植菌昆虫与其共生真菌协同进化分子机制

昆虫菌业（fungiculture）是一种类似于人类种植业的昆虫种植体系,包括种植、耕作、收获和营养依赖4个过程,可分为高级的社会性昆虫如切叶蚂蚁、白蚁等和低级的非社会性昆虫如食菌小蠹虫、卷叶象甲、蜥蜴甲虫、树蜂等,它们均能种植并取食真菌。近年来随着组学及微生物组技术的发展,植菌昆虫与其共生真菌协同进化的分子机制研究方面取得了重要进展。系统发育分析阐明了植菌昆虫的起源与进化历程,并显示出与共生真菌系统发育的一致性;共生真菌细胞核数量也从双核增加到最多17个核,而染色体倍型也从单倍体增加为二倍体甚至多倍体;组学分析则揭示了植菌昆虫与其共生真菌在精氨酸、碳水化合物、木质素及几丁质合成或降解等方面显示出了高度的协同进化。本文系统综述了植菌昆虫及其共生真菌的系统进化、核进化及基因组进化进展,并探讨这种协同进化机制的生物学意义。     

虎杖象甲培植真菌营养特性及对菌丝附属物形成的影响

虎杖象甲培植共生真菌形成的共生体系是植菌昆虫菌业中的典型代表。共生真菌Penicillium herquei如何向虎杖象甲Euops chinensis提供营养尚未明确。本研究发现共生真菌P. herquei的菌丝表面存在大量瘤状凸起物及由凸起物衍生的附属丝等特化结构,该结构可能为虎杖象甲提供营养;对共生真菌的营养研究表明,共生真菌能高效利用山梨醇、蔗糖、海藻糖、葡萄糖等单糖或双糖,以及酪氨酸、甘氨酸、谷氨酰胺等昆虫非必须氨基酸,同时在高碳和最适碳源条件下有利于菌丝特化附属物的产生。研究结果不仅提供了植菌卷叶象甲菌业中共生真菌在营养方面的适应性进化证据,而且为进一步揭示共生真菌适应卷叶象甲的营养机制奠定了基础。     

不同衰退等级蒙古沙冬青根内真菌多样性与群落结构

本研究根据沙冬青现有的生长状况,将内蒙古西鄂尔多斯保护区珍稀濒危植物蒙古沙冬青群落分为轻度衰退、中度衰退和重度衰退3个衰退等级,并选取一个未衰退的沙冬青群落作为对照,应用Illumina Hiseq测序平台,对这4个沙冬青群落的植株根内真菌进行高通量测序分析。测序共获得215个真菌的OTU,不同衰退等级沙冬青根内真菌群落结构在门、科、属水平上都有显著差异。Top 10属中的Agaricus、Tomentella、Tricholoma、Fusarium、Inocybe以及Tuber这6个属真菌在所有衰退等级的沙冬青都有分布。不同衰退等级的沙冬青根内都有腐生或寄生真菌和“共生”真菌分布,不同衰退等级两类群的真菌占比不同且呈现出动态变化,随着衰退等级的增加沙冬青根内腐生真菌或寄生真菌与“共生”真菌比例显著增高。土壤有机质与土壤容重具有协同作用,且对Agaricus、Inocybe、Fusarium、Penicillium、Amphinema具有正相关影响,对Tricholoma、Tomentella、Tuber具有负相关影响。     

