Taming the massive genome of Scots pine with PiSy50k, a new genotyping array for conifer research.

Pinus sylvestris (Scots pine) is the most widespread coniferous tree in the boreal forests of Eurasia, with major economic and ecological importance. However, its large and repetitive genome presents a challenge for conducting genome-wide analyses such as association studies, genetic mapping and genomic selection. We present a new 50K single-nucleotide polymorphism (SNP) genotyping array for Scots pine research, breeding and other applications. To select the SNP set, we first genotyped 480 Scots pine samples on a 407 540 SNP screening array and identified 47 712 high-quality SNPs for the final array (called 'PiSy50k'). Here, we provide details of the design and testing, as well as allele frequency estimates from the discovery panel, functional annotation, tissue-specific expression patterns and expression level information for the SNPs or corresponding genes, when available. We validated the performance of the PiSy50k array using samples from Finland and Scotland. Overall, 39 678 (83.2%) SNPs showed low error rates (mean = 0.9%). Relatedness estimates based on array genotypes were consistent with the expected pedigrees, and the level of Mendelian error was negligible. In addition, array genotypes successfully discriminate between Scots pine populations of Finnish and Scottish origins. The PiSy50k SNP array will be a valuable tool for a wide variety of future genetic studies and forestry applications.

Promotive and risk factors for children's mental health-Finnish municipal policymakers' and leading officeholders' views.

Findings on children's mental health promotion at the policy level are scarce, and the perceptions of the municipal administration on factors affecting children's mental health have not been reported. This study describes the perspectives of policymakers and leading officeholders on promotive and risk factors for children's mental health in a socioecological context. The perspectives of Finnish policymakers (n = 15) and officeholders (n = 10) in municipalities were examined using semi-structured interviews. The data were analyzed using inductive content analysis and were categorized according to the five levels of a socioecological model of health promotion: public policy, community, organizational, interpersonal and individual levels. The public policy level emerged strongly in the findings, specifically strategic planning and implementation challenges related to the promotion of children's mental health in the municipality and state administration. At the community level, environmental factors promoting children's mental health as well as risk factors were described. The organizational level consisted of support, requirements and development needs in children's services. The importance of family and close networks at the interpersonal level, as well as the individual basis of mental health, were also evident. The integration and better collaboration of child and family services, the use of child rights impact assessment in political decision-making, and financial support from the state could contribute to improving strategic planning to support children's mental health at the municipal level.

Maintenance of Adaptive Dynamics and No Detectable Load in a Range-Edge Outcrossing Plant Population.

During range expansion, edge populations are expected to face increased genetic drift, which in turn can alter and potentially compromise adaptive dynamics, preventing the removal of deleterious mutations and slowing down adaptation. Here, we contrast populations of the European subspecies Arabidopsis lyrata ssp. petraea, which expanded its Northern range after the last glaciation. We document a sharp decline in effective population size in the range-edge population and observe that nonsynonymous variants segregate at higher frequencies. We detect a 4.9% excess of derived nonsynonymous variants per individual in the range-edge population, suggesting an increase of the genomic burden of deleterious mutations. Inference of the fitness effects of mutations and modeling of allele frequencies under the explicit demographic history of each population predicts a depletion of rare deleterious variants in the range-edge population, but an enrichment for fixed ones, consistent with the bottleneck effect. However, the demographic history of the range-edge population predicts a small net decrease in per-individual fitness. Consistent with this prediction, the range-edge population is not impaired in its growth and survival measured in a common garden experiment. We further observe that the allelic diversity at the self-incompatibility locus, which ensures strict outcrossing and evolves under negative frequency-dependent selection, has remained unchanged. Genomic footprints indicative of selective sweeps are broader in the Northern population but not less frequent. We conclude that the outcrossing species A. lyrata ssp. petraea shows a strong resilience to the effect of range expansion.

Adaptive responses to temperature and precipitation variation at the early-life stages of Pinus sylvestris.

