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Microbiology and Biotechnology Letters

Genome Report(Note)

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Genome Report  |  Genome Report

Microbiol. Biotechnol. Lett. 2023; 51(4): 538-541

https://doi.org/10.48022/mbl.2311.11003

Received: November 7, 2023; Revised: November 26, 2023; Accepted: November 28, 2023

Draft Genome Sequence of Aureobasidium pullulans Strain MHAU2101, a Biological Control Agent against Fire Blight from Korea

Lin He1, Huan Luo1, Mi-Hyun Lee2, and Jun Myoung Yu1*

1Department of Applied Biology, Chungnam National University, Daejeon 34134, Republic of Korea
2Crop Protection Division, National Institute of Agricultural Sciences, Wanju 55365, Republic of Korea

Correspondence to :
Jun Myoung Yu,        junmyu@gmail.com

Abstract

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In this study, we present the draft genome of Aureobasidium pullulans strain MHAU2101, which is the first strain to effectively control fire blight caused by Erwinia amylovora in Korea. The genome of strain MHAU2101 was composed of 28,669,322 base pairs, with a C+G content of 50.4%. The assembly comprised 17 contigs and had 99.22% completeness. The results of this study will be a valuable resource for future research on the biocontrol mechanism of A. pullulans strain MHAU2101.

Keywords: Aureobasidium pullulans, biocontrol, fire blight, PacBio, Illumina, draft genome

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Aureobasidium pullulans is often referred to as “yeastlike” or “black yeast” due to its melanized yeast-like morphology. This fungus is ubiquitous in a variety of environments and is frequently isolated from the phyllosphere and soil [1]. A. pullulans is well-known for its significant biotechnological potential in medicine, pharmacy, and the food industry, which has been attributed to its capacity to produce polymeric substances, particularly pullulan, and hydrolytic enzymes [2, 3]. Additionally, this fungus has been used as a commercial biocontrol agent against postharvest diseases and fire blight [4, 5]. Notably, strain MHAU2101 was recently isolated from pears in Korea and demonstrated great efficacy in controlling fire blight caused by Erwinia amylovora [6]. The fungal strain MHAU2101 was identified as A. pullulans by morphological and phylogenetic analyses, employing large-subunit 26S ribosomal DNA and the internal transcribed spacer [6].

In this study, we sequenced the genome of A. pullulans MHAU2101, isolated from pear flowers in Wanju, South Korea, in 2021 [6]. The genomic DNA of strain MHAU2101 was extracted using the Quick-DNA Fungal/Bacterial Miniprep Kit (Zymo Research, USA). DNA purity and content were evaluated by spectrophotometry (NanoPhotometer NP80, Implen, Germany), and DNA integrity was assessed with electrophoresis. The quality-controlled DNA was used for genome sequencing using a combination of the PacBio Sequel and Illumina NovaSeq sequencing platforms, and the PacBio SMRTbell prep kit 3.0 (PacBio, USA) and TruSeq Nano DNA Preparation Kit (350) (Illumina, USA) were used to construct two sequencing libraries, respectively. Sequencing was performed by SEEDERS Co. (Republic of Korea; http://www.seeders.co.kr/). The Illumina data were used to polish the assembled genome using PacBio data and Pilon v1.21 software [7]. The benchmarking universal single-copy orthologs (BUSCO v5.1.3) dataset eukaryota_odb10 was used to evolve the completeness of the assembled genome [8]. The MAKER v3.01.03 program [9] was used to predict the protein-coding genes. Additionally, a gene functional analysis was performed using the Gene Ontology (GO), InterPro (v69.0), Pfam (v31.0), Conserved Domains Database (CDD v3.16), TIGRFAM (v15.0), and EggNOG (v4.5.1) databases. In this study, default parameters were utilized for all software and databases.

