转地蜂群病原微生物及肠道共生菌的变化

doi:10.3864/j.issn.0578-1752.2017.05.018

摘要/Abstract

摘要： 【目的】探明转地放蜂过程中常见蜜蜂病毒和寄生虫的流行规律，及不同地区工蜂肠道中两种主要共生菌Gilliamella apicola和Snodgrassella alvi的变化情况。【方法】在转地放蜂过程中，对同一意大利蜜蜂(Apis mellifera ligustica）蜂场的固定蜂群连续取样，采用RT-PCR方法检测蜂群中病毒和寄生虫的感染情况，使用SPSS 17.0软件对不同地区、不同季节样本的病毒和寄生虫感染率进行卡方检验。以蜜蜂β-actin 为内参基因，对不同地区样本中的共生菌G. apicola 和S. alvi进行荧光定量PCR分析，检测转地过程中工蜂肠道中这两种细菌的变化情况，并采用Kendall Rank相关系数对病原物感染率和共生菌含量进行相关性分析。【结果】转地放蜂7个地区的样本中，仅检测出以色列急性麻痹病毒（IAPV）、黑蜂王台病毒（BQCV）和蜜蜂残翅病毒（DWV）3种蜜蜂病毒。其中IAPV和BQCV在所有地区均有检出且感染率较高，不同地区之间感染率差异显著；DWV感染率相对较低，不同地区之间感染率差异极显著。西方蜜蜂微孢子虫（Nosema apis）在各地区样本中均未检出，东方蜜蜂微孢子虫（Nosema ceranae）在4个地区的样本中检出，且不同地区间感染率差异极显著，熊蜂微孢子虫（Nosema bombi）在各个地区均有检出，不同地区间感染率差异显著；季节性差异分析表明，IAPV在不同季节的感染率差异不显著，而BQCV、DWV、N. ceranae和N. bombi在不同季节的感染率差异显著，且春夏季的感染率普遍高于秋冬季；不同转地地区的工蜂肠道内均含有共生菌G. apicola和S. alvi, 且两种共生菌含量在不同地区间均差异极显著；相关性分析表明，S. alvi和IAPV之间呈显著负相关作用。【结论】转地蜂场工蜂病原微生物的检测结果表明IAPV、BQCV、DWV和微孢子虫在蜂群中普遍存在；蜜蜂病原物的感染率和肠道共生菌的含量在不同地理区域间差异显著；部分病原物与肠道共生菌间呈显著负相关关系；转地放蜂方式对蜜蜂的健康状况有一定影响。

关键词: 转地放蜂, 意大利蜜蜂, 病原微生物, 共生菌, 荧光定量PCR

Abstract: 【Objective】The objectives of this study are to examine the occurrence and prevalence of viruses and parasites in migratory honeybees, Apis mellifera ligustica, and to analyze the variation of two main symbiotic bacteria Gilliamella apicola and Snodgrassella alvi.【Method】The virus and parasite infections were detected by RT-PCR in the same colony of A. m. ligustica at different migratory locations. The infection rate of virus and parasite in different regions and seasons were analyzed by chi-square test. The honey bee β-actin gene was selected as the reference gene, the quantitative real-time PCR (qPCR) was used to explore the quantity variation of symbiotic bacteria G. apicola and S. alvi in the different regions, and the Kendall Rank correlation coefficient was used to analyze the correlation between the rate of pathogen infection and the quantity of symbiotic bacteria.【Result】In the seven regions, three viruses were found: Israeli acute paralysis virus (IAPV), black queen cell virus (BQCV), and deformed wing virus (DWV). The infection rates of IAPV and BQCV were higher in all regions, and the difference was significant among different regions. DWV infection rate was relatively low, and the infection rate among different regions was extremely significant. Nosema apis was not detected in all the samples. N. ceranae was detected in four regions and the infection rate was extremely significant among different regions. N. bombi was detected in all regions and the infection rate among different regions was significant. The results showed that the infection rates of IAPV was not significantly different in different seasons, but the DWV, BQCV, N. bombi and N. ceranae infection rates had significant differences in different seasons, and the infection rate in spring and summer was significantly higher than that in autumn and winter. The qPCR result showed that G. apicola and S. alvi were detected in all migratory beekeeping periods, and the difference of the two symbiotic bacteria in different regions was extremely significant. There was a significant negative correlation between the prevalence of S. alvi and IAPV. 【Conclusion】The results showed that IAPV, BQCV, DWV and microsporidia are prevalent in the migratory colonies. The infection rate of bee pathogens and the quantity of symbiotic bacteria are significantly different among different geographical areas; there are negative correlations between the incidence of some pathogens and the quantity of some symbiotic bacteria; migratory beekeeping has negative impacts on worker longevity and colony health.

