狄斯瓦螨和蜜蜂残翅病毒对蜜蜂健康的协同影响

世界各地大范围的西方蜜蜂Apis mellifera蜂群损失现象已引起科学界和公众的持续关注。狄斯瓦螨Varroa destructor和蜜蜂残翅病毒(Deformed wing virus,DWV)是西方蜜蜂群中最主要的两大生物威胁。尽管二者侵害蜜蜂均已有较长历史,但直至近十年来的研究才发现两者间的协同效应对蜂群健康的影响远超过其单独作用时所造成的危害:(1)蜜蜂残翅病毒可在狄斯瓦螨体内大量复制,继而进一步传播;(2)狄斯瓦螨的刺吸行为使病毒粒子跨越寄主的生理屏障而直接进入蜜蜂血淋巴;(3)狄斯瓦螨的寄生促使蜜蜂残翅病毒的高毒力毒株在蜂群中优势扩增和盛行;(4)狄斯瓦螨影响蜜蜂个体发育与免疫系统等生理机能,以致降低了蜂群对病毒的抵抗力;(5)蜜蜂残翅病毒对宿主造成的免疫抑制有利于狄斯瓦螨的寄生与繁殖。狄斯瓦螨、蜜蜂残翅病毒和西方蜜蜂间的关系已经成为昆虫外寄生物、病原体与寄主相互作用研究的一个典型模型。本文对近十年该领域的相关研究进行综述,以期为蜂群损失的原因调查以及昆虫寄生虫、病原微生物与寄主间关系的研究提供参考和借鉴。     

Detection of Deformed wing virus, a honey bee viral pathogen, in bumble bees (Bombus terrestris and Bombus pascuorum) with wing deformities

Honey bees (Apis mellifera) productively infected with Deformed wing virus (DWV) through Varroa destructor (V. destructor) during pupal stages develop into adults showing wing and other morphological deformities. Here, we report for the first time the occurrence of bumble bees (Bombus terrestris, Bombus pascuorum) exhibiting wing deformities resembling those seen in clinically DWV-infected honey bees. Using specific RT-PCR protocols for the detection of DWV followed by sequencing of the PCR products we could demonstrate that the bumble bees were indeed infected with DWV. Since such deformed bumble bees are not viable DWV infection may pose a serious threat to bumble bee populations.     

First come,first served: superinfection exclusion in Deformed wing virus is dependent upon sequence identity and not the order of virus acquisition

Deformed wing virus (DWV) is the most important globally distributed pathogen of honey bees and, when vectored by the ectoparasite Varroa destructor, is associated with high levels of colony losses. Divergent DWV types may differ in their pathogenicity and are reported to exhibit superinfection exclusion upon sequential infections, an inevitability in a Varroa-infested colony. We used a reverse genetic approach to investigate competition and interactions between genetically distinct or related virus strains, analysing viral load over time, tissue distribution with reporter gene-expressing viruses and recombination between virus variants. Transient competition occurred irrespective of the order of virus acquisition, indicating no directionality or dominance. Over longer periods, the ability to compete with a pre-existing infection correlated with the genetic divergence of the inoculae. Genetic recombination was observed throughout the DWV genome with recombinants accounting for ~2% of the population as determined by deep sequencing. We propose that superinfection exclusion, if it occurs at all, is a consequence of a cross-reactive RNAi response to the viruses involved, explaining the lack of dominance of one virus type over another. A better understanding of the consequences of dual- and superinfection will inform development of cross-protective honey bee vaccines and landscape-scale DWV transmission and evolution.Subject terms: Viral genetics, Microbial ecology, Virus-host interactions, Small RNAs     

Deformed wing virus in western honey bees (Apis mellifera) from Atlantic Canada and the first description of an overtly-infected emerging queen

Deformed wing virus (DWV) in western honey bees (Apis mellifera) often remains asymptomatic in workers and drones, and symptoms have never been described from queens. However, intense infections linked to parasitism by the mite Varroa destructor can cause worker wing deformity and death within 67 h of emergence. Ten workers (eight with deformed wings and two with normal wings) and three drones (two with deformed wings and one with normal wings) from two colonies infected with V. destructor from Nova Scotia, Canada, and two newly-emerged queens (one with deformed wings and one with normal wings) from two colonies infected with V. destructor from Prince Edward Island, Canada, were genetically analyzed for DWV. We detected DWV in all workers and drones, regardless of wing morphology, but only in the deformed-winged queen. This is the first report of DWV from Atlantic Canada and the first detection of a symptomatic queen with DWV from anywhere.     

