Erban Tomas, Ledvinka Ondrej, Kamler Martin, Hortova Bronislava, Nesvorna Marta, Tyl Jan, Titera Dalibor, Markovic Martin, Hubert Jan
Crop Research Institute, Prague, Czechia.
Czech Hydrometeorological Institute, Prague, Czechia.
PeerJ. 2017 Sep 25;5:e3816. doi: 10.7717/peerj.3816. eCollection 2017.
is an entomopathogenic bacterium that causes European foulbrood (EFB), a honeybee ( L.) disease that necessitates quarantine in some countries. In Czechia, positive evidence of EFB was absent for almost 40 years, until an outbreak in the Krkonose Mountains National Park in 2015. This occurrence of EFB gave us the opportunity to study the epizootiology of EFB by focusing on the microbiome of honeybee workers, which act as vectors of honeybee diseases within and between colonies.
The study included worker bees collected from brood combs of colonies (i) with no signs of EFB (EFB0), (ii) without clinical symptoms but located at an apiary showing clinical signs of EFB (EFB1), and (iii) with clinical symptoms of EFB (EFB2). In total, 49 samples from 27 honeybee colonies were included in the dataset evaluated in this study. Each biological sample consisted of 10 surface-sterilized worker bees processed for DNA extraction. All subjects were analyzed using conventional PCR and by metabarcoding analysis based on the 16S rRNA gene V1-V3 region, as performed through Illumina MiSeq amplicon sequencing.
The bees from EFB2 colonies with clinical symptoms exhibited a 75-fold-higher incidence of than those from EFB1 asymptomatic colonies. was identified in all EFB1 colonies as well as in some of the control colonies. The proportions of , , , , and were higher in EFB2 than in EFB1, whereas was significantly higher in EFB2 than in EFB0. and , and, exhibited higher proportion in EFB1 than in EFB2 and EFB0. The occurrence of and were higher in EFB0 than in EFB2 and EFB1. incidence was highest in EFB2.
High-throughput Illumina sequencing permitted a semi-quantitative analysis of the presence of within the honeybee worker microbiome. The results of this study indicate that worker bees from EFB-diseased colonies are capable of transmitting due to the greatly increased incidence of the pathogen. The presence of sequences in control colonies supports the hypothesis that this pathogen exists in an enzootic state. The bacterial groups synergic to both the colonies with clinical signs of EFB and the EFB-asymptomatic colonies could be candidates for probiotics. This study confirms that is a secondary invader to ; however, other putative secondary invaders were not identified in this study.
是一种昆虫病原细菌，可引发欧洲幼虫腐臭病（EFB），这是一种蜜蜂（西方蜜蜂）疾病，在一些国家需要进行隔离。在捷克，近40年来一直没有EFB的阳性证据，直到2015年在克尔科诺谢山国家公园爆发疫情。EFB的此次出现让我们有机会通过关注蜜蜂工蜂的微生物群来研究EFB的流行病学，蜜蜂工蜂是蜂群内部和之间蜜蜂疾病的传播媒介。
该研究包括从蜂群的巢脾中采集的工蜂，（i）无EFB迹象（EFB0），（ii）无临床症状但位于有EFB临床症状的蜂场（EFB1），以及（iii）有EFB临床症状（EFB2）。本研究评估的数据集中总共包括来自27个蜂群的49个样本。每个生物样本由10只经过表面消毒的工蜂组成，用于DNA提取。所有样本均通过常规PCR以及基于16S rRNA基因V1 - V3区域的元条形码分析进行分析，通过Illumina MiSeq扩增子测序完成。
有临床症状的EFB2蜂群中的蜜蜂携带该菌的发生率比无症状的EFB1蜂群中的蜜蜂高75倍。在所有EFB1蜂群以及一些对照蜂群中都检测到了该菌。EFB2中[具体细菌种类1]、[具体细菌种类2]、[具体细菌种类3]、[具体细菌种类4]和[具体细菌种类5]的比例高于EFB1，而[具体细菌种类6]在EFB2中显著高于EFB0。[具体细菌种类7]和[具体细菌种类8]、[具体细菌种类9]在EFB1中的比例高于EFB2和EFB0。[具体细菌种类10]和[具体细菌种类11]的发生率在EFB0中高于EFB2和EFB1。[具体细菌种类12]的发生率在EFB2中最高。
高通量Illumina测序允许对蜜蜂工蜂微生物群中该菌的存在进行半定量分析。本研究结果表明，来自患有EFB疾病蜂群的工蜂由于病原体发生率大幅增加而能够传播该菌。对照蜂群中该菌序列的存在支持了这种病原体以地方流行状态存在的假设。与有EFB临床症状的蜂群和无症状的EFB蜂群协同作用的细菌类群可能是益生菌的候选者。本研究证实该菌是[另一种细菌名称]的继发性入侵者；然而，本研究未鉴定出其他假定的继发性入侵者。
