Profiling functions of ectomycorrhizal diversity and root structuring in seedlings of Norway spruce (Picea abies) with fast- and slow-growing phenotypes.

We studied the role of taxonomical and functional ectomycorrhizal (ECM) fungal diversity in root formation and nutrient uptake by Norway spruce (Picea abies) seedlings with fast- and slow-growing phenotypes. Seedlings were grown with an increasing ECM fungal diversity gradient from one to four species and sampled before aboveground growth differences between the two phenotypes were apparent. ECM fungal colonization patterns were determined and functional diversity was assayed via measurements of potential enzyme activities of eight exoenzymes probably involved in nutrient mobilization. Phenotypes did not vary in their receptiveness to different ECM fungal species. However, seedlings of slow-growing phenotypes had higher fine-root density and thus more condensed root systems than fast-growing seedlings, but the potential enzyme activities of ectomycorrhizas did not differ qualitatively or quantitatively. ECM species richness increased host nutrient acquisition potential by diversifying the exoenzyme palette. Needle nitrogen content correlated positively with high chitinase activity of ectomycorrhizas. Rather than fast- and slow-growing phenotypes exhibiting differing receptiveness to ECM fungi, our results suggest that distinctions in fine-root structuring and in the belowground growth strategy already apparent at early stages of seedling development may explain later growth differences between fast- and slow-growing families.

Effects of transcriptional pausing on gene expression dynamics.

Stochasticity in gene expression affects many cellular processes and is a source of phenotypic diversity between genetically identical individuals. Events in elongation, particularly RNA polymerase pausing, are a source of this noise. Since the rate and duration of pausing are sequence-dependent, this regulatory mechanism of transcriptional dynamics is evolvable. The dependency of pause propensity on regulatory molecules makes pausing a response mechanism to external stress. Using a delayed stochastic model of bacterial transcription at the single nucleotide level that includes the promoter open complex formation, pausing, arrest, misincorporation and editing, pyrophosphorolysis, and premature termination, we investigate how RNA polymerase pausing affects a gene's transcriptional dynamics and gene networks. We show that pauses' duration and rate of occurrence affect the bursting in RNA production, transcriptional and translational noise, and the transient to reach mean RNA and protein levels. In a genetic repressilator, increasing the pausing rate and the duration of pausing events increases the period length but does not affect the robustness of the periodicity. We conclude that RNA polymerase pausing might be an important evolvable feature of genetic networks.

Development of a research dedicated archival system (TARAS) in a university hospital.

Recent healthcare policies have influenced the manner in which patient data is handled in research projects, and the regulations concerning protected health information have become significantly tighter. Thus, new procedures are needed to facilitate research while protecting the confidentiality of patient data and ensuring the integrity of clinical work in the expanding environment of electronic files and databases. We have addressed this problem in a university hospital setting by developing the Tampere Research Archival System (TARAS), an extensive data warehouse for research purposes. This dynamic system includes numerous integrated and pseudonymized imaging studies and clinical data. In a pilot study on asthma patients, we tested and improved the functionality of the data archival system. TARAS is feasible to use in retrieving, analyzing, and processing both image and non-image data. In this paper, we present a detailed workflow of the implementation process of the data warehouse, paying special attention to administrative, ethical, practical, and data security concerns. The establishment of TARAS will enhance and accelerate research practice at Tampere University Hospital, while also improving the safety of patient information as well as the prospects for national and international research collaboration. We hope that much can be learned from our experience of planning, designing, and implementing a research data warehouse combining imaging studies and medical records in a university hospital.

Occurrence of similar mycoviruses in pathogenic, saprotrophic and mycorrhizal fungi inhabiting the same forest stand.

Fungal viruses (mycoviruses) are considered to be highly host specific, but phylogenetic analysis supports the occasional occurrence of horizontal transmission between species. We used an extensive sampling strategy to investigate whether similar viruses occur in more than one fungal species of the same forest habitat. Mycelial samples were collected from in-growth mesh bags (N = 259), fruiting bodies (N = 173) and cultured isolates (N = 68) at a forest site where the spatial distribution of viral infections in clonal individuals of the wood decay fungus Heterobasidion parviporum was mapped in detail earlier. The investigation revealed previously known Heterobasidion viruses in ∼2% of the single or pooled mycorrhizal samples from mesh bags, ∼3% of the fruiting body samples and none of the fungal cultures analyzed. Novel virus strains distinct from known Heterobasidion viruses were detected in cultures of ectomycorrhizal fungi (Lactarius tabidus, L. rufus) and saprotrophic fungi (Megacollybia platyphylla, Mucoraceae spp.). Overall, our results support the view that mycoviruses do not readily cross species borders. Regarding potential virocontrol applications, the introduction of Heterobasidion viruses into natural habitats is not expected to cause a major infection pressure towards the indigenous fungal community. However, the ecological consequences of the putative interspecies virus transmission events detected require further investigation.