蜜蜂残翅病研究进展

蜜蜂残翅病病毒(Deformed Wing Virus, DWV)是蜜蜂残翅病的病原体,能够侵染各个发育阶段的蜜蜂,严重时导致蜂蛹发育缓慢,成蜂翅膀畸形、体型萎缩、行动乏力,蜂群早衰;不严重的隐形感染也会缩短蜜蜂寿命。DWV借助于狄斯瓦螨Varroa destructor在全球广泛传播,并产生DWV-B和DWV-C等变异体,它们的共同作用促使蜂群群势急剧衰弱,是导致"越冬蜂群的消失(OCL)"和"蜂群崩溃失调症(CCD)"的主要病因之一,给养蜂业造成巨大损失,而目前尚没有特异性的有效治疗药物。DWV的自身特征、与狄斯瓦螨的关系、以及单独或者和狄斯瓦螨共同作用对蜜蜂的影响等研究都是开发防治技术方法的基础,本文从生物学、流行学、病理学和防治技术等方面对残翅病病毒的最新研究成果进行了综述。     

New evidence that deformed wing virus and black queen cell virus are multi-host pathogens

The host-range breadth of pathogens can have important consequences for pathogens' long term evolution and virulence, and play critical roles in the emergence and spread of the new diseases. Black queen cell virus (BQCV) and Deformed wing virus (DWV) are the two most common and prevalent viruses in European honey bees, Apis mellifera. Here we provide the evidence that BQCV and DWV infect wild species of honey bees, Apis florea and Apis dorsata. Phylogenetic analyses suggest that these viruses might have moved from A. mellifera to wild bee species and that genetic relatedness as well as the geographical proximity of host species likely play an important role in host range of the viruses. The information obtained from this present study can have important implication for understanding the population structure of bee virus as well as host-virus interactions.     

Cross-species infection of deformed wing virus poses a new threat to pollinator conservation

The Deformed wing virus (family Iflaviridae, genus Iflavirus, DWV), one of the most prevalent and common viruses in honey bees, Apis mellifera L., is present in both laboratory-reared and wild populations of bumble bees, Bombus huntii Greene. Our studies showed that DWV infection spreads throughout the entire body of B. huntii and that the concentration of DWV is higher in workers than in males both collected in the field and reared in the laboratory, implying a possible association between the virus infection and foraging activities. Further results showed that gut tissue of B. huntii can support the replication of DWV, suggesting that B. huntii is a biological host for DWV, as are honey bees. Bumble bees and honey bees sometimes share nectar and pollen resources in the same field. The geographical proximity of two host species probably plays an important role in host range breadth of the virus.     

Detection of deformed wing virus in the greenhouse for possible horizontal transmission of virus in honey bee colony

The honey bee Apis mellifera L. is a crucial insect in the agricultural industry and natural ecosystem by being a major pollinator. Nevertheless, honey bee population has been recently facing a decline. Among the several factors responsible for this decline, deformed wing virus (DWV) is considered a primary cause that negatively affects honey bee health. DWV is a cosmopolitan honey bee pathogen and causes morphological disadvantages in individual honey bees and colony collapse. Regarding the horizontal transmission of DWV, in addition to Varroa destructor, a well-known major vector of DWV, flowers have recently been implied as a transmission route. Therefore, in this study, we detected DWV from various substances, including flowers, honey bee feces, pupa, larva, nurse bee, surface of nurse bee, pollen collected by forager bee, and forager bee samples in four strawberry greenhouses, which could suggest the potential for the horizontal transmission of DWV in the semi-field condition. We also detected DWV in pollen collected by DWV-negative forager bees, implying that flowers can serve as a potential source of virus infection. These findings suggest that the surrounding environment such as shared floral sources affects the spread of DWV.     

