Exploring nucleo-cytoplasmic large DNA viruses in Tara Oceans microbial metagenomes

Nucleo-cytoplasmic large DNA viruses (NCLDVs) constitute a group of eukaryotic viruses that can have crucial ecological roles in the sea by accelerating the turnover of their unicellular hosts or by causing diseases in animals. To better characterize the diversity, abundance and biogeography of marine NCLDVs, we analyzed 17 metagenomes derived from microbial samples (0.2–1.6 μm size range) collected during the Tara Oceans Expedition. The sample set includes ecosystems under-represented in previous studies, such as the Arabian Sea oxygen minimum zone (OMZ) and Indian Ocean lagoons. By combining computationally derived relative abundance and direct prokaryote cell counts, the abundance of NCLDVs was found to be in the order of 104–105 genomes ml−1 for the samples from the photic zone and 102–103 genomes ml−1 for the OMZ. The Megaviridae and Phycodnaviridae dominated the NCLDV populations in the metagenomes, although most of the reads classified in these families showed large divergence from known viral genomes. Our taxon co-occurrence analysis revealed a potential association between viruses of the Megaviridae family and eukaryotes related to oomycetes. In support of this predicted association, we identified six cases of lateral gene transfer between Megaviridae and oomycetes. Our results suggest that marine NCLDVs probably outnumber eukaryotic organisms in the photic layer (per given water mass) and that metagenomic sequence analyses promise to shed new light on the biodiversity of marine viruses and their interactions with potential hosts

Ohustatud Euroopa naaritsa (Mustela lutreola) sigimine ja käitumine tehiskeskkonnas

Väitekirja elektrooniline versioon ei sisalda publikatsiooneEuroopa naarits on kriitiliselt ohustatud imetaja, mis on loodusest kadumas. Päästmaks liiki väljasuremisest hakati naaritsaid pidama loomaaedades. Vangistuses on loomade paljundamine keeruline, kuna sealne keskkond erineb looma looduslikust elupaigast. Loomaaias elavatel naaritsatel on täheldatud probleeme sigimisega. Paljud ei saa järglasi, mis võib saada saatuslikuks asurkonna jätkusuutlikkusele. Käesolevas doktoritöös uurisin, miks on naaritsa sigimine loomaaias ebaedukas ja kuidas leida probleemile lahendus. Olukorrast parema ülevaate saamiseks uurisime, mis mõjutab pesakonna suurust ja poegade tõenäosust ellu jääda. Leidsime, et olulised on ema vanus ja kaal, kuid muidu oli emaste sigimisedu sarnane. Lisaks uurisime emase hormonaaltsüklit indlemise ja tiinuse ajal. Tulemused olid ootuspärased, hormonaaltsükkel järgis liigile tüüpilist rada. Me ei leidnud tõendeid, et paaritamiskatsete ebaõnnestumise põhjust tuleks otsida emastest. Leidsime, et ebaedu sigimisel on seotud vangistuses sündinud isastega. Mõni isane on paaritamiskatses emase vastu agressiivne või siis neil puudub indleva emase vastu huvi. Mõistmaks isaste käitumist vaatlesime naaritsate lapsepõlve – uurisime poegade vahelisi suhteid pesakonnas kasvamise ajal. Paljudel imetajatel on tõendatud, et varajane keskkond mõjutab isendi käitumist täiskasvanueas. Me ei leidnud, et naaritsatel oleks pesakonnaperioodil ebanormaalset käitumist. Nagu igati kohane, kulus poegade põhiaeg mängule. Agressiivsust ei olnud palju ja see ei tõusnud ajas. Pesakonnad ei erinenud käitumismustrilt üksteisest, seega ei leidnud me hälbiva poegade käitumisega pesakondi, mis oleksid võinud ebasobiva sigimiskäitumise tekkimist seletada. Töötasime välja metoodika naartisate iseloomutüüpide määramiseks, sest üha enam soovitatakse, et loomadesse tuleks loomaaias suhtuda neist igaühe iseloomu arvestades. Leidsime, et naaritsaid saab eristada julguse, uurivuse ja sotsiaalsuse alusel . Meie töös ilmnes, et probleemid sigimishooajal on seotud eelkõige vangistuses sündinud isastega, nende põhjuseid peaks otsima mujalt kui pesakonnaperioodist, abi võiks olla iga looma iseloomu tundmisest. Saadud tulemused on oluliseks alusteadmisteks töös, mis on suunatud naaritsate tehiskeskkonnas pidamise edukuse tõstmisele.European mink is a critically endangered carnivore which has almost disappeared from nature. To save the species from extinction a captive population was established. Unfortunately, keeping wild animals in captivity always causes problems, just because in a zoo everything is different from the species’ natural habitat. In the case of the European mink, there are problems with breeding in captive conditions. Some animals will fail to produce offspring which may threaten the persistence of the captive population. In this doctorial thesis, I addressed the reasons of breeding failures and how to find solutions to this problem. At first we studied what affects the size and survival of the litter. We found that the weight and age of the mother are most significant. Additionally we studied the hormonal cycle of the females during the mating season and gestation. Results were expectable: the hormonal cycle followed a profile typical of the species. Analysing the diaries kept at Tallinn zoo for 20 years, we did not find that something is wrong on the females’ side. Instead, we found that the breeding problems are caused by males which have been born in captivity. Some males are aggressive toward the female during the mating attempt or remain passive. To understand the reasons for kind of behaviour, we focused on the childhood of the mink: we studied the interactions between the cubs during the litter period. It has often been found that, in mammals, the early environment affects the development of the behaviour of an animal. In our study, however, we did not find abnormalities in cubs’ behaviour during the litter period. As it should be, the most frequent type of behaviour was play. Aggression was low, it did not rise in time and did not differ between the litters. We developed tests to identify personality types in European mink. We found that it is possible to distinguish individual mink in boldness, sociability and exploration. In summary, we found that the main reason why breeding fails in the European mink is in the behaviour of captive born males. The causes of behavioural distortion may not be related to the litter period, an analysis of personality types may help here. The knowledge obtained forms important basis information for the work aimed at improving keeping condition of captive European mink