Early-stage fitness variation has been seldom evaluated at broad scales in forest tree species, despite the long tradition of studying climate-driven intraspecific genetic variation. In this study, we evaluated the role of climate in driving patterns of population differentiation at early-life stages in Pinus sylvestris and explored the fitness and growth consequences of seed transfer within the species range. We monitored seedling emergence, survival and growth over a 2-yr period in a multi-site common garden experiment which included 18 European populations and spanned 25° in latitude and 1700 m in elevation. Climate-fitness functions showed that populations exhibited higher seedling survival and growth at temperatures similar to their home environment, which is consistent with local adaptation. Northern populations experienced lower survival and growth at warmer sites, contrary to previous studies on later life stages. Seed mass was higher in populations from warmer areas and was positively associated with survival and growth at more southern sites. Finally, we did not detect a survival-growth trade-off; on the contrary, bigger seedlings exhibited higher survival probabilities under most climatic conditions. In conclusion, our results reveal that contrasting temperature regimes have played an important role in driving the divergent evolution of P. sylvestris populations at early-life stages.

Selection patterns on early-life phenotypic traits in Pinus sylvestris are associated with precipitation and temperature along a climatic gradient in Europe.

Understanding the dynamics of selection is key to predicting the response of tree species to new environmental conditions in the current context of climate change. However, selection patterns acting on early recruitment stages and their climatic drivers remain largely unknown in most tree species, despite being a critical period of their life cycle. We measured phenotypic selection on Pinus sylvestris seed mass, emergence time and early growth rate over 2 yr in four common garden experiments established along the latitudinal gradient of the species in Europe. Significant phenotypic plasticity and among-population genetic variation were found for all measured phenotypic traits. Heat and drought negatively affected fitness in the southern sites, but heavy rainfalls also decreased early survival in middle latitudes. Climate-driven directional selection was found for higher seed mass and earlier emergence time, while the form of selection on seedling growth rates differed among sites and populations. Evidence of adaptive and maladaptive phenotypic plasticity was found for emergence time and early growth rate, respectively. Seed mass, emergence time and early growth rate have an adaptive role in the early stages of P. sylvestris and climate strongly influences the patterns of selection on these fitness-related traits.

Municipal strategies and meeting minutes' descriptions of the promotion of children's mental health: a document analysis.

AIMS: Little is known about how municipal strategies, programmes and plans pay attention to the promotion of children's mental health and whether it is discussed and reported in the municipal councils, boards and committees. The purpose of this study was to examine how municipalities in one Finnish region promote mental health, with a focus on the promotion of children's mental health. METHODS: Document analysis was used as a research method. Documents were selected for a one-year period (2018) from three municipalities of the North Savo region. Analysed documents (n=269) were municipal strategies, programmes and plans, as well as meeting minutes of municipal councils, boards and committees. Eight domains of the structural indicators of mental health were used as an analysis frame. RESULTS: In total, 1169 mentions related to the structural indicators of mental health were found in the documents. In strategies, programmes and plans, parenting-related mentions were found most often. Regarding the minutes, the issues discussed and reported about the wellbeing of children focused on practical issues, such as the construction of day care buildings. CONCLUSIONS: Document analysis indicated that mental health promotion involved mostly the society and environment and not as much the age and setting. There was a lack of mentions regarding preschool experiences and family support/childcare and the promotion of mental health through schools and education, especially in the meeting minutes of municipal councils, boards and committees.

Public health nurses' perceptions on promotive and risk factors for children's mental health: A qualitative interview study.

AIM: To describe promotive and risk factors for children's mental health at the individual, interpersonal, organizational, community and public policy levels of the socioecological environment. DESIGN: A descriptive qualitative interview study. METHODS: Data were collected in Finland during autumn 2019 via semi-structured interviews. Participants (n = 23) comprised public health nurses who worked in child health clinics and school healthcare. Data were analyzed using inductive content analysis. RESULTS: Several promotive and risk factors were identified to have affected children's mental health. Intra-family factors were the most important in the opinion of public health nurses, and they were most concerned about family related risk factors. In addition, the descriptions of nurses revealed that children are in a socially unequal position in terms of their place of residence. It was also uncovered that there were fewer social and healthcare and leisure services in small municipalities, the distances to services were longer, which was perceived to complicate the use of services, and the fear of stigma was greater. CONCLUSION: According to public health nurses, factors at the individual, interpersonal, organizational, community and public policy levels of the socioecological environment affect children's mental health and overall well-being. Consequently, the promotion of children's mental health should be emphasized at every level of the society, considering the whole family. IMPACT: This study indicated the importance of considering factors that affect children's mental health at all levels of the socioecological environment. The results can be reflected in the relation of the socioecological model of health promotion and used in planning the work of nurses in primary healthcare and other relevant nursing settings to emphasize promotive and preventive work.