竹叶兰根系共生真菌群落结构及生物学功能初探

为研究竹叶兰Arundina graminifolia根系共生真菌的群落结构及对寄主的营养作用,采用高通量测序技术对野生竹叶兰根围土壤、根表、根内3个生态位真菌的种类及生物学功能进行鉴定与预测。所测竹叶兰根系共生真菌的平均辛普森指数为根围0.972、根 表0.905、根内0.703,说明3个位点都具有丰富的共生真菌;根围土壤真菌香农指数7.393显著高于根表和根内香农指数4.728和2.872,根表真菌多样性显著高于根内（P<0.05）。根表、根内有子囊菌门、担子菌门、罗兹壶菌门、壶菌门、毛霉菌门和被孢霉门,较根围土壤缺少球囊菌门和梳霉菌门;分子方差分析（AMOVA）证实3个生态位的真菌群落结构差异极显著（P=0.004）。各生态位丰度最高的20个真菌仅属于子囊菌和担子菌的40个属,其中子囊菌有30个属,种类数量占优势,而担子菌的单个种类丰度高于子囊菌。根围土壤腐生型真菌占优势（44.07%）;根表主要有腐生、病原、共生型真菌;根内真菌营养类型差异大,样品M1的共生型真菌为优势类型（97.21%）,M2、M3以腐生型占优（90.96%）。优势共生型真菌有革菌属Threlephora、根须腹菌属Rhizopogon和Dentiscutata,腐生型真菌主要有长毛孔菌属Funalia和Vuilleminia,均为担子菌。研究结果为揭示竹叶兰与根系真菌的营养关系以及共生真菌的开发提供了依据。     

硬叶兰种子的迁地共生萌发及有效共生真菌的分离和鉴定

兰科植物的种子原地和迁地共生萌发技术是近年发展起来的开展兰科植物种子和共生真菌研究的有效方法。该研究对兰属(Cymbidium)附生植物硬叶兰(C. mannii)开展了种子的迁地共生萌发研究, 试图获得其种子萌发的有效真菌。利用硬叶兰成年植株根部周围的树皮、苔藓、枯枝落叶、腐殖质等作为培养基质, 进行种子的共生培养。在培养133天后, 成功地获得了处于不同阶段的已萌发种子、原球茎和幼苗, 并从原球茎中分离得到一种瘤菌根菌属(Epulorhiza)真菌。用所分离到的FCb4菌株和一种从兜唇石斛(Dendrobium aphyllum)分离到的胶膜菌属(Tulasnella) FDaI7菌株和硬叶兰种子在燕麦琼脂培养基上进行共生萌发, 设置不接菌作为对照处理, 以检验FCb4菌株对硬叶兰种子萌发的有效性。经过58天的培养, 不接菌的对照处理中种子没有萌发, 接种FCb4和FDaI7菌株的处理都有很高的种子萌发率, 两种接菌处理在不同光照条件下的种子萌发率均无显著性差异。但暗培养条件下, 种子萌发形成原球茎后, 表现出生长停滞的趋势, 仅有很少的原球茎继续生长达到幼苗阶段, 说明原球茎发育后期与幼苗发育阶段需要光照。在光照条件下, 接种FCb4菌株处理中达到幼苗阶段种子的比例为(25.67 ± 9.27)%, 显著高于接种FDaI7菌株处理的(3.04 ± 2.27)% (W = 56, p = 0.026, Mann-Whitney U-test), 表明此研究中分离到的瘤菌根菌属真菌能有效地促使硬叶兰种子萌发并生长发育到幼苗阶段。     

AMF和DSE组合菌剂促生防线虫病效应

本试验旨在探究丛枝菌根真菌（AMF）和暗隔内生真菌（DSE）组合菌剂对南方根结线虫Meloidogyne incognita发育、侵染、黄瓜根结线虫病以及黄瓜生长发育的影响,为进一步探索AMF和DSE协同发挥生理生态效应的作用机制奠定基础。试验对“津优35号”黄瓜接种南方根结线虫、Phoma leveillei（DSE）、Funneliformis mosseae（AMF）、Glomus versiforme（AMF）、Acaulospora laevis（AMF）和/或Scutellospora aurigloba（AMF）。结果表明,供试AMF与DSE 能促进黄瓜生长、提高产量和增强抗病性。AMF+DSE组合处理的菌根和DSE的定殖数量、株高、茎粗、地上部和根系干重、单株产量等显著优于单接种AMF或单接种DSE处理。以F. mosseae + P. leveillei组合抑制南方根结线虫的发育、降低线虫繁殖数量、根内定殖数量、发病率和根结指数的效果最好。     

