Seasonal fluctuations in blue-green algae, especially in heterocystous species in polluted coastal waters off Helsinki city, in the northern part of the Gulf of Finland

Tiivistelmä

Nomenclatural corrections in Mesoamerican Plantaginaceae and a new species of Tetranema from Honduras

Study of the Plantaginaceae for the Flora Mesoamericana project has resulted in five lectotypifications, a new combination in Rhodochiton, and the discovery of a new species of Tetranema from Honduras. This species, Tetranema michaelfayanum, is described here, a key to the species of Tetranema is provided, and the T. roseum complex is discussed.

Basic and translational studies on species B adenoviruses

Human adenoviruses (Ads) have been classified into six species (A to F) currently containing 55 serotypes. For almost 2 decades vectors derived from group C serotype Ad5 have been extensively used for gene transfer studies. These Ad5 based vectors are able to efficiently infect many mammalian cell types (including both mitotic and post-mitotic cells) through interaction with a primary attachment receptor, the coxsackie and adenovirus receptor (CAR). Despite the many advantages of Ad5 based vectors a number of limitations have affected their therapeutic application to many diseases. Although they can transduce many tissue types, Ad5 based vectors are unable to efficiently transduce several potential disease target cell types, including hematopoietic stem cells and malignant tumor cells. Therefore, newer vectors have been developed based on Ad serotypes other than Ad5. This thesis focuses on species B Ads. Species B Ads are comprised of three groups based on their receptor usage. Group 1 of species B Ads (Ad16, 21, 35, 50) nearly exclusively utilize CD46 as a receptor; Group 2 (Ad3, Ad7, 14) share a common, unidentified receptor/s, which is not CD46 and which was tentatively named receptor X; Group 3 (Ad11) preferentially interacts with CD46, but also utilizes receptor X if CD46 is blocked. Species B group Ads are important human pathogens. Species B group 2 serotypes are isolated from patients with respiratory tract infections, whereas the Group 1 viruses are described as causing kidney and urinary tract infections. B-group Ad infections often occur in immunocompromised patients, including AIDS patients, recipients of bone marrow transplants, or chemotherapy patients. Recent studies performed in U.S. military training facilities indicate an emergence of diverse species B serotypes at the majority of sites. This included the group 1 serotype 21 and the group 2 serotypes 3, 7, and 14. CD46-targeting vectors derived from Ad35 and Ad11 are important tools for in vitro gene transfer into human stem cells, including hematopoietic stem cells and induced pluripotent stem cells. Ad35 and Ad11 have been used as tools for cancer therapy, because CD46 appears to be uniformely overexpressed on many cancers. Furthermore, receptor X-targeting vectors, i.e vectors derived from Ad3 or vectors containing Ad3 fibers have shown superior in the transduction of tumor cells both in vitro and in vivo and are currently being used clinically in cancer patients. While extensive basic virology studies have been done on Ad5, the information of species B group 1 interaction with CD46 is limited. Furthermore, the receptor for a major subgroup of species B Ads (receptor X) is unknown. The goal of this thesis was it therefore to better understand virological and translational aspects of species B Ads. The specific findings described in this thesis include i) the identification of CD46 binding sites within the Ad35 fiber knob, ii) the study of the in vitro and in vivo properties of Ad vectors with increased affinity to CD46. iii) the study of the receptor usage of a newly emergent Ad14a, iv) the identification of desmoglein 2 as the receptor for Ad3, Ad7, Ad11, and Ad14, v) the delineation of structural details of Ad3 virus interaction with DSG2, and vi) the analysis of functional consequences of Ad3-DSG2 interaction. As a result of these basic virology studies two Ad-derived recombinant proteins have been generated that can be used to enhance cancer therapy by monoclonal antibodies.