Genomic Patterns of Local Adaptation under Gene Flow in Arabidopsis lyrata.

Short-scale local adaptation is a complex process involving selection, migration, and drift. The expected effects on the genome are well grounded in theory but examining these on an empirical level has proven difficult, as it requires information about local selection, demographic history, and recombination rate variation. Here, we use locally adapted and phenotypically differentiated Arabidopsis lyrata populations from two altitudinal gradients in Norway to test these expectations at the whole-genome level. Demography modeling indicates that populations within the gradients diverged <2 kya and that the sites are connected by gene flow. The gene flow estimates are, however, highly asymmetric with migration from high to low altitudes being several times more frequent than vice versa. To detect signatures of selection for local adaptation, we estimate patterns of lineage-specific differentiation among these populations. Theory predicts that gene flow leads to concentration of adaptive loci in areas of low recombination; a pattern we observe in both lowland-alpine comparisons. Although most selected loci display patterns of conditional neutrality, we found indications of genetic trade-offs, with one locus particularly showing high differentiation and signs of selection in both populations. Our results further suggest that resistance to solar radiation is an important adaptation to alpine environments, while vegetative growth and bacterial defense are indicated as selected traits in the lowland habitats. These results provide insights into genetic architectures and evolutionary processes driving local adaptation under gene flow. We also contribute to understanding of traits and biological processes underlying alpine adaptation in northern latitudes.

Inflorescence shoot elongation, but not flower primordia formation, is photoperiodically regulated in Arabidopsis lyrata.

BACKGROUND AND AIMS: Photoperiod contains information about the progress of seasons. Plants use the changing photoperiod as a cue for the correct timing of important life history events, including flowering. Here the effect of photoperiod on flowering in four Arabidopsis lyrata populations originating from different latitudes was studied, as well as expression levels of candidate genes for governing the between-population differences. METHODS: Flowering of plants from four A. lyrata populations was studied in three different photoperiods after vernalization. Flowering development was separated into three steps: flower primordia formation, inflorescence shoot elongation and opening of the first flower. Circadian expression rhythms of the A. lyrata homologues of GIGANTEA (GI), FLAVIN-BINDING, KELCH REPEAT, F-BOX 1 (FKF1), CONSTANS (CO) and FLOWERING LOCUS T (FT) were studied in three of the populations in the intermediate (14 h) photoperiod treatment. KEY RESULTS: Most plants in all populations formed visible flower primordia during vernalization. Further inflorescence development after vernalization was strongly inhibited by short days in the northern European population (latitude 61°N), only slightly in the central European population (49°N) and not at all in the North American populations (36°N and 42°N). In the 14 h daylength, where all plants from the three southernmost populations but only 60 % of the northernmost population flowered, the circadian expression rhythm of the A. lyrata FT was only detected in the southern populations, suggesting differentiation in the critical daylength for activation of the long-day pathway. However, circadian expression rhythms of A. lyrata GI, FKF1 and CO were similar between populations. CONCLUSIONS: The results indicate that in A. lyrata, transition to flowering can occur through pathways independent of long days, but elongation of inflorescences is photoperiodically regulated.

Ecological genomics of local adaptation.

It is increasingly important to improve our understanding of the genetic basis of local adaptation because of its relevance to climate change, crop and animal production, and conservation of genetic resources. Phenotypic patterns that are generated by spatially varying selection have long been observed, and both genetic mapping and field experiments provided initial insights into the genetic architecture of adaptive traits. Genomic tools are now allowing genome-wide studies, and recent theoretical advances can help to design research strategies that combine genomics and field experiments to examine the genetics of local adaptation. These advances are also allowing research in non-model species, the adaptation patterns of which may differ from those of traditional model species.

Genome-Wide Analysis of Colonization History and Concomitant Selection in Arabidopsis lyrata.