In total, 140,079 HiFi reads were obtained from PacBio sequencing, with a HiFi N50 value of 8,264 bp (half of all bases resided in reads of this size or longer), and an average read length of 7,301 bp, consisting of 1,022,750,246 HiFi bases. Illumina sequencing generated approximately 2,022.9 Mb of raw data, and 1,364.2 Mb of clean data were retained after filtering. De novo assembly was performed on the pure PacBio long reads using SMART Link v11.1.0 (https://www.pacb.com/), and an automatic circular check was conducted to ensure there were no overlaps within contigs. Illumina short reads were applied for sequence compensation to construct the contigs more accurately. Read mapping and data polishing were performed using Racon v1.4.13-74c937c program [10]. The classification of the species has been confirmed as A. pullulans MHAU2101 based on the whole-genome data. Upon comparing this data with the strain A. pullulans EXF-150 (Accession: AYEO00000000.1), it was observed that they share an average nucleotide identity (ANI) of 97.28% and a Digital DNA-DNA hybridization (dDDH) coverage of 73.70%.

The assembled draft genome was 28,669,322 bp in size, comprising 17 contigs and 50.4% G+C content (Fig. 1). The N50 of the genome was 2,213,608 bp. The length of the 17 contigs ranged from 18,956 bp to 4,494,408 bp, and the average length of a contig was 1,686,430 bp. The BUSCO results indicated that the genome assembly displayed a high overall completeness of 99.22%; only 0.78% of BUSCOs were present as fragments, and none were missing in the analysis. The genome contained 20,765 introns and 32,338 exons in total. Moreover, 11,573 protein-coding genes, 313 transfer RNAs, and 70 ribosomal RNA coding sequences were predicted. The features of the strain MHAU2101 genome are shown in Table 1. A total of 6,043 proteins were assigned to the GO database, 7,664 proteins were assigned to the InterPro database, 7,864 proteins were assigned to the Pfam database, 3,511 proteins were assigned to the CDD database, 878 proteins were assigned to the TIGRFAM database, and 11,054 proteins were assigned to the EggNOG database.

Table 1 . Genome statistics of Aureobasidium pullulans Strain MHAU2101.

Features StrainMHAU2101
PacBio platform
Number of read140,079
Total length of reads1,022,750,246 bp
N50 length8,264 bp
Average read length7,301 bp
Illumina platform
Raw data of read13,396,436
Clean data of read9,037,346
Clean data Q2099.32%
Clean data Q3096.78%
Clean data GC49.59%
Genome assembly
Contig17
Contig size28,669,322 bp
N50 length2,213,608 bp
GC content50.4%
Number of exons32, 338
Number of introns20,765
Putative protein-coding genes11,573
Number of tRNA313
Number of rRNA70
BUSCOs
Complete and single-copy BUSCOs253 (99.22%)
Complete and duplicated BUSCOs0 (0.00%)
Fragmented BUSCOs2 (0.78%)
Missing BUSCOs0 (0.00%)
Total BUSCO groups searched255 (100%)

Figure 1.Circular representation of the draft genome of Aureobasidium pullulans MHAU2101. From the outside to the inside of the circle graph are the genome contigs, the predicted coding sequence (sense strand and antisense strand), GC skew, positive (green), negative (purple), and the inner circle indicates the GC content. All these contigs are visualized in a single circular representation. In the assembly, contig 16 is the only one that is circular.

In summary, we present a high-quality draft genome assembly of A. pullulans MHAU2101, a biocontrol strain known for its efficacy against fire blight caused by E. amylovora. The insights derived from this genome assembly not only enhance our fundamental understanding but also carry practical implications for developing targeted strategies to combat the devastating plant disease, fire blight. The genomic sequence serves as a pivotal resource for future research on genetic diversity and biocontrol mechanisms of A. pullulans. This will significantly enrich the knowledge base essential for advancing our understanding of biocontrol mechanisms and facilitating the development of practical applications in agriculture.

Nucleotide Sequence Accession Number(s)

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The draft genome sequence of A. pullulans strain MHAU2101 in this article was deposited at DDBJ/ENA/GenBank under accession number JAWJBJ000000000. The version described in this study is JAWJBJ010000000. (BioProject: PRJNA1027605; BioSample: SAMN37801316).

Acknowledgments

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This work was supported by a grant from the Agenda program (PJ015594) of the Rural Development Administration, Republic of Korea.

Conflict of Interest

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The authors have no financial conflicts of interest to declare.

References

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