Mutagenic potential and modulatory effects of the medicinal plant Luehea divaricata (Malvaceae) in somatic cells of Drosophila melanogaster: SMART/wing

Luehea divaricata is a native plant of the Brazilian Cerrado, known as "a?oita-cavalo". It is used as a popular herbal medicine in the treatment of dysentery, bleeding, arthritis, tumors, ulcers, and gangrenous wounds. Considering that herbal medicines sometimes provoke tumors and/or may prevent mutational events, it is important to study the action of these natural drugs on DNA. Aqueous extract of the bark of L. divaricata was evaluated at three different concentrations (0.10, 0.30, 0.50 mg/mL), individually and in combination with the neoplastic drug doxorubicin (DXR), by the somatic mutation and recombination test (SMART/wing) in Drosophila melanogaster. Distilled water was included as a negative control. The mutation frequency in the treatments with L. divaricata extract alone was not significantly higher than in the negative control for standard (ST) and high bioactivation (HB) crosses. When L. divaricata extract was combined with DXR, there was a significant reduction in the frequency of spots when compared to DXR alone, in both crosses. Further studies with other experimental models would be useful to confirm that L. divaricata extract is not harmful and that it could be used in the prevention of cancer.     

Genetic structure of the Atlantic Rainforest tree species Luehea divaricata (Malvaceae)

The Atlantic Rain Forest is one of the most important Brazilian biomes and a hotspot for biodiversity that is characterized by its high level of endemism, where new species are still being described. Luehea divaricata (Malvaceae) is commonly found in riparian forests areas of the Atlantic forest. Because of the importance of this species in reforestation programs, we used nine pairs of microsatellite loci to study the genetic variability of this species along its distribution area and verify if fragmentation is compromising the survival of these populations. A total of 50 alleles were obtained with an average observed and expected heterozygosity of 0.53 and 0.67, respectively. Seven of the nine populations studied showed a heterozygosity deficit. Most of the genetic diversity was found within populations; while the level of genetic differentiation was moderated (6.84) between populations. Different levels of gene flow between the populations were detected. Positive and significant values of Fis were found for seven populations. The signal test for excess of heterozygosity indicated that a recent genetic bottleneck occurred in the fragmented populations. The dendrogram constructed by the UPGMA method revealed the formation of seven clusters, which was confirmed by the Bayesian analysis for number of K clusters. The presence of several pairs of loci in linkage disequilibrium confirms that these populations experienced a loss of genetic diversity caused by genetic drift. The results showed that it is necessary to develop management strategies for the conservation of these populations of L. divaricata as the viability of the next generations are severely compromised.     

Molecular characterization of fungal endophyte diversity isolated from Aconitum heterophyllum: a critically endangered medicinal plant of Kashmir Himalaya

International Microbiology - Aconitum heterophyllum is a rare perennial herb from Kashmir Himalayas. Due to its threatened status and dependence on its environment, the plant was examined for any...     

A foliar endophyte increases the diversity of phosphorus-solubilizing rhizospheric fungi and mycorrhizal colonization in the wild grass Bromus auleticus

Asexual Epichloë endophytes establish mutualistic symbioses with grasses, improve fitness of their hosts and modify the surrounding environment. To test the hypothesis that this symbiotic association increases the abundance and diversity of phosphate-solubilizing fungi (PSF), a pot experiment was conducted combining two endophytic statuses: Epichloë-infected (E+) and non-infected (E−) Bromus auleticus plants, and two soil types collected from agricultural (A) and non-agricultural (NA) fields. Soil fungi were isolated at the beginning of the experiment and 12 months after the introduction of B. auleticus, and tested for their inorganic P (Pi)-solubilizing capability. Arbuscular mycorrhizal colonization in B. auleticus roots of E+ and E− plants was also analyzed. PSF abundance was affected by the endophytic status and by the type of soil; the highest value was detected in the E−NA treatment, followed by the E+A treatment. PSF diversity was higher in NA than in A soils and higher in soils treated with E+ than in those treated with E−. Arbuscular mycorrhizal fungi colonization was higher in E+ plants. We hypothesize that the positive association between Epichloë endophytes and mycorrhizal fungi with an increase in the PSF diversity would generate an increase in the phosphorus (P) available to plants.     