The high climatic variability in the past hundred thousand years has affected the demographic and adaptive processes in many species, especially in boreal and temperate regions undergoing glacial cycles. This has also influenced the patterns of genome-wide nucleotide variation, but the details of these effects are largely unknown. Here we study the patterns of genome-wide variation to infer colonization history and patterns of selection of the perennial herb species Arabidopsis lyrata, in locally adapted populations from different parts of its distribution range (Germany, UK, Norway, Sweden, and USA) representing different environmental conditions. Using site frequency spectra based demographic modeling, we found strong reduction in the effective population size of the species in general within the past 100,000 years, with more pronounced effects in the colonizing populations. We further found that the northwestern European A. lyrata populations (UK and Scandinavian) are more closely related to each other than with the Central European populations, and coalescent based population split modeling suggests that western European and Scandinavian populations became isolated relatively recently after the glacial retreat. We also highlighted loci showing evidence for local selection associated with the Scandinavian colonization. The results presented here give new insights into postglacial Scandinavian colonization history and its genome-wide effects.

Scots pine aminopropyltransferases shed new light on evolution of the polyamine biosynthesis pathway in seed plants.

Background and Aims: Polyamines are small metabolites present in all living cells and play fundamental roles in numerous physiological events in plants. The aminopropyltransferases (APTs), spermidine synthase (SPDS), spermine synthase (SPMS) and thermospermine synthase (ACL5), are essential enzymes in the polyamine biosynthesis pathway. In angiosperms, SPMS has evolved from SPDS via gene duplication, whereas in gymnosperms APTs are mostly unexplored and no SPMS gene has been reported. The present study aimed to investigate the functional properties of the SPDS and ACL5 proteins of Scots pine (Pinus sylvestris L.) in order to elucidate the role and evolution of APTs in higher plants. Methods: Germinating Scots pine seeds and seedlings were analysed for polyamines by high-performance liquid chromatography (HPLC) and the expression of PsSPDS and PsACL5 genes by in situ hybridization. Recombinant proteins of PsSPDS and PsACL5 were produced and investigated for functional properties. Also gene structures, promoter regions and phylogenetic relationships of PsSPDS and PsACL5 genes were analysed. Key Results: Scots pine tissues were found to contain spermidine, spermine and thermospermine. PsSPDS enzyme catalysed synthesis of both spermidine and spermine. PsACL5 was found to produce thermospermine, and PsACL5 gene expression was localized in the developing procambium in embryos and tracheary elements in seedlings. Conclusions: Contrary to previous views, our results demonstrate that SPMS activity is not a novel feature developed solely in the angiosperm lineage of seed plants but also exists as a secondary property in the Scots pine SPDS enzyme. The discovery of bifunctional SPDS from an evolutionarily old conifer reveals the missing link in the evolution of the polyamine biosynthesis pathway. The finding emphasizes the importance of pre-existing secondary functions in the evolution of new enzyme activities via gene duplication. Our results also associate PsACL5 with the development of vascular structures in Scots pine.

Role of seed germination in adaptation and reproductive isolation in Arabidopsis lyrata.

Seed germination is an important developmental and life history stage. Yet, the evolutionary impact of germination has mainly been studied in the context of dormancy, or for its role in reproductive isolation between species. Here, we aim to examine multiple consequences of genetic divergence on germination traits between two Arabidopsis lyrata subspecies: ssp. petraea (Eurasia) and ssp. lyrata (North America). Postdormancy germination time, a potentially adaptive trait, showed differentiation between the populations, and quantitative trait loci (QTL) mapping revealed that the trait variation is mainly controlled by two antagonistic loci. These QTL areas contain several candidate genes with known function in postdormancy germination in A. thaliana. The sequence variation of three genes was consistent with differential selection, and they also included fixed nonsynonymous substitutions with potential to account for the phenotypic differentiation. We further show that the divergence between the subspecies has led to a slight but significant reduction in hybrid germination proportions, indicating incipient reproductive isolation. Comparison of reciprocal F1 and F2 progenies suggests that Bateson-Dobzhansky-Muller incompatibilities likely act through uniparentally inherited factors. Examination of genomewide transmission ratio distortion further revealed that cytonuclear interactions cause substantial pregermination inviability in the hybrids. These results confirm that seed germination has adaptive potential beyond the dormancy stage and that hybrid seed inviability can be one of the first reproductive barriers to arise during divergence.