从细胞内共生到多尺度共生：回顾与展望

基于共生概念的历史变化,目前人们普遍接受了广义共生概念。即共生是包含互利共生（mutualism）、偏利共生（commensalism）和拮抗/寄生（antagonism/parasitism）的共生连续体。本文简述了近20年间,全球9次国际共生学术大会取得的重要成果,对细胞内共生、时间、空间以及多种互作尺度共生关系的研究利用进展进行了评述。同时展望了一些活跃共生领域的研究概况,如共生失调 （dysbiosis）、植物-微生物-昆虫三角共生关系（plant-microbe-insect triangle）、细菌-真菌互作（bacterial- fungal interaction,BFI）、菌根菌-真菌内生细菌-植物多方共生联盟（multipartite symbiosis consortium）以及与共生相关微生物组的集合群落（metacommunity）研究及应用等。共生（symbiosis）正成为当代生物学的核心原则,正以一种与更宏大系统方法相一致的概念,从根本上改变了传统上的一些生物学概念,如孤立性的个体（individuality）概念。基因组测序和高通量RNA技术分析揭示,动、植物与共生微生物的重要互作,打破了迄今为止生物个体的特征边界,挑战了这些学科的定义;共生不仅是一对一的互利共生关系,共生实际是多种共生模式的连续共生体。此外,植物-昆虫-微生物互作的三角关系;菌根-真菌-真菌内生细菌-植物的多方联盟等新关系的发现,更把生命科学推向了快速发展的方向。这些科学进展不仅对生物科学的遗传学、免疫学、进化、发育、解剖学和生理学的研究至关重要,拓宽了新的视野,而且对农业中生物制剂研发,人类微生物组的管理及调控,以及对发酵食品及工业微生物生产的设计和管理将产生积极影响。     

白蚁与动物及微生物的共生类型及其机制

综述了白蚁螱客的主要种类、共生关系及相关机制的研究进展。白蚁螱客中,已报道的动物种类达170种。在与动物的共生关系中存在偏利共生（宾主共栖和异种共栖）、互利共生和无关共生三种;在与微生物的共生关系中,存在与内生菌（原生动物、细菌、真菌和放线菌）和外生菌（蚁巢伞菌等）间的互利关系。指出了白蚁与螱客研究中存在的问题,给出了解决方案,并提出了今后可能的研究热点或方向,为白蚁的综合利用（如纤维素酶）及今后研究物种间的协同进化提供了基础资料。     

摩西管柄囊霉Funneliformis mosseae对连作大豆分枝期根系AM真菌群落结构的影响

本文研究摩西管柄囊霉Funneliformis mosseae对短期连作3个大豆品种分枝期根系AM真菌群落结构的影响,旨在探索不同大豆品种与AM真菌在连作土壤中的互作效应。通过向黑农44（HN44）、黑农48（HN48）和恳丰16（KF16）3个大豆品种盆栽土壤中接种F. mosseae,对接菌连作0年（未连作对照组）、连作1年、连作2年土壤中的大豆根系,采用Nested-Program Control Register-DGGE技术分析。结果表明,接种F. mosseae后,不同大豆品种根系AM真菌多样性指数和丰度值表现为连作2年的土壤>连作1年的土壤>未连作的土壤;其中,球囊霉属Glomus和F. mosseae为3个大豆品种根系中AM真菌的优势菌群。接种F. mosseae对连作1年、连作2年3个大豆品种根系AM真菌群落结构具有显著影响。     

