水稻二化螟内共生菌杀雄菌属和沃尔巴克氏体的检测与分析

应用杀雄菌属(Arsenophonus)23S rDNA基因和沃尔巴克氏体(Wolbachia) wsp基因的特异引物,通过PCR扩增的方法对我国20个水稻二化螟地理种群感染两类内共生菌的情况进行了检测.结果表明: 我国水稻二化螟不同地理种群的Arsenophonus和Wolbachia感染率不一致,哈尔滨、惠水、吉林、南阳和扬州5个地理种群的Arsenophonus感染率为5.0%～50.0%;汉中、南宁和扬州3个地理种群的Wolbachia感染率为25.0%～40.0%,其他地理种群中没有检测到这两种共生菌的存在.水稻二化螟不同地理种群感染的Arsenophonus 23S rDNA基因序列完全一致,将该基因株型命名为csArs.但所检测到的Wolbachia wsp基因序列不一致,分别为wChisup1、wChisup5和wChisup6,其中wChisup1属于A群,其他属于B群.这说明水稻二化螟感染的Arsenophonus为同一株型,而感染的Wolbachia株型较为复杂.通过构建系统发育树发现,水稻二化螟体内的Arsenophonus23S rDNA基因和Wolbachia wsp基因与其他物种体内相关序列完全一致或高度相似.     

Endosymbiont metacommunities,mtDNA diversity and the evolution of the Bemisia tabaci (Hemiptera: Aleyrodidae) species complex

Bemisia tabaci, an invasive pest that causes crop damage worldwide, is a highly differentiated species complex, divided into biotypes that have mainly been defined based on mitochondrial DNA sequences. Although endosymbionts can potentially induce population differentiation, specialization and indirect selection on mtDNA, studies have largely ignored these influential passengers in B. tabaci, despite as many as seven bacterial endosymbionts have been identified. Here, we investigate the composition of the whole bacterial community in worldwide populations of B. tabaci, together with host genetic differentiation, focusing on the invasive B and Q biotypes. Among 653 individuals studied, more than 95% of them harbour at least one secondary endosymbiont, and multiple infections are very common. In addition, sequence analyses reveal a very high diversity of facultative endosymbionts in B. tabaci, with some bacterial genus being represented by more than one strain. In the B and Q biotypes, nine different strains of bacteria have been identified. The mtDNA‐based phylogeny of B. tabaci also reveals a very high nucleotide diversity that partitions the two ITS clades (B and Q) into six CO1 genetic groups. Each genetic group is in linkage disequilibrium with a specific combination of endosymbionts. All together, our results demonstrate the rapid dynamics of the bacterial endosymbiont–host associations at a small evolutionary scale, questioning the role of endosymbiotic communities in the evolution of the Bemisia tabaci species complex and strengthening the need to develop a metacommunity theory of inherited endosymbionts.     

Diversity of secondary endosymbionts among different putative species of the whitefly Bemisia tabaci

Abstract Endosymbionts are important components of arthropod biology. The whitefly Bemisia tabaci (Gennadius) (Hemiptera: Aleyrodidae) is a cryptic species complex composed of ≥ 28 putative species. In addition to the primary endosymbiont Portiera aleyrodidarum, six secondary endosymbionts (S‐endosymbionts), Hamiltonella, Rickettsia, Wolbachia, Cardinium, Arsenophonus and Fritschea, have been identified in B. tabaci thus far. Here, we tested five of the six S‐endosymbiont lineages (excluding Fritschea) from 340 whitely individuals representing six putative species from China. Hamiltonella was detected only in the two exotic invaders, Middle East‐Asia Minor 1 (MEAM1) and Mediterranean (MED). Rickettsia was absent in Asia II 1 and MED, scarce in Asia II 3 (13%), but abundant in Asia II 7 (63.2%), China 1 (84.7%) and MEAM1 (100%). Wolbachia, Cardinium and Arsenophonus were absent in the invasive MEAM1 and MED but mostly abundant in the native putative species. Furthermore, phylogenetic analyses revealed that some S‐endosymbionts have several clades and different B. tabaci putative species can harbor different clades of a given S‐endosymbiont, demonstrating further the complexity of S‐endosymbionts in B. tabaci. All together, our results demonstrate the variation and diversity of S‐endosymbionts in different putative species of B. tabaci, especially between invasive and native whiteflies.     

