Pigmentidel põhinev kemotaksonoomia – tõhus vahend fütoplanktoni koosseisu selgitamiseks järvedes ja rannikumeres

A Thesis for applying for the degree of Doctor of Philosophy in Applied BiologyAll members of the aquatic food web rely either directly on indirectly on the autotrophic phytoplankton. The distribution and dynamics of phytoplankton groups reflects a number of things: rate of eutrophication, biogeochemical cycling and formation of the potentially harmful algal blooms. Therefore we are bound to quantify the community composition of phytoplankton. These measurements are complicated since the population growth of the phytoplankton can be explosive and the distribution is patchy. Traditionally phytoplankton is quantified via microscopy which is expensive, time-consuming, subjective and requires highly skilful specialists. This thesis aims to improve phytoplankton monitoring by using algal pigments. This method relies on a fact that all photosynthetic algae have chlorophyll a in their cells that helps to convert light energy into chemical energy. Additionally all algae have accessory pigments that help to capture the light from different wavelenghts. Since there are hundreds of different accessory pigments they can be used as markers for phytoplankton taxonomic groups. Therefore we can quantify different algal groups by measuring the amount specific marker pigments. These measurements are fast and easily automated. Biggest complication is that the amount of accessory pigments is not constant and depends on several environmental factors – e.g. irradiance, nutrients. My work aims to analyse the relationship of marker pigments and phytoplankton groups in various aquatic envrionments. Our studies in lake Võrtsjärv and coastal areas of Baltic Sea have demonstrated that phytoplankton pigments are a powerful tool in monitoring. Quantification of phytoplankton goups via pigments is faster, cheaper, more objective and precise than with traditional approach. Additional perk of pigment-based chemotaxonomy is that the smallest fraction of phytoplankton – picoplankton, which is left out of microscopy analysis, can be easily studied with this method.Kõikide vees elavate organismide toidulaud sõltub otseselt või kaudselt mikrovetikatest. Mikrovetikad on vees elavad kõige väiksemad fotosünteesivad organismid ja ligi pool hapnikust, mida hingame on toodetud just nende poolt. Siiski on mikrovetikatel ka pahupool – liigsoodsad tingimused põhjustavad vetikate massilise vohamise ehk õitsengu, millel on tihti negatiivsed tagajärjed, näiteks hapniku puudus vees, toksilisus ja kalade suremine. Seetõttu on vaja mikrovetikate hulka ja koosseisu mõõta. Probleem on selles, et mikrovetikate hulk ja koosseis muutub kiiresti. Traditsiooniliselt kasutatakse vetikarühmade hulga hindamiseks mikroskoopiat, mis on kulukas, aeganõudev, subjektiivne ja nõuab väga kõrgeid erialaoskusi süstemaatikas. Minu doktoritöö arendab mikrovetikate seiret neis sisalduvaid pigmente kasutades. Meetodi põhimõte seisneb selles, et kõikides vetikates on pigment nimega klorofüll a, mis päikeseenergia keemiliseks energiaks muudab. Veel on vetikates erinevad lisapigmendid, mis aitavad valgust püüda lainepikkustel, kus klorofüll on ebaefektiivne. Lisapigmente on vetikates sadu ja paljud neist on rühmaspetsiifilised. Seega võime me vetikate hulga hindamiseks mõõta just konkreetsele rühmale iseloomuliku pigmendi koguse. Selline mõõtmine on kiire ja automatiseeritav. Kitsaskohaks on asjaolu, et lisapigmentide hulk sõltub mitmetest faktoritest – näiteks valguse või toiteainete hulgast. Minu töö uurib pigmentide ja vetikarühmade vahelisi seosed erineva iseloomuga veekogudes. Meie uurimused nii Võrtsjärves kui ka rannikumeres (Läänemeri) on näidanud, et pigmendid võimaldavad hinnata kiiremini, odavamalt, objektiivsemalt ja täpsemalt seda, millised mikrovetikad veekogudes tegutsevad. Selliselt on võimalik juba varakult tuvastada ka ohtlikud õitsengud. Olulise lisaväärtusena võimaldab pigmentidel põhinev kemotaksonoomia uurida ka kõige väiksema suurusjärgu vetikaid ehk pikoplanktonit.Publication of this thesis is supported by the Estonian University of Life Sciences

Using Microcystin Gene Copies to Determine Potentially-Toxic Blooms, Example from a Shallow Eutrophic Lake Peipsi

