DNA-Based Enzyme Reactors and Systems

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Bursti-tunnistusmenetelmät ihmisperäisistä monikykyisistä kantasoluista erilaistetuille hermosoluverkostoille

A burst is a set of subsequent action potentials that are fired at a high frequency. Although bursts are a fundamental part of electrical activity of neuronal networks in vitro, no standardized method exists for burst detection. Visual identification of bursts is a widely accepted method, but it is not objective nor time-efficient. Therefore, various algorithms have been developed for burst detection. Burst detection algorithms are typically developed and verified only on one specific type of data. This can be problematic because the bursting activity is highly variable between different cell types. Consequently, the applicability of the algorithms is restricted to a narrow range of activity types. Especially applicability to human neuronal networks is questionable because the algorithms are often developed on rodent neuronal networks, which display distinct activity patterns in comparison to human networks. The aim of this thesis was to produce a test data set, which would well represent bursting and non-bursting activity observed in human pluripotent stem cell (hPSC)-derived neuronal networks, and to identify a single algorithm with optimal parameters that would successfully detect the bursts in this test data set. As rodent neuronal networks are also widely used in neuroscience, the algorithm was desired to function also on activity derived from rodent cultures. To achieve these goals, hESCs were differentiated into functional neuronal networks and cultured on microelectrode array (MEA). Primary rat cortical neurons were similarly cultured on MEA. Electrical activity of the developing networks was recorded twice a week until synchronized bursting emerged. At this point, pharmacological assays were performed in order to record modulated activity. On the MEA recordings, distinct activity patterns were identified, and short recordings representative of the distinct patterns were included to the test data set. The performance of four contemporary burst detection algorithms was evaluated on the test data set. The evaluation was based on visual identification of bursts from the raw MEA signal. For each algorithm, a performance score was determined and sensitivity and specificity were computed. The evaluation was performed in two runs using either 3 or 5 as minimum number of spikes required for a burst. Other algorithm parameters were set to default values suggested by the original authors. The optimization possibilities were not encouraging for other algorithms but logISI, which also provided the highest performance and the most balanced sensitivity and specificity values. Parameters of logISI were optimized for the test data set, which significantly improved its performance. As a result, logISI displayed good or excellent performance on the test data obtained from human and rat neuronal networks during spontaneous and pharmacologically modulated activity. Based on these results, logISI could have the potential to become a standard burst detection algorithm in the field.Bursti (engl. burst) on peräkkäisten korkealla taajuudella esiintyvien toimintapotentiaalien ryhmä. Vaikka burstit ovat olennainen osa maljalla kasvatettujen hermosoluverkostojen sähköistä aktiivisuutta, ei niiden tunnistukseen ole standardimenetelmää. Visuaalinen bursti-tunnistus on laajasti hyväksytty menetelmä, mutta se ei ole objektiivinen eikä ajallisesti tehokas. Tästä syystä bursti-tunnistukseen on kehitetty useita algoritmeja. Tyypillisesti nämä algoritmit on kehitetty ja niiden toiminta on varmennettu vain tietyn tyyppisellä datalla. Tämä voi olla ongelmallista, koska bursti-aktiivisuus eri solutyyppien välillä on vaihtelevaa. Näin ollen algoritmien soveltaminen on rajoitettu vain pieneen osaan aktiivisuustyyppejä. Erityisesti algoritmien soveltaminen ihmisperäisiin hermosoluverkostoihin on kyseenalaista, sillä algoritmit on usein kehitetty jyrsijäperäisillä hermosoluverkostoilla, joiden aktiivisuustyypit eroavat ihmisperäisisten hermosoluverkostojen aktiivisuustyypeistä. Tämän työn tavoitteena oli kerätä testiaineisto, joka sisältäisi monikykyisistä ihmisen kantasoluista erilaistetuissa hermosoluverkostoissa havaittavat burstaavat ja ei-burstaavat aktiviisuustyypit, sekä löytää tällä testiaineistolla toimiva algoritmi ja optimaaliset arvot sen muuttujille. Koska jyrsijäperäiset hermosoluverkostot ovat neurotieteissä paljon käytettyjä, valitun algoritmin haluttiin toimivan myös niistä peräisin olevalla aineistolla. Tavoitteen saavuttamiseksi ihmisperäisistä alkion kantasoluista erilaistettiin toiminnallisia hermosoluverkostoja, joita viljeltiin mikroelektrodihilan (engl. microelectrode array, MEA) päällä. Rotan eristettyjä aivokuoren hermosoluja viljeltiin samoin MEA:lla. Hermosoluverkostojen sähköistä aktiivisuutta mitattiin niiden kehityksen aikana kahdesti viikossa, kunnes havaittiin synkronista bursti-aktiivisuutta. Synkronisen bursti-aktiivisuuden ilmaannuttua suoritettiin farmakologiset testit ja mitattiin näin muunneltua aktiivisuutta. Saaduista MEA-mittauksista etsittiin erilaisia aktiivisuustyyppejä, joista muodostettiin testiaineisto. Neljän nykyaikaisen algoritmin toimintaa arvioitiin tässä testiaineistossa. Arviointi tehtiin vertailemalla algoritmien tuloksia raakasignaalista tehdyn visuaalisen bursti-tunnistuksen tuloksiin. Jokaisen algoritmin suoritus pisteytettiin ja niiden herkkyys ja tarkkuus laskettiin. Suoritusta arvioitiin kahdesti siten, että burstin vähimmäispiikkimäärä asetettiin ensin kolmeen ja sitten viiteen. Muiden muuttujien arvot asetettiin algoritmien kehittäjien alkuperäisten suositusten mukaisesti. Optimointi- mahdollisuudet olivat lupaavat vain logISI-algoritmille, joka myös suoriutui parhaiten ja jonka herkkyys ja tarkkuus olivat parhaassa tasapainossa. LogISI:n muuttujat optimoitiin testiaineistolle, mikä huomattavasti paransi sen suoritusta kyseisessä aineistossa. Optimoidulla logISI:llä saatiin joko hyvä tai erinomainen tulos koko testiaineistolla, joka oli saatu mittaamalla spontaania ja farmakologisesti muunneltua aktiivisuutta sekä ihmisperäisistä kantasoluista erilaistetuista että rotan aivokuoresta eristetyistä hermosolu-verkostoista. Näiden tulosten perusteella logISI on potentiaalinen vaihtoehto bursti-tunnistuksen standardimetodiksi

