Seasonal dynamics of N in two Sphagnum moss species and the underlying peat treated with 15NH415NO3

Uptake of 15N labelled NH4NO3 by two Sphagnum mosses on a raised bog in north east Scotland was measured at different times of the year. In a field experiment, fortnightly additions of NH4NO3 at natural abundance, equivalent to 3 g N m-2 yr-1, were made over 14 months to cores of Sphagnum capillifolium occupying hummocks and S. recurvum colonizing hollows. Pre-harvested cores were treated with 15NH415NO3 two weeks before harvesting and 15N abundance determined for the total N in the moss, inorganic and dissolved organic N (DON) in the moss water and extractable inorganic, organic and microbial N in the underlying peat. The proportion of added 15N taken up by the mosses two weeks after each addition averaged 72% and ranged between 11 and 100%, tending to be least during October when the rising water table reached the surface, particularly for S. recurvum. A small proportion of the 15N was detected in the moss water as NH4+ (0.01%) and as DON (0.03%) and on occasions a large proportion remained unaccounted for. In waters from S. capillifolium, DON was proportional to the amount of inorganic N added, but this was not the case for S. recurvum. Little or no 15N was detected in the underlying peat partly because of the large size and variability of the NH4+, DON and microbial N pools.     

Development of microsatellites for the invasive riparian plant Impatiens glandulifera (Himalayan balsam) using intersimple sequence repeat cloning

Impatiens glandulifera (Himalayan balsam) is an invasive riparian plant species that can outcompete native perennials. Population genetic data on dispersal may aid in the management of invasive species, so we have developed microsatellite markers for this significant invader using an intersimple sequence repeat (ISSR)‐based cloning method. Eight polymorphic markers displayed between two and five alleles, with overall levels of observed and expected heterozygosities ranging from 0.0500 to 0.7500 and from 0.1449 to 0.7692, respectively.     

Development of Sphagnum fallax diaspores on bare peat with implications for the restoration of cut-over bogs

1. The growth of Sphagnum fallax was studied in a glasshouse experiment. Capitulum diaspores of S. fallax were cultivated on five different types of bare peat core, representing a gradient of increasing disturbance. Increases in length and weight were measured. Three microclimates were simulated by protecting the bare peat with a shading mesh and a plastic cover in combination with two water levels.
2. Significant differences in the growth of S. fallax were observed in relation to microclimate changes. Protection techniques such as shading mesh and perforated plastic film allowed a better development of the diaspores compared to bare peat. A plastic cover caused the best growth and compensated for a low water table level.
3. Peat properties are critical when diaspores grow in direct contact with decomposed peat. The porosity, and especially its vertical pattern in the peat profile, proved to be an important factor. A particular combination of microclimate conditions at the surface of the bare peat, and physical properties of the upper peat layers, may favour the growth of S. fallax diaspores even where there is a low water
Table 4 . Restoration of Sphagnum species on cut-over bogs needs to consider both the microclimate conditions at the surface of the bare peat and the peat properties themselves. These factors are important for the diaspores, particularly in periods of climatic and hydrological stress. In such situations, commensalism with some vascular plants may be a useful trigger for Sphagnum growth.     

Recombination and introgression of nuclear and chloroplast genomes between the peat mosses, Sphagnum capillifolium and Sphagnum quinquefarium

Haploid hybrid gametophytes are often present at low frequencies in sympatric populations of Sphagnum capillifolium and Sphagnum quinquefarium. We used intersimple sequence repeat (ISSR) markers and polymerase chain reaction-restriction fragment length polymorphism of the trnL(UAA) intron of the chloroplast genome to reveal the nuclear and chloroplast composition of mature hybrid gametophytes from natural populations and of gametophytes derived from spores of hybrid sporophytes collected in nature. Asymmetrical nuclear inheritance was found in the progeny of the hybrid sporophytes, indicating that only spores with a low level of recombination of parental genomes were viable. A similarly skewed nuclear composition was found among the naturally occurring hybrid gametophytes. All hybrid genomes contained a larger proportion of S. capillifolium ISSR markers, combined with only two to five S. quinquefarium markers together with a chloroplast haplotype derived from S. quinquefarium. In this way, a pattern resembling introgression is created within a single generation. Some individuals possessed nuclear genomes typical for S. capillifolium in combination with the chloroplast haplotype of S. quinquefarium, possibly indicating backcrossing. Our results indicate that hybridization between S. capillifolium and S. quinquefarium is relatively common, but the resistance of large parts of the genome against heterospecific genes maintains the genetic distinctness of the species. Further evolutionary and phylogenetic consequences of restricted interspecific gene exchange are discussed.     