Diversity and antimicrobial activities of the fungal endophyte community associated with the traditional Brazilian medicinal plant Solanum cernuum Vell. (Solanaceae)

The diversity and antimicrobial activity of endophytic fungi associated with the Brazilian medicinal plant Solanum cernuum Vell. were studied during summer and winter seasons. A total of 246 fungal isolates were obtained, including 225 filamentous fungi and 21 yeasts. They were identified by morphological, physiological, and molecular methods. Fifty-five different taxa represented by the phyla Ascomycota (33 taxa), Basidiomycota (21 taxa), and Zygomycota (one taxon) were identified. The most abundant taxa were closely related to Arthrobotrys foliicola , Colletotrichum gloeosporioides , Coprinellus radians , Glomerella acutata , Diatrypella frostii , Phoma glomerata , Mucor sp., Phlebia subserialis , Phoma moricola , Phanerochaete sordida , and Colletotrichum sp. A total of 265 fungal extracts were screened and 64 (26.01%) displayed antimicrobial activities. Among these extracts, 18 (28.12%) presented antibacterial and antifungal activities, 42 (65.62%) displayed selective antibacterial activity, and four (6.25%) exhibited only antifungal activity. The best values of minimum inhibitory concentration were obtained from extracts of Cryptococcus rajasthanensis , Glomerella acutata, Leptosphaeria sp., and Phoma glomerata ranging from 7.8 to 15.62?μg/mL. This study is the first survey of the endophytic fungi community associated with S. cernuum, and our results show that they can represent a promising source of bioactive compounds.     

药用植物绶草(兰科)的菌根真菌群落组成分析

兰科菌根真菌(OMF)被认为是影响兰科植物物种丰度和分布的一个重要因素.对广域分布兰科植物的菌根区系进行研究有助于人们更深入地了解兰科植物分布格局的形成机制.本研究以我国广域分布的兰科药用植物绶草Spiranthes sinensis为材料,采用Illumina Miseq高通量测序技术对北京、上海、江西、广西、云南、...     

Characteristics of foliar fungal endophyte assemblages and host effective components in Salvia miltiorrhiza Bunge

Salvia miltiorrhiza Bunge, a well-known medicinal plant, has more than 20 effective components. The aim of this study was to comprehensively investigate foliar fungal endophyte communities of S. miltiorrhiza and explore the inside relationship between host-specific fungal endophytes and effective components accumulation. Five plant samples were collected from four geological different provinces in China. Foliar fungal endophyte communities were determined by terminal restriction fragment length polymorphism (T-RFLP) of the ITS region. Effective components were analyzed with high-performance liquid chromatography. The results showed that S. miltiorrhiza foliage harbored a large diversity of fungal endophytes. Principal component analysis revealed similar T-RFLP profiles and the characteristics of the 24 effective components among the five samples, which could be clustered into three groups. In foliar T-RFLP profiles derived from the restriction digestion by CfoI, HaeIII, MspI, or TaqI, there were identical 45, 42, 38, and 34 terminal restriction fragments (T-RFs) from the five samples. We consider these T-RFs as host-specific fungal endophytes. Correlation analysis of these T-RFs’ area and 24 effective components contents revealed a significant correlationship between some host-specific fungal endophytes and foliage or root effective components accumulation.     

Clonal diversity and conservation genetics of the medicinal plant Carapichea ipecacuanha (Rubiaceae)

The roots of the understorey shrub Carapichea ipecacuanha (ipecac) have medicinal properties, and the uprooting of wild plants has supplied most of the world demand for this species. Although under severe population decline, C. ipecacuanha lacks legal protection. In the wild, the aerial stems of ipecac clump together to form clusters with well-defined borders. Cluster size may range from several to hundreds of aerial stems. To investigate the extent of clonality among aerial stems in ipecac clusters, we sampled 50 wild clusters (a total of 291 aerial stems) and screened them with 89 inter-simple sequence repeat (ISSR) markers. The 291 aerial stems were grouped into 42 putative clones. The clonal groups generally consisted of aerial stems from the same cluster, and there was little or no genetic differentiation among aerial stems at the cluster level. These findings suggest that strategies designed to conserve ipecac in situ should not rely upon census data, which are based on the number of aerial stems per cluster and the number of clusters per population, because such data greatly underestimate the species effective population size and genetic diversity. Our results also indicate that this species needs protection at a federal level.     