Diversity of bacteria associated with Hormaphidinae aphids (Hemiptera: Aphididae)

Bacteria are ubiquitous inhabitants of animals.Hormaphidinae is a particular aphid group exhibiting very diverse life history traits.However,the microbiota in this group is poorly known.In the present study,using high-throughput sequencing of bacterial 16S ribosomal RNA gene amplicons,we surveyed the bacterial flora in hormaphidine aphids and explored whether the aphid tribe,host plant and geographical distribution are associated with the distribution of secondary symbionts.The most dominant bacteria detected in hormaphidine species are heritable symbionts.As expected,the primary endosymbiont Buchnera aphidicola is the most abundant symbiont across all species and has cospeciated with its host aphids.Six secondary symbionts were detected in Hormaphidinae.Arsenophonus is widespread in Hormaphidinae species,suggesting the possibility of ancient acquisition of this symbiont.Ordination analyses and statistical tests show that the symbiont composition does not seem to relate to any of the aphid tribes,host plants or geographical distributions,which indicate that horizontal transfers might occur for these symbionts in Hormaphidinae.Correlation analysis exhibits negative interference between Buchnera and coexisting secondary symbionts,while the interactions between different secondary symbionts are complicated.These findings display a comprehensive picture of the microbiota in Hormaphidinae and may be helpful in understanding the symbiont diversity within a group of aphids.     

白蜡虫体内杀雄菌属(Arsenophonus)共生菌的分子检测

[目的]研究白蜡虫体内杀雄菌属(Arsenophonus)共生菌的含量与白蜡虫性比之间的关系.[方法]采用16S rDNA文库的方法对白蜡虫雄虫体内的共生菌进行分析,利用杀雄菌属特异的2条16S rDNA引物以及23S rDNA引物进行PCR检测.对昭通、昆明、金口河、杭州、长春、江华6个不同地理种群白蜡虫体内的杀雄菌属共生菌进行半定量分析,并采用荧光定量PCR对昭通、昆明、金口河白蜡虫体内的杀雄菌属共生菌进行绝对定量分析.[结果]在白蜡虫体内首次发现杀雄菌属共生菌.在白蜡虫雌雄虫体内均扩增出杀雄菌属的两条不同长度的16S rDNA序列,分别为445bp和1462bp,并扩增得到长度为582bp的23S rDNA序列.杭州和江华地理种群白蜡虫的一些个体不含杀雄菌属共生菌.昭通地理群的杀雄菌属共生菌含量显著高于金口河和昆明,而昆明和金口河白蜡虫的杀雄菌属含量无显著差异.[结论]白蜡虫体内杀雄菌属共生菌的含量与白蜡虫性比无关.     

我国不同地区草地贪夜蛾内共生菌感染和耐寒能力差异研究

本研究旨在明确我国不同地区草地贪夜蛾Spodoptera frugiperda共生菌感染差异和耐寒能力差异。采用16S或23S rDNA等定性检测方法,鉴定草地贪夜蛾共生菌种类并测定其感染率,通过邻接法（Neighbor-joining）构建系统进化树,进行共生菌系统发育分析。从我国8个地区采集的草地贪夜蛾幼虫中鉴定出2种内共生细菌,分别为Arsenophonus和Cardinium。两种共生菌在不同地理种群中的感染率存在显著差异,其中Arsenophonus感染率总体较低,云南保山种群感染率最高,为20.0%,其次是广西南宁种群(15.5%),贵州贵阳和江苏盐城的种群感染率均为1.7%,而广东广州、江苏无锡以及湖北宜昌和孝感种群中均未发现Arsenophonus的感染;Cardinium感染以江苏无锡种群最高,达96.7%,云南保山和广东广州种群感染率分别为93.3%和86.7%,贵州贵阳种群为53.3%,湖北孝感和宜昌感染率则分别为30.0%和18.3%,江苏盐城种群感染率也为18.3%,而广西南宁种群中未发现该共生菌。对8个地区草地贪夜蛾的耐寒能力比较表明,江苏盐城和湖北孝感种群耐寒能力最弱,-6℃下死亡率分别为76%和69%,显著高于其它6个种群,后6个种群间无显著差异,死亡率在35%~49%之间。我国不同地区草地贪夜蛾在共生菌感染和耐寒能力上均存在差异,但未发现共生菌感染与耐寒能力的相关性。     

Coinfection of the secondary symbionts,Hamiltonella defensa and Arsenophonus sp.contribute to the performance of the major aphid pest,Aphis gossypii(Hemiptera:Aphididae)

Bacterial endosymbionts play important roles in ecological traits of aphids.In this study,we characterize the bacterial endosymbionts of A.gossypii collected in Karaj,Iran and their role in the performance of the aphid.Our results indicated that beside Buchnera aphidicola,A.gossypii,also harbors both Hamiltonella defensa and Arsenophonus sp.Quantitative PCR(qPCR)results revealed that the populations of the endosymbionts increased throughout nymphal development up to adult emergence;thereafter,populations of Buchnera and Arsenophonus were diminished while the density of H.defensa constantly increased.Buchnera reduction caused prolonged development and no progeny production.Furthermore,secondary symbiont reduction led to reduction of the total life span and intrinsic rate of natural increase as well as appearance of the deformed dead offspring in comparison with the control insects.Reduction of the secondary symbionts did not affect parasitism rate of the aphid by the parasitic wasp Aphidius matricariae.Together these findings showed that H.defensa and Arsenophonus contributed to the fitness of A.gossypii by enhancing its performance,but not through parasitoid resistance.     

