海南霸王岭热带雨林常见植物丛枝菌根真菌调查

对海南霸王岭热带雨林的12科16种常见植物的丛枝菌根状况进行了调查,用碱解离-酸性品红染色法进行了真菌鉴定。结果表明,13种植物形成典型的丛枝菌根,占所调查植物的81%;3种植物没有形成丛枝菌根,占所调查植物的19%。用湿筛沉淀法从这些植物根际土壤中共分离鉴定出了3属11种丛枝菌根真菌(AMF),即无梗囊霉属(Acaulos-pora)3种,球囊霉属(Glomus)7种,巨孢囊霉属(Gigaspora)1种;其中,球囊霉属是样地的优势属。在AMF中,孔窝无梗囊霉(A.foveata)分离频率最高,在14种植物的根际土中都有发现;此外,大果球囊霉(G.macrocarpum)的相对多度最大,为59%,具有最强的产孢能力。同时,在11种植物的根中发现了深色有隔内生真菌(DSE),占调查植物的69%;其中,11种植物同时被DSE和AMF感染。     

Community composition and diversity of Neotropical root‐associated fungi in common and rare trees

Interactions between plants and root‐associated fungi can affect the assembly, diversity, and relative abundances of tropical plant species. Host–symbiont compatibility and some degree of host specificity are prerequisites for these processes to occur, and these prerequisites may vary with host abundance. However, direct assessments of whether specificity of root‐associated fungi varies with host abundance are lacking. Here, in a diverse tropical forest in Los Tuxtlas, Mexico, we couple DNA metabarcoding with a sampling design that controls for host phylogeny, host age, and habitat variation, to characterize fungal communities associated with the roots of three confamilial pairs of host species that exhibit contrasting (high and low) relative abundances. We uncovered a functionally and phylogenetically diverse fungal community composed of 1,038 OTUs (operational taxonomic units with 97% genetic similarity), only 14 of which exhibited host specificity. Host species was a significant predictor of fungal community composition only for the subset of OTUs composed of putatively pathogenic fungi. We found no significant difference in the number of specialists associating with common versus rare trees, but we found that host abundance was negatively correlated with the diversity of root fungal communities. This latter result was significant for symbiotrophs (mostly arbuscular mycorrhizal fungi) and, to a lesser extent, for pathotrophs (mostly plant pathogens). Thus, root fungal communities differ between common and rare trees, which may impact the strength of conspecific negative density dependence. Further studies from other tropical sites and host lineages are warranted, given the role of root‐associated fungi in biodiversity maintenance.     

Rapid,economical single‐nucleotide polymorphism and microsatellite discovery based on de novo assembly of a reduced representation genome in a non‐model organism: a case study of Atlantic cod Gadus morhua

By combining next‐generation sequencing technology (454) and reduced representation library (RRL) construction, the rapid and economical isolation of over 25 000 potential single‐nucleotide polymorphisms (SNP) and >6000 putative microsatellite loci from c. 2% of the genome of the non‐model teleost, Atlantic cod Gadus morhua from the Celtic Sea, south of Ireland, was demonstrated. A small‐scale validation of markers indicated that 80% (11 of 14) of SNP loci and 40% (6 of 15) of the microsatellite loci could be amplified and showed variability. The results clearly show that small‐scale next‐generation sequencing of RRL genomes is an economical and rapid approach for simultaneous SNP and microsatellite discovery that is applicable to any species. The low cost and relatively small investment in time allows for positive exploitation of ascertainment bias to design markers applicable to specific populations and study questions.     

Silencing a key gene of the common symbiosis pathway in Nicotiana attenuata specifically impairs arbuscular mycorrhizal infection without influencing the root‐associated microbiome or plant growth

While the biochemical function of calcium and calmodulin‐dependent protein kinase (CCaMK) is well studied, and plants impaired in the expression of CCaMK are known not to be infected by arbuscular mycorrhizal fungi (AMF) in glasshouse studies, the whole‐plant and ecological consequences of CCaMK silencing are not well understood. Here we show that three independently transformed lines of Nicotiana attenuata plants silenced in CCaMK (irCCaMK) are neither infected by Rhizophagus irregularis in the glasshouse nor by native fungal inoculum in the field. The overall fungal community of field‐grown roots did not differ significantly among empty vector (EV) and the transgenic lines, and the bacterial communities only showed minor differences, as revealed by the alpha‐diversity parameters of bacterial OTUs, which were higher in EV plants compared with two of the three transformed lines, while beta‐diversity parameters did not differ. Furthermore, growth and fitness parameters were similar in the glasshouse and field. Herbivory‐inducible and basal levels of salicylic acid, jasmonic acid and abscisic acid did not differ among the genotypes, suggesting that activation of the classical defence pathways are not affected by CCaMK silencing. Based on these results, we conclude that silencing of CCaMK has few, if any, non‐target effects.     

