Carbon dynamics in peatlands under changing hydrology : Effects of water level drawdown on litter quality, microbial enzyme activities and litter decomposition rates

Pristine peatlands are carbon (C) accumulating wetland ecosystems sustained by a high water level (WL) and consequent anoxia that slows down decomposition. Persistent WL drawdown as a response to climate and/or land-use change directly affects decomposition: increased oxygenation stimulates decomposition of the old C (peat) sequestered under prior anoxic conditions. Responses of the new C (plant litter) in terms of quality, production and decomposability, and the consequences for the whole C cycle of peatlands are not fully understood. WL drawdown induces changes in plant community resulting in shift in dominance from Sphagnum and graminoids to shrubs and trees. There is increasing evidence that the indirect effects of WL drawdown via the changes in plant communities will have more impact on the ecosystem C cycling than any direct effects. The aim of this study is to disentangle the direct and indirect effects of WL drawdown on the new C by measuring the relative importance of 1) environmental parameters (WL depth, temperature, soil chemistry) and 2) plant community composition on litter production, microbial activity, litter decomposition rates and, consequently, on the C accumulation. This information is crucial for modelling C cycle under changing climate and/or land-use. The effects of WL drawdown were tested in a large-scale experiment with manipulated WL at two time scales and three nutrient regimes. Furthermore, the effect of climate on litter decomposability was tested along a north-south gradient. Additionally, a novel method for estimating litter chemical quality and decomposability was explored by combining Near infrared spectroscopy with multivariate modelling. WL drawdown had direct effects on litter quality, microbial community composition and activity and litter decomposition rates. However, the direct effects of WL drawdown were overruled by the indirect effects via changes in litter type composition and production. Short-term (years) responses to WL drawdown were small. In long-term (decades), dramatically increased litter inputs resulted in large accumulation of organic matter in spite of increased decomposition rates. Further, the quality of the accumulated matter greatly changed from that accumulated in pristine conditions. The response of a peatland ecosystem to persistent WL drawdown was more pronounced at sites with more nutrients. The study demonstrates that the shift in vegetation composition as a response to climate and/or land-use change is the main factor affecting peatland ecosystem C cycle and thus dynamic vegetation is a necessity in any models applied for estimating responses of C fluxes to changes in the environment. The time scale for vegetation changes caused by hydrological changes needs to extend to decades. This study provides grouping of litter types (plant species and part) into functional types based on their chemical quality and/or decomposability that the models could utilize. Further, the results clearly show a drop in soil temperature as a response to WL drawdown when an initially open peatland converts into a forest ecosystem, which has not yet been considered in the existing models.

Chromatographic separation, fractionation and oxidation of selected beta-lactoglobulin peptides

The literature review elucidates the mechanism of oxidation in proteins and amino acids and gives an overview of the detection and analysis of protein oxidation products as well as information about ?-lactoglobulin and studies carried out on modifications of this protein under certain conditions. The experimental research included the fractionation of the tryptic peptides of ?-lactoglobulin using preparative-HPLC-MS and monitoring the oxidation process of these peptides via reverse phase-HPLC-UV. Peptides chosen to be oxidized were selected with respect to their amino acid content which were susceptible to oxidation and fractionated according to their m/z values. These peptides were: IPAVFK (m/z 674), ALPMHIR (m/z 838), LIVTQTMK (m/z 934) and VLVLDTDYK (m/z 1066). Even though it was not possible to solely isolate the target peptides due to co-elution of various fractions, the percentages of target peptides in the samples were satisfactory to carry out the oxidation procedure. IPAVFK and VLVLDTDYK fractions were found to yield the oxidation products reviewed in literature, however, unoxidized peptides were still present in high amounts after 21 days of oxidation. The UV data at 260 and 280 nm enabled to monitor both the main peptides and the oxidation products due to the absorbance of aromatic side-chains these peptides possess. ALPMHIR and LIVTQTMK fractions were oxidatively consumed rapidly and oxidation products of these peptides were observed even on day 0. High rates of depletion of these peptides were acredited to the presence of His (H) and sulfur-containing side-chains of Met (M). In conclusion, selected peptides hold the potential to be utilized as marker peptides in ?-lactoglobulin oxidation.