Development and characterization of polymorphic microsatellites from Prunus avium‘Napoleon’

Primers were developed for 21 microsatellite loci isolated by enrichment from Prunus avium‘Napoleon’. Twelve loci contained uninterrupted dinucleotide repeats and nine were more complex. Nineteen primer pairs (EMPA001–019) showed single locus polymorphisms in a cultivar survey of 14 sweet cherries, with two to seven alleles per locus. Three primer pairs in combination (EMPA014, 015 and 018) discriminated all cultivars. Two primer pairs for loci monomorphic in P. avium were included: EMPA020 revealed segregation in an interspecific progeny and EMPA021 revealed polymorphism in P. dulcis. Twelve primer pairs reliably amplified products in three peach cultivars of which seven revealed polymorphisms.     

Freezing cytorrhysis and critical temperature thresholds for photosystem II in the peat moss Sphagnum capillifolium

Leaflets of Sphagnum capillifolium were exposed to temperatures from ?5°C to +60°C under controlled conditions while mounted on a microscope stage. The resultant cytological response to these temperature treatments was successfully monitored using a light and fluorescence microscope. In addition to the observable cytological changes during freezing cytorrhysis and heat exposure on the leaflets, the concomitant critical temperature thresholds for inactivation of photosystem II (PS II) were studied using a micro fibre optic and a chlorophyll fluorometer mounted to the microscope stage. Chlorophyllous cells of S. capillifolium showed extended freezing cytorrhysis immediately after ice nucleation at ?1.1°C in the water in which the leaflets were submersed during the measurement. The occurrence of freezing cytorrhysis, which was visually manifested by cell shrinkage, was highly dynamic and was completed within 2 s. A total reduction of the mean projected diameter of the chloroplast containing area during freezing cytorrhysis from 8.9 to 3.8 μm indicates a cell volume reduction of approximately ?82%. Simultaneous measurement of chlorophyll fluorescence of PS II was possible even through the frozen water in which the leaf samples were submersed. Freezing cytorrhysis was accompanied by a sudden rise of basic chlorophyll fluorescence. The critical freezing temperature threshold of PS II was identical to the ice nucleation temperature (?1.1°C). This is significantly above the temperature threshold at which frost damage to S. capillifolium leaflets occurs (?16.1°C; LT₅₀) which is higher than observed in most higher plants from the European Alps during summer. High temperature thresholds of PS II were 44.5°C which is significantly below the heat tolerance of chlorophyllous cells (49.9°C; LT₅₀). It is demonstrated that light and fluorescence microscopic techniques combined with simultaneous chlorophyll fluorescence measurements may act as a useful tool to study heat, low temperature, and ice-encasement effects on the cellular structure and primary photosynthetic processes of intact leaf tissues.     

Hybridization between the peat mosses, Sphagnum capillifolium and S. quinquefarium (Sphagnaceae, Bryophyta) as inferred by morphological characters and isozyme markers

 Isozyme markers and morphological characters were studied in four populations of Sphagnum capillifolium and S. quinquefarium. Recombinant plants were found in three populations, where the two species occur sympatrically. All recombinants possessed different haplotypes and combinations of morphological characters, which show that they are results of independent hybridization events. Strongly male-biased sex ratios were found for Sphagnum capillifolium in all populations where it grew sympatrically with S. quinquefarium. Most of the recombinants were also male fertile. These observations suggest that S. quinquefarium is the female parent in the primary crosses and in subsequent backcrosses. Received September 3, 2001; accepted March 16, 2002 Published online: November 7, 2002 Addresses of the authors: Nils Cronberg (e-mail: Nils.Cronberg@sysbot.lu.se), Department of Systematic Botany, Lund University, S?lvegatan 37, SE-223 62 Lund, Sweden. Rayna Natcheva (e-mail: renimoss@iph.bio.bas.bg), Institute of Botany, Bulgarian Academy of Sciences, 23 Acad. G. Bonchev Str., 1113 Sofia, Bulgaria.     