Cytochrome P450 monooxygenase CYP53 family in fungi : Structural analysis of CYP53A and its redox partner from the thermophilic fungus Thielavia terrestris

ThesisCytochrome P450 monooxygenases (CYPs or P450s) are heme-thiolate proteins distributed across biological kingdoms. P450s show multiple and diverse catalytic activity on a wide range of substrates, and as such the use of these enzymes have been prompted in various areas such as the production of human valuable chemicals, pharmaceutical compounds, antibiotics, fragrances and the degradation of xenobiotic compounds. If P450s are to be used on a large-scale production, it is imperative that they are able to resist extreme industrial conditions, including thermostability. Considering the fact that P450s are weak and less stable enzymes, research has focused on identifying thermostable P450s. Furthermore, P450s have been used as a drug target against pathogens. However, a study revealed that pathogens are developing drug resistance against currently available drugs. To tackle and address this emerging dilemma to drug resistance, novel drug targets need to be discovered and identified. The current study is the first of its kind that focuses on two aspects, which are identifying a common alternative anti-fungal drug target and structural characterization of thermostable P450 CYP53A and its redox partner cytochrome P450 reductase (CPR) from biomass-degrading thermophilic ascomycete Thielavia terrestris. This study also marks the beginning of our understanding on thermostable P450s from eukaryotes. Part of the study has been published as an article in PLoS ONE journal (impact factor 3.5). Below are the details of the manuscript

Kohalike ja invasiivsete röövtoiduliste suurselgrootute elupaigakasutus ja toitumissuhted Läänemere põhjaosas

