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Increasing problems with antibiotic resistance has directed interest towards phages as tools to treat bacterial infections in the aquaculture industry. However, phage resistance evolves rapidly in bacteria posing a challenge for successful phage therapy. To investigate phage resistance in the fish pathogenic bacterium Flavobacterium columnare, two phage-sensitive, virulent wild-type isolates, FCO-F2 and FCO-F9, were exposed to phages and subsequently analyzed for bacterial viability and colony morphology. Twenty-four phage-exposed isolates were further characterized for phage resistance, antibiotic susceptibility, motility, adhesion and biofilm formation on polystyrene surface, protease activity, whole genome sequencing and virulence against rainbow trout fry. Bacterial viability first decreased in the exposure cultures, subsequently increasing after 1-2 days. Simultaneously, the colony morphology of the phage-exposed isolates changed from original rhizoid to rough. The rough isolates arising in phage exposure were phage-resistant with low virulence, whereas rhizoid isolates maintained phage sensitivity, though reduced, and high virulence. Gliding motility and protease activity were also related to the phage sensitivity. Observed genetic mutations in phage-resistant isolates were mostly located in genes coding for type IX secretion system, a component of the flavobacterial gliding motility machinery. However, there were mutational differences between individual isolates, and not all phage-resistant isolates had genetic mutations. This indicates that development of phage resistance in F. columnare probably is a multifactorial process including both genetic mutations and changes in gene expression. Phage resistance may not, however, be a challenge for development of phage therapy against F. columnare infections, since phage resistance is associated with decrease in bacterial virulence.ImportancePhage resistance of infectious bacteria is a common phenomenon posing challenges for development of phage therapy. Along with growing World population and need for increased food production, constantly intensifying animal farming has to face increasing problems of infectious diseases. Columnaris disease, caused by F. columnare, is a worldwide threat for salmonid fry and juvenile farming. Without antibiotic treatments, infections can lead to 100% mortality in a fish stock. Phage therapy of columnaris disease would reduce a development of antibiotic-resistant bacteria and antibiotic loads by the aquaculture industry, but phage-resistant bacterial isolates may become a risk. However, phenotypic and genetic characterization of phage-resistant F. columnare isolates in this study revealed that they are less virulent than phage-sensitive isolates and thus not a challenge for phage therapy against columnaris disease. This is a valuable information for the fish farming industry globally when considering phage-based prevention and curing methods for F. columnare infections.

