# Whole Genome Sequences of Nine Xanthomonas Strains Responsible for Common Bacterial Blight of Bean

Charlotte Gaudin, Christopher Gihaut, Martial Briand, Abi S.A. Marques,

Marisa A. S. V. Ferreira, Marie-Agnès Jacques, Nicolas Chen

► To cite this version:

Charlotte Gaudin, Christopher Gihaut, Martial Briand, Abi S.A. Marques, Marisa A. S. V. Ferreira, et al.. Whole Genome Sequences of Nine Xanthomonas Strains Responsible for Common Bacterial Blight of Bean. Microbiology Resource Announcements, 2023, 12, pp.e01259-22. 10.1128/mra.01259-22. hal-04044008

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# Whole Genome Sequences of Nine *Xanthomonas* Strains Responsible for Common Bacterial Blight of Bean

Charlotte Gaudin,ª Christopher Gihaut,ª Martial Briand,ª Abi S. A. Marques,ᵇ Marisa A. S. V. Ferreira,ᶜ Marie-Agnès Jacques,ª

ᵇNicolas W. G. Chenª

**ABSTRACT** We report the complete and circularized genome sequences of nine strains of *Xanthomonas phaseoli* pv. *phaseoli* and *Xanthomonas citri* pv. *fuscans*, which cause common bacterial blight of bean. These assemblies provide high-quality material for functional and evolutionary studies of these legume pathogens.

Common bean (*Phaseolus vulgaris*) is an important legume crop for direct human consumption that is used worldwide for its edible pods and seeds (1). Common bacterial blight (CBB) is a devastating seed-borne bacterial disease that represents an important economic and agricultural threat to both common bean production and the seed industry worldwide (2). CBB is caused by two phylogenetically distant bacterial species that cause identical symptoms on common bean, i.e., *Xanthomonas phaseoli* pv. *phaseoli* and *Xanthomonas citri* pv. *fuscans* (2). These two species are subdivided into four genetic lineages. *X. phaseoli* pv. *phaseoli* corresponds to lineage GL1, while *X. citri* pv. *fuscans* comprises lineages GL2, GL3, and *fuscans* (2, 3).

Here, we produced Pacific Biosciences (PacBio) high-fidelity (HiFi) genome sequences of nine strains that had been isolated from *Phaseolus vulgaris* plants in different regions of the world and belonged to either GL1, GL2, or *fuscans* lineages (Table 1). Strains were isolated from symptomatic leaflets as described previously (4). The status as a CBB agent was confirmed by pathogenicity tests and sequencing of the housekeeping gene *gyrB* (5). Bacteria were grown on Trypticase soy (TS) agar medium for 2 days at 28°C, and then 5 or 6 clones were scraped, pooled, and cultured overnight in 100% TS broth to obtain fresh cultures. One milliliter of overnight culture was centrifuged for 2 min at 13,000 × g. Genomic DNA was extracted using the Wizard genomic DNA purification kit (Promega) according to the manufacturer's recommendations. DNA was mechanically sheared using g-TUBE columns (Covaris) and purified using PacBio AMPure PB beads. PacBio HiFi SMRTbell libraries were prepared from ~1 µg of genomic DNA using the SMRTbell Express template preparation kit v2.0. Single-molecule real-time (SMRT) sequencing was performed on a PacBio Sequel II system using the Sequel II binding kit v2.2, sequencing primer v5, and Sequel II sequencing plate v2.0.

PacBio reads were filtered and demultiplexed using the ccs v6.3.0 and lima v2.5.1 tools of the PacBio SMRT Tools v11.0.0.146107 toolkit and then assembled and circularized using Flye v2.9 (https://github.com/fenderglass/Flye) (6). For the BB007a strain, assembly and circularization were performed again using preselection of reads with Filtlong (https://github.com/rrwick/Filtlong). Circularization failed for strains BRM48906 and UnB187. For circularized genomes, the sequence start was fixed using the fixstart option of Circlator v1.5.1 (7). Polishing was performed using Flye v2.9 (6).

The genome sequences consisted of 5,101,251 to 5,242,207 bp, forming a chromosome plus 1 to 3 plasmids, with an average G+C content of 64.7% (Table 1). A total of 4,038 to 4,236 predicted protein-coding sequences (CDSs) were identified using the NCBI Prokaryotic

**Editor** David A. Baltrus, University of Arizona

**Copyright** © 2023 Gaudin et al. This is an open-access article distributed under the terms of the Creative Commons Attribution 4.0 International license.

Address correspondence to Nicolas W. G. Chen, nicolas.chen@agrocampus-ouest.fr.
The authors declare no conflict of interest.

[This article was published on 13 February 2023 with information missing from the Acknowledgments section. The section was updated in the current version, posted on 16 March 2023.]

