十种虫生真菌液体培养芽生孢子的形态发生

芽生孢子是许多真菌通过出芽方式产生的无性孢子,在液体发酵中主要以这种形式大量繁殖。真菌杀虫剂活性成分分生孢子的大量生产使用液体发酵得到的芽生孢子作为接种物,另外,它也是当今虫生真菌遗传转化的重要受体,因而研究虫生真菌芽生孢子的形态和发生特点具有重要意义。本研究分别对液体发酵中的球孢白僵菌、粉棒束孢、蝉棒束孢、环链棒束孢、玫烟色棒束孢、细脚棒束孢、斜链棒束孢、金龟子绿僵菌、蝗绿僵菌和蜡蚧霉等10种常见虫生真菌芽生孢子的形成过程进行显微观察,了解其分生孢子产生过程的异同。结果表明,芽生孢子的产生方式有两种类型：1）蝉棒束孢在其整个生活史中以菌丝生长为主,芽生孢子产生的数量很少。2）其他各种真菌芽生孢子产生方式相似,在菌丝体形成后就开始大量以菌丝出芽或缢缩产生芽生孢子,接着还可通过芽生孢子的出芽或缢缩断裂产生新的芽生孢子。芽生孢子的产生分为3个时期：初期先由菌丝形成芽生孢子;指数期芽生孢子大量增殖,菌丝和芽生孢子都可产生芽生孢子;后期以芽生孢子产新芽生孢子为主要方式。     

丝孢真菌无性与有性进化分析

绝大多数丝孢真菌属、种主要以无性型存在,但该类真菌绝大多数属、种含有若干调控有性分化、发育及进化的基因信息,如交配型、信息素及G-蛋白α-亚基基因等,胁迫环境诱导这些真菌有性基因有效表达乃至发育为有性型,一直倍受真菌学界的关注。文中系统介绍了丝孢真菌无性型与有性型属种分类研究现状、进化分析及性别调控基因,1)分析属、种三类有性基因的多样性、异型交配基因比率、系统发育关系及有性进化潜力与趋势;2)分析属、种不同有性基因调控特性胁迫环境适应性,界定靶标基因及适宜生态因子,结合地球生境演化特性,预测属、种自然演化性别变化动态;3)分析靶标基因调控生态,阐明两性种有性基因调控生态遗传机制。综上有望解析绝大多数丝孢真菌属、种主要为无性型的本质原因,丰富丝孢真菌分类研究的理论,为真菌分类研究提供科学依据。     

苎麻疫霉有性生殖导致菌落形态和生长速率在无性后代持续变异的研究

研究了苎麻疫霉菌株ＪＳ－５自交Ｓ１代单卵孢株的菌落形态和生长速率在其单游动孢子无性系后代的遗传与变异。结果表明,亲本菌株ＪＳ－５的菌落形态和生长速率在连续４代单游动孢子无性系后代稳定遗传,而该菌株的自交Ｓ１代的约２／３单卵孢株的上述性状在其单孢无性后代中发生分离,且上述性状的分离在无性单孢后代中至少可连续保持３￣４代。本研究结果提示,苎麻疫霉有性生殖导致上述性状的变异,除有性生殖过程中的基因重组外     

捕食线虫真菌的微循环产孢

<正> 微循环产孢(microcycle conidiation),特别是虫生真菌的微循环产孢在理论和实际中均有着极为重要的意义,现已被用于提高液体培养的分生孢子产量,缩短达到稳定产孢的时间,降低孢子收获时营养体含量比例,以利于孢子分离和分散,以及能大量获得生理条件一致的孢子等。近年来,微循环产孢还被用于全型真菌的研究,例如证明虫草与无性型的真正联系。 微循环产孢这一重要领域的研究在我国仅处于起步阶段,捕食线虫真菌的微循环产孢更是从未涉     

梯棱羊肚菌无性孢子形态及显微结构观察

借助于扫描电子显微镜和光学显微镜观察了梯棱羊肚菌(Morchella importuna)无性孢子的形态,并采用激光共聚焦显微镜观察分析了无性孢子细胞核染色后的细胞核行为。显微观察结果表明,梯棱羊肚菌无性孢子大小均一,无色,球形,无隔,光滑,直径3.5~5.2μm;无性孢子梗纺锤状,呈轻微的S型弯曲,2.1~6.2×14.1~18.5μm,4~6个轮状环绕生长在特化的菌丝四周。细胞核染色结果显示,无性孢子大多为单核,少数2核、3核或4核。观察到正在进行核分裂的无性孢子,推测双核或多核无性孢子是由单核孢子经过2次或多次有丝分裂发育而来,梯棱羊肚菌的无性孢子应为"同核体"。梯棱羊肚菌无性孢子在形态发生等方面类似于真菌的小分生孢子,但功能还不甚明了。     