Teaching and supporting students with special educational needs at a distance during the COVID-19 school closures in Finland : special needs teachers’ experiences

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Photochemical transformation of dissolved organic matter in aquatic environment : From a boreal lake and Baltic Sea to global coastal ocean

As dissolved organic matter (DOM) constitutes a vast reservoir of carbon and nutrients in lakes, rivers and ocean, it plays an important role in the global carbon and nutrient cycles. Only a part of DOM is directly biologically utilizable by bacteria, but solar radiation induced photochemical reactions may mineralize DOM to inorganic forms such as dissolved inorganic carbon (DIC) and ammonium (NH4+), and also degrade non-labile DOM molecules to labile organic substrates available for bacteria. This dissertation examined the photochemical transformation of DOM in the surface water in different scales of aquatic environments; from a Finnish boreal lake and the Baltic Sea to the coastal areas of ten globally big rivers. This dissertation studied photochemical reactions such as photoproduction of DIC, NH4+, and labile substrates supporting bacterial growth, and determined the photoreactivity of DOM, i.e., apparent quantum yields for the photoreactions in each environment. To consider the relevance of photochemistry of DOM, an optical model was used to quantify the photoreaction rates taking into account for the determined photoreactivity of DOM and solar radiation incidental to each environment studied. In the Baltic Sea, the pelagic heterotrophic bacterioplankton was carbon-limited indicating low bioavailability of DOM in the surface water. Irradiations of the waters with natural or simulated solar light resulted in photobleaching of chromophoric dissolved organic matter (CDOM) and phototransformation of DOM to DIC and NH4+ as well as to labile DOM substrates. This dissertation showed that across the entire Baltic Sea, the annual photoproduction of NH4+ corresponded to 9 18% of the annual river loading of DON, but the photoproduction of DIC exceeded the annual river loading of photoreactive terrigenous DOC. Furthermore, the studies along the salinity transects indicated that terrigenous DOC was more photoreactive than marine DOC but the marine DON was more reactive than terrigenous DON. The photoproduced labile substrates supported bacterial production and biomass leading to a 3-level trophic transfer of non-labile DOM and a simultaneous stimulation of autotrophic algae and primary production. The annual amount of photostimulated bacterial biomass corresponded to 3 5% of total bacterial biomass across the entire Baltic Sea. In the Baltic Sea as well as in the mesohumic lake studied, the photolytic water layer was shallow and limited the phototransformation of DOM to the top 30 cm. In the global scale, the annual DIC photoproduction from terrigenous CDOM in front of the ten rivers studied (12.5 ± 2.1 Tg C y-1) corresponded to 18 ± 4% of annual flux of terrigenous DOC of the rivers. When extrapolated to a global estimate, 44.5 ± 10.6 Tg of terrigenous DOC was annually mineralized to DIC by solar radiation in coastal waters. Globally, the amount of photomineralization of terrigenous DOC was larger in coastal ocean than in lakes and reservoirs. However, the areal rates of DIC photoproduction were larger in the lakes and reservoirs than in the coastal waters indicating that phototransformation of terrigenous DOC was