Global warming, paired with eutrophication processes, is shifting phytoplankton communities towards the dominance of bloom-forming and potentially toxic cyanobacteria. The ecosystems of shallow lakes are especially vulnerable to these changes. Traditional monitoring via microscopy is not able to quantify the dynamics of toxin-producing cyanobacteria on a proper spatio-temporal scale. Molecular tools are highly sensitive and can be useful as an early warning tool for lake managers. We quantified the potential microcystin (MC) producers in Lake Peipsi using microscopy and quantitative polymerase chain reaction (qPCR) and analysed the relationship between the abundance of the mcyE genes, MC concentration, MC variants and toxin quota per mcyE gene. We also linked environmental factors to the cyanobacteria community composition. In Lake Peipsi, we found rather moderate MC concentrations, but microcystins and microcystin-producing cyanobacteria were widespread across the lake. Nitrate (NO3−) was a main driver behind the cyanobacterial community at the beginning of the growing season, while in late summer it was primarily associated with the soluble reactive phosphorus (SRP) concentration. A positive relationship was found between the MC quota per mcyE gene and water temperature. The most abundant variant—MC-RR—was associated with MC quota per mcyE gene, while other MC variants did not show any significant impact

Using Microcystin Gene Copies to Determine Potentially-Toxic Blooms, Example from a Shallow Eutrophic Lake Peipsi

Global warming, paired with eutrophication processes, is shifting phytoplankton communities towards the dominance of bloom-forming and potentially toxic cyanobacteria. The ecosystems of shallow lakes are especially vulnerable to these changes. Traditional monitoring via microscopy is not able to quantify the dynamics of toxin-producing cyanobacteria on a proper spatio-temporal scale. Molecular tools are highly sensitive and can be useful as an early warning tool for lake managers. We quantified the potential microcystin (MC) producers in Lake Peipsi using microscopy and quantitative polymerase chain reaction (qPCR) and analysed the relationship between the abundance of the mcyE genes, MC concentration, MC variants and toxin quota per mcyE gene. We also linked environmental factors to the cyanobacteria community composition. In Lake Peipsi, we found rather moderate MC concentrations, but microcystins and microcystin-producing cyanobacteria were widespread across the lake. Nitrate (NO3−) was a main driver behind the cyanobacterial community at the beginning of the growing season, while in late summer it was primarily associated with the soluble reactive phosphorus (SRP) concentration. A positive relationship was found between the MC quota per mcyE gene and water temperature. The most abundant variant—MC-RR—was associated with MC quota per mcyE gene, while other MC variants did not show any significant impact

A Future Tale of Two Winters? Sediment-water interface nitrogen dynamics in Lake Võrtsjärv (Estonia) during the ice-free winter 2019/2020 : [presentation]

The presentation took place at the Lahti Lakes 2021 Symposium.This project has received funding from the European Union’s Horizon 2020 research and innovation programme under grant agreement No 95196. Estonian University of Life Sciences ASTRA project “Value-chain based bio-economy”.This project has received funding from the European Union’s Horizon 2020 research and innovation programme under grant agreement No 95196. Estonian University of Life Sciences ASTRA project “Value-chain based bio-economy”

Multiple Myeloma Treatment in Real-world Clinical Practice : Results of a Prospective, Multinational, Noninterventional Study