昆虫病原线虫共生细菌共生性的分子生物学研究进展

嗜线虫致病杆菌属Xenorhabdus和发光杆菌属Photorhabdus细菌隶属肠杆菌科Enterobacteriaceae,对多种害虫致病能力强,分别与斯氏属Steinernema和异小杆属Heterorhabditis昆虫病原线虫互惠共生。该两属共生细菌既存在对昆虫寄主的病原性,又存在与线虫寄主的共生性。共生细菌与其线虫寄主的共生性主要表现以下4方面：（1）细菌产生食物信号诱导滞育不取食的感染期线虫恢复;（2）细菌为线虫生长与繁殖提供营养;（3）细菌能于感染期线虫的肠道定殖与生长;（4）细菌产生杀线虫毒素杀死非共生线虫。本文综述了共生菌以上4方面的共生性及其相关的分子机制。     

Contribution of symbiotic mycangial fungi to larval nutrition of a leaf-rolling weevil

Some phytophagous insects have been known to inoculate certain fungi on plant substrates. In many cases of such insect–fungi relationships it has been considered that fungi contribute to insects by decomposing lignin or polysaccharides, and that the insects feed on the decomposition products or fungi themselves. Females of the leaf-rolling weevil in the genus Euops (Attelabidae) store spores of symbiotic fungi in the mycangia and inoculate them on leaf rolls. To determine the effect of mycangial fungi on larval nutrition in E. lespedezae, the nutritional value was compared between leaves with and without mycangial fungi. Two Penicillium species were isolated from the mycangia. These mycangial fungi showed little effect on the decomposition of lignin and polysaccharides, and showed little effect on enhancement of soluble sugars within leaves. Thus, the mutualism between Euops and its mycangial fungi contrasts with the mainly nutritional mutualisms between wood-infesting insects (termites, bark/ambrosia beetles, and wood wasps) and lignin/polysaccharide-decomposing fungi.     

Fungi associated with the southern pine beetle: avoidance of induced defense response in loblolly pine

Summary Loblolly pines (Pinus taeda) are rapidly killed by colonizing southern pine beetle (Dendroctonus frontalis). The female beetles carry two species of fungi (Ceratocystis minor var. barrasii and an unnamed basidiomycete) within a mycangium. The insects are also frequently associated with a blue-staining form of C. minor. These fungi are inoculated into the tree during colonization. The tree has an induced defensive response that involves resin soaking and necrosis of affected tissue isolating the invading organlsms. The blue-staining fungus stimulates formation of this response in the tree, but the two mycangial fungi do not. These results suggest that the beetles are closely associated with two highly pathogenic fungi that do not stimulate one of the critical components of tree defense.     

Mycangial fungus benefits the development of a leaf-rolling weevil, Euops chinesis

While a wide array of insects form symbiotic relationships with microbes, the underlying mechanisms of these relationships are various and complex. In this study, we investigated the role that the mycangial fungus Penicillium herquei plays in the development of the leaf-rolling weevil Euops chinesis, which feeds on the knotweed Fallopia japonica. The weevil inoculates the fungus during oviposition into a leaf-roll that it creates for its larvae. We found that removal of P. herquei inocula from leaf-rolls significantly decreased the weevil's survival rate especially in the larval stage. Although inoculation with P. herquei had no effect on the plant's lignin content, it significantly decreased the cellulose content of the knotweed leaves. P. herquei also showed antibiotic properties against two fungi (Rhizopus sp.) that attack the weevil's leaf-rolls. Our results suggest that the mycangial fungus may help alter leaf chemical components and protect against pathogens thus improve leaf-rolls for the development of E. chinesis.     

Strong indirect interactions of Tarsonemus mites (Acarina: Tarsonemidae) and Dendroctonus frontalis (Coleoptera: Scolytidae)