Väitekirja elektrooniline versioon ei sisalda publikatsiooneRöövtoidulised suurselgrootud, peamiselt vähilaadsed, on rannikumere toitumisvõrgustikes oluliseks lüliks põhjaelustiku ja kalade vahel ning ühtlasi nad reguleerivad väiksemate selgrootute arvukust. Läänemere põhjaosas elas selle rühma esindajaid seni ainult kaks liiki – läänemere krevett (Palaemon adspersus) ja põhjamere garneel (Crangon crangon). Hiljuti lisandusid neile kohalikele liikidele kaks võõrliiki – elegantne krevett (Palaemon elegans) ja rändkrabi (Rhithropanopeus harrisii). Kohalike ja võõrliikidest röövtoiduliste suurselgrootute leviku, elupaigaeelistuste ja toitumise uurimiseks kasutati olemasolevaid põhjaelustiku leviku andmeid, spetsiaalselt uuritavatele liikidele suunatud proovide kogumist loodusest ja laborikatseid, kus rakendati muuhulgas loomade raadiomärgistamist. Võõrliigist kreveti levik oli ulatuslikum kui kohalike krevettide levik. Võõrliik oli kohalike krevettidega võrreldes enam seotud elupaikadega, millele on iseloomulikud etrofeerumise tunnused (kõrge toitainete kontsentratsioon, lühiealiste niitjate vetikate suur hulk). Krevettide toitumisuuringud näitasid, et uuritud Palaemon liikide toitumisintensiivsus ja toidu kooseis ei erinenud. Seega on kohaliku ja võõrkreveti roll rannikumere toiduvõrkudes sarnane, ent võõrliik võib troofilisi suhteid ümber kujundada piirkondades, kus kohalikud krevetid puuduvad. Kõige kitsama elupaigakasutusega oli põhjamere garneel ja Palaemon liikidest oli võõrliigi P. elegans spetsialiseerumise tase mõnevõrra kõrgem kui kohalikul liigil. R. harrisii eelistas põisadruga (Fucus vesiculosus) elupaika mis viitab sellele, et põisadruga elupaik pakub krabile aastaringset stabiilset elupaika ja et mitmekesine põhjakooslus võib saada seetõttu krabidest oluliselt mõjutatud. Doktoritöö tulemused näitasid, et elupaikade iseloom ja seisund mõjutab kohalike ja võõrliikide levikumustreid ning võõrliikidest selgrootute kiskjate saabumine ja kiire levila laienemine Läänemere põhjaosas toovad kaasa täiesti uue ökoloogilise funktsiooni (suuremõõtmeline kiskja) või juba varem regionaalselt esinenud funktsiooni leviku piirkondadesse, kus see varem puudus.Decapod crustaceans, such as crabs and shrimps play an important role in coastal ecosystems as they prey on small benthic invertebrates and at the same time they are an important food item for fishes. Until recently, this group of macroinvertebrates consisted of only a few species in the northern Baltic Sea, including only two species of native shrimps – Crangon crangon and Palaemon adspersus. However, very recently two non-native crustacean predators arrived – the shrimp Palaemon elegans and the crab Rhithropanopeus harrisii. Data from available databases were used together with field sampling and laboratory experiments (incl. radio frequency positioning technology) to describe the geographical distribution, habitat selection, and feeding of native and invasive predatory crustaceans in the coastal areas of the northern Baltic Sea. The distribution of the non-native P. elegans was wider than that of native species. P. elegans was associated with lower salinity, higher concentrations of nutrients and higher proportions of ephemeral filamentous algae, relative to P. adspersus. According to results of the experiment on feeding activity, P. elegans performs a similar role in the coastal food web as the native congener, although it may rearrange trophic interactions and introduce new ecological function in the most eutrophicated areas previously lacking any native shrimp. Among the studied shrimps, C. crangon had the highest habitat specialization. The non-native P. elegans showed slightly higher habitat specialization compared to the native P. adspersus. The invasive crab R. harrisii preferred habitats with bladder wrack where the diverse native community may become heavily impacted by this novel large-bodied predator. This thesis showed that the successful establishment of non-native predatory invertebrates can introduce new ecological functions (large-bodied predators) or considerably strengthen already existed ones, while the heterogeneity of habitats has varying effects on the distribution patterns of native and invasive species

The Origin of GPCRs: Identification of Mammalian like Rhodopsin, Adhesion, Glutamate and Frizzled GPCRs in Fungi