Funding Information: The authors would like to thank all patients and their families and all the EMMOS investigators for their valuable contributions to the study. The authors would like to acknowledge Robert Olie for his significant contribution to the EMMOS study. Writing support during the development of our report was provided by Laura Mulcahy and Catherine Crookes of FireKite, an Ashfield company, a part of UDG Healthcare plc, which was funded by Millennium Pharmaceuticals, Inc, and Janssen Global Services, LLC. The EMMOS study was supported by research funding from Janssen Pharmaceutical NV and Millennium Pharmaceuticals, Inc. Funding Information: The authors would like to thank all patients and their families and all the EMMOS investigators for their valuable contributions to the study. The authors would like to acknowledge Robert Olie for his significant contribution to the EMMOS study. Writing support during the development of our report was provided by Laura Mulcahy and Catherine Crookes of FireKite, an Ashfield company, a part of UDG Healthcare plc, which was funded by Millennium Pharmaceuticals, Inc, and Janssen Global Services, LLC. The EMMOS study was supported by research funding from Janssen Pharmaceutical NV and Millennium Pharmaceuticals, Inc. Funding Information: M.M. has received personal fees from Janssen, Celgene, Amgen, Bristol-Myers Squibb, Sanofi, Novartis, and Takeda and grants from Janssen and Sanofi during the conduct of the study. E.T. has received grants from Janssen and personal fees from Janssen and Takeda during the conduct of the study, and grants from Amgen, Celgene/Genesis, personal fees from Amgen, Celgene/Genesis, Bristol-Myers Squibb, Novartis, and Glaxo-Smith Kline outside the submitted work. M.V.M. has received personal fees from Janssen, Celgene, Amgen, and Takeda outside the submitted work. M.C. reports honoraria from Janssen, outside the submitted work. M. B. reports grants from Janssen Cilag during the conduct of the study. M.D. has received honoraria for participation on advisory boards for Janssen, Celgene, Takeda, Amgen, and Novartis. H.S. has received honoraria from Janssen-Cilag, Celgene, Amgen, Bristol-Myers Squibb, Novartis, and Takeda outside the submitted work. V.P. reports personal fees from Janssen during the conduct of the study and grants, personal fees, and nonfinancial support from Amgen, grants and personal fees from Sanofi, and personal fees from Takeda outside the submitted work. W.W. has received personal fees and grants from Amgen, Celgene, Novartis, Roche, Takeda, Gilead, and Janssen and nonfinancial support from Roche outside the submitted work. J.S. reports grants and nonfinancial support from Janssen Pharmaceutical during the conduct of the study. V.L. reports funding from Janssen Global Services LLC during the conduct of the study and study support from Janssen-Cilag and Pharmion outside the submitted work. A.P. reports employment and shareholding of Janssen (Johnson & Johnson) during the conduct of the study. C.C. reports employment at Janssen-Cilag during the conduct of the study. C.F. reports employment at Janssen Research and Development during the conduct of the study. F.T.B. reports employment at Janssen-Cilag during the conduct of the study. The remaining authors have stated that they have no conflicts of interest. Publisher Copyright: © 2018 The AuthorsMultiple myeloma (MM) remains an incurable disease, with little information available on its management in real-world clinical practice. The results of the present prospective, noninterventional observational study revealed great diversity in the treatment regimens used to treat MM. Our results also provide data to inform health economic, pharmacoepidemiologic, and outcomes research, providing a framework for the design of protocols to improve the outcomes of patients with MM. Background: The present prospective, multinational, noninterventional study aimed to document and describe real-world treatment regimens and disease progression in multiple myeloma (MM) patients. Patients and Methods: Adult patients initiating any new MM therapy from October 2010 to October 2012 were eligible. A multistage patient/site recruitment model was applied to minimize the selection bias; enrollment was stratified by country, region, and practice type. The patient medical and disease features, treatment history, and remission status were recorded at baseline, and prospective data on treatment, efficacy, and safety were collected electronically every 3 months. Results: A total of 2358 patients were enrolled. Of these patients, 775 and 1583 did and did not undergo stem cell transplantation (SCT) at any time during treatment, respectively. Of the patients in the SCT and non-SCT groups, 49%, 21%, 14%, and 15% and 57%, 20%, 12% and 10% were enrolled at treatment line 1, 2, 3, and ≥ 4, respectively. In the SCT and non-SCT groups, 45% and 54% of the patients had received bortezomib-based therapy without thalidomide/lenalidomide, 12% and 18% had received thalidomide/lenalidomide-based therapy without bortezomib, and 30% and 4% had received bortezomib plus thalidomide/lenalidomide-based therapy as frontline treatment, respectively. The corresponding proportions of SCT and non-SCT patients in lines 2, 3, and ≥ 4 were 45% and 37%, 30% and 37%, and 12% and 3%, 33% and 27%, 35% and 32%, and 8% and 2%, and 27% and 27%, 27% and 23%, and 6% and 4%, respectively. In the SCT and non-SCT patients, the overall response rate was 86% to 97% and 64% to 85% in line 1, 74% to 78% and 59% to 68% in line 2, 55% to 83% and 48% to 60% in line 3, and 49% to 65% and 36% and 45% in line 4, respectively, for regimens that included bortezomib and/or thalidomide/lenalidomide. Conclusion: The results of our prospective study have revealed great diversity in the treatment regimens used to manage MM in real-life practice. This diversity was linked to factors such as novel agent accessibility and evolving treatment recommendations. Our results provide insight into associated clinical benefits.publishersversionPeer reviewe

Parallel assessment of marine autotrophic picoplankton using flow cytometry and chemotaxonomy