Impacts of long‐term elevated atmospheric CO2 concentrations on communities of arbuscular mycorrhizal fungi

The ecological impacts of long‐term elevated atmospheric CO₂ (eCO₂) levels on soil microbiota remain largely unknown. This is particularly true for the arbuscular mycorrhizal (AM) fungi, which form mutualistic associations with over two‐thirds of terrestrial plant species and are entirely dependent on their plant hosts for carbon. Here, we use high‐resolution amplicon sequencing (Illumina, HiSeq) to quantify the response of AM fungal communities to the longest running (>15 years) free‐air carbon dioxide enrichment (FACE) experiment in the Northern Hemisphere (GiFACE); providing the first evaluation of these responses from old‐growth (>100 years) semi‐natural grasslands subjected to a 20% increase in atmospheric CO₂. eCO₂ significantly increased AM fungal richness but had a less‐pronounced impact on the composition of their communities. However, while broader changes in community composition were not observed, more subtle responses of specific AM fungal taxa were with populations both increasing and decreasing in abundance in response to eCO₂. Most population‐level responses to eCO₂ were not consistent through time, with a significant interaction between sampling time and eCO₂ treatment being observed. This suggests that the temporal dynamics of AM fungal populations may be disturbed by anthropogenic stressors. As AM fungi are functionally differentiated, with different taxa providing different benefits to host plants, changes in population densities in response to eCO₂ may significantly impact terrestrial plant communities and their productivity. Thus, predictions regarding future terrestrial ecosystems must consider changes both aboveground and belowground, but avoid relying on broad‐scale community‐level responses of soil microbes observed on single occasions.     

“Active” and “passive” ecological restoration strategies in meta‐analysis

One of the means of creating a more robust methodology for ecological restoration involves reducing the gap between ecological theory and restoration practices. A common strategy to do so is using meta‐analysis to understand key drivers of restoration outcomes. “Active” and “passive” is a dichotomy often used to separate restoration strategies in such meta‐analyses. We investigate previously raised concerns about selection bias and subjective categorization of strategies. We promote a paired experimental design in future empirical research and propose the use of three categories of restoration strategy in lieu of “passive” and “active” to alleviate inconsistency in definitions and categorization.     

Sex‐specific interaction between arbuscular mycorrhizal and dark septate fungi in the dioecious plant Antennaria dioica (Asteraceae)

Male and female plants of dioecious species often differ in their resource demands and this has been linked to secondary sexual dimorphism, including sex‐specific interactions with other organisms such as herbivores and pollinators. However, little is known about the interaction between dioecious plants and fungal root endophytes. Plants may be simultaneously colonised by arbuscular mycorrhizal (AM) and dark septate (DS) fungi. While it is well established that AM mutualism involves reciprocal transfer of photosynthates and mineral nutrients between roots of host plants and these fungi, the role of DS fungi remains controversial. Here, we report the temporal and spatial variation in AM and DS fungi in female, male and non‐reproductive Antennaria dioica plants in three natural populations in Finland during flowering and after seed production. Females had higher colonisation by AM fungi, but lower colonisation by DS fungi than male and non‐reproductive plants. The higher AM colonisation was observed during flowering, and this difference varied among populations. Our results suggest that females and males of A. dioica interact with AM and DS fungi differently and that this relationship is dependent on soil fertility.     

