球囊菌黑色素样色素的亚细胞定位及感染功能初探  

Subcellular Localization of Melanin-Like Pigment on the Spore Wall of Pathogenic Fungus Ascosphaera apis and Its Role in Infection

作　　者：衡惠 何彪 王悦弟 董玉萍 李治[1] HENG Hui;HE Biao;WANG Yuedi;DONG Yuping;LI Zhi(Collage of Life sciences,Chongqing Normal University,Chongqing 401331,China)

机构地区：[1]重庆师范大学生命科学学院,重庆401331

出　　处：《重庆师范大学学报（自然科学版）》2020年第2期120-125,F0003,共7页Journal of Chongqing Normal University:Natural Science

基　　金：现代农业产业技术体系建设专项资金(No.CARS-44-KXJ21);国家自然科学基金(No.31101770,No.31302037);重庆市教育委员会科学技术研究项目(No.KJ1703051);重庆市基础科学与前沿技术研究项目(No.cstc2014jcyjA80039,No.cstc2015jcyj A80014,No.cstc2017jcyjAX0184);重庆师范大学自然科学基金(No.13XLB009)。

摘　　要：【目的】鉴定黑色素样色素在球囊菌(Ascosphaera apis)孢子的定位情况,分析它在球囊菌感染意大利蜜蜂(Apis mellifera)(以下简称意蜂)幼虫中的作用。【方法】以黑色素提取法提取和纯化球囊菌孢子的黑色素样色素,制备多克隆抗血清。通过间接免疫荧光定位技术分析类黑色素在孢子中的定位特征。分别以含有孢子、黑色素样色素、孢子及黑色素样色素、磷酸缓冲液的饲料喂食意蜂3龄幼虫,统计幼虫存活率,探讨黑色素样色素在孢子感染幼虫中的作用。【结果】经分离和纯化得到了球囊菌黑色素样色素,并将它注射入小鼠(Mus musculus)体内后成功制备了多克隆抗血清。间接免疫荧光亚细胞定位结果显示黑色素样色素主要分布在球囊菌孢子壁上。感染分析结果表明:与正常摄食对照组和磷酸缓冲液对照组相比,添食球囊菌孢子使意蜂幼虫存活率有统计学意义上的下降(p<0.05),单独添食黑色素样色素对意蜂幼虫存活率没有统计学意义上的影响;较单独添食孢子处理而言,同时添食球囊菌孢子和黑色素样色素使意蜂幼虫存活率有统计学意义上的下降(p<0.05)。【结论】球囊菌黑色素样色素定位于球囊菌孢子的细胞壁上,潜在的参与了球囊菌孢子对蜜蜂幼虫的感染。[Purposes]This study aims to characterize the subcellular localization of melanin-like pigment from Ascosphaera apis,to analyze its role in A.apisinfecting the honeybee Apis melliferalarvae.[Methods]The melanin-like pigment was isolated and purified from A.apis spores using the melanin extraction method.The polyclonal antiserum against melanin-like was produced through immunized mice.The indirect immunofluorescence localization technique was used to analyze the subcellular localization characteristics of melanin-like in the spores.The mortality rate of A.mellifera was statistical analysis after 3rd instar larvae fed with spores,melanin-like,spore-melanin-like,and phosphate buffer solution(PBS)respectively,for exploring the role of melanin-like pigment in spore infecting the bee larvae.[Findings]Melanin-like pigment fromA.apis spore was isolated and purified,and polyclonal antiserum was successfully produced after melanin-like was injected into mice(Mus musculus).Indirect immunofluorescence subcellular localization showed that melanin-like was mainly located on the spore wall of A.apis.Compared to the normal feeding control group and PBS control group,the infection analysis showed that the survival of bee larvae significantly(p<0.05)decreased after fed with spores,and there was no significant different after fed with the melanin-like.Compared with the group fed only spores,feeding with both spores and melanin-like decreased the survival of bee larvae significantly(p<0.05).[Conclusions]The melanin-like pigment is located on the spore wall of A.apis,potentially plays an important role in spore infecting the bee larvae.

关 键 词：黑色素 球囊菌 亚细胞定位 感染 

分 类 号：Q93[生物学—微生物学]