Dangxiong, Tibet in 1962. This strain exhibits a 50% lethal dose of less than 5 CFU by intraperitoneal infection of either mice or guinea pigs. Genotyping studies demonstrated that this strain belongs to an intermediate genotype between subsp. holarctica biovar japonica and other subsp. holarctica strains in the world. Identification of this intermediate genotype provides evidence for an Asian origin and radiation of holarctica-type tularemia.

The strain 410108 was grown on a glucose-cysteine blood agar plate with 5% CO₂ at 37°C. Cells were harvested by scraping and suspension in PBS followed by treatment with protease K. DNA was extracted using phenol-chloroform-isoamyl alcohol followed by isopropanol precipitation as previously described (7). The DNA was then used for full-length genome sequencing using an Illumina GA genome analyzer (Life Technologies). Two different DNA libraries (400-bp paired end and 3,000-bp mate pair) were sequenced, yielding 26,713,598 and 26,881,845 reads, respectively. De novo assembly was performed using SOAPdenovo version 1.05 (8). A total of 302 contigs were generated with an average length of 311,187 bp (N₅₀ size of 655,460 bp and total size of 1,867,125 bp), and were further assembled into 6 scaffolds. The gaps within the scaffolds were resolved using PCR and Sanger sequencing.

The draft chromosome of strain 410108 is 1,899,676 bp in size. The G+C content is 31.97%. The protein coding sequences (CDSs) were analyzed using FgenesB (SoftBerry), predicting a total of 1,508 CDSs (>90 amino acids). The putative functions of these CDSs were annotated using BLASTp as well as KEGG, COG, and GO protein databases (9, 10). Interestingly, strain 410108 has an intact FTT1080c homolog, which encodes a hypothetical membrane protein and has been recognized as subsp. tularensis-specific (11, 12). We surmise that this FTT1080c homolog might be a virulence gene candidate.

This is the first genome report of an F. tularensis strain in China. It is also the first genome of an intermediate genotype of F. tularensis subsp. holarctica between biovar japonica and other non-japonica strains in the world. Next we will perform genomic comparisons between more genomes of China isolates and genomes of other areas, which will likely provide new insights into the evolution and radiation of tularemia.

Nucleotide sequence accession number.

The draft genome sequence has been deposited in GenBank under the accession no. CP012092.