A critique of the “ultra‐high risk” and “transition” paradigm

The transdiagnostic expression of psychotic experiences in common mental disorder (anxiety/depression/substance use disorder) is associated with a poorer prognosis, and a small minority of people may indeed develop a clinical picture that meets criteria for schizophrenia. However, it appears neither useful nor valid to observe early states of multidimensional psychopathology in young people through the “schizo”‐prism, and apply misleadingly simple, unnecessary and inefficient binary concepts of “risk” and “transition”. A review of the “ultra‐high risk” (UHR) or “clinical high risk” (CHR) literature indicates that UHR/CHR samples are highly heterogeneous and represent individuals diagnosed with common mental disorder (anxiety/depression/substance use disorder) and a degree of psychotic experiences. Epidemiological research has shown that psychotic experiences are a (possibly non‐causal) marker of the severity of multidimensional psychopathology, driving poor outcome, yet notions of “risk” and “transition” in UHR/CHR research are restrictively defined on the basis of positive psychotic phenomena alone, ignoring how baseline differences in multidimensional psychopathology may differentially impact course and outcome. The concepts of “risk” and “transition” in UHR/CHR research are measured on the same dimensional scale, yet are used to produce artificial diagnostic shifts. In fact, “transition” in UHR/CHR research occurs mainly as a function of variable sample enrichment strategies rather than the UHR/CHR “criteria” themselves. Furthermore, transition rates in UHR/CHR research are inflated as they do not exclude false positives associated with the natural fluctuation of dimensional expression of psychosis. Biological associations with “transition” thus likely represent false positive findings, as was the initial claim of strong effects of omega‐3 polyunsatured fatty acids in UHR samples. A large body of UHR/CHR intervention research has focused on the questionable outcome of “transition”, which shows lack of correlation with functional outcome. It may be more productive to consider the full range of person‐specific psychopathology in all young individuals who seek help for mental health problems, instead of “policing” youngsters for the transdiagnostic dimension of psychosis. Instead of the relatively inefficient medical high‐risk approach, a public health perspective, focusing on improved access to a low‐stigma, high‐hope, small scale and youth‐specific environment with acceptable language and interventions may represent a more useful and efficient strategy.     

Presence and distribution of insect-associated and entomopathogenic fungi in a temperate pine forest soil: An integrated approach

For decades entomopathogenic fungi have garnered interest as possible alternatives to chemical pesticides. However, their ecology outside of agroecosystems demands further study. We assessed the diversity and abundance of entomopathogenic and insect-associated fungi at a loblolly pine forest in North Carolina, USA using culture-dependent and next-generation sequencing libraries. Fungi were isolated using Galleria mellonella larvae, as well as from soil dilutions plated on a selective medium. Isolates were identified using Sanger sequencing of the ITS and LSU rRNA gene regions, and represented 36 OTUs including Metarhizium, Lecanicillium, and Paecilomyces. Additionally, we assessed the chitinolytic potential of isolates and found widespread, variable ability to degrade chitin within and between genera. Phylogenetic analyses resolved several isolates to genus, with some forming clades with other insect-associated taxa, as well as with fungi associated with plant tissues. Saprophytes were widely distributed in soil, while entomopathogens were less abundant and present primarily in the top two cm of the soil. The similarity between culture-dependent and next-generation sequencing results demonstrates that both methods can be used concurrently in this system to study the ecology of entomopathogenic fungi.     

Assessing the “Short Mental Distance” in Eco‐Industrial Networks

Like many economic exchanges, industrial symbiosis (IS) is thought to be influenced by social relationships and shared norms among actors in a network. While many implicit references to social characteristics exist throughout the literature, there have been few explicit attempts to operationalize and measure the concepts. The “short mental distance,”“trust,”“openness,” and “communication” recorded among managers in Kalundborg, Denmark, set a precedent for examining and encouraging social interactions among key personnel in the dozens of eco‐industrial networks around the world. In this article we explore the relationships among various aspects of social embeddedness, social capital, and IS. We develop a conceptual framework and an approach using quantitative and qualitative methods to identify and measure these social characteristics, including social network structure, communication, and similarities in norms and conceptions of waste, and apply them in an industrial network in Nanjangud, South India. The findings suggest that there is a fairly high level of shared norms about dealing with waste—the “short mental distance”—in this network, but by‐product transactions are only weakly correlated with the structure and content of communication among managers. Replication of this approach can increase the understanding and comparability of the role of social characteristics in eco‐industrial activities around the world.     