Key words: migratory beekeeping, Apis melliferaligustica, pathogens, symbiotic bacteria, quantitative PCR

刘珊，王刘豪，郭军，李继莲，徐龙龙. 转地蜂群病原微生物及肠道共生菌的变化[J]. 中国农业科学, 2017, 50(5): 951-958.

LIU Shan, WANG LiuHao, GUO Jun, LI JiLian, XU LongLong. The Variation of Pathogens, Parasites and Symbionts in Migratory Honeybees (Apis mellifera ligustica)[J]. Scientia Agricultura Sinica, 2017, 50(5): 951-958.

0
/ / 推荐

导出引用管理器 EndNote|Reference Manager|ProCite|BibTeX|RefWorks

链接本文: https://www.chinaagrisci.com/CN/10.3864/j.issn.0578-1752.2017.05.018

https://www.chinaagrisci.com/CN/Y2017/V50/I5/951

参考文献

[1] 李易谷. 转地蜂场蜂群损失分析. 蜜蜂杂志, 2013(11): 33.

Li Y G. Analysis on the loss of bee colony. Journal of Bee, 2013(11): 33. (in Chinese)

[2] 刁青云, 吴杰, 姜秋玲, 谢文闻. 中国蜂业现状及存在问题. 世界农业, 2008(10): 59-61.

Diao Q Y, Wu J, Jiang Q L, Xie W W. Current situation of Chinese apiculture. World Agriculture, 2008(10): 59-61. (in Chinese)

[3] Welch A, Drummond F, Tewari S, Averill A, Burand J P. Presence and prevalence of viruses in local and migratory honeybees (Apis mellifera) in Massachusetts. Applied and environmental microbiology, 2009, 75(24): 7862-7865.

[4] Moritz R F, Pirk C W, Hepburn H R, Neumann P. Short-sighted evolution of virulence in parasitic honeybee workers (Apis mellifera capensis Esch.). Naturwissenschaften, 2008, 95(6): 507-513.

[5] Dillon R J, Vennard C T, Buckling A, Charnley A K. Diversity of locust gut bacteria protects against pathogen invasion. Ecology Letters, 2005, 8(12): 1291-1298.

[6] Fraune S, Bosch T C. Why bacteria matter in animal development and evolution. Bioessays, 2010, 32(7): 571-580.

[7] Engel P, Martinson V G, Moran N A. Functional diversity within the simple gut microbiota of the honey bee. Proceedings of the National Academy of Sciences of the United States of America, 2012, 109(27): 11002-11007.

[8] Dainat B, Ken T, Berthoud H, Neumann P. The ectoparasitic mite Tropilaelaps mercedesae (Acari, Laelapidae) as a vector of honeybee viruses. Insectes sociaux, 2009, 56(1): 40-43.

[9] Eyer M, Chen Y P, Schäfer M O, Pettis J, Neumann P. Small hive beetle, Aethina tumida, as a potential biological vector of honeybee viruses. Apidologie, 2009, 40(4): 419-428.

[10] Boecking O, Genersch E. Varroosis–the ongoing crisis in bee keeping. Journal für Verbraucherschutz und Lebensmittelsicherheit, 2008, 3(2): 221-228.

[11] Sammataro D, Gerson U, Needham G. Parasitic mites of honey bees: life history, implications, and impact. Annual review of entomology, 2000, 45(1): 519-548.

[12] De Miranda J R, Genersch E. Deformed wing virus. Journal of invertebrate pathology, 2010, 103: S48-S61.

