中华蜜蜂和意大利蜜蜂工蜂幼虫肠道中蜜蜂球囊菌ass-milR0037-3p的调控作用与表达模式

doi:10.16380/j.kcxb.2025.09.009

昆虫学报 ›› 2025, Vol. 68 ›› Issue (9): 1251-1260.doi: 10.16380/j.kcxb.2025.09.009

中华蜜蜂和意大利蜜蜂工蜂幼虫肠道中蜜蜂球囊菌ass-milR0037-3p的调控作用与表达模式

叶道有^1,#, 臧贺^1,2,3,#, 王梦怡¹, 樊念¹, 吴陶¹, 郑可心¹, 严提珍^2,4, 卢兆辉⁴, 谢润桂⁴, 陈大福^1,2,3, 郭睿^1,2,3,*, 邱剑丰^1,2,3,*

(1. 福建农林大学蜂学与生物医药学院, 福州 350002; 2. 天然生物毒素国家地方联合工程实验室, 福州 350002; 3. 福建农林大学蜂疗研究所, 福州 350002; 4. 东莞市妇幼保健院, 东莞 523000)

出版日期:2024-09-20 发布日期:2025-10-28

Regulatory role and expression pattern of Ascosphaera apis ass-milR0037-3p in the guts of Apis cerana cerana and Apis mellifera ligustica worker larvae

YE Dao-You^1,#, ZANG He^1,2,3,#, WANG Meng-Yi¹, FAN Nian¹, WU Tao¹, ZHENG Ke-Xin¹, YAN Ti-Zhen^2,4, LU Zhao-Hui⁴, XIE Run-Gui⁴, CHEN Da^Fu^1,2,3, GUO Rui^1,2,3,*, QIU Jian-Feng^1,2,3,*

(1. College of Bee Science and Biomedicine, Fujian Agriculture and Forestry University, Fuzhou 350002, China; 2. National & Local United Engineering Laboratory of Natural Biotoxin, Fuzhou 350002, China; 3. Apitherapy Research Institute of Fujian Agriculture and Forestry University, Fuzhou 350002, China; 4. Dongguan Maternal and Child Health Care Hospital, Dongguan 523000, China)

Online:2024-09-20 Published:2025-10-28

摘要/Abstract

摘要： 【目的】本研究旨在检测ass-milR0037-3p及其关键靶基因在蜜蜂球囊菌Ascosphaera apis侵染中华蜜蜂Apis cerana cerana和意大利蜜蜂A. mellifera ligustica工蜂幼虫过程中的表达模式，为深入探究ass-milR0037-3p调控蜜蜂球囊菌侵染机制提供基础。【方法】使用相关软件预测蜜蜂球囊菌ass-milR0037-3p的靶基因并进行GO和KEGG数据库注释。中华蜜蜂和意大利蜜蜂3日龄工蜂幼虫饲喂含1×10⁷孢子/mL蜜蜂球囊菌的饲料后采用stem-loop RT-PCR和RT-qPCR分别验证中华蜜蜂和意大利蜜蜂工蜂6日龄幼虫肠道中ass-milR0037-3p的表达和检测中华蜜蜂和意大利蜜蜂工蜂4-6日龄幼虫肠道中ass-milR0037-3p及其2个关键靶基因组蛋白乙酰转移酶(histone acetyltransferase)基因ESA1和黄素胺氧化还原酶(flavin containing amine oxidase)基因FAO的表达量。【结果】ass-milR0037-3p靶向225个基因，可注释到繁殖、结合和细胞等28个GO条目以及剪接体、次生代谢产物的生物合成及氨基糖和核苷酸糖代谢等105条KEGG通路。蜜蜂球囊菌接种后，中华蜜蜂和意大利蜜蜂工蜂6日龄幼虫肠道中均扩增出ass-milR0037-3p的目的片段；相较于中华蜜蜂4日龄工蜂幼虫，5和6日龄工蜂幼虫肠道中ass-milR0037-3p的表达量均显著上调并且ESA1和FAO的表达量均显著下调；意大利蜜蜂5日龄工蜂幼虫肠道中ass-milR0037-3p的表达量比4日龄工蜂幼虫肠道中的上调，6日龄工蜂幼虫肠道中的比4日龄工蜂幼虫肠道中的显著上调；靶基因ESA1和FAO的表达量在意大利蜜蜂5日龄工蜂幼虫肠道中均比4日龄工蜂幼虫肠道中的上调，在6日龄工蜂幼虫肠道中比4日龄工蜂幼虫肠道中的显著上调。【结论】ass-milR0037-3p通过负调控FAO表达潜在调节蜜蜂球囊菌侵染中华蜜蜂工蜂幼虫的过程，通过正调控ESA1表达潜在影响蜜蜂球囊菌侵染意大利蜜蜂工蜂幼虫的过程。研究结果为阐明milRNA通过调控靶基因的表达来响应蜜蜂球囊菌侵染蜜蜂幼虫肠道的机制提供了基础。

