Rubén Javier López

GENOME SEQUENCES Draft Genome Sequence of Agrobacterium radiobacter Strain MD22b, Isolated from a Grape Plant in Tajikistan Munavvara Dzhuraeva,a,b,c Khursheda Bobodzhanova,a Rubén Javier-Lopez,c Marina Tediashvili,b Ekaterine Jaiani,b,d Nils-Kåre Birkelandc Center of Biotechnology of the Tajik National University, Dushanbe, Tajikistan a G. Eliava Institute of Bacteriophages, Microbiology and Virology, Tbilisi, Georgia b c Department of Biological Sciences, University of Bergen, Bergen, Norway School of Medicine, New Vision University, Tbilisi, Georgia d ABSTRACT A grobacterium radiobacter (Beijerinck and van Delden 1902) Conn 1942 (formerly Agrobacterium tumefaciens) (1) was first isolated from grapevine galls in 1897 as the causative agent of crown gall disease (2). Despite their great importance in agriculture, strains of Agrobacterium from Tajikistan have not been studied previously. Within the framework of a project aiming at development of phage therapy for treatment of Agrobacteriuminfected grapevines in Tajikistan, strain MD22b was isolated from an infected fruit collected in September 2019 at Vatan Farm, a dekhkan farm in Yangibog (Tursunzade, Tajikistan). The fruit was rinsed three times in sterile distilled water and homogenized. Aliquots were spread onto Roy-Sasser agar (3) plates and incubated at 30°C for 3 days. Colonies were picked and streaked onto fresh Roy-Sasser agar plates for purification. One dark-red Agrobacterium-like colony was picked and identified as A. radiobacter by its API 20 NE (bioMérieux) profile -). DNA was extracted from cells cultivated in LB for 24 h at 30°C with shaking using the GenElute bacterial genomic DNA kit (Sigma-Aldrich). Sequencing of the 16S rRNA gene using the primer combination 59-AGRGTTTGATYHTGGCTCAG-39 (27f_mod) and 59TASGGHTACCTTGTTACGACTT-39 (1492r_mod) (4) as previously described (5) yielded a 1,275-nucleotide sequence identical to that of A. radiobacter strain ATCC 23308 (GenBank accession number MT-) using BLASTn v. 2.13.0 searches against the GenBank v. 252 nonredundant nucleotide database (https://blast.ncbi.nlm.nih.gov/Blast.cgi?PROGRAM= blastn&PAGE_TYPE=BlastSearch&LINK_LOC=blasthome). For genome sequencing by Eurofins Genomics, a NEBNext Ultra II DNA preparation kit was used, and Illumina NovaSeq 6000 S2 paired-end genomic sequencing was performed with a read length of 2 151 bp, resulting in 10,125,972 reads with a Phred score of $28 and a total of 1,518,897,000 sequenced bases. Additional quality control was performed to remove any remaining adapter sequences using the Trim Reads tool in the CLC Genomics Workbench v. 20.1. Assembly was performed using the CLC de novo assembly tool, resulting in a total sequence length of 5,736,602 bp and a total gapped length of 5,736,406 bp, distributed in 38 contigs, with an N50 value of 267,803 bp, coverage of 265, and GC content of 59.44%. Unless otherwise noted, default parameters were used for all software. Annotation of the draft genome was done using the NCBI Prokaryotic Genome Annotation Pipeline (PGAP) v. 6.1 (https://www.ncbi.nlm.nih.gov/genome/annotation_prok). The genome completeness was estimated at 100% using CheckM v. 1.0.18 (6). A phylogenomic analysis revealed clustering within the genus Agrobacterium with pairwise average nucleotide identity (ANI) and digital April 2023 Volume 12 Issue 4 Editor Simon Roux, DOE Joint Genome Institute Copyright © 2023 Dzhuraeva et al. This is an open-access article distributed under the terms of the Creative Commons Attribution 4.0 International license. Address correspondence to Nils-Kåre Birkeland,- The authors declare no conflict of interest. Received 16 November 2022 Accepted 22 February 2023 Published 20 March 2023 10.1128/mra- 1 Downloaded from