likely limited by relatively short residence times in inland waters. To conclude, the phototransformation in coastal waters formed the final sink for riverine terrigenous CDOM and DOC, but was restricted in general, to a few hundred kilometres from river mouths to the ocean with the exception of largest discharging rivers.Liuennutta orgaanista ainetta (dissolved organic matter, DOM), ml. humusaineet, kulkeutuu vesistöihin valuma-alueelta mm. luonnonhuuhtouman mukana, minkä lisäksi sitä syntyy vesieliöiden toiminnan seurauksena vesistöissä. Maailmanlaajuisesti tarkastellen merien, järvien ja jokivesien DOM muodostaa merkittävän orgaanisen hiilen ja typen varaston, ja siten sen dynamiikan ja hajoamisen ymmärtäminen on olennaista hiilen ja ravinteiden kierron kannalta. Tämän väitöskirjatyön tarkoituksena oli selvittää ja mitata prosesseja, jotka hajottavat DOMia luonnonvesissä (valokemiallinen ja mikrobiologinen hajoaminen), sekä mallinnuksen avulla arvioida DOMin hajoamisen merkitystä erilaisissa vesiympäristöissä. Tutkimusalueina olivat Lammin Pääjärvi, Itämeri sekä jokisuualueet kymmenestä suuresta joesta (Mississippi, Amazon, Kongo, Paraná, Yantgze, Mekong, Ganges-Brahmaputra, Lena, Tonava ja St. Lawrence). Liuennut orgaaninen aine voi hajota vesistöissä joko biologisesti bakteeritoiminnan seurauksena tai valokemiallisesti auringon säteilyn, erityisesti UV-säteilyn, vaikutuksesta. Useimmiten DOM-yhdisteet ovat niin monimutkaisia ja heikosti biologisesti käyttökelpoisia, että ainoastaan valokemialliset reaktiot voivat hajottaa niitä. Toisaalta valokemiallisia hajotusreaktioita voi tapahtua vain esimerkiksi järven tai meren valaistussa pintavesikerroksessa, joka väitöskirjassa tutkitussa järvessä sekä Itämerellä rajoittui ylimpään 30 cm:iin. Auringon säteily voi muuntaa DOMin sisältämän orgaanisen typen esim. leville käyttökelpoiseen muotoon (ammoniumtypeksi) ja orgaanisen hiilen joko hiilidioksidiksi tai pienemmiksi orgaanisiksi yhdisteiksi, jotka ovat käyttökelpoisia bakteereille. Osa bakteerien käyttämästä hiilestä muuntuu bakteerihengityksen kautta edelleen hiilidioksidiksi ja osa bakteeribiomassaksi, joka siirtyy bakteereja syövien siimaeliöiden kautta osaksi ylempää ravintoverkkoa. Tässä väitöskirjassa havaittiin, että valokemiallisten reaktioiden tuottamat bakteereille käyttökelpoiset hiiliyhdisteet lisäsivät bakteerien kasvun lisäksi myös siimaeliöiden sekä niitä laiduntavien ripsieläinten biomassaa Itämerellä. Valokemiallisesti tuotettu typpi puolestaan lisäsi planktonlevien kasvua. Väitöskirjassa arvioitiin, että koko Itämeren alueella valokemia hajotti vuodessa orgaanista hiiltä hiilidioksidiksi määrällisesti enemmän kuin joet vuosittain kuljettavat DOMia valuma-alueelta Itämereen. Orgaanisen typen valokemiallinen hajoaminen puolestaan vastasi 9-18 % vuosittaisesta liuenneen orgaanisen typen jokikuormituksesta ja saattaa siten muodostaa merkittävän typenlähteen typpirajoitteisille leville Itämerellä. Väitöskirjassa tarkasteltujen suurten jokien mereen kuljettamasta DOM-kuormasta 14-22 % hajosi hiilidioksidiksi valokemian seurauksena. Maailmanlaajuinen arvio jokivesien mereen kuljettaman orgaanisen hiilen kohtalosta osoitti, että maalta mereen kulkeutunut aines hajoaa jo rannikkovesialueella. Jokivesien vaikutus ulottuu keskimäärin korkeintaan noin 500 km etäisyydelle rannasta, lukuun ottamatta suurimpia DOMia kuljettavia jokia (Amazon ja Kongo) sekä Jäämerelle laskevaa jokea (Lena), jossa valaistusolot rajoittavat DOMin valokemiallista hajoamista