Molecular profiling of indoor microbial communities in moisture damaged and non-damaged buildings

Epidemiological studies have shown an elevation in the incidence of asthma, allergic symptoms and respiratory infections among people living or working in buildings with moisture and mould problems. Microbial growth is suspected to have a key role, since the severity of microbial contamination and symptoms show a positive correlation, while the removal of contaminated materials relieves the symptoms. However, the cause-and-effect relationship has not been well established and knowledge of the causative agents is incomplete. The present consensus of indoor microbes relies on culture-based methods. Microbial cultivation and identification is known to provide qualitatively and quantitatively biased results, which is suspected to be one of the reasons behind the often inconsistent findings between objectively measured microbiological attributes and health. In the present study the indoor microbial communities were assessed using culture-independent, DNA based methods. Fungal and bacterial diversity was determined by amplifying and sequencing the nucITS- and16S-gene regions, correspondingly. In addition, the cell equivalent numbers of 69 mould species or groups were determined by quantitative PCR (qPCR). The results from molecular analyses were compared with results obtained using traditional plate cultivation for fungi. Using DNA-based tools, the indoor microbial diversity was found to be consistently higher and taxonomically wider than viable diversity. The dominant sequence types of fungi, and also of bacteria were mainly affiliated with well-known microbial species. However, in each building they were accompanied by various rare, uncultivable and unknown species. In both moisture-damaged and undamaged buildings the dominant fungal sequence phylotypes were affiliated with the classes Dothideomycetes (mould-like filamentous ascomycetes); Agaricomycetes (mushroom- and polypore-like filamentous basidiomycetes); Urediniomycetes (rust-like basidiomycetes); Tremellomycetes and the family Malasseziales (both yeast-like basidiomycetes). The most probable source for the majority of fungal types was the outdoor environment. In contrast, the dominant bacterial phylotypes in both damaged and undamaged buildings were affiliated with human-associated members within the phyla Actinobacteria and Firmicutes. Indications of elevated fungal diversity within potentially moisture-damage-associated fungal groups were recorded in two of the damaged buildings, while one of the buildings was characterized by an abundance of members of the Penicillium chrysogenum and P. commune species complexes. However, due to the small sample number and strong normal variation firm conclusions concerning the effect of moisture damage on the species diversity could not be made. The fungal communities in dust samples showed seasonal variation, which reflected the seasonal fluctuation of outdoor fungi. Seasonal variation of bacterial communities was less clear but to some extent attributable to the outdoor sources as well. The comparison of methods showed that clone library sequencing was a feasible method for describing the total microbial diversity, indicated a moderate quantitative correlation between sequencing and qPCR results and confirmed that culture based methods give both a qualitative and quantitative underestimate of microbial diversity in the indoor environment. However, certain important indoor fungi such as Penicillium spp. were clearly underrepresented in the sequence material, probably due to their physiological and genetic properties. Species specific qPCR was a more efficient and sensitive method for detecting and quantitating individual species than sequencing, but in order to exploit the full advantage of the method in building investigations more information is needed about the microbial species growing on damaged materials. In the present study, a new method was also developed for enhanced screening of the marker gene clone libraries. The suitability of the screening method to different kinds of microbial environments including biowaste compost material and indoor settled dusts was evaluated. The usability was found to be restricted to environments that support the growth and subsequent dominance of a small number microbial species, such as compost material.