关键词: 中华蜜蜂, 意大利蜜蜂, 微小RNA, 类微小RNA的RNA, 组蛋白乙酰转移酶基因, 黄素胺氧化还原酶基因

Abstract: 【Aim】The purpose of this study is to detect the expression patterns of ass-milR0037-3p and its key target genes in worker larvae of Apis cerana cerana and A. mellifera ligustica in the process of Ascosphaera apis infection, so as to offer a foundation for further exploring the mechanism of ass-milR0037-3p regulating the As. apis infection. 【Methods】 The target genes of ass-milR0037-3p of As. apis were predicted using related software and then annotated to GO and KEGG databases. Stem-loop RT-PCR and RT-qPCR were employed to verify the expression of ass-milR0037-3p in the guts of the 6-day-old worker larvae of Ap. c. cerana and Ap. m. ligustica and to detect the expression levels of ass-milR0037-3p and its two key target genes (genomic protein acetyltransferase gene ESA1 and flavin containing amine oxidase gene FAO) in the guts of the 4-6-day-old worker larvae after feeding the 3-day-old worker larvae of Ap. c. cerana and Ap. m. ligustica with diets containing 1×10⁷ spores/mL As. apis, respectively.【Results】 ass-milR0037-3p could target 225 genes annotated to 28 GO terms such as reproduction, binding and cells, as well as 105 KEGG pathways such as splicing, biosynthesis of secondary metabolites, and amino sugar and nucleotide sugar metabolism. The target fragments of ass-milR0037-3p were amplified in the guts of the 6-day-old worker larvae of both Ap. c. cerana and Ap. m. ligustica infected with As. apis. The expression levels of ass-milR0037-3p in the guts of the 5- and 6-day-old worker larvae of Ap. c. cerana infected with As. apis were significantly up-regulated and the expression levels of ESA1 and FAO in the guts of the 5- and 6-day-old worker larvae of Ap. c. cerana infected with As. apis were significantly down-regulated, as compared with those of the 4-day-old worker larvae of Ap. c. cerana infected with As. apis. The expression level of ass-milR0037-3p in the gut of the 5-day-old worker larvae of Ap. m. ligustica infected with As. apis was up-regulated as compared with that of the 4-day-old worker larvae of Ap. m. ligustica infected with As. apis and that of the 6-day-old worker larvae of Ap. c. cerana infected with As. apis was significantly up-regulated as compared with that of the 4-day-old worker larvae of Ap. m. ligustica infected with As. apis. The expression levels of the target genes ESA1 and FAO in the gut of the 5-day-old worker larvae of Ap. m. ligustica infected with As. apis were up-regulated and those of the 6-day-old worker larvae of Ap. m. ligustica infected with As. apis were significantly up-regulated as compared with those of the 4-day-old worker larvae of Ap. m. ligustica infected with As. apis.【Conclusion】 ass-milR0037-3p potentially modulates the process of As. apis infecting worker larvae of Ap. c. cerana by negatively regulating the FAO expression, while potentially affects the process of As. apis infecting worker larvae of Ap. m. ligustica through positively regulating the ESA1 expression. The results provide a basis for elucidating the mechanism by which milRNAs respond to the As. apis infection of honeybee larval guts via regulating the expression of target genes.

Key words: Apis cerana cerana, Apis mellifera ligustica, microRNA, microRNA-like RNA, histone acetyltransferase gene, flavin-containing amine oxidase gene

叶道有, 臧贺, 王梦怡, 樊念, 吴陶, 郑可心, 严提珍, 卢兆辉, 谢润桂, 陈大福, 郭睿, 邱剑丰. 中华蜜蜂和意大利蜜蜂工蜂幼虫肠道中蜜蜂球囊菌ass-milR0037-3p的调控作用与表达模式[J]. 昆虫学报, 2025, 68(9): 1251-1260.

YE Dao-You, ZANG He, WANG Meng-Yi, FAN Nian, WU Tao, ZHENG Ke-Xin, YAN Ti-Zhen, LU Zhao-Hui, XIE Run-Gui, CHEN Da-Fu, GUO Rui, QIU Jian-Feng. Regulatory role and expression pattern of Ascosphaera apis ass-milR0037-3p in the guts of Apis cerana cerana and Apis mellifera ligustica worker larvae[J]. Acta Entomologica Sinica, 2025, 68(9): 1251-1260.

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中华蜜蜂和意大利蜜蜂工蜂幼虫肠道中蜜蜂球囊菌ass-milR0037-3p的调控作用与表达模式

Regulatory role and expression pattern of Ascosphaera apis ass-milR0037-3p in the guts of Apis cerana cerana and Apis mellifera ligustica worker larvae

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