Identification and characterization of nuclear microsatellite loci in the aquatic moss Platyhypnidium riparioides (Brachytheciaceae)

Eight microsatellite loci from the aquatic moss Platyhypnidium riparioides were identified using the method of microsatellite‐enriched libraries. Polymorphism was assessed in a sample of four populations of 20 individuals each from four streams of the Meuse hydrographic basin in southern Belgium. The markers amplified three to seven alleles per locus. Comparison of observed and expected heterozygosities as well as F‐statistics (F_ST = 0.62) reveals a significant genetic differentiation among populations. These markers will be useful for further investigation of population genetic structure and diversity at different nested spatial scales.     

Universal primers for the amplification of chloroplast microsatellites in grasses (Poaceae)

Attempts to design truly universal primers to amplify chloroplast microsatellites have met with limited success due to nonconservation of repeat loci across widely divergent taxa. We have used the complete chloroplast genome sequences of rice, maize and wheat to design five pairs of primers that amplify homologous mononucleotide repeats across the Poaceae (grasses). Sequencing confirmed conservation of repeat motifs across subfamilies and a preliminary study in Anthoxanthum odoratum revealed polymorphism at two loci with a haplotype diversity value of 0.495. These primers provide a valuable tool to study cytoplasmic diversity in this extensively studied and economically important range of taxa.     

Development and mapping of SSR markers for maize

Microsatellite or simple sequence repeat (SSR) markers have wide applicability for genetic analysis in crop plant improvement strategies. The objectives of this project were to isolate, characterize, and map a comprehensive set of SSR markers for maize (Zea mays L.). We developed 1051 novel SSR markers for maize from microsatellite-enriched libraries and by identification of microsatellite-containing sequences in public and private databases. Three mapping populations were used to derive map positions for 978 of these markers. The main mapping population was the intermated B73 × Mo17 (IBM) population. In mapping this intermated recombinant inbred line population, we have contributed to development of a new high-resolution map resource for maize. The primer sequences, original sequence sources, data on polymorphisms across 11 inbred lines, and map positions have been integrated with information on other public SSR markers and released through MaizeDB at URL:www.agron.missouri.edu. The maize research community now has the most detailed and comprehensive SSR marker set of any plant species.     

Expressed sequence tag‐derived microsatellites for the cool‐water marine copepod Calanus finmarchicus

The copepod Calanus finmarchicus is the major contributor to zooplankton biomass in the North Atlantic and Norwegian Sea, but recent studies have shown a 70% decrease in abundance as well as a northward shift in the species’ range. Insights into dispersal capabilities gained from population genetic studies will be crucial in predicting the response of C. finmarchicus communities to climate change and, consequently, we have developed a set of expressed sequence tag‐derived microsatellite markers to allow fine‐scale elucidation of population structuring and dispersal. Ten polymorphic markers displayed between two and 19 alleles, with levels of expected heterozygosity ranging from 0.044 to 0.924.     

Climate drivers alter nitrogen availability in surface peat and decouple N2 fixation from CH4 oxidation in the Sphagnum moss microbiome