Teaching and supporting students with special educational needs at a distance during the COVID-19 school closures in Finland : special needs teachers’ experiences

Peer reviewedPublisher PD

Loadings of dissolved organic matter and nutrients from the Neva River into the Gulf of Finland - Biogeochemical composition and spatial distribution within the salinity gradient

We studied the loadings of dissolved organic matter (DOM) and nutrients from the Neva River into the Eastern Gulf of Finland, as well as their distribution within the salinity gradient. Concentrations of dissolved organic carbon (DOC) ranged from 390 to 840 mu M, and were related to absorption of colored DOM (CDOM) at 350 nm, a(CDOM)(350), ranging from 2.70 to 17.8 m(-1). With increasing salinity both DOC and a(CDOM) decreased, whereas the slope of a(CDOM) spectra, S-CDOM(300-700), ranging from 14.3 to 21.2 mu m(-1), increased with salinity. Deviations of these properties from conservative mixing models were occasionally observed within the salinity range of approximately 1-4, corresponding to the region between 27 and 29 degrees E. These patterns are suggested to mostly reflect seasonal changes in properties of river end-member and hydrodynamics of the estuary, rather than non-conservative processes. On the other hand, observed nonlinear relationships observed between a(CDOM)*(350) and S-CDOM(275-295) emphasized the importance of photochemistry among various transformation processes of DOM. Dissolved inorganic nitrogen was effectively transformed in the estuary into particulate organic nitrogen (PON) and dissolved organic nitrogen (DON), of which DON was mostly exported from the estuary, enhancing productivity in nitrogen limited parts of the Gulf of Finland. DON concentrations ranged from 12.4 to 23.5 mu M and its estuarine dynamics were clearly uncoupled from DOC. In contrast to DOC, estuarine DON dynamics suggest that its production exceeds losses in the estuary. Total nitrogen (TN) and phosphorus (TP) loadings from the Neva River and St. Petersburg were estimated as 73.5 Gg N yr(-1) and 4.2 Gg P yr(-1), respectively. Approximately 59% of TN and 53% of TP loads were in organic forms. DOC and DON loadings were estimated as 741.4 Gg C yr(-1) and 19.0 Gg N yr(-1), respectively. Our estimate for DOC loading was evaluated against a previously published carbon budget of the Baltic Sea. According to the updated model, the Baltic Sea could be identified as a weak source of carbon into the atmosphere. (C) 2016 The Authors. Published by Elsevier B.V.Peer reviewe