G protein-coupled receptors (GPCRs) in humans are classified into the five main families named Glutamate, Rhodopsin, Adhesion, Frizzled and Secretin according to the GRAFS classification. Previous results show that these mammalian GRAFS families are well represented in the Metazoan lineages, but they have not been shown to be present in Fungi. Here, we systematically mined 79 fungal genomes and provide the first evidence that four of the five main mammalian families of GPCRs, namely Rhodopsin, Adhesion, Glutamate and Frizzled, are present in Fungi and found 142 novel sequences between them. Significantly, we provide strong evidence that the Rhodopsin family emerged from the cAMP receptor family in an event close to the split of Opisthokonts and not in Placozoa, as earlier assumed. The Rhodopsin family then expanded greatly in Metazoans while the cAMP receptor family is found in 3 invertebrate species and lost in the vertebrates. We estimate that the Adhesion and Frizzled families evolved before the split of Unikonts from a common ancestor of all major eukaryotic lineages. Also, the study highlights that the fungal Adhesion receptors do not have N-terminal domains whereas the fungal Glutamate receptors have a broad repertoire of mammalian-like N-terminal domains. Further, mining of the close unicellular relatives of the Metazoan lineage, Salpingoeca rosetta and Capsaspora owczarzaki, obtained a rich group of both the Adhesion and Glutamate families, which in particular provided insight to the early emergence of the N-terminal domains of the Adhesion family. We identified 619 Fungi specific GPCRs across 79 genomes and revealed that Blastocladiomycota and Chytridiomycota phylum have Metazoan-like GPCRs rather than the GPCRs specific for Fungi. Overall, this study provides the first evidence of the presence of four of the five main GRAFS families in Fungi and clarifies the early evolutionary history of the GPCR superfamily

Fungal regulatory evolution: cis and trans in the balance

available in PMC 2010 June 17.Regulatory divergence is likely a major driving force in evolution. Comparative genomics is being increasingly used to infer the evolution of gene regulation. Ascomycota fungi are uniquely suited among eukaryotes for regulatory evolution studies, due to broad phylogenetic scope, many sequenced genomes, and tractability of genomic analysis. Here we review recent advances in the identification of the contribution of cis- and trans-factors to expression divergence. Whereas current strategies have led to the discovery of surprising signatures and mechanisms, we still understand very little about the adaptive role of regulatory evolution. Empirical studies including experimental evolution, comparative functional genomics and hybrid and engineered strains are showing early promise toward deciphering the contribution of regulatory divergence to adaptation.Human Frontier Science Program (Strasbourg, France)Howard Hughes Medical InstituteBurroughs Wellcome Fund (Career Award)Alfred P. Sloan Foundatio

Small RNA pathway genes identified by patterns of phylogenetic conservation and divergence

Genetic and biochemical analyses of RNA interference (RNAi) and microRNA (miRNA) pathways have revealed proteins such as Argonaute/PIWI and Dicer that process and present small RNAs to their targets. Well validated small RNA pathway cofactors, such as the Argonaute/PIWI proteins show distinctive patterns of conservation or divergence in particular animal, plant, fungal, and protist species. We compared 86 divergent eukaryotic genome sequences to discern sets of proteins that show similar phylogenetic profiles with known small RNA cofactors. A large set of additional candidate small RNA cofactors have emerged from functional genomic screens for defects in miRNA- or siRNA-mediated repression in C. elegans and D. melanogaster1,2 and from proteomic analyses of proteins co-purifying with validated small RNA pathway proteins3,4. The phylogenetic profiles of many of these candidate small RNA pathway proteins are similar to those of known small RNA cofactor proteins. We used a Bayesian approach to integrate the phylogenetic profile analysis with predictions from diverse transcriptional coregulation and proteome interaction datasets to assign a probability for each protein for a role in a small RNA pathway. Testing high-confidence candidates from this analysis for defects in RNAi silencing, we found that about half of the predicted small RNA cofactors are required for RNAi silencing. Many of the newly identified small RNA pathway proteins are orthologues of proteins implicated in RNA splicing. In support of a deep connection between the mechanism of RNA splicing and small RNA-mediated gene silencing, the presence of the Argonaute proteins and other small RNA components in the many species analysed strongly correlates with the number of introns in that species