In this PhD research, restrictive and disciplinary practices refer to procedures which differ from the usual regulative practices of everyday life in a children’s residential care institution in child welfare. These practices restrict child’s possibilities to act and participate or they violate the privacy of a child. In this study restrictive and disciplinary practices are interpreted as ways to govern children and young people within child welfare. The two main research questions are as follows: 1) What kind of restrictive and disciplinary practices have there been in the children’s residential care institution at different times and how have these practices been organized in the social relations of specific time? 2) How has their regulation changed? Theoretically and methodologically, the study adheres to the approach developed by Dorothy E. Smith. This approach has been used to explore the organization of restrictive and disciplinary practices as well as the distinctive forms of relations of ruling mediating their implementation. Practices under examination are interpreted as the intersection of the everyday life of residential care and the social relations governing it. The most important social relation is the generational relation. Furthermore, Michel Foucault’s idea of a history of the present is applied heuristically. In this qualitative social work research, a single case study design is applied as a research strategy. One children’s residential care institution forms the case. Data collected from the case study institution and the archive contains textual documentary data such as the journals of punishments, the decisions of restraints, the memos of meetings, daily reports and the extracts of children’s case files, produced by residential care workers. Regulative relations are examined by using the municipal guidelines and legislation as well as professional texts such as committee reports. In the analysis, three key points of transformation are first located by exploring the continuities and discontinuities of practices under examination. Secondly, these moments are analysed by applying the method of textual inquiry and close reading, which takes into account definite places and times. Thirdly, the transformation of the practices under examination and their regulation are analysed during the time period of this research. Restrictive and disciplinary practices have changed during the period under examination from the practices of punishment and disciplinary power to practices emphasizing care and caring power. For example, corporal punishments were replaced by various methods of isolation, and drug testing was implemented as a new practice. The municipal guidelines formed the regulative basis for practices under examination until 1983. Since then they have been regulated by child welfare legislation. Both practices under examination as well as the generational relations of children and residential care workers have been re-shaped by the professionalization, democratization and legalization of children’s residential care. Residential care workers have been expected to integrate the events of everyday life and the formal regulative organization of practices. They have also been expected to adjust their professional activities accordingly. Key words: Children’s residential care, restriction, discipline, history of child welfare, relation of ruling, generational relation Väitöskirjatutkimuksessa rajoittamis- ja kurinpitokäytännöillä tarkoitetaan laitosarjen järjestyksen tyypillisestä ylläpitämisestä poikkeavia toimenpiteitä, joilla rajataan lastensuojelulaitokseen sijoitetun lapsen osallistumis- ja toimintamahdollisuuksia tai kajotaan lapsen yksityisyyteen. Tässä tutkimuksessa niitä tulkitaan lastensuojelun piirissä olevien lasten ja nuorten hallinnan tapoina. Tutkimuskysymyksinä ovat 1) millaisia rajoittamisen ja kurinpitämisen käytäntöjä lastensuojelulaitoksessa on eri aikoina ollut ja miten ne ovat organisoituneet oman aikansa yhteiskunnallisissa suhteissa sekä 2) miten niiden sääntely on muuttunut. Tutkimus kiinnittyy Dorothy E. Smithin teoreettismetodologiseen lähestymistapaan, jota käytetään rajoittamisen ja kurinpidon organisoitumisen sekä tutkittujen käytäntöjen toimeenpanoa ehdollistavien vallanomaisten hallinnan suhteiden tarkasteluun. Tutkittuja käytäntöjä tulkitaan lastensuojelulaitoksen toimijoiden arkipäiväisen elämän ja sitä määrittävien yhteiskunnallisten suhteiden kohtaamisena. Keskeisin yhteiskunnallinen suhde on sukupolvisuhde. Lisäksi sovelletaan heuristisesti Michel Foucault’n nykyisen historian ideaa. Sosiaalityön oppialaan kuuluvan laadullisen tutkimuksen tutkimusstrategiana käytetään yhden tapauksen tutkimusasetelmaa, jossa yksi lastensuojelulaitos muodostaa tutkittavan tapauksen. Tapaustutkimuslaitoksesta ja arkistosta kerättyyn ensisijaiseen aineistoon sisältyy laitostyöntekijöiden tuottamia tekstejä, kuten rangaistuspäiväkirjoja, rajoituspäätöksiä, kokousmuistioita, päivittäisraportteja ja asiakasasiakirjaotteita. Sääntelysuhteita tarkastellaan ohjesääntö- ja lainsäädäntöaineiston sekä asiantuntijatekstien, kuten komiteamietintöjen, avulla. Analyysissa paikannetaan ensin kolme keskeisintä murroskohtaa erittelemällä tutkittujen käytäntöjen jatkuvuuksia ja katkoksia. Toiseksi ne analysoidaan ajan ja paikan huomioivan tekstianalyysin ja kriittisen lähiluvun keinoin. Kolmanneksi käytäntöjen ja niiden sääntelyn muuntumista tarkastellaan koko tutkimusajanjaksolla. Lastensuojelulaitoksen rajoittamis- ja kurinpitokäytännöt ovat muuntuneet lähihistoriassa rangaistuspainotteisista kurinpitovallan käytännöistä huolenpitoa korostavan hoivaavan vallan toimenpiteisiin. Yksittäisistä käytännöistä esimerkiksi kurittaminen korvattiin eristämiskäytännöillä ja huumetestaus otettiin käyttöön uutena keinona. Kunnalliset ohjesäännöt toimivat tutkittujen käytäntöjen sääntelyperustana vuoteen 1983 asti, jonka jälkeen niistä on säädetty lastensuojelulaissa. Rajoittamis- ja kurinpitokäytännöt sekä lasten ja työntekijöiden sukupolvisuhteet ovat muokkaantuneet uudelleen laitostoiminnan yleisen ammatillistumisen, demokratisoitumisen ja oikeudellistumisen myötä. Lastensuojelulaitoksen työntekijöiltä on edellytetty taitoa sovittaa yhteen arkiset tilanteet ja käytäntöjä organisoiva muodollinen sääntely sekä taitoa sopeuttaa ammatillista toimintaa uudelleen muuttuneissa tilanteissa. Avainsanat: Lastensuojelun laitoshoito, rajoittaminen, kurinpito, lastensuojelun historia, hallinnan suhde, sukupolvisuhde