丝虫草及其无性型关系的分子确证

目的对丝虫草（Ophiocordyceps filiform is）及从子囊孢子分离所得的无性型进行测序和比较分析,确证二者之间的对应关系。方法采用PCR技术,以rDNA-ITS区为分子指标。结果系统进化树显示丝虫草的无性型归为被毛孢属较为合理,同时通过形态特征比较的结果表明,其无性型为荔波被毛孢（Hirsutella liboensis）。结论本研究首次确立了丝虫草的有性型与无性型的对应关系,并且成功培育出了与天然丝虫草相同的虫草子实体。     

苎麻疫霉有性生殖导致菌落形态和生长速率在无性后代持续变异的研究

研究了苎麻疫霉（PhytophthoraboehmeriaeSawada）菌株JS-5自交S1代单卵孢株的菌落形态和生长速率在其单游动孢子无性系后代的遗传与变异。结果表明,亲本菌株JS-5的菌落形态和生长速率在连续4代单游动孢子无性系后代稳定遗传,而该菌株的自交S1代的约2／3单卵孢株的上述性状在其单拘无性后代中发生分离,且上述性状的分离在无性单孢后代中至少可连续保持3～4代。本研究结果提示,苎麻疫霉有性生殖导致上述性状的变异,除有性生殖过程中的基因重组外,有性生殖还诱导产生了一种新的机制参与上述性状遗传与变异的调控。本研究还观察到供试菌株及其后代所产生的游动孢子绝大多数细胞内只含有1个细胞核,表明上述机制与异核现象无关。有性生殖导致生物学性状在随后的无性后代中发生持续变异可能是苎麻疫霉生物学性状遗传多样性的重要机制之一。     

苎麻疫霉有性生殖导致菌落形态和生长速率在无性后代持续变异的研究*

研究了苎麻疫霉（PhytophthoraboehmeriaeSawada）菌株JS-5自交S1代单卵孢株的菌落形态和生长速率在其单游动孢子无性系后代的遗传与变异。结果表明,亲本菌株JS-5的菌落形态和生长速率在连续4代单游动孢子无性系后代稳定遗传,而该菌株的自交S1代的约2／3单卵孢株的上述性状在其单拘无性后代中发生分离,且上述性状的分离在无性单孢后代中至少可连续保持3～4代。本研究结果提示,苎麻疫霉有性生殖导致上述性状的变异,除有性生殖过程中的基因重组外,有性生殖还诱导产生了一种新的机制参与上述性状遗传与变异的调控。本研究还观察到供试菌株及其后代所产生的游动孢子绝大多数细胞内只含有1个细胞核,表明上述机制与异核现象无关。有性生殖导致生物学性状在随后的无性后代中发生持续变异可能是苎麻疫霉生物学性状遗传多样性的重要机制之一。     

蛹虫草(Cordyceps militaris)无性型的多型现象*

在萨氏琼脂和PDA上,蛹虫草Cordycepsmilitaris（Vuill）Fr.无性型,蛹草拟青霉Paecilomycesmilitaris(Kob）Br.＆Sm．的一些单孢子株可自发产生突变,基于多型现象及其它形态特征可分三种类型：（1）具野生型菌株特征;产孢结构拟青霉型（Paecilomyces－type）,稳定,菌落通常不自发产生角变,大多数单孢子株属此类群。（2）属此类群的单孢子株,在PDA上可形成亮褐色至橙黄色的角变,它们典型的产孢结构为瓶梗轮生,分生孢子常聚集成头状的轮枝孢型（Verticillium－type）。在查氏培养基上则相反,拟青霉型的产孢结构占优势。单孢子株Cm-71是这一类群的代表。（3）单孢子株Cm－42是在所观察单孢子株中形态特征独特的菌株。在PDA上菌丝生长纤细,蛛网状,产孢结构轮枝孢型。蛹草拟青霉对氯酸钾（KClO3）不敏感,在浓度为6％（W／V）的KMM培养基上,23～24℃14天菌落的直径仅1～2cm,无气生菌丝生长。各单孢子株形成的不利用硝酸盐突变株（nit突变株）的途径和形态特征,也可分成类似于粉被虫草CordycepspruinosaPeteh不利用硝酸盐突变株（nit突变株）的三种类型。     