药用植物防风(Saposhnikovia divaricata)不同时期光合作用特性比较

通过对防风光合特性及光合有效辐射(PAR)、大气二氧化碳浓度(Ca)、气温(Ta)、叶温(Tl)和相对湿度(RH)等环境因子的同期观测,系统地研究了防风净光合速率(Pn)等在不同时期的日变化特征及其与环境因子之间的关系,为吉林省防风规范化种植提供理论依据.结果表明,不同时期防风净光合速率日变化均呈双峰型曲线,具有明显的光合"午休"现象.7月份防风光合速率中午降低主要由非气孔限制引起,8月份、9月份防风光合速率中午降低则主要由气孔限制引起,7月份高温低湿是限制防风Pn的主要环境因子.光合速率和蒸腾速率的不同步是引起水分利用效率变化的主要原因.光合有效辐射直接影响防风叶片的光合速率,从而影响防风有机物质的合成与转化,对防风生产力起到直接的限制作用.不同时期防风Pn与PAR呈显著正相关,不同时期防风叶片对不同环境因子的反映有所不同,环境因子中对Pn直接作用由大到小为7月份 Ta＞PAR＞Tl＞Ca＞RH,8月份为Tl＞PAR＞Ta＞RH＞Ca,9月份为Ta＞PAR＞Tl＞Ca＞RH.     

Effects of host plant environment and Ustilago maydis infection on the fungal endophyte community of maize (Zea mays)

The focus of many fungal endophyte studies has been how plants benefit from endophyte infection. Few studies have investigated the role of the host plant as an environment in shaping endophyte community diversity and composition. The effects that different attributes of the host plant, that is, host genetic variation, host variation in resistance to the fungal pathogen Ustilago maydis and U. maydis infection, have on the fungal endophyte communities in maize (Zea mays) was examined. The internal transcribed spacer (ITS) region of the rDNA was sequenced to identify fungi and the endophyte communities were compared in six maize lines that varied in their resistance to U. maydis. It was found that host genetic variation, as determined by maize line, had significant effects on species richness, while the interactions between line and U. maydis infection and line and field plot had significant effects on endophyte community composition. However, the effects of maize line were not dependent on whether lines were resistant or susceptible to U. maydis. Almost 3000 clones obtained from 58 plants were sequenced to characterize the maize endophyte community. These results suggest that the endophyte community is shaped by complex interactions and factors, such as inoculum pool and microclimate, may be important.     

Host associations and beta diversity of fungal endophyte communities in New Guinea rainforest trees

Processes shaping the distribution of foliar fungal endophyte species remain poorly understood. Despite increasing evidence that these cryptic fungal symbionts of plants mediate interactions with pathogens and herbivores, there remain basic questions regarding the extent to which dispersal limitation and host specificity might shape fungal endophyte community composition in rainforests. To assess the relative importance of spatial pattern and host specificity, we isolated fungi from a sample of mapped trees in lowland Papua New Guinea. Sequences of the internal transcribed spacer (ITS) region were obtained for 2079 fungal endophytes from three sites and clustered into molecular operational taxonomic units (MOTUs) at 95% similarity. Multivariate analyses suggest that host affinity plays a significant role in structuring endophyte community composition whereas there was no evidence of endophyte spatial pattern at the scale of tens to hundreds of metres. Differences in endophyte communities between sampled trees were weakly correlated with variation in foliar traits but not with tree species relatedness. The dominance of relatively few generalist endophytes and the presence of a large number of rare MOTUs was a consistent observation at three sites separated by hundreds of kilometres and regional turnover was low. Host specificity appears to play a relatively weak but more important role than dispersal limitation in shaping the distribution of fungal endophyte communities in New Guinea forests. Our results suggest that in the absence of strong ecological gradients and host turnover, beta diversity of endophyte communities could be low in large areas of contiguous forest.     