[13] Hails R S, Ball B V, Genersch E. Infection strategies of insect viruses//Virology and the Honey Bee. European Commission, 2008: 255-276.

[14] Genersch E, Von Der Ohe W, Kaatz H, Schroeder A, Otten C, Büchler R, Berg S, Ritter W, Mühlen W, Gisder S, Meixner M, Liebig G, Rosenkranz P. The German bee monitoring project: a long term study to understand periodically high winter losses of honey bee colonies. Apidologie, 2010, 41(3): 332-352.

[15] Kwong W K, Moran N A. Gut microbial communities of social bees. Nature Reviews Microbiology, 2016, 14(6): 374-384.

[16] Hamdi C, Balloi A, Essanaa J, Crotti E, Gonella E, Raddadi N, Ricci I, Boudabous A, Borin S, Manino A, Bandi C, Alma A, Daffonchio D, Cherif A. Gut microbiome dysbiosis and honeybee health. Journal of Applied Entomology, 2011, 135(7): 524-533.

[17] Martinson V G, Moy J, Moran N A. Establishment of characteristic gut bacteria during development of the honeybee worker. Applied and environmental microbiology, 2012, 78(8): 2830-2840.

[18] Li J, Qin H, Wu J, Sadd B M, Wang X, Evans J D, Peng W, Chen Y. The prevalence of parasites and pathogens in Asian honeybees Apis cerana in China. Plos one, 2012, 7(11): e47955.

[19] Ai H, Yan X, Han R. Occurrence and prevalence of seven bee viruses in Apis mellifera and Apis cerana apiaries in China. Journal of invertebrate pathology, 2012, 109(1): 160-164.

[20] Yang B, Peng G, Li T, Kadowaki T. Molecular and phylogenetic characterization of honey bee viruses, Nosema microsporidia, protozoan parasites, and parasitic mites in China. Ecology and evolution, 2013, 3(2): 298-311.

[21] 贾慧茹, 刘进祖, 王星, 吴艳艳, 周婷. 北京地区六种蜜蜂病毒病的流行病学研究. 应用昆虫学报, 2014, 51(3): 772-780.

Jia H R, Liu J Z, Wang X, Wu Y Y, Zhou T. Occurrence and prevalence of six bee viruses in Beijing. Chinese Journal of Applied Entomology, 2014, 51(3): 772-780. (in Chinese)

[22] Benjeddou M, Leat N, Allsopp M, Davison S. Detection of acute bee paralysis virus and black queen cell virus from honeybees by reverse transcriptase PCR. Applied and Environmental Microbiology, 2001, 67(5): 2384-2387.

[23] Ribière M, Triboulot C, Mathieu L, Aurières C, Faucon J P, Pépin M. Molecular diagnosis of chronic bee paralysis virus infection.Apidologie, 2002, 33(3): 339-351.

[24] Chen Y, Smith I B, Collins A M, Pettis J S, Feldlaufer M F. Detection of deformed wing virus infection in honey bees, Apis mellifera L., in the United States. American Bee Journal, 2004, 144(7): 557-559.

[25] Di Prisco G, Pennacchio F, Caprio E, Boncristiani Jr H F, Evans J D, Chen Y. Varroa destructor is an effective vector of Israeli acute paralysis virus in the honeybee, Apis mellifera. Journal of General Virology, 2011, 92(1): 151-155.

[26] Stoltz D, Shen X R, Boggis C, Sisson G. Molecular diagnosis of Kashmir bee virus infection. Journal of Apicultural Research, 1995, 34(3): 153-160.

[27] Chen Y, Pettis J S, Feldlaufer M F. Detection of multiple viruses in queens of the honey bee Apis mellifera L.. Journal of invertebrate pathology, 2005, 90(2): 118-121.

[28] Chen Y, Evans J D, Smith I B, Pettis J S. Nosema ceranae is a long-present and wide-spread microsporidian infection of the European honey bee (Apis mellifera) in the United States. Journal of invertebrate pathology, 2008, 97(2): 186-188.