On formation, growth and concentrations of air ions

Aerosol particles have effect on climate, visibility, air quality and human health. However, the strength of which aerosol particles affect our everyday life is not well described or entirely understood. Therefore, investigations of different processes and phenomena including e.g. primary particle sources, initial steps of secondary particle formation and growth, significance of charged particles in particle formation, as well as redistribution mechanisms in the atmosphere are required. In this work sources, sinks and concentrations of air ions (charged molecules, cluster and particles) were investigated directly by measuring air molecule ionising components (i.e. radon activity concentrations and external radiation dose rates) and charged particle size distributions, as well as based on literature review. The obtained results gave comprehensive and valuable picture of the spatial and temporal variation of the air ion sources, sinks and concentrations to use as input parameters in local and global scale climate models. Newly developed air ion spectrometers (Airel Ltd.) offered a possibility to investigate atmospheric (charged) particle formation and growth at sub-3 nm sizes. Therefore, new visual classification schemes for charged particle formation events were developed, and a newly developed particle growth rate method was tested with over one year dataset. These data analysis methods have been widely utilised by other researchers since introducing them. This thesis resulted interesting characteristics of atmospheric particle formation and growth: e.g. particle growth may sometimes be suppressed before detection limit (~ 3 nm) of traditional aerosol instruments, particle formation may take place during daytime as well as in the evening, growth rates of sub-3 nm particles were quite constant throughout the year while growth rates of larger particles (3-20 nm in diameter) were higher during summer compared to winter. These observations were thought to be a consequence of availability of condensing vapours. The observations of this thesis offered new understanding of the particle formation in the atmosphere. However, the role of ions in particle formation, which is not well understood with current knowledge, requires further research in future.

Detection, epidemiology and host spectrum of cowpox and Borna disease virus infections

Several orthopoxviruses (OPV) and Borna disease virus (BDV) are enveloped, zoonotic viruses with a wide geographical distribution. OPV antibodies cross-react, and former smallpox vaccination has therefore protected human populations from another OPV infection, rodent-borne cowpox virus (CPXV). Cowpox in humans and cats usually manifests as a mild, self-limiting dermatitis and constitutional symptoms, but it can be severe and even life-threatening in the immunocompromised. Classical Borna disease is a progressive meningoencephalomyelitis in horses and sheep known in central Europe for centuries. Nowadays the virus or its close relative infects humans and also several other species in central Europe and elsewhere, but the existence of human Borna disease with its suspected neuropsychiatric symptoms is controversial. The epidemiology of BDV is largely unknown, and the present situation is even more intriguing following the recent detection of several-million-year-old, endogenized BDV genes in primate and various other vertebrate genomes. The aims of this study were to elucidate the importance of CPXV and BDV in Finland and in possible host species, and particularly to 1) establish relevant methods for the detection of CPXV and other OPVs as well as BDV in Finland, 2) determine whether CPXV and BDV exist in Finland, 3) discover how common OPV immunity is in different age groups in Finland, 4) characterize possible disease cases and clarify their epidemiological context, 5) establish the hosts and possible reservoir species of these viruses and their geographical distribution in wild rodents, and 6) elucidate the infection kinetics of BDV in the bank vole. An indirect immunofluorescence assay and avidity measurement were established for the detection, timing and verification of OPV or BDV antibodies in thousands of blood samples from humans, horses, ruminants, lynxes, gallinaceous birds, dogs, cats and rodents. The mostly vaccine-derived OPV seroprevalence was found to decrease gradually according to the year of birth of the sampled human subjects from 100% to 10% in those born after 1977. On the other hand, OPV antibodies indicating natural contact with CPXV or other OPVs were commonly found in domestic and wild animals: the horse, cow, lynx, dog, cat and, with a prevalence occasionally even as high as 92%, in wild rodents, including some previously undetected species and new regions. Antibodies to BDV were detected in humans, horses, a dog, cats, and for the first time in wild rodents, such as bank voles (Myodes glareolus). Because of the controversy within the human Borna disease field, extra verification methods were established for BDV antibody findings: recombinant nucleocapsid and phosphoproteins were produced in Escherichia coli and in a baculovirus system, and peptide arrays were additionally applied. With these verification assays, Finnish human, equine, feline and rodent BDV infections were confirmed. Taken together, wide host spectra were evident for both OPV and BDV infections based on the antibody findings, and OPV infections were found to be geographically broadly distributed. PCR amplification methods were utilised for hundreds of blood and tissue samples. The methods included conventional, nested and real-time PCRs with or without the reverse transcription step and detecting four or two genes of OPVs and BDV, respectively. OPV DNA could be amplified from two human patients and three bank voles, whereas no BDV RNA was detected in naturally infected individuals. Based on the phylogenetic analyses, the Finnish OPV sequences were closely related although not identical to a Russian CPXV isolate, and clearly different from other CPXV strains. Moreover, the Finnish sequences only equalled each other, but the short amplicons obtained from German rodents were identical to monkeypox virus, in addition to German CPXV variants. This reflects the close relationship of all OPVs. In summary, RNA of the Finnish BDV variant could not be detected with the available PCR methods, but OPV DNA infrequently could. The OPV species infecting the patients of this study was proven to be CPXV, which is most probably also responsible for the rodent infections. Multiple cell lines and some newborn rodents were utilised in the isolation of CPXV and BDV from patient and wildlife samples. CPXV could be isolated from a child with severe, generalised cowpox. BDV isolation attempts from rodents were unsuccessful in this study. However, in parallel studies, a transient BDV infection of cells inoculated with equine brain material was detected, and BDV antigens discovered in archival animal brains using established immunohistology. Thus, based on several independent methods, both CPXV and BDV (or a closely related agent) were shown to be present in Finland. Bank voles could be productively infected with BDV. This experimental infection did not result in notable pathological findings or symptoms, despite the intense spread of the virus in the central and peripheral nervous system. Infected voles commonly excreted the virus in urine and faeces, which emphasises their possible role as a BDV reservoir. Moreover, BDV RNA was regularly reverse transcribed into DNA in bank voles, which was detected by amplifying DNA by PCR without reverse transcription, and verified with nuclease treatments. This finding indicates that BDV genes could be endogenized during an acute infection. Although further transmission studies are needed, this experimental infection demonstrated that the bank vole can function as a potential BDV reservoir. In summary, multiple methods were established and applied in large panels to detect two zoonoses novel to Finland: cowpox virus and Borna disease virus. Moreover, new information was obtained on their geographical distribution, host spectrum, epidemiology and infection kinetics.

