Production of a chimeric enzyme tool associating the Trichoderma reesei swollenin with the Aspergillus niger feruloyl esterase A for release of ferulic acid

The main goals of this work were to produce the fusion protein of the Trichoderma reesei swollenin I (SWOI) and Aspergillus niger feruloyl esterase A (FAEA) and to study the effect of the physical association of the fusion partners on the efficiency of the enzyme. The fusion protein was produced up to 25 mg l⁻¹ in the T. reesei strains Rut-C30 and CL847. In parallel, FAEA alone was produced for use as a control protein in application tests. Recombinant FAEA and SWOI–FAEA were purified to homogeneity and characterized. The biochemical and kinetic characteristics of the two recombinant proteins were found to be similar to those of native FAEA, except for the temperature stability and specific activity of the SWOI–FAEA. Finally, the SWOI–FAEA protein was tested for release of ferulic acid from wheat bran. A period of 24 h of enzymatic hydrolysis with the SWOI–FAEA improved the efficiency of ferulic acid release by 50% compared with the results obtained using the free FAEA and SWOI. Ferulic acid is used as an antioxidant and flavor precursor in the food and pharmaceutical industries. This is the first report of a potential application of the SWOI protein fused with an enzyme of industrial interest.     

The OSBP-related proteins: a novel protein family involved in vesicle transport,cellular lipid metabolism,and cell signalling

Proteins/genes showing high sequence homology to the mammalian oxysterol binding protein (OSBP) have been identified in a variety of eukaryotic organisms from yeast to man. The unifying feature of the gene products denoted as OSBP-related proteins (ORPs) is the presence of an OSBP-type ligand binding (LB) domain. The LB domains of OSBP and its closest homologue bind oxysterols, while data on certain other family members suggest interaction with phospholipids. Many ORPs also have a pleckstrin homology (PH) domain in the amino-terminal region. The PH domains of the family members studied in detail are known to interact with membrane phosphoinositides and play an important role in the intracellular targeting of the proteins. It is plausible that the ORPs constitute a regulatory apparatus that senses the status of specific lipid ligands in membranes, using the PH and/or LB domains, and mediates information to yet poorly known downstream machineries. Functional studies carried out on the ORP proteins in different organisms indicate roles of the gene family in diverse cellular processes including control of lipid metabolism, regulation of vesicle transport, and cell signalling events.     

Environmental Variation Generates Environmental Opportunist Pathogen Outbreaks

Many socio-economically important pathogens persist and grow in the outside host environment and opportunistically invade host individuals. The environmental growth and opportunistic nature of these pathogens has received only little attention in epidemiology. Environmental reservoirs are, however, an important source of novel diseases. Thus, attempts to control these diseases require different approaches than in traditional epidemiology focusing on obligatory parasites. Conditions in the outside-host environment are prone to fluctuate over time. This variation is a potentially important driver of epidemiological dynamics and affect the evolution of novel diseases. Using a modelling approach combining the traditional SIRS models to environmental opportunist pathogens and environmental variability, we show that epidemiological dynamics of opportunist diseases are profoundly driven by the quality of environmental variability, such as the long-term predictability and magnitude of fluctuations. When comparing periodic and stochastic environmental factors, for a given variance, stochastic variation is more likely to cause outbreaks than periodic variation. This is due to the extreme values being further away from the mean. Moreover, the effects of variability depend on the underlying biology of the epidemiological system, and which part of the system is being affected. Variation in host susceptibility leads to more severe pathogen outbreaks than variation in pathogen growth rate in the environment. Positive correlation in variation on both targets can cancel the effect of variation altogether. Moreover, the severity of outbreaks is significantly reduced by increase in the duration of immunity. Uncovering these issues helps in understanding and controlling diseases caused by environmental pathogens.     