[29] Chaimanee V, Chen Y, Pettis J S, Cornman R S, Chantawannakul P. Phylogenetic analysis of Nosema ceranae isolated from European and Asian honeybees in Northern Thailand. Journal of invertebrate pathology, 2011, 107(3): 229-233.

[30] Tay W T, O’MAHONY E M, Paxton R J. Complete rRNA gene sequences reveal that the microsporidium Nosema bombi infects diverse bumblebee (Bombus spp.) hosts and contains multiple polymorphic sites. Journal of Eukaryotic Microbiology, 2005, 52(6): 505-513.

[31] Chen Y P, Higgins J A, Feldlaufer M F. Quantitative real-time reverse transcription-PCR analysis of deformed wing virus infection in the honeybee (Apis mellifera L.). Applied and Environmental Microbiology, 2005, 71(1): 436-441.

[32] 徐龙龙, 吴杰, 郭军, 李继莲. 共生菌群在熊蜂生长发育过程中的动态变化. 中国农业科学, 2014, 47(10): 2030-2037.

Xu L L, Wu J, Guo J, Li J L. Dynamic variation of symbionts in bumblebees during hosts growth and development. Scientia Agricultura Sinica, 2014, 47(10): 2030-2037. (in Chinese)

[33] Cox-Foster D L, Conlan S, Holmes E C, Palacios G, Evans J D, Moran N A, Quan P L, Briese T, Hornig M, Geiser D M, Martinson V, vanEngelsdorp D, Kalkstein A L, Drysdale A, Hui J, Zhai J H, Cui L W, Hutchison S K, Simons J F, Egholm M, Pettis J S, Lipkin W I. A metagenomic survey of microbes in honey bee colony collapse disorder. Science, 2007, 318(5848): 283-287.

[34] 李志国. 蜜蜂以色列急性麻痹病毒的分子检测、流行传播及其对蜜蜂行为的影响[D]. 杭州: 浙江大学, 2014.

Li Z G. Molecular detection, prevalence and transmission of Israeli acute paralysis virus and its effects on honey bee behaviors[D]. Hangzhou: Zhejiang University, 2014. (in Chinese)

[35] Ribière M, Ball B, Aubert M. Natural history and geographical distribution of honey bee viruses//Virology and the honey bee. European Commission, 2008: 15-84.

[36] Allen M, Ball B. The incidence and world distribution of honey bee viruses. Bee world, 1996, 77(3): 141-162.

[37] Shen M, Yang X, Cox-Foster D, Cui L. The role of varroa mites in infections of Kashmir bee virus (KBV) and deformed wing virus (DWV) in honey bees. Virology, 2005, 342(1): 141-149.

[38] Botías C, Martín-Hernández R, Garrido-Bailón E, González-Porto A, Martínez-Salvador A, De La Rúa P, Meana A, Higes M. The growing prevalence of Nosema ceranae in honey bees in Spain, an emerging problem for the last decade. Research in veterinary science, 2012, 93(1): 150-155.

[39] Maiolino P, Iafigliola L, Rinaldi L, De Leva G, Restucci B, Martano M. Histopathological findings of the midgut in European honey bee (Apis mellifera L.) naturally infected by Nosema spp. Veterinary Medicine and Animal Sciences, 2014, 2(1): Article 4.

[40] McIvor C A, Malone L A. Nosema bombi, a microsporidian pathogen of the bumble bee Bombus terrestris (L.). New Zealand Journal of Zoology, 1995, 22(1): 25-31.

[41] 郭军. 蜜蜂肠道菌群多样性及其影响因素研究[D]. 北京: 中国农业科学院 2015.

Guo J. Diversity and influencing factors of gut microbiota in honey bees[D]. Beijing: Chinese academy of agricultural sciences, 2015. (in Chinese)

[42] Koch H, Cisarovsky G, Schmid-Hempel P. Ecological effects on gut bacterial communities in wild bumblebee colonies. Journal of Animal Ecology, 2012, 81(6): 1202-1210.

[43] 吴杰. 蜜蜂学. 北京: 中国农业出版社, 2012.

Wu J. Apiology. Beijing: China Agriculture Press, 2012. (in Chinese)