Testing genotyping strategies for ultra‐deep sequencing of a co‐amplifying gene family: MHC class I in a passerine bird

Characterization of highly duplicated genes, such as genes of the major histocompatibility complex (MHC), where multiple loci often co‐amplify, has until recently been hindered by insufficient read depths per amplicon. Here, we used ultra‐deep Illumina sequencing to resolve genotypes at exon 3 of MHC class I genes in the sedge warbler (Acrocephalus schoenobaenus). We sequenced 24 individuals in two replicates and used this data, as well as a simulated data set, to test the effect of amplicon coverage (range: 500–20 000 reads per amplicon) on the repeatability of genotyping using four different genotyping approaches. A third replicate employed unique barcoding to assess the extent of tag jumping, that is swapping of individual tag identifiers, which may confound genotyping. The reliability of MHC genotyping increased with coverage and approached or exceeded 90% within‐method repeatability of allele calling at coverages of >5000 reads per amplicon. We found generally high agreement between genotyping methods, especially at high coverages. High reliability of the tested genotyping approaches was further supported by our analysis of the simulated data set, although the genotyping approach relying primarily on replication of variants in independent amplicons proved sensitive to repeatable errors. According to the most repeatable genotyping method, the number of co‐amplifying variants per individual ranged from 19 to 42. Tag jumping was detectable, but at such low frequencies that it did not affect the reliability of genotyping. We thus demonstrate that gene families with many co‐amplifying genes can be reliably genotyped using HTS, provided that there is sufficient per amplicon coverage.     

Decrease in diversity and changes in community composition of arbuscular mycorrhizal fungi in roots of apple trees with increasing orchard management intensity across a regional scale

Understanding which factors drive the diversity and community composition of arbuscular mycorrhizal fungi (AMF) is important due to the role of these soil micro‐organisms in ecosystem functioning and current environmental threats to AMF biodiversity. Additionally, in agro‐ecosystems, this knowledge may help to evaluate their use in making agriculture more sustainable. Here, we used 454‐pyrosequencing of small subunit rRNA gene amplicons to quantify AMF diversity and community composition in the roots of cultivated apple trees across 24 orchards in central Belgium. We aimed at identifying the factors (soil chemical variables, organic vs. conventional farming, and geographical location) that affect AMF diversity and community composition. In total, 110 AMF OTUs were detected, of which the majority belonged to the Glomeraceae (73%) and the Claroideoglomeraceae (19%). We show that soil characteristics and farming system, rather than the geographical location of the orchards, shape AMF communities on apple trees. Particularly, plant‐available P content of the soil was associated with lower AMF diversity. In orchards with a lower plant‐available P content of the soil (P < 100 mg/kg soil), we also found a significantly higher AMF diversity in organically managed orchards as compared to conventionally managed orchards. Finally, the degree of nestedness of the AMF communities was related to plant‐available P and N content of the soil, pointing at a progressive loss of AMF taxa with increasing fertilization. Overall, we conclude that a combination of organic orchard management and moderate fertilization may preserve diverse AMF communities on apple trees and that AMF in the roots of apple trees appear not to be dispersal limited at the scale of central Belgium.     

Neighboring trees regulate the root‐associated pathogenic fungi on the host plant in a subtropical forest

Root‐associated fungi and host‐specific pathogens are major determinants of species coexistence in forests. Phylogenetically related neighboring trees can strongly affect the fungal community structure of the host plant, which, in turn, will affect the ecological processes. Unfortunately, our understanding of the factors influencing fungal community composition in forests is still limited. In particular, investigation of the relationship between the phytopathogenic fungal community and neighboring trees is incomplete. In the current study, we tested the host specificity of members of the root‐associated fungal community collected from seven tree species and determined the influence of neighboring trees and habitat variation on the composition of the phytopathogenic fungal community of the focal plant in a subtropical evergreen forest. Using high‐throughput sequencing data with respect to the internal transcribed spacer (ITS) region, we characterized the community composition of the root‐associated fungi and found significant differences with respect to fungal groups among the seven tree species. The density of conspecific neighboring trees had a significantly positive influence on the relative abundance of phytopathogens, especially host‐specific pathogens, while the heterospecific neighbor density had a significant negative impact on the species richness of host‐specific pathogens, as well as phytopathogens. Our work provides evidence that the root‐associated phytopathogenic fungi of a host plant depend greatly on the tree neighbors of the host plant.     