Objectives The aim of this study was to analyze the behavior of different bioactive glass fillers (BAGs) embedded in a polydimethylsiloxane matrix of an endodontic sealer. Methods Three different endodontic sealers were fabricated using S53P4, 45S5 and 18−06 glass fillers. Endodontic sealer Guttaflow Bioseal consisting of polydimethylsiloxane (PDMS) matrix was used as base of the experimental sealers. Behaviors of different glass fillers leaching from polymer matrix was studied in vitro for 14 days by measuring static ion dissolution profiles of Si, Na, Ca and P -ions. In addition, pH of the simulated bodyfluid (SBF) was monitored during the 14 days and all the sealer samples was examined with SEM/EDX analysis on the surface. Identical but non-glass filler containing polydimethylsiloxane-based sealer was used as a control material. Results By the time point of 24 h sealer with 45S5 had released twice as much of Si-ions compared to sealer with S53P4. No statistical differences of Na, Ca and P -ions dissolution were observed in the first 168 h for any groups whereas concentrations of Ca and P -ions decreased with 45S5 significantly after 336 h. Highest pH was measured for sealers with glass filler 45S5 and S53P4 (7.64–7.65). Visible mineral precipitation was observed only on sealer surfaces after 336 h’ time period with groups of 45S5 and S53P4. However, presence of calcium and phosphorus oxides was confirmed only with 45S5. Significance Bioactive glass type 45S5 outperforms S53P4 and 18−06 by acting more dynamically in vitro set-up

Funding: UK Natural Environment Research Council (Grant Number(s): NE/L501852/1, NE/P000592/1); Academy of Finland (GrantNumber(s): 267244, 268214, 322980), Ella ja Georg Ehrnroothin Säätiö. Detecting signatures of ecological adaptation in comparative genomics is challenging, but analysing population samples with characterised geographic distributions, such as clinal variation, can help identify genes showing covariation with important ecological variation. Here, we analysed patterns of geographic variation in the cold-adapted species Drosophila montana across phenotypes, genotypes and environmental conditions and tested for signatures of cold adaptation in population genomic divergence. We first derived the climatic variables associated with the geographic distribution of 24 populations across two continents to trace the scale of environmental variation experienced by the species, and measured variation in the cold tolerance of the flies of six populations from different geographic contexts. We then performed pooled whole genome sequencing of these six populations, and used Bayesian methods to identify SNPs where genetic differentiation is associated with both climatic variables and the population phenotypic measurements, while controlling for effects of demography and population structure. The top candidate SNPs were enriched on the X and fourth chromosomes, and they also lay near genes implicated in other studies of cold tolerance and population divergence in this species and its close relatives. We conclude that ecological adaptation has contributed to the divergence of D. montana populations throughout the genome and in particular on the X and fourth chromosomes, which also showed highest interpopulation FST. This study demonstrates that ecological selection can drive genomic divergence at different scales, from candidate genes to chromosome-wide effects. Publisher PDF

Identifying elements that enhance insect adaptation into changing environmental conditions is challenging, especially for traits affected by multiple factors. We studied latitudinal variation in the basal cold tolerance and body colour of two highly cold-tolerant Drosophila virilis group species, D. montana and D. flavomontana , in climatically diverse locations in North America. D. montana , which is generally found on higher latitudes and altitudes than D. flavomontana , was darker and more cold-tolerant than D. flavomontana . In D. montana only fly cold tolerance showed latitudinal variation, while in D. flavomontana both traits varied according to latitude and local climatic conditions, but showed no correlation with each other. We also examined the role of a circadian clock gene vrille and an insulin-signalling pathway gene Yolk protein 3 ( Yp3 ) in the basal cold tolerance and cold acclimation ability of D. montana females using RNA interference. Silencing of vrille induced expression changes in the period , but not clock , and decreased flies’ basal cold tolerance and cold acclimation ability, while silencing of Yp3 affected only cold acclimation. Our study demonstrates that the dependence of insect cold tolerance on latitudinally varying factors and local climatic conditions may vary even between closely-related species. Furthermore, we propose that a functional circadian clock system plays an essential role both in insect basal cold tolerance and cold acclimation ability, and that Yp3 affects cold acclimation likely through its interactions with other genes in the insulin-signalling pathway.

CRISPR-Cas immune systems adapt to new threats by acquiring spacers from invading nucleic acids such as phage genomes. However, some CRISPR-Cas loci lack genes necessary for spacer acquisition, despite apparent variation in spacer content between strains. It has been suggested that such loci may use acquisition machinery from co-occurring CRISPR-Cas systems. Here, using a lytic dsDNA phage, we observe spacer acquisition in the native host Flavobacterium columnare that carries an acquisition-deficient subtype VI-B locus and a complete subtype II-C locus. We characterize acquisition events in both loci and show that the RNA-targeting VI-B locus acquires spacers in trans using acquisition machinery from the DNA-targeting II-C locus. Our observations reinforce the concept of modularity in CRISPR-Cas systems and raise further questions regarding plasticity of adaptation modules.