蛹虫草(Cordycepsmilitaris)无性型的多型现象

在萨氏琼脂和PDA上,蛹虫草Cordycepsmilitaris（Vuill）Fr.无性型,蛹草拟青霉Paecilomycesmilitaris(Kob）Br.＆Sm．的一些单孢子株可自发产生突变,基于多型现象及其它形态特征可分三种类型：（1）具野生型菌株特征;产孢结构拟青霉型（Paecilomyces－type）,稳定,菌落通常不自发产生角变,大多数单孢子株属此类群。（2）属此类群的单孢子株,在PDA上可形成亮褐色至橙黄色的角变,它们典型的产孢结构为瓶梗轮生,分生孢子常聚集成头状的轮枝孢型（Verticillium－type）。在查氏培养基上则相反,拟青霉型的产孢结构占优势。单孢子株Cm-71是这一类群的代表。（3）单孢子株Cm－42是在所观察单孢子株中形态特征独特的菌株。在PDA上菌丝生长纤细,蛛网状,产孢结构轮枝孢型。蛹草拟青霉对氯酸钾（KClO3）不敏感,在浓度为6％（W／V）的KMM培养基上,23～24℃14天菌落的直径仅1～2cm,无气生菌丝生长。各单孢子株形成的不利用硝酸盐突变株（nit突变株）的途径和形态特征,也可分成类似于粉被虫草CordycepspruinosaPeteh不利用硝酸盐突变株（nit突变株）的三种类型。     

Marine isolates of Aspergillus flavus: denizens of the deep or lost at sea?

Most fungal species from marine environments also live on land. It is not clear whether these fungi reach the sea from terrestrial sources as spores or other propagules, or if there are separate ecotypes that live and reproduce in the sea. The emergence of marine diseases has created an urgency to understand the distribution of these fungi. Aspergillus flavus is ubiquitous in both terrestrial and marine environments. This species is an opportunistic pathogen in many hosts, making it a good model to study the relationship between genetic diversity and specificity of marine fungi. In this study, an intraspecific phylogeny of A. flavus isolates based on Amplified Fragment Length Polymorphisms (AFLPs) was used to determine if terrestrial and marine isolates form discrete populations, and to determine if phylogeny predicts substratum specificity. Results suggest lack of population structure in A. flavus. All isolates may compose a single population, with no clade particular to marine environments.     

Trichoderma matsushimae and T. aeroaquaticum: two aero-aquatic species with Pseudaegerita-like propagules

Four isolates tentatively identified as Pseudaegerita matsushimae on the basis of the morphology of bulbil-like propagules were collected from substrates submerged in water in Thailand and Japan. In culture studies the two Thai isolates were found to produce phialoconidia on conidiogenous cells and phialoconidiophores whose morphology was similar to that of Trichoderma. Phylogenetic analysis based on D1/D2 regions of LSU rDNA sequences showed that the four isolates were nested in Hypocrea/Trichoderma (Hypocreales) while P. corticalis, the type species of Pseudaegerita, belongs to Hyaloscypha (Helotiales). Preliminary analysis by ISTH Web tools based on 5.8S-ITS rDNA and phylogenetic analysis based on rpb2 and tef1-int4 genes showed that the isolates have specific sequences of Trichoderma (anchors 1-5) and belong to the Hamatum clade but they grouped apart from any known species of Trichoderma. The sequences of the tef1-int4 gene, which were amplified from the authentic specimen of P. matsushimae (IMI 266915), also showed that it belongs to the Hamatum clade closely clustering with T. yunnanense but separate from our four isolates. The morphology of P. matsushimae (IMI 266915), especially the sizes of phialides and phialoconidia, were different from T. yunnanense. Thus, we conclude that IMI 266915 and our isolates are to be assigned to two different species in the Hamatum clade of Trichoderma, although both species have similar morphology of bulbils and phialoconidia. Morphology and molecular data revealed that P. matsushimae should be assigned to the genus Trichoderma as T. matsushimae and the Thai and Japanese isolates are placed in T. aeroaquaticum sp. nov. This finding supports the interpretation that aero-aquatic fungi have evolved from terrestrial fungi. We assume that these fungi probably were derived from typically soil-inhabiting species of Trichoderma; an adaptation to aquatic environments is shown by formation of bulbil-like propagules floating on water.     