Identification of dissolved organic matter size components in freshwater and marine environments

Dissolved organic matter (DOM) in the transition zone from freshwater to marine systems was analyzed with a new approach for parameterizing the size distribution of organic compounds. We used size-exclusion chromatography for molecular size analysis and quantified colored DOM (CDOM) on samples from two coastal environments in the Baltic Sea (Roskilde Fjord, Denmark and Gulf of Gdansk, Poland). We applied a Gaussian decomposition method to identify peaks from the chromatograms, providing information beyond bulk size properties. This approach complements methods where DOM is separated into size classes with pre-defined filtering cutoffs, or methods where chromatograms are used only to infer average molecular weight. With this decomposition method, we extracted between three and five peaks from each chromatogram and clustered these into three size groups. To test the applicability of our method, we linked our decomposed peaks with salinity, a major environmental driver in the freshwater-marine continuum. Our results show that when moving from freshwater to low-salinity coastal waters, the observed steep decrease of apparent molecular weight is mostly due to loss of the high-molecular-weight fraction (HMW; >2 kDa) of CDOM. Furthermore, most of the CDOM absorbance in freshwater originates from HMW DOM, whereas the absorbing moieties are more equally distributed along the smaller size range (<2 kDa) in marine samples.Peer reviewe

Photochemical Mineralization of Terrigenous DOC to Dissolved Inorganic Carbon in Ocean

When terrigenous dissolved organic carbon (tDOC) rich in chromophoric dissolved organic matter (tCDOM) enters the ocean, solar radiation mineralizes it partially into dissolved inorganic carbon (DIC). This study addresses the amount and the rates of DIC photoproduction from tDOC and the area of ocean required to photomineralize tDOC. We collected water samples from 10 major rivers, mixed them with artificial seawater, and irradiated them with simulated solar radiation to measure DIC photoproduction and the photobleaching of tCDOM. The linear relationship between DIC photoproduction and tCDOM photobleaching was used to estimate the amount of photoproduced DIC from the tCDOM fluxes of the study rivers. Solar radiation was estimated to mineralize 12.5 +/- 3.7 Tg C yr(-1) (10 rivers)(-1) or 18 +/- 8% of tDOC flux. The irradiation experiments also approximated typical apparent spectral quantum yields for DIC photoproduction (phi(lambda)) over the entire lifetime of the tCDOM. Based on phi(lambda)s and the local solar irradiances in river plumes, the annual areal DIC photoproduction rates from tDOC were calculated to range from 52 +/- 4 (Lena River) to 157 +/- 2 mmol C m(-2) yr(-1) (Mississippi River). When the amount of photoproduced DIC was divided by the areal rate, 9.6 +/- 2.5 x 10(6) km(2) of ocean was required for the photomineralization of tDOC from the study rivers. Extrapolation to the global tDOC flux yields 45 (31-58) Tg of photoproduced DIC per year in the river plumes that cover 34 (25-43) x 10(6) km(2) of the ocean.Peer reviewe

Learning environment, attitudes and anxiety across the transition from primary to secondary school mathematics

Past research has revealed that, relative to primary-school students, high-school students have less-positive attitudes to mathematics and perceive their classroom environments and teacher–student relationships less favourably. This study involved the transition experience of 541 students in 47 classes in 15 primary (year 7) and secondary (year 8) government and Catholic schools in metropolitan and regional South Australia. Scales were adapted from three established instruments, namely, the What Is Happening In this Class?, Test of Mathematics Related Attitudes and Revised Mathematics Anxiety Ratings Scale, to identify changes across the transition from primary to secondary school in terms of the classroom learning environment and students’ attitude/anxiety towards mathematics. Relative to year 7 students, year 8 students reported less Involvement, less positive Attitude to Mathematical Inquiry, less Enjoyment of Mathematics and greater Mathematics Anxiety. Differences between students in Years 7 and 8 were very similar for male and female students, although the magnitude of sex differences in attitudes was slightly different in Years 7 and 8