Phoretic mites of bark beetles are classic examples of commensal ectosymbionts. However, many such mites appear to have mutualisms with fungi that could themselves interact with beetles. We tested for indirect effects of phoretic mites on Dendroctonus frontalis, which attacks and kills pine trees in North America. Tarsonemus mites are known to carry ascospores of Ophiostoma minus, which tends to outcompete the mutualistic fungi carried by D. frontalis. Experimental additions and removals of mites from beetles demonstrated that Tarsonemus propagate O. minus in beetle oviposition galleries. Furthermore, the abundance of Tarsonemus and O. minus tended to covary in nature. These results verified a strong mutualism between Tarsonemus and O. minus. Results also indicated that O. minus is an antagonist of D. frontalis: beetle larvae seldom survived in the presence of O. minus (compared to 83% survival elsewhere). Apparently, this is an indirect result of O. minus outcompeting the two species of mycangial fungi that are critical to beetle nutrition. Thus, Tarsonemus mites close a loop of species interactions that includes a commensalism (mites and beetles), a mutualism (mites and O. minus), asymmetric competition (O. minus and mycangial fungi), and another mutualism (mycangial fungi and beetles). This interaction system produces negative feedback that could contribute to the endogenous population dynamics of D. frontalis. Reproductive rate of Tarsonemus was more temperature‐sensitive than beetle generation time (which constrains the time for mite reproduction within a tree). This differential temperature sensitivity produces a narrow range of temperatures (centred at 27°C) in which mite reproduction per D. frontalis generation can attain its maximum of 100 mites/beetle. Consequently, seasonal oscillations in temperature are predicted to produce oscillations in the D. frontalis community, and climatic differences between regions could influence the community to dampen or exacerbate the cyclical outbreak dynamics of D. frontalis.     

Origin and evolution of fungus farming in wood-boring Coleoptera – a palaeontological perspective

Insect–fungus mutualism is one of the better-studied symbiotic interactions in nature. Ambrosia fungi are an ecological assemblage of unrelated fungi that are cultivated by ambrosia beetles in their galleries as obligate food for larvae. Despite recently increased research interest, it remains unclear which ecological factors facilitated the origin of fungus farming, and how it transformed into a symbiotic relationship with obligate dependency. It is clear from phylogenetic analyses that this symbiosis evolved independently many times in several beetle and fungus lineages. However, there is a mismatch between palaeontological and phylogenetic data. Herein we review, for the first time, the ambrosia system from a palaeontological perspective. Although largely ignored, families such as Lymexylidae and Bostrichidae should be included in the list of ambrosia beetles because some of their species cultivate ambrosia fungi. The estimated origin for some groups of ambrosia fungi during the Cretaceous concurs with a known high diversity of Lymexylidae and Bostrichidae at that time. Although potentially older, the greatest radiation of various ambrosia beetle lineages occurred in the weevil subfamilies Scolytinae and Platypodinae during the Eocene. In this review we explore the evolutionary relationship between ambrosia beetles, fungi and their host trees, which is likely to have persisted for longer than previously supposed.     

Paths to last in mixed crop–livestock farming: lessons from an assessment of farm trajectories of change

Mixed crop–livestock systems, combining livestock and cash crops at farm level, are considered to be suitable for sustainable intensification of agriculture. Ensuring the survival of mixed crop–livestock systems is a challenge for European agriculture: the number of European mixed crop–livestock farms has been decreasing since 1970. Analysis of farming system dynamics may elucidate past changes and the forces driving this decline. The objectives of this study were (i) to identify the diversity of paths that allowed the survival of mixed crop–livestock farming and (ii) to elucidate the driving forces behind such survival. We analysed the variety of farm trajectories from 1950 to 2005. We studied the entire farm population of a case study site, located in the ‘Coteaux de Gascogne’ region. In this less favoured area of south-western France, farmers have limited specialisation. Currently, half of the farms use mixed crop–livestock systems. The data set of 20 variables for 50 farms on the basis of six 10-year time steps was collected through retrospective surveys. We used a two-step analysis including (i) a visual assessment of the whole population of individual farm trajectories and (ii) a computer-based typology of farm trajectories on the basis of a series of multivariate analyses followed by automatic clustering. The European Common Agricultural Policy, market globalisation and decreasing workforce availability were identified as drivers of change that favoured the specialisation process. Nevertheless, farmers’ choices and values have opposed against these driving forces, ensuring the survival of some mixed crop–livestock farming systems. The trajectories were clustered into five types, four of which were compatible with mixed crop–livestock systems. The first type was the maximisation of autonomy by combining crops and livestock. The second type was diversification of production to exploit economies of scope and protect the farm against market fluctuations. The other two types involved enlargement and progressive adaptation of the farm to the familial workforce. The survival of mixed crop–livestock systems in these two types is largely dependent on workforce availability. Only one type of trajectory, on the basis of enlargement and economies of scale, did not lead to mixed crop–livestock systems. In view of the current evolution of the driving forces, maximising autonomy and diversification appear to be suitable paths to deal with current challenges and maintain mixed crop–livestock systems in Europe.     