Methyl CpG Binding Domain Ultra‐Sequencing: a novel method for identifying inter‐individual and cell‐type‐specific variation in DNA methylation

Experience‐dependent changes in DNA methylation can exert profound effects on neuronal function and behaviour. A single learning event can induce a variety of DNA modifications within the neuronal genome, some of which may be common to all individuals experiencing the event, whereas others may occur in a subset of individuals. Variations in experience‐induced DNA methylation may subsequently confer increased vulnerability or resilience to the development of neuropsychiatric disorders. However, the detection of experience‐dependent changes in DNA methylation in the brain has been hindered by the interrogation of heterogeneous cell populations, regional differences in epigenetic states and the use of pooled tissue obtained from multiple individuals. Methyl CpG Binding Domain Ultra‐Sequencing (MBD Ultra‐Seq) overcomes current limitations on genome‐wide epigenetic profiling by incorporating fluorescence‐activated cell sorting and sample‐specific barcoding to examine cell‐type‐specific CpG methylation in discrete brain regions of individuals. We demonstrate the value of this method by characterizing differences in 5‐methylcytosine (5mC) in neurons and non‐neurons of the ventromedial prefrontal cortex of individual adult C57BL/6 mice, using as little as 50 ng of genomic DNA per sample. We find that the neuronal methylome is characterized by greater CpG methylation as well as the enrichment of 5mC within intergenic loci. In conclusion, MBD Ultra‐Seq is a robust method for detecting DNA methylation in neurons derived from discrete brain regions of individual animals. This protocol will facilitate the detection of experience‐dependent changes in DNA methylation in a variety of behavioural paradigms and help identify aberrant experience‐induced DNA methylation that may underlie risk and resiliency to neuropsychiatric disease.     

Recent progress and challenges in population genetics of polyploid organisms: an overview of current state‐of‐the‐art molecular and statistical tools

Despite the importance of polyploidy and the increasing availability of new genomic data, there remain important gaps in our knowledge of polyploid population genetics. These gaps arise from the complex nature of polyploid data (e.g. multiple alleles and loci, mixed inheritance patterns, association between ploidy and mating system variation). Furthermore, many of the standard tools for population genetics that have been developed for diploids are often not feasible for polyploids. This review aims to provide an overview of the state‐of‐the‐art in polyploid population genetics and to identify the main areas where further development of molecular techniques and statistical theory is required. We review commonly used molecular tools (amplified fragment length polymorphism, microsatellites, Sanger sequencing, next‐generation sequencing and derived technologies) and their challenges associated with their use in polyploid populations: that is, allele dosage determination, null alleles, difficulty of distinguishing orthologues from paralogues and copy number variation. In addition, we review the approaches that have been used for population genetic analysis in polyploids and their specific problems. These problems are in most cases directly associated with dosage uncertainty and the problem of inferring allele frequencies and assumptions regarding inheritance. This leads us to conclude that for advancing the field of polyploid population genetics, most priority should be given to development of new molecular approaches that allow efficient dosage determination, and to further development of analytical approaches to circumvent dosage uncertainty and to accommodate ‘flexible’ modes of inheritance. In addition, there is a need for more simulation‐based studies that test what kinds of biases could result from both existing and novel approaches.     

Visualizing the “sandwich” structure of osteocytes in their native environment deep in bone in vivo

Osteocytes are the most abundant cells in bone and always the focus of bone research. They are embedded in the highly scattering mineralized bone matrix. Consequently, visualizing osteocytes deep in bone with subcellular resolution poses a major challenge for in vivo bone research. Here we overcome this challenge by demonstrating 3‐photon imaging of osteocytes through the intact mouse skull in vivo. Through broadband transmittance characterization, we establish that the excitation at the 1700‐nm window enables the highest optical transmittance through the skull. Using label‐free third‐harmonic generation (THG) imaging excited at this window, we visualize osteocytes through the whole 140‐μm mouse skull and 155 μm into the brain in vivo. By developing selective labeling technique for the interstitial space, we visualize the “sandwich” structure of osteocytes in their native environment. Our work provides novel imaging methodology for bone research in vivo.