Peat mosses (Sphagnum spp.) are keystone species in boreal peatlands, where they dominate net primary productivity and facilitate the accumulation of carbon in thick peat deposits. Sphagnum mosses harbor a diverse assemblage of microbial partners, including N₂-fixing (diazotrophic) and CH₄-oxidizing (methanotrophic) taxa that support ecosystem function by regulating transformations of carbon and nitrogen. Here, we investigate the response of the Sphagnum phytobiome (plant + constituent microbiome + environment) to a gradient of experimental warming (+0°C to +9°C) and elevated CO₂ (+500 ppm) in an ombrotrophic peatland in northern Minnesota (USA). By tracking changes in carbon (CH₄, CO₂) and nitrogen (NH₄-N) cycling from the belowground environment up to Sphagnum and its associated microbiome, we identified a series of cascading impacts to the Sphagnum phytobiome triggered by warming and elevated CO₂. Under ambient CO₂, warming increased plant-available NH₄-N in surface peat, excess N accumulated in Sphagnum tissue, and N₂ fixation activity decreased. Elevated CO₂ offset the effects of warming, disrupting the accumulation of N in peat and Sphagnum tissue. Methane concentrations in porewater increased with warming irrespective of CO₂ treatment, resulting in a ~10× rise in methanotrophic activity within Sphagnum from the +9°C enclosures. Warming's divergent impacts on diazotrophy and methanotrophy caused these processes to become decoupled at warmer temperatures, as evidenced by declining rates of methane-induced N₂ fixation and significant losses of keystone microbial taxa. In addition to changes in the Sphagnum microbiome, we observed ~94% mortality of Sphagnum between the +0°C and +9°C treatments, possibly due to the interactive effects of warming on N-availability and competition from vascular plant species. Collectively, these results highlight the vulnerability of the Sphagnum phytobiome to rising temperatures and atmospheric CO₂ concentrations, with significant implications for carbon and nitrogen cycling in boreal peatlands.     

Esterified long-chain hydroxy acids in green and senescent parts of the peat moss Sphagnum fuscum

The bifunctional long-chain acids in extracts and alkaline hydrolysates of extractive-free tissues from the green and senescent parts of Sphagnum fuscum were investigated by GC and GC/MS. A series of C₁₄– C₂₆hydroxy acids was liberated by alkaline hydrolysis of senescent tissue demonstrating the presence of insoluble polymeric lipid esters in the moss. In the corresponding hydrolysates of the topmost green parts of the shoot the amounts of these acids were significantly smaller. It is suggested that the presence of such polymeric esters in S. fuscum contributes to the cell-wall resistance of this peat-forming moss.     

Microclimate and production of peat moss Sphagnum palustre L. in the warm‐temperate zone

Sphagnum palustre L. is one of the few Sphagnum species distributed in the warm‐temperate zone. To elucidate the mechanisms that enable S. palustre to maintain its productivity under warm climatic conditions, we examined the temperature conditions and photosynthetic characteristics of this species in a lowland wetland in western Japan. Moss temperatures during the daytime were much lower than the air temperature, particularly during summer. The optimum temperature for the net photosynthetic rate was approximately 20°C, irrespective of the season, but summer and autumn samples maintained high rates at higher temperatures as well. The net photosynthetic rate at near light saturation was much higher during summer–autumn than during spring–winter. A model estimation in which net production was calculated from the photosynthetic characteristics and microclimatic data showed that both the low temperature of the moss colony and the seasonal shift in photosynthetic characteristics are among the mechanisms that enable this species to maintain its productivity under warm climatic conditions.     

Motif mismatches in microsatellites: insights from genome-wide investigation among 20 insect species

We present a detailed genome-wide comparative study of motif mismatches of microsatellites among 20 insect species representing five taxonomic orders. The results show that varying proportions (∼15–46%) of microsatellites identified in these species are imperfect in motif structure, and that they also vary in chromosomal distribution within genomes. It was observed that the genomic abundance of imperfect repeats is significantly associated with the length and number of motif mismatches of microsatellites. Furthermore, microsatellites with a higher number of mismatches tend to have lower abundance in the genome, suggesting that sequence heterogeneity of repeat motifs is a key determinant of genomic abundance of microsatellites. This relationship seems to be a general feature of microsatellites even in unrelated species such as yeast, roundworm, mouse and human. We provide a mechanistic explanation of the evolutionary link between motif heterogeneity and genomic abundance of microsatellites by examining the patterns of motif mismatches and allele sequences of single-nucleotide polymorphisms identified within microsatellite loci. Using Drosophila Reference Genetic Panel data, we further show that pattern of allelic variation modulates motif heterogeneity of microsatellites, and provide estimates of allele age of specific imperfect microsatellites found within protein-coding genes.     