Patterns and regulation of mycorrhizal plant and fungal diversity

The diversity of mycorrhizal fungi does not follow patterns of plant diversity, and the type of mycorrhiza may regulate plant species diversity. For instance, coniferous forests of northern latitudes may have more than 1000 species of ectomycorrhizal (EM) fungi where only a few ectomycorrhizal plant species dominate, but there are fewer than 25 species of arbuscular mycorrhizal (AM) fungi in tropical deciduous forest in Mexico with 1000 plant species. AM and EM fungi are distributed according to biome, with AM fungi predominant in arid and semiarid biomes, and EM fungi predominant in mesic biomes. In addition, AM fungi tend to be more abundant in soils of low organic matter, perhaps explaining their predominance in moist tropical forest, and EM fungi generally occur in soils with higher surface organic matter.EM fungi are relatively selective of host plant species, while AM tend to be generalists. Similar morphotypes of AM fungi collected from different sites confer different physiological benefits to the same plant species. While the EM fungi have taxonomic diversity, the AM fungi must have physiological diversity for individual species to be so widespread, as supported by existing studies. The environmental adaptations of mycorrhizal fungi are often thought to be determined by their host plant, but we suggest that the physiology and genetics of the fungi themselves, along with their responses to the plant and the environment, regulates their diversity. We observed that one AM plant species,Artemisia tridentata, was associated with different fungal species across its range, indicating that the fungi can respond to the environment directly and must not do so indirectly via the host. Different species of fungi were also active during different times of the growing season on the same host, again suggesting a direct response to the environment.These patterns suggest that even within a single functional group of microorganisms, mycorrhizal fungi, considerable diversity exists. A number of researchers have expressed the concept of functional redundancy within functional groups of microorganisms, implying that the loss of a few species would not be detectable in ecosystem functioning. However, there may be high functional diversity of AM fungi within and across habitats, and high species diversity as well for EM fungi. If one species of mycorrhizal fungus becomes extinct in a habitat, field experimental data on AM fungi suggest there may be significant shifts in how plants acquire resources and grown in that habitat.     

菌根真菌多样性与植物多样性的相互作用研究进展

菌根共生双方多样性影响着生态系统的过程与功能。菌根真菌-寄主植物之间的共生组合存在偏好性或特异性,这导致菌根真菌对寄主植物的效益差异和寄主植物对菌根真菌的利益差别：两者在互利共生过程中不仅相互选择,还存在相互促进与制约的关系(如互补与选择效应、竞争),从而在一定程度上决定生态系统的演化与发展。本文概述了植物多样性与菌根真菌多样性的相互影响,探讨了两者互作可能存在的调控因素与机制,对存在的问题和争议进行了总结,并提出了进一步研究的方向。深入阐明植物多样性与菌根真菌多样性之间的互作关系,将丰富生物共生学理论,增强菌根应用潜力及生物多样性的维持。     

Species delimitation in fungal endophyte diversity studies and its implications in ecological and biogeographic inferences

The estimation of species diversity in fungal endophyte communities is based either on species counts or on the assignment of operational taxonomic units (OTUs). Consequently, the application of different species recognition criteria affects not only diversity estimates but also the ecological hypotheses that arise from those observations. The main objective of the study was to examine how the choice and number of genetic markers and species delimitation criteria influence biodiversity estimates. Here, we compare approaches to defining species boundaries in three dominant species complexes of tropical endophytes, specially Colletotrichum gloeosporioides agg., Pestalotiopsis microspora agg. and Trichoderma harzianum agg., from two Amazonian trees: Hevea brasiliensis and H. guianensis. Molecular tools were used to describe and compare the diversity of the different assemblages. Multilocus phylogenetic analyses [gpd, internal transcribed spacer (ITS) and tef1] and modern techniques for phylogenetic species delimitation were overlaid with ecological data to recognize putative species or OTUs. The results demonstrate that ITS alone generally underestimates the number of species predicted by other nuclear loci. These results question the use of ITS and arbitrary divergence thresholds for species delimitation.     

Evaluation of genetic diversity in wild populations of Peganum harmala L., a medicinal plant

Peganum harmala L. is a perennial herbaceous plant and can be a future drug due to its wide medicinal purposes. Despite its economic importance, the molecular genetics of P. harmal have not yet been studied in detail. Genetic diversity of 12 P. harmala genotypes were investigated by using Inter-Simple Sequence Repeats (ISSR), PCR-RFLP of rDNA-ITS, PCR-SSCP of rDNA-ITS and Simple Sequence Repeat (SSR) markers. The level of polymorphism revealed by ITS-SSCP is the lowest, followed by ITS-RFLP then ISSR and the highest polymorphism level was reported for SSR marker. The AMOVA analysis implied that most of the variation occurred within the Populations. A value of inbreeding coefficient F_is estimated by the three co-dominant markers was nearly equal and offer an indication of the partial out-crossing reproductive system of P. harmala. Principal Coordinate Analysis (PCOA) plot revealed a clear pattern of clustering based on the locations of collected plants which coincide with the isolation by distance. The study revealed that ITS-SSCP and ISSR markers respectively were more informative than the other used markers in the assessment of genetic diversity of P. harmala. The results reflect the great diversity of P. harmala and data obtained from this study can be used for future collecting missions.     