**Received** 1 December 2022

**Accepted** 13 January 2023

**Published** 13 February 2023

<footnote>ªUniversité Angers, Institut Agro, INRAE, IRHS, SFR QUASAV, Angers, France</footnote>

<footnote>ᵇPlant Quarantine Station, Embrapa Recursos Genéticos e Biotecnologia, Brasília, Brazil</footnote>

<footnote>ᶜDepartamento de Fitopatologia, Universidade de Brasília, Brasília, Brazil</footnote>
<table><caption>TABLE 1 Accession numbers and statistics</caption><thead><tr><th>Pathovar</th><th>Lineage</th><th>Strain</th><th>Country (date) of collection</th><th>No. of reads<sup>a</sup></th><th>Coverage (x)</th><th>N<sub>50</sub> (bp)<sup>a</sup></th><th>SRA accession no.</th><th>GenBank assembly no.</th><th>Size (bp)</th><th>G+C content (%)</th><th>No. of plasmids</th><th>No. of CDs</th><th>Source or collection</th></tr></thead><tbody><tr><td>X. phaseoli pv. phaseoli</td><td>GL1</td><td>UnB187</td><td>Brazil (1981)</td><td>40,024</td><td>61</td><td>10,755</td><td>SRR2209457</td><td>GCA_025987765.1</td><td>5,217,655</td><td>64.9</td><td>2</td><td>4,187</td><td>This study</td></tr><tr><td>X. phaseoli pv. phaseoli</td><td>GL1</td><td>BB007a</td><td>USA (2007)</td><td>94,609</td><td>141</td><td>10,404</td><td>SRR2209456</td><td>GCA_025987745.1</td><td>5,242,207</td><td>64.8</td><td>1</td><td>4,236</td><td>This study</td></tr><tr><td>X. phaseoli pv. phaseoli</td><td>GL1</td><td>CFBP 7423</td><td>Iran (2006)</td><td>73,335</td><td>111</td><td>10,510</td><td>SRR2209455</td><td>GCA_025987785.1</td><td>5,214,828</td><td>64.8</td><td>3</td><td>4,209</td><td>CIRM-CFBP<sup>b</sup></td></tr><tr><td>X. phaseoli pv. phaseoli</td><td>GL1</td><td>BB075a</td><td>USA (2014)</td><td>86,101</td><td>130</td><td>10,373</td><td>SRR2209454</td><td>GCA_025987725.1</td><td>5,161,803</td><td>64.9</td><td>2</td><td>4,145</td><td>This study</td></tr><tr><td>X. phaseoli pv. phaseoli</td><td>GL1</td><td>BB013a</td><td>USA (2007)</td><td>118,064</td><td>177</td><td>10,200</td><td>SRR2209453</td><td>GCA_025987685.1</td><td>5,199,960</td><td>64.8</td><td>2</td><td>4,194</td><td>This study</td></tr><tr><td>X. citri pv. fuscans</td><td>GL2</td><td>CFBP 6533</td><td>Tunisia (1989)</td><td>71,377</td><td>103</td><td>10,250</td><td>SRR2209452</td><td>GCA_025987705.1</td><td>5,154,616</td><td>64.6</td><td>1</td><td>4,209</td><td>CIRM-CFBP<sup>b</sup></td></tr><tr><td>X. citri pv. fuscans</td><td>GL2</td><td>BRM48906</td><td>Brazil (2014)</td><td>42,986</td><td>64</td><td>10,574</td><td>SRR2209451</td><td>GCA_025987665.1</td><td>5,230,237</td><td>64.6</td><td>2</td><td>4,234</td><td>This study</td></tr><tr><td>X. citri pv. fuscans</td><td>GL2</td><td>LNPV 30.30</td><td>China (2006)</td><td>93,288</td><td>146</td><td>10,736</td><td>SRR2209450</td><td>GCA_025987645.1</td><td>5,156,952</td><td>64.6</td><td>1</td><td>4,212</td><td>This study</td></tr><tr><td>X. citri pv. fuscans</td><td>Fuscans</td><td>X1008017</td><td>France (2010)</td><td>144,222</td><td>211</td><td>10,204</td><td>SRR2209459</td><td>GCA_025987485.1</td><td>5,101,251</td><td>64.7</td><td>2</td><td>4,038</td><td>This study</td></tr></tbody></table>

<footnote><sup>a</sup> Calculated with the n50 tool (https://github.com/quadram-institute-bioscience/seqfu/wiki/n50).</footnote>

<footnote><sup>b</sup> CIRM-CFBP, International Centre for Microbial Resources-French Collection for Plant-Associated Bacteria, INRAE (https://doi.org/10.15454/E8XX-4Z18).</footnote>
Genome Annotation Pipeline (PGAP) (8). Genome sequences were estimated to be >99.3% complete and <0.02% contaminated using CheckM v1.1.6 (9). Chromosomes were assembled in 1 contig except for strain X1008017, with 2 contigs. Every strain contained plasmid A, which is consistent with previous data for CBB agents (10) and reinforced the idea that plasmid A was involved in the pathological convergence on bean by these two species (11). Each genome sequence included the usual arsenal of genes involved in pathogenicity, compared to the reference strain CFBP 4834-R (12), although the GL1 and GL2 lineages showed polymorphism in important components such as the lipopolysaccharide and secretion systems. These assemblies provide high-quality material for functional and evolutionary studies of these common bean pathogens.

**Data availability.** The reads and assemblies were all deposited in GenBank under the accession numbers listed in Table 1.

## ACKNOWLEDGMENTS

Sequencing was performed at the GENTYANE sequencing platform (INRAE, Clermont-Ferrand, France). We thank the Limagrain group for funding the sequencing costs and providing strains isolated on bean seeds. C. Gaudin was funded by a Ph.D. grant (CRISTAL project) cofunded by L'Institut Agro Rennes-Angers and the French Pays de Loire region.

We thank Killian Grilloton for his help in DNA extractions and the CIRM-CFBP (https://doi.org/10.15454/E8XX-4Z18) for the handling of strains CFBP 7423 and CFBP 6533. Strain LNPV 30.30 was provided by ANSES—Laboratoire de santé des végétaux, Angers, France.

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