Enabling Low Cost Biopharmaceuticals: A Systematic Approach to Delete Proteases from a Well-Known Protein Production Host Trichoderma reesei

The filamentous fungus Trichoderma reesei has tremendous capability to secrete proteins. Therefore, it would be an excellent host for producing high levels of therapeutic proteins at low cost. Developing a filamentous fungus to produce sensitive therapeutic proteins requires that protease secretion is drastically reduced. We have identified 13 major secreted proteases that are related to degradation of therapeutic antibodies, interferon alpha 2b, and insulin like growth factor. The major proteases observed were aspartic, glutamic, subtilisin-like, and trypsin-like proteases. The seven most problematic proteases were sequentially removed from a strain to develop it for producing therapeutic proteins. After this the protease activity in the supernatant was dramatically reduced down to 4% of the original level based upon a casein substrate. When antibody was incubated in the six protease deletion strain supernatant, the heavy chain remained fully intact and no degradation products were observed. Interferon alpha 2b and insulin like growth factor were less stable in the same supernatant, but full length proteins remained when incubated overnight, in contrast to the original strain. As additional benefits, the multiple protease deletions have led to faster strain growth and higher levels of total protein in the culture supernatant.     

Climatic Factors Influencing the Anthrax Outbreak of 2016 in Siberia,Russia

In 2016, an outbreak of anthrax killing thousands of reindeer and affecting dozens of humans occurred on the Yamal peninsula, Northwest Siberia, after 70 years of epidemiological situation without outbreaks. The trigger of the outbreak has been ascribed to the activation of spores due to permafrost thaw that was accelerated during the summer heat wave. The focus of our study is on the dynamics of local environmental factors in connection with the observed anthrax revival. We show that permafrost was thawing rapidly for already 6 years before the outbreak. During 2011–2016, relatively warm years were followed by cold years with a thick snow cover, preventing freezing of the soil. Furthermore, the spread of anthrax was likely intensified by an extremely dry summer of 2016. Concurrent with the long-term decreasing trend in the regional annual precipitation, the rainfall in July 2016 was less than 10% of its 30-year mean value. We conclude that epidemiological situation of anthrax in the previously contaminated Arctic regions requires monitoring of climatic factors such as warming and precipitation extremes.

     

Daily and annual variations of free fatty acid, glycerol and leptin plasma concentrations in goats (Capra hircus) under different photoperiods

The aim of the study was to differentiate the impact of lighting conditions and feeding times on the regulation of lipid metabolism of goats under different photoperiods throughout the year. Seven Finnish landrace goats were kept under artificial lighting that simulated the annual changes of photoperiod at 60 degrees N (the longest light period 18 h, the shortest 6 h). Ambient temperature and feeding regime were kept constant. Blood samples were collected six times a year at 2-h intervals for 2 days, first in light/dark (LD) conditions and then after 3 days in constant darkness (DD). Significant daily variations were detected in the concentrations of plasma free fatty acids (FFA) and glycerol throughout the year. The nocturnal decrease and morning rise of FFA levels were related to the photoperiod, while the trough levels of glycerol were associated with the concentrate meal times. In DD conditions, FFA and glycerol rhythms were unstable. A significant seasonal variation was detected in the overall FFA and glycerol levels suggesting decreased lipogenesis in winter, increased lipolysis in spring and high lipogenesis in summer and fall. There was no significant daily rhythm in serum leptin levels, nor did the profiles in LD and DD conditions differ. The leptin level was slightly lower in early fall than in the other seasons, paralleling a small decrease of body mass in the goats after the grazing season. The daily or annual variations of FFA and glycerol levels were not clearly related to leptin concentrations. The results suggest that lipid metabolism of goats is regulated by light even in constant temperature and feeding conditions; however, no significant contribution of leptin levels could be shown.     

Brood rearing costs affect future reproduction in the precocial common goldeneye Bucephala clangula

Life-history theory assumes a trade-off between current reproductive effort and future reproductive success. There are a large number of studies demonstrating reproductive trade-offs in different animal taxa, particularly in birds. Most bird studies have focused on the costs of chick rearing in altricial species. These costs have been assumed to be low in precocial species, but this aspect has been little studied. We used long-term individual reproductive data from the common goldeneye Bucephala clangula , an iteroparous precocial duck with uniparental female care, to examine whether brood rearing carries costs that affect future reproductive performance. All females were experienced breeders, and possible differences in female quality were ruled out. We compared within-individual (between-year) changes in clutch size, hatching date and body mass between females that had reared a brood in the previous year and females that had not. It turned out that brood rearing involved a cost in terms of clutch size and hatching date the next year, but not in terms of body mass: females that had reared a brood in the previous year laid relatively smaller clutches and laid relatively later than females that had not reared a brood. Our results show that normal brood rearing in a precocial species involves costs that affect future reproduction.