Detection of clinical mastitis with the help of a thermal camera

Increasing dairy farm size and increase in automation in livestock production require that new methods are used to monitor animal health. In this study, a thermal camera was tested for its capacity to detect clinical mastitis. Mastitis was experimentally induced in 6 cows with 10 mu g of Escherichia coli lipopolysaccharide (LPS). The LPS was infused into the left forequarter of each cow, and the right forequarters served as controls. Clinical examination for systemic and local signs and sampling for indicators of inflammation in milk were carried out before morning and evening milking throughout the 5-d experimental period and more frequently on the challenge day. Thermal images of experimental and control quarters were taken at each sampling time from lateral and medial angles. The first signs of clinical mastitis were noted in all cows 2 h postchallenge and included changes in general appearance of the cows and local clinical signs in the affected udder quarter. Rectal temperature, milk somatic cell count, and electrical conductivity were increased 4 h postchallenge and milk N-acetyl-beta-D-glucosaminidase activity 8 h postchallenge. The thermal camera was successful in detecting the 1 to 1.5 degrees C temperature change on udder skin associated with clinical mastitis in all cows because temperature of the udder skin of the experimental and control quarters increased in line with the rectal temperature. Yet, local signs on the udder were seen before the rise in udder skin and body temperature. The udder represents a sensitive site for detection of any febrile disease using a noninvasive method. A thermal camera mounted in a milking or feeding parlor could detect temperature changes associated with clinical mastitis or other diseases in a dairy herd.

Detection and molecular epidemiology of tick-borne encephalitis virus infections

Pdf-file, link above

Change detection and controlling forest information using multi-temporal Landsat TM imagery.

A method was developed for relative radiometric calibration of single multitemporal Landsat TM image, several multitemporal images covering each others, and several multitemporal images covering different geographic locations. The radiometricly calibrated difference images were used for detecting rapid changes on forest stands. The nonparametric Kernel method was applied for change detection. The accuracy of the change detection was estimated by inspecting the image analysis results in field. The change classification was applied for controlling the quality of the continuously updated forest stand information. The aim was to ensure that all the manmade changes and any forest damages were correctly updated including the attribute and stand delineation information. The image analysis results were compared with the registered treatments and the stand information base. The stands with discrepancies between these two information sources were recommended to be field inspected.

