Fungal and plant gene expression during the colonization of cacao seedlings by endophytic isolates of four Trichoderma species

Endophytic isolates of Trichoderma species are being considered as biocontrol agents for diseases of Theobroma cacao (cacao). Gene expression was studied during the interaction between cacao seedlings and four endophytic Trichoderma isolates, T. ovalisporum-DIS 70a, T. hamatum-DIS 219b, T. harzianum-DIS 219f, and Trichoderma sp.-DIS 172ai. Isolates DIS 70a, DIS 219b, and DIS 219f were mycoparasitic on the pathogen Moniliophthora roreri, and DIS 172ai produced metabolites that inhibited growth of M. roreri in culture. ESTs (116) responsive to endophytic colonization of cacao were identified using differential display and their expression analyzed using macroarrays. Nineteen cacao ESTs and 17 Trichoderma ESTs were chosen for real-time quantitative PCR analysis. Seven cacao ESTs were induced during colonization by the Trichoderma isolates. These included putative genes for ornithine decarboxylase (P1), GST-like proteins (P4), zinc finger protein (P13), wound-induced protein (P26), EF-calcium-binding protein (P29), carbohydrate oxidase (P59), and an unknown protein (U4). Two plant ESTs, extensin-like protein (P12) and major intrinsic protein (P31), were repressed due to colonization. The plant gene expression profile was dependent on the Trichoderma isolate colonizing the cacao seedling. The fungal ESTs induced in colonized cacao seedlings also varied with the Trichoderma isolate used. The most highly induced fungal ESTs were putative glucosyl hydrolase family 2 (F3), glucosyl hydrolase family 7 (F7), serine protease (F11), and alcohol oxidase (F19). The pattern of altered gene expression suggests a complex system of genetic cross talk occurs between the cacao tree and Trichoderma isolates during the establishment of the endophytic association.Electronic Supplementary Material Supplementary material is available to authorised users in the online version of this article at .     

Proteomic analysis of a filamentous fungal endophyte using EST datasets

Proteomic analysis of many species of fungi, particularly filamentous fungi, is difficult due to the lack of publicly available genome sequence data and the problems associated with cross‐species comparisons. Furthermore, the detection of fungal proteins in biological systems where there are a greater number of proteins present from other eukaryote species provides additional challenges. We present an EST‐based approach for identifying proteins from a fungal endophyte of temperate grasses and demonstrate that this method is well suited for fungi with minimal sequence data.     

Localization of the insect pathogenic fungal plant symbionts Metarhizium robertsii and Metarhizium brunneum in bean and corn roots

Metarhizium is an insect pathogenic fungus and a plant root symbiont. Here the root association patterns (rhizoplane or endophytic colonization) were analyzed in common beans (Phaseolus vulgaris) and sweet corn (Zea mays) using M. robertsii and M. brunneum under various vermiculite treatments (control, with sucrose, with an insect) at two time points of plant growth (10 and 20 days). We observed that M. brunneum and M. robertsii preferentially endophytically colonized the hypocotyl, however, greater rhizoplane colonization was observed at the regions proximal to the hypocotyl in both plants. Vermiculite amended with an infected insect resulted in greater endophytic and rhizoplane colonization at 20 days compared to 10 days, for both plants as well as for both Metarhizium species. Regardless of the vermiculite treatment, corn was preferentially colonized compared to bean. Sucrose amendment in the vermiculite and infected insect amended vermiculite only showed differences in rhizoplane colonization. The greatest root association occurred with M. brunneum with an infected insect and that in corn after 20 days.     

Altered allocation to roots and shoots in the endophyte‐infected seedlings of Puccinellia distans (Poaceae)

Endophytes play an important role in ecological and evolutionary processes in plants and have marked economic value. Seed‐transmitted fungal endophytes are conventionally regarded as mutualistic symbionts, but their fitness consequences for the offspring of the host are not clear. Puccinellia distans infected with the fungus Epichloë typhina (E+) produces seeds that are several times smaller than normal (E?). This observation suggests that the E+ seedlings face a developmental disadvantage. Our growth chamber experiments compared the germination rates of the small E+ and large E? seeds of P. distans and examined the biomass allocation of seedlings to roots and shoots. The E+ seedlings germinated more slowly and maintained shorter shoots and a smaller root biomass for 30–50 days after sowing. Despite this disadvantage, the E+ plants more quickly increased their total size, attaining a larger shoot and whole‐plant biomass. The shoot:root biomass ratio increased more rapidly through time in the E+ seedlings, attaining a value nine times higher in the E+ than the E? group 50 days after sowing. Such differences between the E+ and E? seedlings were not explained by the growth allometry between shoots and roots. The seedlings of P. distans infected with the Epichloë endophyte were initially handicapped by their postponed emergence, but this disadvantage was quickly overcome by their superior growth capacity. The decrease in the relative allocation to roots may indicate that endophytes increase the performance of roots as resource‐acquiring organs and/or reduce the role of roots in protection against herbivores.     

