Initial fungal colonizer affects mass loss and fungal community development in Picea abies logs 6 yr after inoculation

Picea abies logs were inoculated with Resinicium bicolor, Fomitopsis pinicola or left un-inoculated and placed in an old-growth boreal forest. Mass loss and fungal community data were collected after 6 yr to test whether simplification of the fungal community via inoculation affects mass loss and fungal community development. Three techniques were used to survey communities: (1) observation of fruiting structures; (2) culturing on media; and (3) cloning and sequencing of ITS rDNA. Fruit body surveys detected the smallest number of species (18, 3.8 per log), DNA-based methods detected the most species (72, 31.7 per log), and culturing detected an intermediate number (23, 7.2 per log). Initial colonizer affected community development and inoculation with F. pinicola led to significantly greater mass loss. Relationships among fungal community composition, community richness and mass loss are complex and further work is needed to determine whether simplification of fungal communities affects carbon sequestration in forests.     

Identification of rheological properties of human body surface tissue

According to World Health Organization obesity is one of the greatest public health challenges of the 21st century. It has tripled since the 1980s and the numbers of those affected continue to rise at an alarming rate, especially among children. There are number of devices that act as a prevention measure to boost person?s motivation for physical activity and its levels. The placement of these devices is not restricted thus the measurement errors that appear because of the body rheology, clothes, etc. cannot be eliminated. The main objective of this work is to introduce a tool that can be applied directly to process measured accelerations so human body surface tissue induced errors can be reduced. Both the modeling and experimental techniques are proposed to identify body tissue rheological properties and prelate them to body mass index. Multi-level computational model composed from measurement device model and human body surface tissue rheological model is developed. Human body surface tissue induced inaccuracies can increase the magnitude of measured accelerations up to 34% when accelerations of the magnitude of up to 27 m/s² are measured. Although the timeframe of those disruptions are short – up to 0.2 s – they still result in increased overall measurement error.     

Body size and craniometry of the herb field mouse from Lithuania in the context of species range

We present morphometric and craniometric measurements of the herb field mouse (Apodemus uralensis) from Lithuania and analyze variation of body and skull size across species range. We suppose species is characterized by high size variability, not following Bergmann’s or Murphy’s rules. Preliminary, distinct size differences have been registered in the eastern and southern edges of the distribution range, with these populations having largest individuals according to average body and skull size. In terms of tail length and condylobasal length of the skull, Lithuanian mice on the north-western edge of the species range are among the largest, but in terms of body weight, body length, zygomatic skull width and the length of maxillary toothrow, adult A. uralensis from Lithuania are small and correspond to those from populations on the western edge of the range. The relative skull width (ratio of zygomatic skull width to condylobasal length) of Lithuanian A. uralensis is the smallest across the entire range. In A. uralensis from Lithuania, sex dimorphism is weakly expressed, with hind foot length and postorbital constriction larger in adult males, while the height of the mandibula and length of the mandibular diastema is larger in adult females. Juvenile and subadult A. uralensis from Lithuania differ in body weight, but not in size.     

Impact of <Emphasis Type="Italic">Heterobasidion</Emphasis> root-rot on fine root morphology and associated fungi in <Emphasis Type="Italic">Picea abies</Emphasis> stands on peat soils

We examined differences in fine root morphology, mycorrhizal colonisation and root-inhabiting fungal communities between Picea abies individuals infected by Heterobasidion root-rot compared with healthy individuals in four stands on peat soils in Latvia. We hypothesised that decreased tree vitality and alteration in supply of photosynthates belowground due to root-rot infection might lead to changes in fungal communities of tree roots. Plots were established in places where trees were infected and in places where they were healthy. Within each stand, five replicate soil cores with roots were taken to 20 cm depth in each root-rot infected and uninfected plot. Root morphological parameters, mycorrhizal colonisation and associated fungal communities, and soil chemical properties were analysed. In three stands root morphological parameters and in all stands root mycorrhizal colonisation were similar between root-rot infected and uninfected plots. In one stand, there were significant differences in root morphological parameters between root-rot infected versus uninfected plots, but these were likely due to significant differences in soil chemical properties between the plots. Sequencing of the internal transcribed spacer of fungal nuclear rDNA from ectomycorrhizal (ECM) root morphotypes of P. abies revealed the presence of 42 fungal species, among which ECM basidiomycetes Tylospora asterophora (24.6 % of fine roots examined), Amphinema byssoides (14.5 %) and Russula sapinea (9.7 %) were most common. Within each stand, the richness of fungal species and the composition of fungal communities in root-rot infected versus uninfected plots were similar. In conclusion, Heterobasidion root-rot had little or no effect on fine root morphology, mycorrhizal colonisation and composition of fungal communities in fine roots of P. abies growing on peat soils.     

Population genetics of the wood-decay fungus Phlebia centrifuga P. Karst. in fragmented and continuous habitats

The basidiomycete Phlebia centrifuga is a wood-decay fungus characteristic for unmanaged old-growth forests of spruce, a habitat that has become increasingly fragmented due to forest management. The aim of this study was to investigate the genetic population structures of P. centrifuga in both continuous and fragmented habitats, and estimate the potential impact of fragmentation on the genetic diversity of the fungus. Three hundred fifteen single spore isolates (representing 47 spore families and 33 single isolates) from eight populations across northern Europe (Russia, Finland, and Sweden) were screened with seven microsatellite markers and arbitrary primed polymerase chain reaction with the M13 minisatellite. The two molecular methods generally gave the same pattern for the genetic population structure. There were no significant differences between the observed and the expected heterozygosities, and the inbreeding coefficient (FIS) did not indicate any inbreeding. The fixation index (FST) revealed a general pattern with little to moderate genetic differentiation for the majority of populations, while the southernmost Swedish population Norra Kvill was the only one showing high differentiation from about half of the other populations. Swedish population Fiby with the shortest distance to the continuous habitat was moderately differentiated from most of the others and to the largest extent differed from geographically closest population of Norra Kvill. The results indicate that the fragmentation of old-growth forest in Russia and Finland is more recent than the fragmentation in Sweden, and the genetic population structures of P. centrifuga in northern Europe might be related to differences in forest landscape dynamics between the two areas.     