Interactions between soil- and dead wood-inhabiting fungal communities during the decay of Norway spruce logs.

We investigated the interaction between fungal communities of soil and dead wood substrates. For this, we applied molecular species identification and stable isotope tracking to both soil and decaying wood in an unmanaged boreal Norway spruce-dominated stand. Altogether, we recorded 1990 operational taxonomic units, out of which more than 600 were shared by both substrates and 589 were found to exclusively inhabit wood. On average the soil was more species-rich than the decaying wood, but the species richness in dead wood increased monotonically along the decay gradient, reaching the same species richness and community composition as soil in the late stages. Decaying logs at all decay stages locally influenced the fungal communities from soil, some fungal species occurring in soil only under decaying wood. Stable isotope analyses suggest that mycorrhizal species colonising dead wood in the late decay stages actively transfer nitrogen and carbon between soil and host plants. Most importantly, Piloderma sphaerosporum and Tylospora sp. mycorrhizal species were highly abundant in decayed wood. Soil- and wood-inhabiting fungal communities interact at all decay phases of wood that has important implications in fungal community dynamics and thus nutrient transportation.

The community of needle endophytes reflects the current physiological state of Norway spruce.

This study investigated fungal endophytes in the needles of Norway spruce (Picea abies) cuttings in relation to host tree growth. We also determined the prevalence of endophytes in needles incubated for six months. The cuttings originated from clonal origins showing slow- and fast-growth in long-term field trials but the heritable differences in growth rate were not yet detected among the studied cutting. Endophytes were isolated from surface-sterilized needles with culture-free DNA techniques. No significant differences were observed between endophyte communities of slow- and fast-growing clonal origins. However, the endophyte community correlated with the current growth rate of cuttings suggesting that endophytes reflect short- rather than long-term performance of a host. The concentration of condensed tannins was similar in slow- and fast-growing clonal origins but it showed a negative relationship with endophyte species richness, implying that these secondary compounds may play an important role in spruce tolerance against fungal infections. More than a third of endophyte species were detected in both fresh and decomposing needles, indicating that many needle endophytes are facultative saprotrophs. Several potentially pathogenic fungal species were also found within the community of saprotrophic endophytes.

Loss of diversity in wood-inhabiting fungal communities affects decomposition activity in Norway spruce wood.

Hundreds of wood-inhabiting fungal species are now threatened, principally due to a lack of dead wood in intensively managed forests, but the consequences of reduced fungal diversity on ecosystem functioning are not known. Several experiments have shown that primary productivity is negatively affected by a loss of species, but the effects of microbial diversity on decomposition are less studied. We studied the relationship between fungal diversity and the in vitro decomposition rate of slightly, moderately and heavily decayed Picea abies wood with indigenous fungal communities that were diluted to examine the influence of diversity. Respiration rate, wood-degrading hydrolytic enzymes and fungal community structure were assessed during a 16-week incubation. The number of observed OTUs in DGGE was used as a measure of fungal diversity. Respiration rate increased between early- and late-decay stages. Reduced fungal diversity was associated with lower respiration rates during intermediate stages of decay, but no effects were detected at later stages. The activity of hydrolytic enzymes varied among decay stages and fungal dilutions. Our results suggest that functioning of highly diverse communities of the late-decay stage were more resistant to the loss of diversity than less diverse communities of early decomposers. This indicates the accumulation of functional redundancy during the succession of the fungal community in decomposing substrates.

Endophyte communities vary in the needles of Norway spruce clones.

Endophytic fungi show no symptoms of their presence but can influence the performance and vitality of host trees. The potential use of endophytes to indicate vitality has been previously realized, but a standard protocol has yet to be developed due to an incomplete understanding of the factors that regulate endophyte communities. Using a culture-free molecular approach, we examined the extent to which host genotype influences the abundance, species richness, and community composition of endophytic fungi in Norway spruce needles. Briefly, total DNA was extracted from the surface-sterilized needles of 30 clones grown in a nursery field and the copy number of the fungal internal transcribed spacer (ITS) region of ribosomal DNA was estimated by quantitative PCR. Fungal species richness and community composition were determined by denaturing gradient gel electrophoresis and DNA sequencing. We found that community structure and ITS copy number varied among spruce clones, whereas species richness did not. Host traits interacting with endophyte communities included needle surface area and the location of cuttings in the experimental area. Although Lophodermium piceae is considered the dominant needle endophyte of Norway spruce, we detected this species in only 33% of samples. The most frequently observed fungus (66%) was the potentially pathogenic Phoma herbarum. Interestingly, ITS copy number of endophytic fungi correlated negatively with the richness of ectomycorrhizal fungi and thus potential interactions between fungal communities and their influence on the host tree are discussed. Our results suggest that in addition to environmental factors, endophyte communities of spruce needles are determined by host tree identity and needle surface area.

