the genus afipia was first proposed by brenner et al . in 1991 , containing afipia felis sp . this genus comprises of gram - negative , motile , oxidase - positive bacteria belonging to the alpha-2 subgroup of the class proteobacteria . subsequently , the isolation of afipia birgiae , afipia massiliensis and a. felis genospecies a from hospital water supply using amoeba co - cultivation was described in 2002 . more recently , our lab isolated a novel species a. septicemium ohsu_i and ohsu_ii from the blood cultures of 2 patients with poorly defined clinical illnesses . the bacteria were originally living in an inactive state in sp4 broth medium designed to culture fastidious mycoplasmas . the bacteria gradually adapted to grow in modified sp4 medium supplemented with 5% fetal bovine serum , vitamin b12 , ckm ( cacl2/kno3/mgso4 ) and nad / nadp ( -nicotinamide adenine dinucleotide/-nicotinamide adenine dinucleotide phosphate , 10  mg / l ) and could form microscopic colonies on 1.5% noble agar plates prepared using modified sp4 medium kept in room temperature . the subsequent single - colony derived microbes could grow in ym ( yeast malt ) and bhi ( brain heart infusion ) broth without serum supplement . whole - genome sequencing was performed on dna extracted from single - colony clones of a. septicemium ohsu_ii grown in sp4 broth cultures . briefly , 50  ng of purified dna of a bacterial culture derived from a single colony were subjected to dna library construction using the nextera dna sample prep kit ( illumina ) . a mixture of 4 to 6 separate dna libraries was usually analyzed for each run involving 2    250  bp pair - end sequencing . the sequencing data , sufficient to cover bacterial genomes of ~  5  million  bps 378-fold , was obtained by illumina miseq platform . the raw reads generated from genomic dna sequencing were first assembled into 42 contigs using clc genomics workbench ( clc bio version 6.0 ) using the de novo assembly after filtering out and trimming low - quality reads . the 16s rrna gene sequence of a. septicemium was extracted from genome data and compared using ncbi blast under default settings ( e.g. considering only the high - scoring segment pairs ( hsp ) from the best 250 hits ) with the most recent database release in genbank . the analysis showed that among all the matches for the 16s rrna gene sequences , the species with the highest bit score of 2700 was uncultured afipia sp . clone c-12 ( ay568503 ) , corresponding to an identity of 99.7% , followed by 2 uncultured afipia clones and 2 uncultured bacterial clones , with bit scores ranging from 2676 to 2641 . the highest scoring bacteria of currently recognized known species were afipia lausannensis ( bit score 2638 ) and a. broomeae ( bit score 2630 ) . 1 shows the phylogenetic relatedness of a. septicemium with other closely related bacteria species based on the 16s rrna sequences . the phylogenetic relatedness of difficult to culture a. septicemium with several other uncultured afipia species using both neighbor - joining   and minimal evolution   criteria inferred by mega4 program   strongly supports the assigning of the newly identified microbes to the genus afipia . it is important to note that the microbes in this genus , or even in this family , are known to have very high levels of homology in their 16s rrna gene sequences , . thus , what presently classified as afipia bacteria based on 16s rrna sequences are likely to be a heterogeneous group of bradyrhizobiaceae . genomic content differences between a. septicemium and other established afipia species of bacteria including a. broomeae are more than 22% , which strongly supports that a. septicemium is a distinct afipia species . a working draft genome of the a. septicemium ohsu_ii - c1 was constructed using the complete genome of bradyrhizobiaceae bacterium strain sg-6c ( genbank accession number cm001195 ) and the 3 supercontigs ( 1.1 , 1.2 and 1.3 without including supercontig 1.4 of 128  kb ) of a. broomeae ( genbank accession numbers nz_kb375282.1 , nz_kb375283.1 and nz_kb375284 )   as reference genomes using contigutor . sequence of supercontig 1.4 of a. broomeae could not be aligned with bradyrhizobiaceae bacterium sg-6c genome sequence and revealed significant homology with several plasmids of alph-2 proteobacteria , and therefore was not used as a reference for draft genome construction . the draft genome of a. septicemium ohsu_ii - c1 consists of a 5,087,893  bp circular chromosome with no identifiable autonomous plasmid with a g  +  c content of 61.09% and contains 49 rna genes including 3 rrna genes and 46 trna genes predicted by rnammer 1.2   and trnascan - se-1.23 . the functional annotation was carried out by rast ( rapid annotation using subsystem technology ) hosted by fellowship for interpretation of genomes . the functional comparison of genome sequences available on the rast server revealed the closest neighbors of a. septicemium as afipia sp . 1nls2 ( score 537 ) followed by nitrobacter winogradskyi nb-255 ( score 517 ) , a. clevelandensis atcc 49720 ( score 496 ) and oligotropha carboxidovorans om4 ( score 486 ) . a total of 5061 coding regions ( 2337 from a positive strand and 2724 from a negative strand ) were found in the genome of which 3487 ( 69% ) could be functionally annotated . the genome coding density is 87% with an average gene length of 875  bp . 2 revealed that the annotated genome has 135 genes potentially involved in virulence , disease and defense including 109 genes for resistance to antibiotics and toxic compounds such as cobalt additionally , the annotated genome has 18 genes involved in phages , prophages , transposable elements and plasmids ( fig .  2 ) . further examination of the 18 genes revealed that there is a cluster of 14 gene transfer agent ( gta ) genes from the genomic position 3,629,808 to 3,652,259 ( fig .  3 ) . these gta genes have a similar organization of the first gta described in the purple photosynthetic alpha-2 proteobacteria rhodobacter capsulatus ( rcgta ) . these gta genes include terminase , the phage portal protein , the prohead protease , the major capsid protein , the head  tail adaptor protein , the major tail protein , the tail tape measure protein , and host specificity proteins ( fig .  3 ) . this particular region identified in a. septicemium genome revealed the highest similarity with that of brucella abortus , a known pathogenic member of rhizobiales , when compared using the rast database ( fig . several potentially pathogenic afipia bacteria such as a. felis , afipia sp . 1nls2 and a. birgiae are also found to possess similar numbers of rcgta genes in their genomes . however , only the terminase gene is found in genomes of a. clevelandensis and a. broomeae . alphaproteobacteria are known to transfer their genomes into their eukaryotic hosts and to receive dna from other organisms by bidirectional lateral sequence transfer through mechanisms such as transformation , transduction , conjugation , membrane vesicles , intracellular nanotube or gtas . recently a novel putative gta ( bagta ) was recently discovered in the mouse - infecting bacterium bartonella grahamii , another species of alphaproteobacteria within the order of rhizobiales . it was suggested that bagta contributed to the genomic plasticity of bartonella to allow the microbe to evolve adaptively towards infecting a broader range of different hosts . it is presently unclear if bagtas or any novel gtas other than the rcgta - like genes are present in a. septicemium genome . a. septicemium draft genome shotgun ( wgs ) project has been deposited at ddbj / embl / genbank under the project accession apji00000000 . it has a bioproject i d of prjna191806 and a gold i d of gi51905 . the authors declare that there is no conflict of interest on any work published in this paper .
we report the 5.1  mb noncontiguous draft genome of afipia septicemium strain ohsu_ii , isolated from blood of a female patient . 
 the genome consists of 5,087,893  bp circular chromosome with no identifiable autonomous plasmid with a g  +  c content of 61.09% and contains 4898 protein - coding genes and 49 rna genes including 3 rrna genes and 46 trna genes .