Genetic diversity of the traditional Chinese medicinal plant Ypsilandra thibetica (Melanthiaceae): Applications for conservation

Twelve microsatellite markers were developed to determine the genetic diversity and genetic structure of Ypsilandra thibetica, represented by a total of 90 individuals from six natural populations. All twelve microsatellite loci were polymorphic, and the results indicated that a high genetic diversity was present within populations (mean R_S = 4.996; mean H_E = 0.615), with high levels of genetic structure (mean F_ST = 0.165; mean F_IS = 0.692) among populations. This pattern is likely attributable to consanguineous mating, and this hypothesis is supported by a low relatedness coefficient. Our study suggested that environment factors might restrict gene flow among populations. In addition, physical distances between populations were not related to genetic distances, implying that ancestral populations might have been distributed over a wider area. These results suggest that Y. thibetica should be a high priority for conservation managers.     

Does peptide-nucleic acid (PNA) clamping of host plant DNA benefit ITS1 amplicon-based characterization of the fungal endophyte community?

Fungal endophyte community amplicon sequencing can lose a significant number of informative reads due to host-plant co-amplification. Blocking of plant-specific sequences with peptide nucleic acid (PNA) clamps has been shown to improve metrics of detected microbial diversity in studies targeting 16S and 18S regions of rRNA genes. However, PNA clamping has not been applied to the plant ITS region of rRNA gene – a widely accepted fungal marker. By applying PNA clamping technique to ITS amplicon sequencing of the endophytic fungal community of elderberry this study shows that PNA clamping significantly reduces host-plant co-amplification with the universal ITS1/ITS4 primer set. However, PNA clamping in combination with the discriminatory ITS1F/ITS2 primer set did not improve the metrics of fungal endophyte community ITS amplicon Illumina sequencing. This study shows that PNA clamping does not add practical benefit to taxonomic profiling of plant-associated fungal communities if the primers are already specific enough to exclude amplification of host DNA.     

Exopolysaccharide production is required for biofilm formation and plant colonization by the nitrogen-fixing endophyte Gluconacetobacter diazotrophicus

The genome of the endophytic diazotrophic bacterial species Gluconacetobacter diazotrophicus PAL5 (PAL5) revealed the presence of a gum gene cluster. In this study, the gumD gene homologue, which is predicted to be responsible for the first step in exopolysaccharide (EPS) production, was insertionally inactivated and the resultant mutant (MGD) was functionally studied. The mutant MGD presented normal growth and nitrogen (N(2)) fixation levels but did not produce EPS when grown on different carbon sources. MGD presented altered colony morphology on soft agar plates (0.3% agar) and was defective in biofilm formation on glass wool. Most interestingly, MGD was defective in rice root surface attachment and in root surface and endophytic colonization. Genetic complementation reverted all mutant phenotypes. Also, the addition of EPS purified from culture supernatants of the wild-type strain PAL5 to the mutant MGD was effective in partially restoring wild-type biofilm formation and plant colonization. These data provide strong evidence that the PAL5 gumD gene is involved in EPS biosynthesis and that EPS biosynthesis is required for biofilm formation and plant colonization. To our knowledge, this is the first report of a role of EPS in the endophytic colonization of graminaceous plants by a nitrogen-fixing bacterium.     

Effects of elevated CO2, nitrogen deposition, and decreased species diversity on foliar fungal plant disease

Three components of global change, elevated CO₂, nitrogen addition, and decreased plant species richness (‘diversity’), increased the percent leaf area infected by fungi (pathogen load) for much to all of the plant community in one year of a factorial grassland experiment. Decreased plant diversity had the broadest effect, increasing pathogen load across the plant community. Decreased diversity increased pathogen load primarily by allowing remaining plant species to increase in abundance, facilitating spread of foliar fungal pathogens specific to each plant species. Changes in plant species composition also strongly influenced community pathogen load, with communities that lost less disease prone plant species increasing more in pathogen load. Elevated CO₂ increased pathogen load of C₃ grasses, perhaps by decreasing water stress, increasing leaf longevity, and increasing photosynthetic rate, all of which can promote foliar fungal disease. Decreased plant diversity further magnified the increase in C₃ grass pathogen load under elevated CO₂. Nitrogen addition increased pathogen load of C₄ grasses by increasing foliar nitrogen concentration, which can enhance pathogen infection, growth, and reproduction. Because changes in foliar fungal pathogen load can strongly influence grassland ecosystem processes, our study suggests that increased pathogen load can be an important mechanism by which global change affects grassland ecosystems.