Detection of representatives of the Planctomycetes in Sphagnum peat bogs by molecular and cultivation approaches

By means of fluorescence in situ hybridization with 16S rRNA-targeted oligonucleotide probes (FISH), it has been shown that members of the phylum Planctomycetes represent a numerically significant bacterial group in boreal Sphagnum peat bogs. The population size of planctomycetes in oxic layers of the peat bog profile was in the range of 0.4–2.0 × 10⁷ cells per g of wet peat, comprising 4 to 13% of the total bacterial cell number. A novel effective approach that combined a traditional cultivation technique with FISH-mediated monitoring of the target organism during the isolation procedure has been developed for the isolation of planctomycetes. Using this approach, we succeeded in isolating several peat-inhabiting planctomycetes in a pure culture. Sequencing of the 16S rRNA genes from two of these isolates, strains A10 and MPL7, showed that they belonged to the planctomycete lineages defined by the genera Gemmata and Planctomyces, respectively. The 16S rRNA gene sequence similarity between strains A10 and MPL7 and the phylogenetically closest organisms, namely, Gemmata obscuriglobus and Planctomyces limnophilus, was only 90%. These results suggest that the indigenous planctomycetes inhabiting Sphagnum peat bogs are so far unknown organisms.     

A climatic threshold triggers the die‐off of peat mosses during an extreme heat wave

Heat waves, which are projected to be more frequent and intense in a warmer climate, could become a serious threat to plants that rely on water surplus availability, such as bryophytes. Here, I take the advantage of the European summer 2003 climate anomaly to assess the impact of an extreme heat wave on peat mosses of the genus Sphagnum, a group of bryophytes forming the bulk of living and dead biomass in peatlands. With this aim, 20 selected bogs in the Italian Alps were checked for Sphagnum survival in the years following the heat wave. Over the study area, the period May–September 2003 was characterized by higher mean monthly air temperature (13.5 °C) and lower mean monthly precipitation (87 mm) compared with normal climatic conditions (11.5 °C and 117 mm, respectively) so that the heat wave coincided with a drought spell. As a consequence of the unusual water stress, I documented an increased mortality of peat mosses forming high hummocks. In particular, at habitat scale, the distribution of desiccated peat mosses was restricted to the hummock face receiving the greatest amount of solar irradiation. However, at regional scale, the present study identified a climatic threshold, simply defined by the ratio of precipitation to temperature (P : T), which triggered an irreversible desiccation of peat mosses when mean monthly P : T dropped below 6.5 (mm : °C) during May–September 2003. The absence of any sign of recovery after 4 years since the drought must be seen as a harbinger of the deleterious effects of extreme heat waves on organisms not adapted to cope with abrupt climate anomaly.     

Habitat-specific genetic effects on growth rate and morphology across pH and water-level gradients within a population of the moss Sphagnum angustifolium (Sphagnaceae)

To study genetic adaptations in bryophytes on small ecological and spatial scales and to assess the adaptive significance of morphological trait variation, genotypes of Sphagnum angustifolium originating from habitats characterized by different pH and height above water table were clonally propagated and grown along the same gradients that exist in the field. Clones from ombrotrophic habitats grew consistently better ombrotrophically than clones from minerotrophic habitats and vice versa, suggesting that the genotypes were adapted to different pH levels. Genetic variation was found in several morphological traits, but habitat-specific genetic effects were detected only in length of spreading branches. Covariation between morphology and growth was generally environmentally induced. Positive and negative cross-environment genetic correlations suggested the presence of constraints on adaptive reaction norm evolution. The indications of small-scale genetic adaptations suggest either selective establishment of genotypes adapted to specific habitats, strong selective forces operating at the later stages of the life cycle, restricted gene flow over short distances, or a combination of these. In contrast to prevailing views, these results indicate that bryophytes are likely to respond genetically to small-scale environmental gradients.