真菌病毒BbOCuV1对寄主球孢白僵菌生长发育和亚洲玉米螟致病力的影响

doi:10.13560/j.cnki.biotech.bull.1985.2025-0053

摘要/Abstract

摘要：

目的通过明确真菌病毒beauveria bassiana orthocurvula virus 1（BbOCuV1）对寄主球孢白僵菌（Beauveria bassiana）生长发育和对害虫的致病力的影响，研究真菌病毒是否是导致球孢白僵菌致病力下降的关键因素。方法采用脱毒和水平传毒方法比较研究BbOCuV1对寄主球孢白僵菌菌落生长速率、产孢量和生物量的影响，并测定了寄主球孢白僵菌对亚洲玉米螟（Ostrinia furnacalis）二龄幼虫的致病力。利用转录组分析，揭示了真菌病毒影响寄主菌株的分子机制。结果感染BbOCuV1后，寄主菌株生长速率、产孢量和生物量均显著提高，但对亚洲玉米螟二龄幼虫的致病力显著降低。转录组分析显示，感染病毒后的白僵菌菌株生长发育相关通路和基因表达量显著提高，但与昆虫表皮穿透和毒素代谢相关的基因表达降低。结论感染BbOCuV1病毒后的球孢白僵菌，其生长发育及生物量明显增强，但对害虫致病力降低。说明真菌病毒感染是引起球孢白僵菌对害虫致病力下降的重要因素。

关键词: 球孢白僵菌, 真菌病毒, BbOCuV1, 生物学性状, 生物防治

Abstract:

Objective By clarifying the effects of the mycovirus Beauveria bassiana orthocurvula virus 1(BbOCuV1) on the growth and development of the host Beauveria bassiana and on the pathogenicity of insect pests, we proved whether the mycovirus is a key factor that cause the decline of the pathogenicity of B. bassiana. Method Detoxification and horizontal transmission were used to study the effects of BbOCuV1 on the growth rate, spore production and biomass of B. bassiana colonies. The pathogenicity of B. bassiana on the second instar larvae of the Ostrinia furnacalis was determined. Transcriptome analysis was used to define the molecular mechanism of fungal viruses affecting the host strains. Result The growth rate, spore production and biomass of host strain significantly increased after virus BbOCuV1 infection, while the pathogenicity towards the insect pest significantly decreased. Transcriptome analysis showed that growth and development-related pathways and genes expression significantly upretulated, after virus infection, while the expressions of genes related to insect epidermis penetration and toxin metabolism significantly downregulated. Conclusion The growth and development as well as biomass of B. bassiana infected with virus BbOCuV1 increase, but pathogenicity against pests reduces, indicating that mycovirus infection is an important factor in the pathogenicity decline of B. bassiana to pests.

Key words: Beauveria bassiana, mycovirus, BbOCuV1, biological character, biological control

贾雪, 隋丽, 邹晓威, 路杨, 张正坤, 李启云. 真菌病毒BbOCuV1对寄主球孢白僵菌生长发育和亚洲玉米螟致病力的影响[J]. 生物技术通报, 2025, 41(7): 312-325.

JIA Xue, SUI Li, ZOU Xiao-wei, LU Yang, ZHANG Zheng-kun, LI Qi-yun. Effects of the Mycovirus BbOCuV1 on the Growth and Development of Host Beauveria bassiana and Its Pathogenicity to Ostrinia furnacalis Larvae[J]. Biotechnology Bulletin, 2025, 41(7): 312-325.

图/表 11

表1 序列扩增引物

Table 1 Primers for sequence amplification

引物名称 Primer name	序列 Sequence （5′-3′）	片段大小 Fragment size （bp）
dsRNA1-F1	CCTTCCTCCGGTCACTGAACC	1 419
dsRNA1-F2	GGTGGTGCTAAAGCTAAGGTGTCG	1 419
dsRNA1-R1	CGCGCTCAGTCTTGCCAT	1 085
dsRNA1-R2	GTCAACAGACCTACAACCGACACC	1 085
dsRNA2-F1	GCGCAGGTGGGAGGACA	1 019
dsRNA2-F2	ATCCAGCCATCATCGCGC	1 019
dsRNA2-R1	CCTGACCGAACAAATGCTCACC	935
dsRNA2-R2	CCATCGGTGCGGACGG	935

图1 球孢白僵菌BbOCuV1和BbOCuV1-F菌株dsRNA 提取及 RT-PCR 检测结果A：各泳道为BbOCuV1菌株dsRNA提取检测结果，“+”为DNase I和S1核酸酶处理的dsRNA；“-”为未经过酶处理的dsRNA。B：各泳道为BbOCuV1菌株RT-PCR检测；“+”为RT-PCR检测阳性对照；“-”为 RT-PCR检测阴性对照

Fig. 1 dsRNA extraction and RT-PCR detection of Beauveria bassiana strain BbOCuV1A: Each lane indicates the test result of dsRNA extraction in strain BbOCuV1; "+" is dsRNA treated with DNase I and S1 nuclease; "-" is the dsRNA without enzymatic treatment. B: Each lane indicates the RT-PCR detection for strain BbOCuV1; "+" is a positive control for RT-PCR; and "-" is a negative control for RT-PCR and BbOCuV1-F

