Due to the highly fastidious nature of the genus Liberibacter, research on these organisms has traditionally been limited to electron microscopy and genomic analysis [3,7,10]. However, one species of the genus, Liberibacter crescens, has recently been cultured sellectchem and characterized [11], and the relationship between its genome and close relatives will be the focus here. In order to gain insight on both the virulence and metabolism of the genus Liberibacter, all available genomes of the Liberibacter spp. were compared to Liberibacter crescens. To date, the genomes of Candidatus L. asiaticus and Candidatus L. solanacearum are publicly available. The differences between these species may be responsible for the fastidious nature of the Liberibacter spp. Sequencing, assembly, and annotation of L.
crescens were performed in order to proceed with the investigation. Classification and features Figure 1 and Table 1 summarize the phylogenetic position and characteristics of Liberibacter crescens BT-1, respectively. Figure 2 shows transmission electron microscopy of L. crescens BT-1. Figure 1 Maximum likelihood phylogenetic tree constructed using 16S rRNA genes of Liberibacter crescens BT-1 and related members of the Alphaproteobacteria. Branch supports are provided above branches. Sequences were aligned using MUSCLE [12]. Overhanging regions … Table 1 Classification and general features of Liberibacter crescens BT-1 according to the MIGS recommendations [18] Figure 2 Transmission electron microscopy of L. crescens BT-1. Negative stain. Scale bar represents 500 nm.
Genome sequencing and annotation Three sequencing platforms were used to obtain the data necessary to close the genome sequence (Table 2). In addition, other project information and its association with MIGS version 2.0 compliance [32] is provided (Table 2). Table 2 Project information Growth conditions and DNA isolation The initial culture of BT-1 was obtained in 1995 and was isolated from the peduncle of the tropical Babaco plant, also known as the hybrid mountain papaya (Carica stipulata x C. pubescens). Babaco was provided by the Lajas Experiment station in Puerto Rico because it showed signs of Papaya bunchy top (PBT), a disease of papaya in the American tropics. The sap of Babaco expressed an extremely high titer of small, rod-shaped bacteria [1]. Despite being fastidious, the bacterium was able to be grown on BM7 media, a modified form of BBM [1,11]. Cells were grown in BM7 liquid culture at 27��C for 4 days in a shaking incubator with a speed of 120 rpm. DNA was extracted GSK-3 using the UltraClean Microbial DNA Isolation Kit and the manufacturer��s protocol (M0-BIO, Carlsbad, CA).