The many ways to delimit species: hairs, genes and surface chemistry

Species identification forms the basis for understanding the diversity of the living world, but it is also a prerequisite for understanding many evolutionary patterns and processes. The most promising approach for correctly delimiting and identifying species is to integrate many types of information in the same study. Our aim was to test how cuticular hydro- carbons, traditional morphometrics, genetic polymorphisms in nuclear markers (allozymes and DNA microsatellites) and DNA barcoding (partial mitochondrial COI gene) perform in delimiting species. As an example, we used two closely related Formica ants, F. fusca and F. lemani, sampled from a sympatric population in the northern part of their distribu- tion. Morphological characters vary and overlap in different parts of their distribution areas, but cuticular hydrocarbons include a strong taxonomic signal and our aim is to test the degree to which morphological and genetic data correspond to the chemical data. In the morphological analysis, species were best separated by the combined number of hairs on pro- notum and mesonotum, but individual workers overlapped in hair numbers, as previously noted by several authors. Nests of the two species were separated but not clustered according to species in a Principal Component Analysis made on nuclear genetic data. However, model-based Bayesian clustering resulted in perfect separation of the species and gave no indication of hybridization. Furthermore, F. lemani and F. fusca did not share any mitochondrial haplotypes, and the species were perfectly separated in a phylogenetic tree. We conclude that F. fusca and F. lemani are valid species that can be separated in our study area relatively well with all methods employed. However, the unusually small genetic differen- tiation in nuclear markers (FST = 0.12) shows that they are closely related, and occasional hybridization between F. fusca and F. lemani cannot be ruled out.

Taxonomy and phylogeny of the manna lichens and allied species (Megasporaceae)

This dissertation is focused on the taxonomy, phylogeny, and ecology of the vagrant, erratic and allied terricolous and saxicolous species of the genera Aspicilia A. Massal. and Circinaria Link (Megasporaceae), particularly those traditionally referred to as manna lichens . The group has previously been defined on the basis of few morphological characters. The phylogeny of the family Megasporaceae is inferred from the combined dataset of nuLSU and mtSSU sequences. Five genera Aspicilia, Circinaria, Lobothallia, Megaspora, and Sagedia are recognized. Lobothallia is sister of the four other genera, while Aspicilia and Sagedia form the next clade. All these genera have small asci with eight spores. Circinaria is a sister genus of Megaspora, and these two have in common asci with (1 4) 6 8 large spores. Circinaria forms a monophyletic group and sphaerothallioid species form a monophyletic group within Circinaria. The presence of certain morphological characters such as pseudocyphellae, thickness of cortex and medulla layers, as well as ecological differences in sphaerothallioid species distinguish it from some other crustose species, especially those containing aspicilin and characterised by thin cortex and medulla layers, conidium length c. 6 12 µm and absence of pseudocyphellae. If sphaerothallioid species are accepted as a distinct genus, the rest of the Circinaria species would remain as a paraphyletic assemblage. Currently, the genus Circinaria includes all the sphaerothallioid species and its generic position is confirmed and accepted. Thus, it is proposed as a correct generic name also for the manna lichens described originally in other genera. Phylogeny at the species level was studied using nrITS sequence data. Traditionally, morphological characters have been used for the recognition of species. They were re-evaluated in the light of molecular data. Since characters such as vagrant, erratic and crustose growth forms proved to be misleading for the recognition of some species, a combination of several characters (including molecular data) is recommended. Vagrant growth form seems to have evolved several times among the distantly related lineages and even within a single population. The reasons behind the high plasticity in the external morphology of the sphaerothallioid Circinaria remain, however, unknown. Six new species are recognized: Aspicilia tibetica, Circinaria arida, C. digitata nom provis., C. gyrosa nom. provis., C. rogeri nom. provis., and C. rostamii nom. provis. Based on an analysis of nrITS dataset, three new erratic, vagrant and crustose species were also recognized, but these require additional study. The results also reveal that C. elmorei and C. hispida are not monophyletic as currently understood. In addition, 13 new combinations in the genus Circinaria are proposed.