Cowpea aphid (Aphis craccivora) associated with different host plants has different facultative endosymbionts

Maternally inherited facultative endosymbiotic bacteria are common among insects, including many polyphagous insect herbivores. To investigate whether symbiont infection is structured by host plant in the polyphagous aphid Aphis craccivora Koch, pyrosequencing and diagnostic PCR were performed on 26 populations from two different host plants, alfalfa (Medicago sativa) or black locust (Robinia pseudoacacia). Results indicated that Aphis craccivora harbours distinctly different microbial communities in alfalfa versus locust. The facultative symbiont Hamiltonella was found only in aphids collected from alfalfa, and the facultative symbiont Arsenophonus was found only in aphids from locust. Hamiltonella is known to protect aphids against hymenopteran parasitoids, whereas the phenotypic effects of Arsenophonus in aphids are unknown. Correspondingly, a screen of the aphid samples for hymenopteran DNA indicated that Hamiltonella‐bearing alfalfa populations of A. craccivora experienced lower parasitism than Arsenophonus‐bearing locust populations. This study contributes to the growing body of evidence that correlative associations between bacterial endosymbionts and host plants may be a common phenomenon in polyphagous herbivores, and suggests that microbial symbionts have the potential to act as drivers for observed ecological differences among host‐associated populations of polyphagous insects.     

Microbial communities and interactions in the lone star tick,Amblyomma americanum

To quantify microbial composition and interactions, we identified prokaryotic communities in the lone star tick (Amblyomma americanum) based on 16S rRNA gene sequences and direct probing. The lone star tick is the vector of emerging diseases and host to additional symbionts of unknown activity, and is representative of other blood‐sucking arthropods. We evaluated the potential for vertical (transovarial) transmission by molecular analysis of microbial symbionts from egg and larval clutches. Direct probing of adults (N = 8 populations from the southeastern and midwestern USA, 900 ticks total) revealed three vertically transmitted symbionts: a Coxiella symbiont occurred at 100% frequency, Rickettsia species occurred in 45–61% of all ticks in every population and an Arsenophonus symbiont occurred in 0–90% of ticks per population. Arsenophonus and Rickettsia exhibited significant heterogeneity in frequency among populations. The human pathogens Ehrlichia chafeensis and Borrelia lonestari were rare in most populations. Additional microbes were detected sporadically. Most ticks (78%) were co‐infected by two or three microbes but statistical analysis indicated no significant deviation from random co‐occurrence. Our findings indicate that microbial communities within lone star ticks are diverse, and suggest that direct probing for a wider range of prokaryotes and application of quantitative polymerase chain reaction (PCR) may provide further insights into microbial interactions within disease vectors. Our results also emphasize the close phylogenetic relationship between tick symbionts and human pathogens, and consistent differences in their prevalence.     

The natural occurrence of secondary bacterial symbionts in aphids

1. Many insects host secondary bacterial symbionts that are known to have wide‐ranging effects on their hosts, from host‐plant use to resistance against natural enemies. This has been most widely studied in aphids, which have become a model system to study insect–bacteria interactions. 2. While there is an increasing understanding of the role of symbionts in aphids from controlled laboratory studies, we are only beginning to explore the impact of hosting these symbionts on eco‐evolutionary dynamics in natural systems. To date, many research groups have identified bacterial symbionts from various aphid species, providing us with a bank of literature on aphid–symbiont associations in natural populations. 3. The role of secondary symbionts in aphids is discussed, and the taxonomic and geographical distribution of symbionts among aphids are summarised, and the potential reasons for the patterns observed. The need to test for multiple symbiont species (and co‐infections) across many individuals and the whole distribution range of an aphid is highlighted, including sampling on all known host‐plant species. 4. It is further important also to consider variation within the symbiont, the aphid‐host and the surrounding community, e.g. host‐plants or the natural enemies, to understand how these have the potential to mediate aphid–symbiont interactions. 5. Finally, the knowledge gained from experimental work should now be used to understand the role of aphid secondary symbionts in field systems, to fully understand the potentially far‐reaching consequences of aphid endosymbionts on community and ecosystem processes.