Additional file 1. Additional protocols tables. Tables S1-S4 show detailed data related to the methods of the study.

Reproductive diapause is a primary mechanism used by arthropods to synchronize their life cycle with seasonal changes in temperate regions. Our study species, Drosophila montana, represents the northern insect species where flies enter reproductive diapause under short day conditions and where the precise timing of diapause is crucial for both survival and offspring production. We have studied clinal variation in the critical day length for female diapause induction (CDL) and their overall susceptibility to enter diapause (diapause incidence), as well as the temperature sensitivity of these traits. The study was performed using multiple strains from four latitudinal clines of the species – short clines in Finland and Alaska and long clines in the Rocky Mountains and the western coast of North America – and from one population in Kamchatka, Russia. CDL showed strong latitudinal clines on both continents, decreasing by one hour per five degrees decline in latitude, on average. CDL also decreased in all populations along with an increase in fly rearing temperature postponing the diapause to later calendar time, the effects of temperature being stronger in southern than in northern population. Female diapause incidence was close to 100% under short day/low temperature conditions in all populations, but decreased below 50% even under short days in 19°C in the southern North American western coast populations and in 22°C in most populations. Comparing a diversity of climatic data for the studied populations showed that while CDL is under a tight photoperiodic regulation linked with latitude, its length depends also on climatic factors determining the growing season length. Overall, the study deepens our understanding of how spatial and environmental parameters affect the seasonal timing of an important biological event, reproductive diapause and helps to estimate the evolutionary potential of insect populations to survive in changing climatic conditions. peerReviewed

Additional file 2. Additional results tables. Tables S5-S7 show additional data related to the results shown in the main figures.

Additional file 3 Additional results figures. Figures S1-S12 show additional data related to the results shown in the main figures.

Environmental cues, mainly photoperiod and temperature, are known to control female adult reproductive diapause in several insect species. Diapause enhances female survival during adverse conditions and postpones progeny production to the favorable season. Male diapause (a reversible inability to inseminate receptive females) has been studied much less than female diapause. However, if the males maximized their chances to fertilize females while minimizing their energy expenditure, they would be expected to be in diapause at the same time as females. We investigated Drosophila montana male mating behavior under short‐day conditions that induce diapause in females and found the males to be reproductively inactive. We also found that males reared under long‐day conditions (reproducing individuals) court reproducing postdiapause females, but not diapausing ones. The diapausing flies of both sexes had more long‐chain and less short‐chain hydrocarbons on their cuticle than the reproducing ones, which presumably increase their survival under stressful conditions, but at the same time decrease their attractiveness. Our study shows that the mating behavior of females and males is well coordinated during and after overwintering and it also gives support to the dual role of insect cuticular hydrocarbons in adaptation and mate choice. peerReviewed

In recent decades, the phytoplankton community in parts of the Baltic Sea has shifted from diatom dominance to co-occurrence of diatoms and dinoflagellates during the spring bloom. We investigated whether this shift affects bacterial production (BP), abundance and community composition (BCC). Two mesocosm experiments were carried out with water from the SW coast of Finland during the winters of 2012 and 2013. The water was collected before the onset of the spring bloom. Natural seawater was used as a control, and various inocula of diatom and dino flagellate cultures were used as treatments. After the phytoplankton bloom development, BP (thymidine: BPT; leucine: BPL) was significantly higher in the diatom treatments than in the controls and dinoflagellate treatments (BPT and BPL in 2012 and BPL in 2013). In 2013, the BCC was significantly different between the diatom and dinoflagellate treatments and there was a temporal shift in both experiments. Alphaproteobacteria predominated in all treatments at the beginning of the experiments and shifted to flavobacterial (2012) and betaproteobacterial predominance (2013) during the chlorophyll a peak. Towards the end of the experiment, Actinobacteria and Betaproteo bacteria predominated in the diatom treatment in 2012, whereas in 2013 Flavobacteriia (all treatments) predominated together with Gammaproteobacteria and Cytophagia (diatom treatments). The results demonstrated that bacterial physiology and community structure are affected by relatively small changes in the phytoplankton community. Thus, the ongoing changes in the phytoplankton community resulting from co-occurrence of diatoms and dinoflagellates may decrease pelagic remineralization of carbon and reduce organic matter fluxes through the microbial loop.