https://journals.asm.org/journal/mra on 10 November 2025 by 2a01:799:ba7:7200:71c8:c3b1:f219:93f2. Agrobacterium radiobacter strain MD22b was isolated from infected fruit from Vatan Farm, a dekhkan farm in Yangibog (Tursunzade, Tajikistan). The 5.7-Mbp draft genome sequence presented here shares homology with chromosomes 1 and 2, as well as with the Ti plasmid from agrobacteria. Announcement Microbiology Resource Announcements DNA-DNA hybridization (dDDH) values of 97.97% and 83.8%, respectively, against the type strain A. radiobacter NCPPB 3001 using the ANI calculator (http://enve-omics.ce.gatech.edu) (7) and the Genome-to-Genome Distance Calculator v. 3.0 (https://ggdc.dsmz.de/ggdc.php#) (8, 9) with default settings. A BLAST Ring Image Generator (BRIG) v. 0.95 (10) comparison with A. radiobacter K84 was performed, revealing significant homology with both chromosomes of strain K84 (Fig. 1). A complete set of virulence genes guiding the transfer of transfer DNA (T-DNA) from bacteria to plant cells were identified, as well as the plasmid Ti partitioning genes, repA and repB, and the plasmid Ti replication initiator gene, repC. The reported data will be useful for future understanding of the genetic diversity and virulence potential of A. radiobacter in Central Asia. Data availability. The partial 16S rRNA gene and whole-genome shotgun sequences of A. radiobacter strain MD22b have been deposited in DDBJ/ENA/GenBank under accession numbers OP364099 and JANDHV-, respectively. The associated BioProject, SRA, and BioSample accession numbers are PRJNA856860, SRR-, and SAMN-, respectively. ACKNOWLEDGMENT This work was funded by the Eurasia Program of the Norwegian Directorate for Higher Education and Skills (HK-dir) (CPEA-LT-2017/10061). REFERENCES 1. Zhang L, Li X, Zhang F, Wang G. 2014. Genomic analysis of Agrobacterium radiobacter DSM 30147T and emended description of A-radiobacter (Beijerinck and van Delden 1902) Conn 1942 (approved lists 1980) emend. Sawada et al. 1993. Stand Genomic Sci 9:574–584. https://doi.org/10 .4056/sigs-. Kado CI. 2014. Historical account on gaining insights on the mechanism of crown gall tumorigenesis induced by Agrobacterium tumefaciens. Front Microbiol 5:340. https://doi.org/10.3389/fmicb-. Roy MA, Sasser M. 1983. A medium selective for Agrobacterium tumefaciens biotype-3. Phytopathology 73:-. Zeng YH, Koblížek M, Li YX, Liu YP, Feng FY, Ji JD, Jian JC, Wu ZH. 2012. April 2023 Volume 12 Issue 4 Long PCR-RFLP of 16S-ITS-23S rRNA genes: a high-resolution molecular tool for bacterial genotyping. J Appl Microbiol 114:433–447. https://doi .org/10.1111/jam.12057. 5. Dzhuraeva M, Shokirova M, Azaryan A, Panosyan H, Bobodzhanova K, Birkeland N-K. 2020. Draft genome sequence of Escherichia coli strain Tj, isolated from the Varzob River in Tajikistan. Microbiol Resour Announc 9: e-. https://doi.org/10.1128/MRA-. Parks DH, Imelfort M, Skennerton CT, Hugenholtz P, Tyson GW. 2015. CheckM: assessing the quality of microbial genomes recovered from isolates, single cells, and metagenomes. Genome Res 25:-. https:// doi.org/10.1101/gr-/mra- 2 Downloaded from https://journals.asm.org/journal/mra on 10 November 2025 by 2a01:799:ba7:7200:71c8:c3b1:f219:93f2. FIG 1 Circular representation of the MD22b genome compared with chromosomes 1 (left) and 2 (right) of the reference strain A. radiobacter K84, using the BLAST Ring Image Generator (BRIG) (10). The locations of genomic islands (GIs), prophage elements (Phage), a CRISPR/Cas element, and the rRNA genes in strain K84 are indicated. 7. 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BMC Genomics 12:402. https://doi.org/10.1186/-. 10.1128/mra- 3 Downloaded from https://journals.asm.org/journal/mra on 10 November 2025 by 2a01:799:ba7:7200:71c8:c3b1:f219:93f2. Announcement