High rate of adaptive evolution in two widespread European pines.

Comparing related organisms with differing ecological requirements and evolutionary histories can shed light on the mechanisms and drivers underlying genetic adaptation. Here, by examining a common set of hundreds of loci, we compare patterns of nucleotide diversity and molecular adaptation of two European conifers (Scots pine and maritime pine) living in contrasted environments and characterized by distinct population genetic structure (low and clinal in Scots pine, high and ecotypic in maritime pine) and demographic histories. We found higher nucleotide diversity in Scots pine than in maritime pine, whereas rates of new adaptive substitutions (ωa ), as estimated from the distribution of fitness effects, were similar across species and among the highest found in plants. Sample size and population genetic structure did not appear to have resulted in significant bias in estimates of ωa . Moreover, population contraction-expansion dynamics for each species did not affect differentially the rate of adaptive substitution in these two pines. Several methodological and biological factors may underlie the unusually high rate of adaptive evolution of Scots pine and maritime pine. By providing two new case studies with contrasting evolutionary histories, we contribute to disentangling the multiple factors potentially affecting adaptive evolution in natural plant populations.

Selection for population-specific adaptation shaped patterns of variation in the photoperiod pathway genes in Arabidopsis lyrata during post-glacial colonization.

Spatially varying selection can lead to population-specific adaptation, which is often recognized at the phenotypic level; however, the genetic evidence is weaker in many groups of organisms. In plants, environmental shifts that occur due to colonization of a novel environment may require adaptive changes in the timing of growth and flowering, which are often governed by location-specific environmental cues such as day length. We studied locally varying selection in 19 flowering time loci in nine populations of the perennial herb Arabidopsis lyrata, which has a wide but patchy distribution in temperate and boreal regions of the northern hemisphere. The populations differ in their recent population demographic and colonization histories and current environmental conditions, especially in the growing season length. We searched for population-specific molecular signatures of directional selection by comparing a set of candidate flowering time loci with a genomic reference set within each population using multiple approaches and contrasted the patterns of different populations. The candidate loci possessed approximately 20% of the diversity of the reference loci. On average the flowering time loci had more rare alleles (a smaller Tajima's D) and an excess of highly differentiated sites relative to the reference, suggesting positive selection. The strongest signal of selection was detected in photoperiodic pathway loci in the colonizing populations of Northwestern Europe, whereas no evidence of positive selection was detected in the Central European populations. These findings emphasized the population-specific nature of selection and suggested that photoperiodic adaptation was important during postglacial colonization of the species.

Association of FLOWERING LOCUS T/TERMINAL FLOWER 1-like gene FTL2 expression with growth rhythm in Scots pine (Pinus sylvestris).

Understanding the genetic basis of the timing of bud set, an important trait in conifers, is relevant for adaptation and forestry practice. In common garden experiments, both Scots pine (Pinus sylvestris) and Norway spruce (Picea abies) show a latitudinal cline in the trait. We compared the regulation of their bud set biology by examining the expression of PsFTL2, a Pinus sylvestris homolog to PaFTL2, a FLOWERING LOCUS T/TERMINAL FLOWER 1 (FT/TFL1)-like gene, the expression levels of which have been found previously to be associated with the timing of bud set in Norway spruce. In a common garden study, we analyzed the relationship of bud phenology under natural and artificial photoperiods and the expression of PsFTL2 in a set of Scots pine populations from different latitudes. The expression of PsFTL2 increased in the needles preceding bud set and decreased during bud burst. In the northernmost population, even short night periods were efficient to trigger this expression, which also increased earlier under all photoperiodic regimes compared with the southern populations. Despite the different biology, with few limitations, the two conifers that diverged 140 million yr ago probably share an association of FTL2 with bud set, pointing to a common mechanism for the timing of growth cessation in conifers.

A recent local sweep at the PHYA locus in the Northern European Spiterstulen population of Arabidopsis lyrata.

