云南省马铃薯早疫病病原及毒性特点分析

Analysis of pathogens and pathogenicity characteristics of potato early blight in Yunnan Province

摘要:

【背景】早疫病是马铃薯主要病害之一，其病原为链格孢属(Alternaria)真菌，组成具有种复杂性和毒性差异性。【目的】明确云南省马铃薯早疫病两个致病种毒性特点和功能基因差异。【方法】以云南省大理州鹤庆县马铃薯主产区采集、分离和纯化的Alternaria solani (TA-0410)和Alternaria alternata (TB-1129)两株菌为材料，进行孢子形态观察、毒性测定、全基因组测序和比较分析研究。【结果】发现TA-0410为大孢子种，分生孢子褐色或黄色，孢子大小为[37.4-151.9(±28.1)] μm× [4.3-22.9(±4.1)]μm，喙长；TB-1129为小孢子种，分生孢子灰褐色，大小为[18.6-42.6(±9.3)]μm× [6.1-15.3(±2.3)] μm，喙短。毒性测定表明TA-0410为唯一致病种，TB-1129不能直接侵染引起马铃薯早疫病，但A. alternata在有伤接种条件下能产生并扩大病斑。两株菌测序分析发现TA-0410基因组大小为32.26 Mb，contig N₅₀为1 158 607 bp，含177个特有基因，TB-1129基因组大小为33.30 Mb，scaffold N₅₀为2 338 721 bp，含600个特有基因。此外，对唯一致病菌TA-0410进行全基因组分泌蛋白特征RxLx[EDQ]挖掘，共得到12个候选分泌蛋白，同源注释均为假定蛋白。结构域分析发现2个分泌蛋白ASR_2859和ASR_19579中分别包含和致病相关的BID_1和UBA结构域。【结论】A. solani和A. alternata均能引起马铃薯早疫病，A. alternata仅在有伤条件下侵染；两个菌株侵染和病斑扩展阶段的表型主要由孢子活力及生物毒性相关蛋白控制。

Abstract:

[Background] Early blight is one of the main potato diseases and the pathogen Alternaria has complex species composition and different pathogenicity. [Objective] To clarify the virulence characteristics and functional gene differences of two pathogenic species of potato early blight in Yunnan Province. [Methods] In this study, Alternaria solani (TA-0410) and Alternaria alternata (TB-1129) were collected, isolated, and purified from the main potato production area in Heqing County, Dali Prefecture, Yunnan Province, with the aim of clarifying the virulence characteristics and functional gene differences between the two species through spore morphological observation, pathogenicity verification, whole genome sequencing, and comparative analysis. [Results] TA-0410 was a large-spored species with brown or yellow conidia, spore size of (37.4–151.9(±28.1)) μm×(4.3–22.9(±4.1)) μm, and long beak. TB-1129 was a small-spored species with grayish-brown conidia, spore size of (18.6–42.6(±9.3)) μm× (6.1–15.3(±2.3)) μm, and short beak. The pathogenicity verification showed that TA-0410 was the only pathogenic species, as TB-1129 failed to induce early blight in the case of direct inoculation. However, in the instance of wounded-leaf inoculation, TB-1129 successfully infected the leaves and the lesions were expanded. The genome of TA-0410 was 32.26 Mb, with contig N₅₀=1 158 607 bp, and 177 unique genes, and the genome of TB-1129 was 33.30 Mb, with scaffold N₅₀=2 338 721 bp, and 600 unique genes. In addition, the secretory proteins of TA-0410 were analyzed and 12 candidate secretory proteins containing the RxLx[EDQ] motif were screened out, all of which were annotated as putative proteins. The secretory proteins ASR_2859 and ASR_19579 contained the pathogenic BID_1 and UBA domain, respectively. [Conclusion] Both A. solani and A. alternata can cause potato early blight, with A. alternata only in the case of wounded-leaf inoculation. The phenotypes of the two in the infection and expansion stages are mainly controlled by spore viability and biotoxicity-associated proteins.