Phytophthora in Finnish nurseries

Human-mediated movement of plants and plant products is now generally accepted to be the primary mode of introduction of plant pathogens. Species of the genus Phytophthora are commonly spread in this way and have caused severe epidemics in silviculture, horticulture as well as natural systems all over the world. The aims of the study were to gather information on the occurrence of Phytophthora spp. in Finnish nurseries, to produce information for risk assessments for these Phytophthora spp. by determining their host ranges and tolerance of cold temperatures, and to establish molecular means for their detection. Phytophthora cactorum was found to persist in natural waterbodies and results suggest that irrigation water might be a source of inoculum in nurseries. In addition to P. cactorum, isolates from ornamental nursery Rhododendron yielded three species new to Finland: P. ramorum, P. plurivora and P. pini. The only species with quarantine status, P. ramorum, was most adapted to growth in cold temperatures and able to persist in the nursery in spite of an annual sanitation protocol. Phytophthora plurivora and the closely related P. pini had more hosts among Nordic tree and plant species than P. ramorum and P. cactorum, and also had higher infectivity rates. All four species survived two weeks in -5 °C , and thus soil survival of these Phytophthoras in Finland is likely under current climatic conditions. The most common tree species in Finnish nurseries, Picea abies, was highly susceptible to P. plurivora and P. pini in pathogenicity trials. In a histological examination of P. plurivora in P. abies shoot tissues, fast necrotrophic growth was observed in nearly all tissues. The production of propagules in P. abies shoot tissue was only weakly indicated. In this study, a PCR DGGE technique was developed for simultaneous detection and identification of Phytophthora spp. It reliably detected Phytophthora in plant tissues and could discriminate most test species as well as indicate instances of multiple-species infections. It proved to be a useful detection and identification tool either applied alone or in concert with traditional isolation culture techniques. All of the introduced species of Phytophthora had properties that promote a high risk of establishment and spread in Finland. It is probable that more pathogens of this genus will be introduced and become established in Finland and other Nordic countries unless efficient phytosanitary control becomes standard practice in the international plant trade.

Microbial identification by detection of ligation probes on DNA microarray

Microbes in natural and artificial environments as well as in the human body are a key part of the functional properties of these complex systems. The presence or absence of certain microbial taxa is a correlate of functional status like risk of disease or course of metabolic processes of a microbial community. As microbes are highly diverse and mostly notcultivable, molecular markers like gene sequences are a potential basis for detection and identification of key types. The goal of this thesis was to study molecular methods for identification of microbial DNA in order to develop a tool for analysis of environmental and clinical DNA samples. Particular emphasis was placed on specificity of detection which is a major challenge when analyzing complex microbial communities. The approach taken in this study was the application and optimization of enzymatic ligation of DNA probes coupled with microarray read-out for high-throughput microbial profiling. The results show that fungal phylotypes and human papillomavirus genotypes could be accurately identified from pools of PCR amplicons generated from purified sample DNA. Approximately 1 ng/μl of sample DNA was needed for representative PCR amplification as measured by comparisons between clone sequencing and microarray. A minimum of 0,25 amol/μl of PCR amplicons was detectable from amongst 5 ng/μl of background DNA, suggesting that the detection limit of the test comprising of ligation reaction followed by microarray read-out was approximately 0,04%. Detection from sample DNA directly was shown to be feasible with probes forming a circular molecule upon ligation followed by PCR amplification of the probe. In this approach, the minimum detectable relative amount of target genome was found to be 1% of all genomes in the sample as estimated from 454 deep sequencing results. Signal-to-noise of contact printed microarrays could be improved by using an internal microarray hybridization control oligonucleotide probe together with a computational algorithm. The algorithm was based on identification of a bias in the microarray data and correction of the bias as shown by simulated and real data. The results further suggest semiquantitative detection to be possible by ligation detection, allowing estimation of target abundance in a sample. However, in practise, comprehensive sequence information of full length rRNA genes is needed to support probe design with complex samples. This study shows that DNA microarray has the potential for an accurate microbial diagnostic platform to take advantage of increasing sequence data and to replace traditional, less efficient methods that still dominate routine testing in laboratories. The data suggests that ligation reaction based microarray assay can be optimized to a degree that allows good signal-tonoise and semiquantitative detection.