Northern and central European Arabidopsis lyrata ssp. petraea populations are locally adapted to prevailing climatic conditions through differences in timing of life history events. The timing of flowering and, in perennials, the timing of growth cessation influence fitness. Phytochrome A may have an important role in regulating these life history traits as it perceives changes in daylength. We asked whether PHYA has contributed to local adaptation to the northern conditions in A. l. petraea. To search for signals of directional selection at the PHYA locus, we resequenced PHYA and 9 short fragments around PHYA from a 57-kb region from a German (Plech) and a Norwegian (Spiterstulen) population and compared patterns of differentiation and diversity to a set of 19 reference loci around the genome. First, we found that the populations were highly differentiated: there were three nonsynonymous fixed differences at the PHYA locus, which was in stark contrast with the total four fixed differences in the 19 reference loci. Compatible with a sweep hypothesis, variation was almost completely removed from the 9.4-kb region around PHYA in the northern Spiterstulen population. The overall level of linkage disequilibrium (LD) was higher in Spiterstulen, but there was no LD across the PHYA locus in the population, which is also a known consequence of a selective sweep. The sweep has likely occurred after the last glacial maximum, which suggests that it has contributed to adaptation to the northern conditions.

Climatic adaptation and ecological divergence between two closely related pine species in Southeast China.

Climate is one of the most important drivers for adaptive evolution in forest trees. Climatic selection contributes greatly to local adaptation and intraspecific differentiation, but this kind of selection could also have promoted interspecific divergence through ecological speciation. To test this hypothesis, we examined intra- and interspecific genetic variation at 25 climate-related candidate genes and 12 reference loci in two closely related pine species, Pinus massoniana Lamb. and Pinus hwangshanensis Hisa, using population genetic and landscape genetic approaches. These two species occur in Southeast China but have contrasting ecological preferences in terms of several environmental variables, notably altitude, although hybrids form where their distributions overlap. One or more robust tests detected signals of recent and/or ancient selection at two-thirds (17) of the 25 candidate genes, at varying evolutionary timescales, but only three of the 12 reference loci. The signals of recent selection were species specific, but signals of ancient selection were mostly shared by the two species likely because of the shared evolutionary history. FST outlier analysis identified six SNPs in five climate-related candidate genes under divergent selection between the two species. In addition, a total of 24 candidate SNPs representing nine candidate genes showed significant correlation with altitudinal divergence in the two species based on the covariance matrix of population history derived from reference SNPs. Genetic differentiation between these two species was higher at the candidate genes than at the reference loci. Moreover, analysis using the isolation-with-migration model indicated that gene flow between the species has been more restricted for climate-related candidate genes than the reference loci, in both directions. Taken together, our results suggest that species-specific and divergent climatic selection at the candidate genes might have counteracted interspecific gene flow and played a key role in the ecological divergence of these two closely related pine species.

Epidemiology of genital herpes simplex virus type 1 and 2 infections in southwestern Finland during a 10-year period (2003-2012).

BACKGROUND: The objective of this study was to analyze the proportion of herpes simplex virus types 1 and 2 (HSV-1 and HSV-2) in genital infections during a 10-year period (2003-2012) among outpatients of a clinic of sexually transmitted disease in Southwestern Finland. METHODS: We analyzed prospectively the proportion of HSV-1- or HSV-2-positive culture samples from our sexually transmitted disease clinic outpatients with genital herpes infection during the years 2003 to 2012 and compared the proportions of positive HSV-1 and HSV-2 findings with the age and sex of the patients. RESULTS: Herpes simplex virus type 2 was typed in 66.4% (557/839) and HSV-1 in 33.6% (282/839) of the patients during the entire study period. The mean age of male patients (26.3 years) with a laboratory-confirmed HSV-1 infection was significantly lower than that in male patients with an HSV-2 infection in 2003 to 2007 (26.3 vs. 32.9 years), with P < 0.0001, and (28.6 vs. 34.0) P < 0.0019 in 2008 to 2012, respectively. More female than male patients had HSV-1 infection; in the first study period (2003-2007), 58% were female, and in 2008 to 2012, it was again 63.6%. There was no significant difference between the age of female patients with HSV-1 and those with HSV-2. CONCLUSIONS: Herpes simplex virus type 2 was still the most common causative agent of genital herpes in Southwestern Finland, but the proportion of HSV-1 was increasingly high. The age difference between male patients with HSV-1 and HSV-2 narrowed during the years studied.