Selective expression of the soluble product fraction in Escherichia coli cultures employed in recombinant protein production processes

Recombinant proteins produced in Escherichia coli hosts may appear within the cells’ cytoplasm in form of insoluble inclusion bodies (IB’s) and/or as dissolved functional protein molecules. If no efficient refolding procedure is available, one is interested in obtaining as much product as possible in its soluble form. Here, we present a process engineering approach to maximizing the soluble target protein fraction. For that purpose, a dynamic process model was developed. Its essential kinetic component, the specific soluble product formation rate, if represented as a function of the specific growth rate and the culture temperature, depicts a clear maximum. Based on the dynamic model, optimal specific growth rate and temperature profiles for the fed-batch fermentation were determined. In the course of the study reported, the mass of desired soluble protein was increased by about 25%. At the same time, the formation of inclusion bodies was essentially avoided. As the optimal cultivation procedure is rather susceptible to distortions, control measures are necessary to guarantee that the real process can be kept on its desired path. This was possible with robust closed loop control. Experimental process validation revealed that, in this way, high dissolved product fractions could be obtained at an excellent batch-to-batch reproducibility.     

Data-based optimization of protein production processes

While data-based modeling is possible in various ways, data-based optimization has not been previously described. Here we present such an optimization technique. It is based on dynamic programming principles and uses data directly from exploratory experiments where the influence of the adjustable variables u were tested at various values. Instead of formulating the performance index J as a function of time t within a cultivation process it is formulated as a function of the biomass x. The advantage of this representation is that in most biochemical production processes J(x) only depends of the vector u of the adjustable variables. This given, mathematical programming techniques allow determining the desired optimal paths u _opt (x) from the x-derivatives of J(x). The resulting u _opt (x) can easily be transformed back to the u(t) profiles that can then be used in an improved fermentation run. The optimization technique can easily be explained graphically. With numerical experiments the feasibility of the method is demonstrated. Then, two optimization runs for recombinant protein formations in E. coli are discussed and experimental validation results are presented.     

The influence of high nestbox density on the common dormouseMuscardinus avellanarius population

The response of common dormouseMuscardinus avellanarius Linnaeus, 1758 population to availability of nest sites was studied by manipulating the nestbox grid and ring-marking dormice. Abundance of adult dormice more than doubled in the 25 × 25 m nestbox grid in comparison to the 50 × 50 m grid, as a result of increased nestbox density from four to 16 boxes/ha. This effect already became apparent in the first year after additional nestboxes were made available and resulted from dormouse immigration, mostly from adjacent areas without nestboxes. In the second and third years, the number of two-year-old and older resident dormice, which had their home ranges in this plot, increased considerably. The average size of dormouse home range decreased by approximately half both in males and females in the 25 × 25 m grid compared to the 50 × 50 m grid. The proportion of breeding adult females did not differ between the two grids in spite of different adult dormouse density. Shortage of secure nest sites was a limiting factor for the common dormouse population abundance in the forest where natural tree hollows were absent, and high nestbox density increased environmental carrying capacity.     

Increasing batch-to-batch reproducibility of CHO-cell cultures using a model predictive control approach

By means of a model predictive control strategy it was possible to ensure a high batch-to-batch reproducibility in animal cell (CHO-cell) suspensions cultured for a recombinant therapeutic protein (EPO) production. The general control objective was derived by identifying an optimal specific growth rate taking productivity, protein quality and process controllability into account. This goal was approached indirectly by controlling the oxygen mass consumed by the cells which is related to specific biomass growth rate and cell concentration profile by manipulating the glutamine feed rate. Process knowledge represented by a classical model was incorporated into the model predictive control algorithm. The controller was employed in several cultivation experiments. During these cultivations, the model parameters were adapted after each sampling event to cope with changes in the process’ dynamics. The ability to predict the state variables, particularly for the oxygen consumption, led to only moderate changes in the desired optimal operational trajectories. Hence, nearly identical oxygen consumption profiles, cell and protein titers as well as sialylation patterns were obtained for all cultivation runs.     

Improving the batch-to-batch reproducibility of microbial cultures during recombinant protein production by regulation of the total carbon dioxide production

Batch-to-batch reproducibility of fermentation processes performed during the manufacturing processes of biologics can be increased by operating the cultures at feed rate profiles that are robust against typically arising disturbances. Remaining randomly appearing deviations from the desired path should be suppressed automatically by manipulating the feed rate. With respect to the cells' physiology it is best guiding the cultivations along an optimal profile of the specific biomass growth rate mu(t). However, there are two problems that speak for further investigations: Upon severe disturbances that may happen during the fermentation, the biomass concentration X may significantly deviate from its desired value, then a fixed mu-profile leads to a diminished batch-to-batch reproducibility. Second, the specific growth rate cannot easily be estimated online to a favourably high accuracy, hence it is difficult to determine the deviations in mu from the desired profile. The alternative discussed here solves both problems by keeping the process at the corresponding total cumulative carbon dioxide production-profile: it is robust against distortions in X and the controlled variable can accurately be measured online during cultivations of all relevant sizes. As compared to the fermentation practice currently used in industry, the experimental results, presented at the example of a recombinant protein production with Escherichia coli cells, show that CPR-based corrections lead to a considerably improved batch-to-batch reproducibility.