Can microbial-based insecticides replace chemical pesticides in agricultural production?

Extensive use of chemical insecticides to control insect pests in agriculture has improved yields and production of high-quality food products. However, chemical insecticides have been shown to be harmful also to beneficial insects and many other organisms like vertebrates. Thus, there is a need to replace those chemical insecticides by other control methods in order to protect the environment. Insect pest pathogens, like bacteria, viruses or fungi, are interesting alternatives for production of microbial-based insecticides to replace the use of chemical products in agriculture. Organic farming, which does not use chemical pesticides for pest control, relies on integrated pest management techniques and in the use of microbial-based insecticides for pest control. Microbial-based insecticides require precise formulation and extensive monitoring of insect pests, since they are highly specific for certain insect pests and in general are more effective for larval young instars. Here, we analyse the possibility of using microbial-based insecticides to replace chemical pesticides in agricultural production.     

Host plant effects on the behavioural phenotype of a Chrysomelid

1. The food plant quality influences feeding preferences and various life history traits of herbivorous insects. However, the effects of different host plant qualities on the behavioural phenotype have rarely been studied in behavioural ecology, especially in a pest‐crop‐framework. 2. Behavioural phenotypes of insects may not only be affected by external environmental factors, such as the host plant quality but are also shaped by internal factors, such as the sex and the age of individuals. 3. To study host plant effects on behavioural phenotypes, we reared mustard leaf beetles (Phaedon cochleariae Fabricius) either on their natural host watercress or on the crop cabbage, on which this beetle can be a pest. The behavioural phenotype was characterised twice in the adult lifetime by measuring six behavioural traits tested in distinct contexts. 4. Depending on the context, different behavioural traits were specifically affected by the host plant, the sex and/or the age. Beetles fed on cabbage became more active with age. Furthermore, the boldness tested in an unprotected environment context was influenced by the host, with beetles fed on cabbage being bolder, whereas the boldness in a hiding or predator attack context was affected by the age and/or the interaction of host plant × sex. 5. In conclusion, beetles fed on the crop cabbage develop a different behavioural phenotype compared to beetles fed on watercress. Previous results showed that beetles reared on cabbage have a higher reproductive output. Thus, beetles fed on the crop potentially express a faster pace‐of‐life.     

Atomic Force Microscopy Study of Nano-Physiological Response of Ladybird Beetles to Photostimuli

Background

Insects are of interest not only as the most numerous and diverse group of animals but also as highly efficient bio-machines varying greatly in size. They are the main human competitors for crop, can transmit various diseases, etc. However, little study of insects with modern nanotechnology tools has been done.

Methodology/Principal Findings

Here we applied an atomic force microscopy (AFM) method to study stimulation of ladybird beetles with light. This method allows for measuring of the internal physiological responses of insects by recording surface oscillations in different parts of the insect at sub-nanometer amplitude level and sub-millisecond time. Specifically, we studied the sensitivity of ladybird beetles to light of different wavelengths. We demonstrated previously unknown blindness of ladybird beetles to emerald color (∼500nm) light, while being able to see UV-blue and green light. Furthermore, we showed how one could study the speed of the beetle adaptation to repetitive flashing light and its relaxation back to the initial stage.

Conclusions

The results show the potential of the method in studying insects. We see this research as a part of what might be a new emerging area of “nanophysiology” of insects.