Complete genome sequence of Paracidovorax avenae causing red stripe in sugarcane

Abstract

Objective

Paracidovorax avenae (Pa) is the causative agent of red stripe disease in sugarcane and belongs to the Gram-negative β-Proteobacteria. Red stripe is a major bacterial disease of sugarcane worldwide. Limited genome sequences of Pa can be used for exploring the phylogenetic and genetic diversity analysis in this pathogen at the complete genome level. In this study, a whole genome sequence of Pa CNGX08 strain isolated from sugarcane in China was assembled and annotated.

Data description

Genome assembly data from second- and third-generation sequencing revealed that the entire genomic sequence of Pa CNGX08 strain causing red stripe in sugarcane, consisted of a 5,625,582 bp circular chromosome with a GC content of 68.97%. In total, 4,915 protein-coding genes were annotated. Additionally, 9 ribosomal RNAs and 52 transfer RNAs were identified. This genomic resource will facilitate the genome-based taxonomic classification of the genus Paracidovorax and the exploration of pathogenic mechanisms underlying sugarcane red stripe disease caused by Pa.

Objective

Red stripe is one of the three major bacterial diseases in sugarcane and is prevalent in many sugarcane-growing regions worldwide [1]. This disease was first reported in the 1920s in Hawaii [2] and now has commonly occurred in Argentina [3] and China [4]. Two symptoms of this disease are leaf stripe and top rot, appearing separately or simultaneously in the field [2]. Economic losses are caused by red stripe, especially the top rot form [5]. Red stripe is caused by Acidovorax avenae subsp. avenae belonging to the Gram-negative β-Proteobacteria. Recently, Du et al. (2023) [6] proposed the division of the genus Acidovorax into the two novel genera Paracidovorax gen. nov. and Paenacidovorax gen. nov. Thus, the Acidovorax avenae subsp. avenae was renamed as Paracidovorax avenae (Pa). Here, we report the complete and annotated genome sequence of Pa strain CNGX08 causing sugarcane red stripe in Guangxi province, China. This work provides valuable insights into the comparative genome analysis among global Pa strains and the molecular mechanisms of interaction between Pa and host sugarcane.

Data description

The Pa CNGX08 was originally isolated from sugarcane leaf showing red stripe symptoms in the Guangxi province, China (24.5285°N, 109.2613°E). The CNGX08 forms white-cream colored colonies on nutrient agar (NA) medium [7]. Bacterial genome DNA was extracted using the SDS method, then detected by agarose gel electrophoresis and quantified using a Qubit^® 2.0 Fluorometer (Thermo Scientific, China). The Blue Pippin system was utilized to automatically isolate large DNA fragments, which were subsequently subjected to a repair process. The incorporation of a unique barcode was facilitated by the PCR-free EXP-NBD104 kit from Oxford Nanopore Technologies (ONT), ensuring minimal bias and maintaining the integrity of the DNA. Following this, the Advanced Analytical Technologies, Inc. (AATI) capillary electrophoresis device was employed to precisely measure the fragment sizes, ensuring they met the required specifications. The samples were then mixed to achieve an isomolar concentration, allowing for an equal representation of each DNA fragment. The final step involved the use of the SQK-LSK109 kit (Oxford Nanopore Technologies, Oxford, UK) to attach the necessary adapters, culminating in the successful assembly of the 10 K sequencing library.

Sequencing libraries were prepared using the NEBNext^® Ultra™ DNA Library Prep Kit for Illumina (NEB, USA), with index codes added to attribute sequences to each sample. The whole genome of CNGX08 was sequenced using the Nanopore PromethION platform and the Illumina NovaSeq PE150 at the Beijing Novogene Bioinformatics Technology Co., Ltd. The genome was assembled with all processed long-reads and short-reads using Unicycler (version 0.4.8) [8]. Briefly, the 150-bp paired-end reads were filtered and trimmed using fastp (v0.23.0). A frame diagram genome was assembled using SPAdes, and long-reads were added to the frame map using miniasm and Racon. A complete genome of CNGX08 was 5,625,582 bp long with a GC content of 68.97% (Data file 1, Table 1) [9] and was generated using Unicycler [8].

Subsequently, the BUSCO v4.0.2 tool (http://busco.ezlab.org/) was used to assess the final genome assembly. BUSCO (98.4%) analyses suggest that our genome assembly exhibits a high degree of completeness. The CNGX08 genome was generated into one circular chromosome harboring a length of 5,625,582 bp with 68.97% GC content (Data file 1) [9]. As shown in Table 1, circos (version 0.64) was used to display the genome structure and annotation of CNGX08 (Data file 2) [10]. The genomic sequence was uploaded to the National Center for Biotechnology Information (NCBI) GenBank database with the accession number CP156079 under the Bioproject number PRJNA1006162, as shown in Data file 3 [11].

Moreover, the coding genes were predicted using GeneMarkS (version 4.17). The signal peptides and transmembrane structures of proteins were analyzed by the SignalP (version 4.1) [12], while the secreted protein was comprehensively predicted with the TMHMM (version 2.0c). Among these annotation results of the protein sequence function database, the secretion system proteins and the type III secretion system (T3SS) effector proteins were predicted using EffectiveT3 (version 1.0.1) [13] (Data file 1) [9]. A total of 310 T3SS effective proteins were found. The genome of CNGX08 contains 4,915 protein-coding genes, 41 repeat regions, 8 genomic islands, 9 bacteriophages, and 4 clustered regularly spaced short palindromic repeats (CRISPR) (Data file 1) [9]. Gene functions were predicted with GO (Gene Ontology), KEGG (Kyoto Encyclopedia of Genes and Genomes), COG (Clusters of Orthologous Groups), NR (Non-Redundant Protein Database), TCDB (Transporter Classification Database), and Swiss-Prot. A BLAST search (E-value less than 1 × 10^− 5, minimal alignment length percentage larger than 40%) was performed against these databases. Pathogenicity and drug resistance analyses were carried out using the PHI (Pathogen Host Interactions) database. These results are shown in Supplementary Data file 4 [14].

The genomic sequences of CNGX08 and 19 worldwide strains of Pa along with P. facilis strain DSM 649 (as an outgroup) were used for collinearity analysis, population evolution analysis, SNP/INDEL analysis, and Average nucleotide identity (ANI) analysis. A phylogenetic tree was constructed using TreeBeST with the 1,000 bootstraps based on the core pan-genome level. All tested Pa strains were clustered into several clades, while the CNGX08 was clustered in a unique clade. ANI analysis showed that the CNGX08 were 93.32-94.04% identities with other Pa strains (Data file 5) [15]. Notably, the CNGX08 shared the sequence identities of 93.43% and 93.45% with the T10_61 strain infecting sugarcane in Argentina and the reference strain of ATCC 19860, respectively. These results suggested that CNGX08 is a distinct Pa strain from T10_61 and ATCC 19860 strains. However, it is necessary that more genomic sequences of Pa strains from various plant hosts and geographical origins were sequenced and assembly for further illustrating the divergence and taxonomic classification of this pathogen.

In conclusion, this study demonstrated genomic information of the Pa strain CNGX08 infecting sugarcane in China, providing an understanding of the evolutionary history of the Paracidovorax genus. The availability of this data serves as a reference for comparative genomics of Pa across different hosts.

Table 1 Overview of data files/data sets

Limitations

This Data Note presents the whole-genome sequencing of the Pa CNGX08 strain based on the combination of the Nanopore PromethION (third-generation) and the Illumina NovaSeq PE150 (second-generation) platforms, providing a high-quality and gap-free reference genome. Therefore, the authors believe that there are no limitations in the data.