图2 球孢白僵菌BbOFDHOCuV和BbOFDH1-5-GFP菌株dsRNA提取及RT-PCR检测结果A：BbOFDHOCuV菌株dsRNA提取检测结果；“+”为DNase I和S1核酸酶处理的dsRNA；“-”为未经过酶处理的dsRNA。B：BbOFDHOCuV菌株RT-PCR检测；“+”为RT-PCR检测阳性对照；“-”为 RT-PCR检测阴性对照

Fig. 2 dsRNA extraction and RT-PCR detection of B. bassiana strain BbOFDHOCuV and BbOFDH1-5-GFPA: dsRNA extraction test result in strain BbOFDHOCuV; "+"is dsRNA treated with DNase I and S1 nuclease; "-" is the dsRNA without enzymatic treatment. B: RT-PCR detection for strain BbOFDHOCuV; "+" is a positive control for RT-PCR, and "-" is a negative control for RT-PCR

图3 球孢白僵菌接种在PDA培养基上的菌落形态及生长速度A：BbOFDHOCuV和BbOFDH菌株菌落形态；B：BbOCuV1和BbOCuV1-F菌株菌落形态；C：BbOFDHOCuV和BbOFDH菌株生长速度；D：BbOCuV1和BbOCuV1-F菌株生长速度；**P＜0.01；***P＜0.001，下同

Fig. 3 Colony morphologies and growth rates of B. bassiana inoculated on PDA mediumA: Colony morphology of strain BbOFDHOCuV and BbOFDH; B: colony morphology of strain BbOCuV1and BbOCuV1-F strains; C: growth rates of BbOFDHOCuV and BbOFDH strains; D: growth rates of BbOCuV1 and BbOCuV1-F strains. ** indicates P＜0.01; *** indicates P＜0.001, the same below

图4 球孢白僵菌菌丝尖端形态

Fig. 4 Morphology of mycelium tip of B. bassiana

图5 菌株单位面积产孢量A：获毒菌株单位面积产孢量；B：脱毒菌株单位面积产孢量

Fig. 5 Strain's sporulation yield per unit areaA: Sporulation yield per unit area of the toxified strain. B: Sporulation yield per unit area of the detoxified strain

图6 球孢白僵菌菌株生物量的测定A：获毒菌株生物量测定；B：脱毒菌株生物量测定

Fig. 6 Determination of the biomass of B. bassiana strainsA: Biomass measurement of the toxified strains. B: Biomass measurement of detoxified strains

图7 2龄亚洲玉米螟8 d内存活率A：获毒菌株2龄亚洲玉米螟8 d内存活率；B：脱毒菌株2龄亚洲玉米螟8 d内存活率；*P<0.05

Fig. 7 Survival rates of 2-instar O. furnacalis within 8 dA: Survival rate of 2-instar Ostrinia furnacalis larvae infected with toxified strains B: Survival rate of 2-instar O. furnacalis larvae infected with detoxified strains;*P<0.05

图8 RNA-Seq数据的转录组谱A：无病毒组和BbOCuV1组的主成分分析；B：使用log2倍变化和log10p值绘制RNA-Seq数据的火山图。红点和绿点分别表示上调和下调的基因，蓝点表示没有显著表达的基因

Fig. 8 Transcriptome profile of RNA-Seq dataA: Principal component analysis of virus-free and BbOCuV1 groups; B: Volcano plot of RNA-Seq data using log2 fold change and log10p-value. Red and green dots denote up- and down-regulated genes, respectively, and blue dots indicate genes with no significant expression

图9 DEGs的GO和KEGG富集分析A：上调基因的GO富集分析；B：下调基因的GO富集分析；C：上调基因的KEGG富集分析；D：下调基因的KEGG富集分析

Fig. 9 GO and KEGG enrichment analyses of DEGsA: GO enrichment analysis of upregulated genes. B: GO enrichment analysis of downregulated genes. C: KEGG enrichment analysis of upregulated genes. D: KEGG enrichment analysis of downregulated genes

表2 差异基因Padj值

Table 2 Padj values of differential genes

Gene ID	Padj
BBA_00890	5.05031204527094e-37
BBA_01635	5.19645125134483e-05
BBA_02155	7.19194419245941e-06
BBA_01808	4.57389790420523e-12
BBA_02822	1.45371968945862e-07
BBA_04028	0.00497034986646022
BBA_08424	2.58045865685601e-06
BBA_08699	0.00121775638944784
BBA_03616	0.00170577409824362
BBA_04374	8.74503508574553e-20
BBA_07338	1.08247806189268e-12
BBA_09043	5.05031204527094e-37
BBA_00611	4.4575588085647e-05
BBA_00635	0.000102945242727946
BBA_00702	1.47055069946437e-23
BBA_02264	0.00174335718176381
BBA_09043	5.05031204527094e-37

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