Palatability of the Antarctic rhodophyte Palmaria decipiens (Reinsch) RW Ricker and its endo/epiphyte Elachista antarctica Skottsberg to sympatric amphipods

Although many macroalgae that occur throughout Western Antarctic Peninsular waters are known to produce defensive secondary metabolites that deter grazing, the rhodophyte, Palmaria decipiens is palatable to several sympatric meso- and macro-grazers. It has been hypothesized that high levels of mesoherbivory by amphipods may account for the conspicuous lack of filamentous epiphytes emerging from the thalli of marcophytes in this region. Nonetheless, Elachista antarctica is a filamentous phaeophyte found growing within, and emerging from the thallus of the rhodophyte P. decipiens. It is surprising that E. antarctica occurs exclusively in association with a palatable species of macroalgae considering the standing biomass of other chemically defended unpalatable species is very high. We tested the hypothesis that E. antarctica grows on P. decipiens due to the host's overwhelming palatability compared to that of the epiphyte. That is, the hypothesis that mesograzers prefer the host over the epiphyte, grazing around emerging filaments. Choice and no choice feeding assays with live tissues of E. antarctica and P. decipiens were conducted in three different trials with four sympatric amphipod species (Prostebbingia gracilis, Gondogeneia antarctica, Oradarea bidentata, and Paraphimedia integricauda) commonly found in association with P. decipiens. G. antarctica consumed both species but ate P. decipiens at a faster rate than the epiphyte in two of three trials. P. gracilis, O. bidentata, and P. integricauda fed on the epiphyte, E. antarctica at faster rates than upon P. decipiens. Aggressive grazing of the epiphyte by this suite of amphipods indicates that differences in palatability and differences in grazing pressure on host and epiphyte do not explain the exclusive epiphytism of E. antarctica on P. decipiens.     

Conjugation of antifungal benzoic acid derivatives as a path for detoxification in Penicillium brasilianum,an endophyte from Melia azedarach

In this study, the consumption of 4-bromobenzoic acid and 4-chlorobenzoic acid by the fungus Penicillium brasilianum, an endophyte from Melia azedarach is evaluated. This fungus metabolizes these halobenzoic acids to produce three new brominated compounds, which have been isolated and characterized, and three new chlorinated derivatives identified by HRMS. Among these products, (4-bromobenzoyl)proline has been also chemically synthesized and employed in biological assays, thus providing insights for the elucidation of the defense mechanism of P. brasilianum towards these halobenzoic acids.     

Molecular characterization of endophytic strains of Fusarium verticillioides (= Fusarium moniliforme) from maize (Zea mays. L)

The species Fusarium verticillioides (= F. moniliforme) is often found in maize seeds, constituting an important source of inoculum in the field. Fusarium spp., associated with symptomatic and asymptomatic plants, may be a primary causal agent of disease, a secondary invader or an endophyte. In the present work, endophytic fungi were isolated from two populations of Zea mays (BR-105 and BR-106) and their respective inbred lines. Within different inbred lines of maize, Fusarium was found at a frequency of 0 to 100% relative to the number of total isolated fungi. The frequency with which the genus occurred was practically the same in the two field sites (around 60%). Twenty-one F. verticillioides strains were analysed using the random amplified polymorphic DNA (RAPD) technique, employing 10 random primers. Variability analysis of endophytic isolates via RAPD showed genome polymorphism taxa of species around 60%. Endophytic isolates were clustered by their sites of origin. RAPD analysis clustered the endophytic isolates by their maize inbred lines hosts (Mil-01 to Mil-06), whereas at site A they clustered into two major groups related to the maize gene pool (BR-105 or BR-106 population). All strains isolated from seeds collected in Site A, except strains L9 and L10, were sub-grouped according to maize inbred lines. The analysis showed a discrete sub-grouping at site B. Results obtained here could be explained by a co-evolution process involving endophytic isolates of F. verticillioides and maize inbred lines.     

内生真菌侵染对黑麦草种子萌发、幼苗生长及渗透胁迫抗性的影响

 以含有内生真菌的黑麦草(Lolium perenne L.)种子为材料,采用4 ℃冰箱内和20 ℃培养箱内保存18个月的方式分别构建内生真菌侵染（EI）和内生真菌非侵染（EF）的黑麦草种群,通过比较EI和EF种群在正常条件下（对照）和渗透胁迫条件下种子发芽、幼苗生长等方面的差异,探讨内生真菌对其宿主植物的直接和间接影响。结果表明：在对照和胁迫条件下,EI种子的发芽势及发芽率均明显高于EF种子,而在重度胁迫下EI植株的叶延伸速率、根系总长度高于EF植株。内生真菌对宿主植物分蘖数和生物量的变化没有促进作用,但