蜜蜂残翅病毒在中华蜜蜂及其蜂箱奇露尾甲的发生与进化分析

蜜蜂残翅病毒(Deformed wing virus,DWV)广泛存在于我国规模化养殖的蜂群中,具有明显的季节流行特征.近年来,DWV不断向野生授粉昆虫及其它昆虫传播,包括蜜蜂新的寄生性病害蜂箱奇露尾甲Aethina tumida.前期本实验室已鉴定来源于中华蜜蜂Apis cerana与意大利蜜蜂Apis mellifera的蜂箱奇露尾甲均感染了蜜蜂病毒,但未调查来源于同一群的蜜蜂与蜂箱奇露尾甲是否感染了同一病毒,且在进化上有无差异.本研究调查了来源于同一群的中蜂及蜂箱奇露尾甲感染病毒情况,发现只感染了蜜蜂残翅病毒A型(DWV-A),未发现其它病毒.对DWV-A的3个片段进化分析表明,虽然DWV-A同时感染了中华蜜蜂和蜂箱奇露尾甲,但是VP3基因在不同的寄主中呈现明显不同的亲缘关系.这一结果为蜜蜂残翅病毒在蜜蜂及寄生病害中的进化提供了数据,为进一步研究病毒在不同寄主间的传播与进化奠定了基础.     

应用等温环介导扩增方法检测蜜蜂残翅病毒的研究

本文的目的在于建立用于临床检测残翅病病毒(Deformed wing virus,DWV)的等温环介导扩增技术(Loopmediated isothermal amplification,LAMP),为该疾病的预防和控制提供理论依据。在DWV基因保守序列设计4条引物,探究LAMP扩增的最优条件,并与常规的PCR(polymerase chain reaction)检测方法进行比较。建立的LAMP方法检测下限为0.89 pg,灵敏度比PCR高100倍而且特异性好。临床检测显示建立的LAMP方法可行、准确、方便、灵敏。针对DWV的LAMP建立的检测方法为养蜂生产第一线检测和预防DWV提供了技术支持,有一定的应用价值。     

Molecular and biological characterization of deformed wing virus of honeybees (Apis mellifera L.)

Deformed wing virus (DWV) of honeybees (Apis mellifera) is closely associated with characteristic wing deformities, abdominal bloating, paralysis, and rapid mortality of emerging adult bees. The virus was purified from diseased insects, and its genome was cloned and sequenced. The genomic RNA of DWV is 10,140 nucleotides in length and contains a single large open reading frame encoding a 328-kDa polyprotein. The coding sequence is flanked by a 1,144-nucleotide 5' nontranslated leader sequence and a 317-nucleotide 3' nontranslated region, followed by a poly(A) tail. The three major structural proteins, VP1 (44 kDa), VP2 (32 kDa), and VP3 (28 kDa), were identified, and their genes were mapped to the N-terminal section of the polyprotein. The C-terminal part of the polyprotein contains sequence motifs typical of well-characterized picornavirus nonstructural proteins: an RNA helicase, a chymotrypsin-like 3C protease, and an RNA-dependent RNA polymerase. The genome organization, capsid morphology, and sequence comparison data indicate that DWV is a member of the recently established genus Iflavirus.     