病原真菌生态学研究现状及方法

通过形态学、生理生化学及分子生物学等方法鉴定由自然界分离的真菌菌种,构建系统发生树,了解菌种间亲缘关系,评估种间和种内的进化规律,了解真菌与其所处环境的相互关系,是真菌生态学的主要研究目的之一。本文就真菌与自然界和动物宿主的相互关系,环境中真菌的分离和鉴定方法及系统发生学研究及现状进行综述,以期了解致病真菌的生存环境、种类发生、感染途径等,达到尽早控制感染源、探讨致病性及防治疾病传播的目的。     

Oribatid Mites as Potential Vectors for Soil Microfungi: Study of Mite-Associated Fungal Species

The ability of soil-living oribatid mites to disperse fungal propagules on their bodies was investigated. Classical plating methods were applied to cultivate these fungi and to study their morphology. Molecular markers were used for further determination. The nuclear ribosomal large subunit and the nuclear ribosomal internal transcribed spacer of DNA extracts of the cultured fungi as well as total DNA extracts of the mites themselves, also containing fungal DNA, were amplified and sequenced. Based on phylogenetic analysis, a total of 31 fungal species from major fungal groups were found to be associated with oribatid mites, indicating that mites do not selectively disperse specific species or species groups. The detected taxa were mainly saprobiontic, cosmopolitan (e.g., Alternaria tenuissima), but also parasitic fungi (Beauveria bassiana) for whose dispersal oribatid mites might play an important role. In contrast, no mycorrhizal fungi were detected in association with oribatid mites, indicating that their propagules are dispersed in a different way. In addition, fungi that are known to be a preferred food for oribatid mites such as the Dematiacea were not detected in high numbers. Results of this study point to the potential of oribatid mites to disperse fungal taxa in soil and indicate that co-evolutionary patterns between oribatid mites and their associated fungi might be rare or even missing in most cases, since we only detected ubiquitous taxa attached to the mites.     

Evolution of ketosynthase domains of polyketide synthase genes in the Cladonia chlorophaea species complex (Cladoniaceae)

Lichen-forming fungi synthesize a diversity of polyketides, but only a few non-reducing polyketide synthase (PKS) genes from a lichen-forming fungus have been linked with a specific polyketide. While it is a challenge to link the large number of PKS paralogs in fungi with specific products, it might be expected that the PKS paralogs from closely related species would be similar because of recent evolutionary divergence. The objectives of this study were to reconstruct a PKS gene phylogeny of the Cladonia chlorophaea species complex based on the ketosynthase domain, a species phylogeny of the complex, and to explore the presence of PKS gene paralogs among members of the species complex. DNA was isolated from 51 individuals of C. chlorophaea and allies to screen for the presence of 13 PKS paralogs. A 128 sequence PKS gene phylogeny using deduced amino acid sequences estimated from the 13 PKS paralogs and sequences subjected to BLASTx comparisons showed losses of each of two PKS domains (reducing and methylation). This research provided insight into the evolution of PKS genes in the C. chlorophaea group, species evolution in the group, and it identified potential directions for further investigation of polyketide synthesis in the C. chlorophaea species complex.     

Reconstructing the evolution of agarics from nuclear gene sequences and basidiospore ultrastructure

Traditional classifications of agaric fungi involve gross morphology of their fruit bodies and meiospore print-colour. However, the phylogeny of these fungi and the evolution of their morphological and ecological traits are poorly understood. Phylogenetic analyses have already demonstrated that characters used in traditional classifications of basidiomycetes may be heavily affected by homoplasy, and that non-gilled taxa evolved within the agarics several times. By integrating molecular phylogenetic analyses including domains D1–D3 and D7–D8 of nucLSU rDNA and domains A–C of the RPB1 gene with morphological and chemical data from representative species of 88 genera, we were able to resolve higher groups of agarics. We found that the species with thick-walled and pigmented basidiospores constitute a derived group, and hypothesize that this specific combination of characters represents an evolutionary advantage by increasing the tolerance of the basidiospores to dehydration and solar radiation and so opened up new ecological niches, e.g. the colonization of dung substrates by enabling basidiospores to survive gut passages through herbivores. Our results confirm the validity of basidiospore morphology as a phylogenetic marker in the agarics.     