Fungal community dynamics in relation to substrate quality of decaying Norway spruce ( Picea abies [L.] Karst.) logs in boreal forests.

Decaying wood plays an important role in forest biodiversity, nutrient cycling and carbon balance. Community structure of wood-inhabiting fungi changes with mass loss of wood, but the relationship between substrate quality and decomposers is poorly understood. This limits the extent to which these ecosystem services can be effectively managed. We studied the fungal community and physico-chemical quality (stage of decay, dimensions, density, moisture, C : N ratio, lignin and water or ethanol extractives) of 543 Norway spruce logs in five unmanaged boreal forest sites of southern Finland. Fungi were identified using denaturing gradient gel electrophoresis and sequencing of DNA extracted directly from wood samples. Macroscopic fruiting bodies were also recorded. Results showed a fungal community succession with decreasing wood density and C : N ratio, and increasing moisture and lignin content. Fungal diversity peaked in the most decayed substrates. Ascomycetes typically colonized recently fallen wood. Brown-rot fungi preferred the intermediate decay stages. White-rot fungi represented approximately one-fifth of sequenced species in all decay phases excluding the final phase, where ectomycorrhizal (ECM) fungi became dominant. Lignin content of logs with white-rot fungi was low, and ECM fungi were associated with substrates containing abundant nitrogen. Macroscopic fruiting bodies were observed for only a small number of species detected with molecular techniques.

Delayed stochastic model of transcription at the single nucleotide level.

We present a delayed stochastic model of transcription at the single nucleotide level. The model accounts for the promoter open complex formation and includes alternative pathways to elongation, namely pausing, arrest, misincorporation and editing, pyrophosphorolysis, and premature termination. We confront the dynamics of this detailed model with a single-step multi-delayed stochastic model and with measurements of expression of a repressed gene at the single molecule level. At low expression rates both models match the experiments but, at higher rates the two models differ significantly, with consequences to cell-to-cell phenotypic variability. The alternative pathway reactions, due to, for example, causing polymerases to collide more often on the template, are the cause for the difference in dynamical behaviors. Next, we confront the model with measurements of the transcriptional dynamics at the single RNA level of an induced gene and show that RNA production, besides its bursting dynamics, also exhibits pulses (2 or more RNAs produced in intervals smaller than the smallest interval between initiations). The distribution of occurrences and amplitudes of pulses match the experimental measurements. This pulsing and the noise at the elongation stage are shown to play a role in the dynamics of a genetic switch.

Decomposition and fungi of needle litter from slow- and fast-growing Norway spruce (Picea abies) clones.

The fungal species involved in the decomposition of needle litter and their response to intraspecific genetic variation of trees are poorly known. First, we compared the needle decomposition and fungal decomposers underneath eight different Norway spruce clones in situ. This experiment revealed 60-70% loss of needle mass in two years. Although spruce clones differed considerably in growth (twofold height difference) and their needles differed in chemical composition, no significant difference was found for loss of needle mass under the spruce clones. Furthermore, the spruce clones did not affect the community structure of the fungal decomposers. Fungi inhabiting needle litter were identified by extracting ribosomal RNA (rRNA) and sequencing complementary DNA (cDNA) of internal trascribed spacer 1 (ITS1) region. The most frequent identifications were Lophodermium, Pezizales, Mycena, and Marasmius, suggesting that endophytic fungi were involved in the decomposition process. Second, we evaluated the potential of endophytes to decompose needle material in a microcosm experiment in which all other fungi than endophytes were excluded. Within 2 years, the endophytes had decomposed 35-45% of the needle mass. Sequences of Mollisia, Lophodermium, Lachnum, and Phialocephala were most frequently found in rRNA and rDNA extracted from the needles at the end of the microcosm experiment. The dominant needle endophyte in fresh, green needles was Lophodermium piceae, and this species was also found frequently in the needle material after 2 years of decay both in the field and laboratory experiments. Moreover, the relative abundance of Lophodermium-derived denaturing gradient gel electrophoresis (DGGE) bands correlated positively with the decomposition in the microcosm experiment. Hence, our results suggest a significant role of endophytic fungi, and particularly L. piceae, in the process of needle decomposition in boreal forests.