Characterization of the Tribolium Deformed ortholog and its ability to directly regulate Deformed target genes in the rescue of a Drosophila Deformed null mutant

 We have analyzed the Tribolium castaneum ortholog of the Drosophila homeotic gene Deformed (Dfd) and determined its expression pattern during embryogenesis in this beetle. Tc Deformed (Tc Dfd) is expressed in the blastoderm and the condensing germ rudiment in a region that gives rise to gnathal segments. During germ band extension Tc Dfd is expressed in the mandibular and maxillary segments, their appendages, and the dorsal ridge. Comparison of insect Dfd protein sequences reveals several highly conserved regions. To determine whether common molecular features reflect conserved regulatory functions we used the Gal4 system to express the Tribolium protein in Drosophila embryos. When Tc Dfd is expressed throughout embryonic ectoderm under the control of P69B, the beetle protein autoregulates the endogenous Dfd gene. In addition, the Drosophila proboscipedia gene (a normal target of Dfd) is ectopically activated in the antennal and thoracic segments. We also compared the ability of the beetle and fly proteins to rescue defects in Dfd ^– mutants by expressing each throughout the embryonic during embryogenesis. Both proteins rescued Dfd ^– defects to the same extent in that they each restore the development of mouth hooks and cirri, as well as cause gain-of-function abnormalities of posterior mouth parts. As before, pb was ectopically activated in the antennal segment. This is the first demonstration of the ability of a heterologous homeotic selector protein to directly regulate a target gene independent of an endogenous Drosophila autoregulatory loop. Received: 11 December 1998 / Accepted: 8 March 1999     

Venereal and vertical transmission of deformed wing virus in honeybees (Apis mellifera L.)

Deformed wing virus (DWV) infected semen was used for artificial insemination of DWV-free virgin queens. High titres of DWV could subsequently be detected not only in the spermatheca, but also in the ovaries, demonstrating venereal transmission of DWV in honey bees. Subsequent vertical transmission of the virus to the progeny of DWV infected queens was also demonstrated. Neither transmission route is 100% effective. Whether venereal transmission of DWV occurs during natural mating remains to be determined. The implications for the use, sale and transport of semen samples for artificial insemination are discussed.     

Deformed wing virus associated with <Emphasis Type="Italic">Tropilaelaps mercedesae</Emphasis> infesting European honey bees (<Emphasis Type="Italic">Apis mellifera</Emphasis>)

Mites in the genus Tropilaelaps (Acari: Laelapidae) are ectoparasites of the brood of honey bees (Apis spp.). Different Tropilaelaps subspecies were originally described from Apis dorsata, but a host switch occurred to the Western honey bee, Apis mellifera, for which infestations can rapidly lead to colony death. Tropilaelaps is hence considered more dangerous to A. mellifera than the parasitic mite Varroa destructor. Honey bees are also infected by many different viruses, some of them associated with and vectored by V. destructor. In recent years, deformed wing virus (DWV) has become the most prevalent virus infection in honey bees associated with V. destructor. DWV is distributed world-wide, and found wherever the Varroa mite is found, although low levels of the virus can also be found in Varroa free colonies. The Varroa mite transmits viral particles when feeding on the haemolymph of pupae or adult bees. Both the Tropilaelaps mite and the Varroa mite feed on honey bee brood, but no observations of DWV in Tropilaelaps have so far been reported. In this study, quantitative real-time RT-PCR was used to show the presence of DWV in infested brood and Tropilaelaps mercedesae mites collected in China, and to demonstrate a close quantitative association between mite-infested pupae of A. mellifera and DWV infections. Phylogenetic analysis of the DWV sequences recovered from matching pupae and mites revealed considerable DWV sequence heterogeneity and polymorphism. These polymorphisms appeared to be associated with the individual brood cell, rather than with a particular host.     

Butterfly wing patterns

This paper integrates genetical studies of variation in the wing patterns of Lepidoptera with experimental investigations of developmental mechanisms. Research on the tropical butterfly,Bicyclus anynana, is described. This work includes artificial selection of lines with different patterns of wing eyespots followed by grafting experiments on the lines to examine the phenotypic and genetic differences in terms of developmental mechanisms. The results are used to show how constraints on the evolution of this wing pattern may be related to the developmental organisation. The eyespot pattrn can be envisaged as a set of developmental homologues; a common developmental mechanism is associated with a quantitative genetic system involving high genetic correlations. However, individual genes which influence only subsets of the eyespots, thus uncoupling the interdependence of the eyespots, may be important in evolutionary change. The postulated evolutionary constraints are illustrated with respect to differences in wing pattern found among other species ofBicyclus.