An ecological view of the formation of VA mycorrhizas

In spite of the major advances in understanding the functioning of symbioses between plants and arbuscular mycorrhizal fungi, details of the ecology of mycorrhizal fungi are not well documented. The benefits of the association are related to the timing and extent of colonization of roots, and fungi differ in their contribution to plant growth and presumably to soil aggregation. Knowledge of the processes that lead to successful colonization of roots by beneficial fungi at appropriate times for the host plants will form the basis of guidelines for soil management to maximize the benefits from the symbiosis. Fungi differ in the manner and extent to which they colonize roots. They also differ in their capacity to form propagules. The importance of hyphae, spores and propagules within living or dead mycorrhizal roots also differs among species and for the same species in different habitats. The relationships between colonization of roots and propagule formation, and between propagule distribution and abundance and subsequent mycorrhiza formation, for different fungi in field environments, are not well understood. Methods for quantifying mycorrhizal fungi are not especially suitable for distinguishing among different fungi within roots. Consequenctly, the dynamics of colonization of roots by different fungi, within and between seasons, have been little studied. Research is required that focuses on the dynamics of fungi within roots as well as on changes in the abundance of propagules of different fungi within soil. Interactions between fungi during the colonization of roots, the colonization of soil by hyphae and sporulation are all poorly understood. Without knowledge of these processes, it will by difficult to predict the likely success of inoculation with introduced fungi. Such knowledge is also required for selecting soil management procedures to enhance growth and survival of key species within the population. The relative tolerance of various fungi to perturbations in their surroundings will provide a basis for identifying those fungi that are likely to persist in specific environments. The processes that influence mycorrhizal fungi in field soils can be identified in controlled studies. However, greater emphasis is required on studying these processes with mixed populations of fungi. The role played by diversity within populations of mycorrhizal fungi is virtually unexplored.     

Keratinophilic fungi associated with starlings Sturnus vulgaris in Brittany,France

The occurrence of keratinophilic fungi on the feathers of 110 starlings (Sturnus vulgaris) was investigated in order to assess the part that these birds may have in spreading of fungi of medical interest. Thirty eight birds (26%) yielded keratinophilic fungi, primarily Chrysosporium species, but also Scopulariopsis brevicaulis and Trichophyton terrestre. We reviewed the pathogenicity of the isolated fungi. Starlings may spread the propagules of keratinophilic fungi but it is difficult to determine if their increasing numbers in cities can be of significance in the epidemiology of today's rare mycoses. This revised version was published online in June 2006 with corrections to the Cover Date.     

A phylogeny of Cenozoic macroperforate planktonic foraminifera from fossil data

We present a complete phylogeny of macroperforate planktonic foraminifer species of the Cenozoic Era (∼65 million years ago to present). The phylogeny is developed from a large body of palaeontological work that details the evolutionary relationships and stratigraphic (time) distributions of species‐level taxa identified from morphology (‘morphospecies’). Morphospecies are assigned to morphogroups and ecogroups depending on test morphology and inferred habitat, respectively. Because gradual evolution is well documented in this clade, we have identified many instances of morphospecies intergrading over time, allowing us to eliminate ‘pseudospeciation’ and ‘pseudoextinction’ from the record and thereby permit the construction of a more natural phylogeny based on inferred biological lineages. Each cladogenetic event is determined as either budding or bifurcating depending on the pattern of morphological change at the time of branching. This lineage phylogeny provides palaeontologically calibrated ages for each divergence that are entirely independent of molecular data. The tree provides a model system for macroevolutionary studies in the fossil record addressing questions of speciation, extinction, and rates and patterns of evolution.     

Geographic and phylogenetic evidence for dispersed nuclear introgression in a daphniid with sexual propagules

The role of among-species gene flow in eukaryotic evolution remains controversial. Putative hybrid lineages are common in water fleas, but their ecological success is often associated with polyploidy and the production of asexual propagules. Advanced hybrid lineages with sexual propagules are expected to be geographically restricted because their successful dispersal is contingent on overcoming fertility complications, assimilation by parent taxa, and competition with parent taxa. Here we provide evidence that a diploid lineage of Daphnia has been formed by introgression between distantly related species and attained a broad distribution (Nearctic) despite its requirement for sexual propagules. The evidence is based on geographical discordance, phylogenetic discordance, recombinant genotypes and additive genotypes of the nuclear internal transcribed spacer regions (ITS) and mitochondrial DNA. Additive genotypes also provided evidence of hybridization between introduced European Daphnia and North American Daphnia. We argue that the unique biology of Holarctic lacustrine water fleas and the spatial separation of lineages during Pleistocene glaciation have promoted hybridization and its evolutionary consequences.