Autotrophic picoplankton (0.2–2 μm) can be a significant contributor to primary production and hence play an important role in carbon flow. The phytoplankton community structure in the Baltic Sea is very region specific and the understanding of the composition and dynamics of pico-size phytoplankton is generally poor. The main objective of this study was to determine the contribution of picoeukaryotic algae and their taxonomic com- position in late summer phytoplankton community of the West-Estonian Archipelago Sea. We found that about 20% of total chlorophyll a (Chl a) in this area belongs to autotrophic picoplankton. With increasing total Chl a, the Chl a of autotrophic picoplankton increased while its contribution in total Chl a decreased. Picoeukaryotes play an important role in the coastal area of the Baltic Sea where they constituted around 50% of the total autotrophic picoplankton biomass. The most abundant groups of picoeukaryotic algae were cryptophytes (16%), chlorophytes (13%) and diatoms (9%). Picocyanobacteria were clearly dominated by phycoerythrin containing Synechococcus. The parallel use of different assessment methods (CHEMTAX and flow cytometry) revealed the share of eukaryotic and prokaryotic part of autotrophic picoplankton.We gratefully thank Dr. Tiit Kutser (Estonian Marine Institute, Uni- versity of Tartu) for organizing the cruises and helping with the sample collection and Karolin Teeveer for providing microscopy counts. This work was supported by Estonian Ministry of Education and Research (IUT 21-02); Estonian Science Foundation (ETF9102, ETF8576); Swiss Grant for Programme “Enhancing public environmental monitoring ca- pacities” and Estonian Doctoral School of Earth Sciences and Ecology. We are thankful for the two reviewers who provided excellent advice to improve this manscript.We gratefully thank Dr. Tiit Kutser (Estonian Marine Institute, Uni- versity of Tartu) for organizing the cruises and helping with the sample collection and Karolin Teeveer for providing microscopy counts. This work was supported by Estonian Ministry of Education and Research (IUT 21-02); Estonian Science Foundation (ETF9102, ETF8576); Swiss Grant for Programme “Enhancing public environmental monitoring ca- pacities” and Estonian Doctoral School of Earth Sciences and Ecology. We are thankful for the two reviewers who provided excellent advice to improve this manscript

Pigment-based chemotaxonomy--a quick alternative to determine algal assemblages in large shallow eutrophic lake?

Pigment-based chemotaxonomy and CHEMTAX software have proven to be a valuable phytoplankton monitoring tool in marine environments, but are yet underdeveloped to determine algal assemblages in freshwater ecosystems. The main objectives of this study were (1) to compare the results of direct microscopy and CHEMTAX in describing phytoplankton community composition dynamics in a large, shallow and eutrophic lake; (2) to analyze the efficiency of the pigment-based method to detect changes in phytoplankton seasonal dynamics and during rapid bloom periods; (3) to assess the suitability of specific marker pigments and available marker pigment:chlorophyll a ratios to follow seasonal changes in eutrophic freshwater environment. A 5-year (2009-2013) parallel phytoplankton assessment by direct microscopy and by CHEMTAX was conducted using published marker pigment:chlorophyll a ratios. Despite displaying some differences from microscopy results, the pigment-based method successfully described the overall pattern of phytoplankton community dynamics during seasonal cycle in a eutrophic lake. Good agreement between the methods was achieved for most phytoplankton groups - cyanobacteria, chlorophytes, diatoms and cryptophytes. The agreement was poor in case of chrysophytes and dinoflagellates. Our study shows clearly that published marker pigment:chlorophyll a ratios can be used to describe algal class abundances, but they need to be calibrated for specific freshwater environment. Broader use of this method would enable to expand monitoring networks and increase measurement frequencies of freshwater ecosystems to meet the goals of the Water Framework Directive

Dissolved O₂ and N₂ gas dynamics in experimental lake mesocosms experiencing forced stratification : [presentation]

The presentation took place at the 11th International Shallow Lakes Conference.This project has received funding from the European Union’s Horizon 2020 research and innovation programme under grant agreement No 951963.This project has received funding from the European Union’s Horizon 2020 research and innovation programme under grant agreement No 951963

Lake Peipsi 2020 (Phytoplankton samples)

Method: Phytoplankton samples were preserved in Lugol’s (acidified iodine) solution and counted under an inverted microscope (Utermöhl, 1958). 3-10 ml of preserved sample was settled overnight and counted in random fields or transects. Biovolumes of algal cells, colonies and/or filaments were calculated using assigned geometric shapes dimensions, and converted to biomass assuming the specific density of 1 g cm-3 in accordance with Edler (1979). Approved by CEN on 14 July 2006 “Water quality - Guidance standard on the enumeration of phytoplankton using inverted microscopy (Utermöhl technique)” (CEN 15204, 2006) European Standard EN 15204:2006 Utermöhl, H., 1958. Zur Vervollkommnung der quantitativen Phytoplankton-Methodik. Mitteilungen der Internationale Vereinigung für Theoretische und Angewandte Limnologie 9, 1- 38. Edler, L. (ed.), 1979. Recommendations on methods for marine biological studies in the Baltic Sea. Phytoplankton and chlorophyll. Baltic Marine Biologists WG 9. Leg: K. Blank, K. Palmik-Das, M. Sepp, M. Tamm; det K. Piirso

Dynamics of phytoplankton communities in Lake Võrtsjärv (2009–2013) according to CHEMTAX (upper) and microscopy (lower).

<p>Dynamics of phytoplankton communities in Lake Võrtsjärv (2009–2013) according to CHEMTAX (upper) and microscopy (lower).</p