Onttojen puiden lahopuukovakuoriaiset — kolmen pyydystyypin vertailu

Vanhat ja ontot puut ovat tärkeä elinympäristö monelle lahopuusta riippuvaiselle eliölajille. Onttoihin puihin on erikoistunut suuri määrä myös vaarantuneita ja harvinaisia hyönteislajeja, jotka elävät puun onkalon seinämillä tai onkalon pohjalle kerääntyvässä orgaanisessa aineksessa, ns. mulmissa. Tutkimuksen tavoitteena oli selvittää, mikä kolmesta pyydystyypistä (ikkuna-, vuoka- ja kuoppapyydys) soveltuu parhaiten onttojen puiden lahopuukovakuoriaisten pyyntiin. Lisäksi tavoitteena oli kartoittaa hyönteisnäytteiden ensimmäiseen laboratoriokäsittelyyn vaadittua aikaa. Tutkimuksessa oli mukana vanhoja rungostaan onttoutuneita lehmuksia, tammia ja vaahteroita pääkaupunkiseudun puisto- ja kartanoalueilta. Puiden onkaloiden sisään aseteltiin ikkuna-, vuoka- ja kuoppapyydyksiä, kaksi kutakin tyyppiä ja ne tyhjennettiin kolmen viikon välein touko-heinäkuussa 2006. Pyydyksiä oli siis yhteensä 90 per pyyntijakso. Kun näytteistä eroteltiin halutut hyönteislahkot (mukaanlukien kovakuoriaiset) niiden käsittelyyn käytetty aika kirjattiin ylös. Aineistosta tunnistettiin yhteensä 3825 kovakuoriaisyksilöä ja 212 lajia, joista lahopuusta riippuvaisia oli yhteensä 3398 yksilöä ja 121 lajia. Ikkunapyydyksissä esiintyi yhteensä 1639 yksilöä ja 140 lajia, vuokapyydyksissä 1506 yksilöä ja 134 lajia, kuoppapyydyksissä 680 yksilöä ja 111 lajia. Näytteiden käsittelyaikojen keskiarvot olivat 48,3 minuuttia ikkunapyydykselle, 65,5 minuuttia vuokapyydykselle ja 34,1 kuoppapyydykselle. Lajistokoostumuksen huomioiva ?-diversiteetti erosi huomattavasti pyydysten välillä, se oli 36,5 % ikkuna- ja vuokapyydysten välillä, 13,1 % ikkuna- ja kuoppapyydysten välillä ja 14,2 % vuoka- ja kuoppapyydysten välillä. Ikkuna- ja vuokapyydysten välillä ei havaittu tilastollisesti merkitsevää eroa saproksyylilajien (p<0,05), -yksilöiden (p<0,05) tai käsittelyaikojen (p<0,05) keskiarvoissa. Ikkuna- ja vuokapyydyksillä saatiin keskimäärin selvästi enemmän saproksyylilajeja ja –yksilöitä kuoppapyydykseen verrattuna. Kuoppapyydyksellä saatiin kokonaisyksilömäärään verrattuna suhteellisesti vähemmän saproksyylejä (59 %) kuin ikkuna- (69 %) ja vuokapyydyksillä (71 %). Ikkunapyydykset olivat tehokkain pyydystyyppi vertailtaessa pyydysten keräämää saproksyyliyksilömäärää suhteessa aineiston käsittelyn vaatimaan aikaan. Tehokkuus (yksilöä minuutissa) ikkunapyydykselle oli 0,74, vuokapyydykselle 0,43 ja kuoppapyydykselle 0,21. Ikkunapyydyksiä ei ole aikaisemmin käytetty puun onkalon sisällä hyönteisiä pyydettäessä vaan ne ovat aikaisemmissa tutkimuksissa roikkuneet onkalon ulkopuolella. Ikkunapyydykset kuitenkin toimivat erinomaisesti myös onkaloiden sisällä. Ikkuna- sekä vuokapyydys toimivatkin selkeästi paremmin lahopuukovakuoriaisten pyynnissä verrattuna kuoppapyydykseen, jonka poisjättäminen olisi kuitenkin tuottanut huomattavasti lajiköyhemmän aineiston. Mahdollisimman monimuotoisen onttojen puiden lahopuukovakuoriaislajiston keräämiseksi tulisi käyttää ikkuna- tai vuokapyydyksiä yhdessä kuoppapyydysten kanssa.