Honeybees affect floral microbiome composition in a central food source for wild pollinators in boreal ecosystems.

Basic knowledge on dispersal of microbes in pollinator networks is essential for plant, insect, and microbial ecology. Thorough understanding of the ecological consequences of honeybee farming on these complex plant-pollinator-microbe interactions is a prerequisite for sustainable honeybee keeping. Most research on plant-pollinator-microbe interactions have focused on temperate agricultural systems. Therefore, information on a wild plant that is a seasonal bottleneck for pollinators in cold climate such as Salix phylicifolia is of specific importance. We investigated how floral visitation by insects influences the community structure of bacteria and fungi in Salix phylicifolia inflorescences under natural conditions. Insect visitors were experimentally excluded with net bags. We analyzed the microbiome and measured pollen removal in open and bagged inflorescences in sites where honeybees were foraging and in sites without honeybees. Site and plant individual explained most of the variation in floral microbial communities. Insect visitation and honeybees had a smaller but significant effect on the community composition of microbes. Honeybees had a specific effect on the inflorescence microbiome and, e.g., increased the relative abundance of operational taxonomic units (OTUs) from the bacterial order Lactobacillales. Site had a significant effect on the amount of pollen removed from inflorescences but this was not due to honeybees. Insect visitors increased bacterial and especially fungal OTU richness in the inflorescences. Pollinator visits explained 38% variation in fungal richness, but only 10% in bacterial richness. Our work shows that honeybee farming affects the floral microbiome in a wild plant in rural boreal ecosystems.

Archaea in boreal Swedish lakes are diverse, dominated by Woesearchaeota and follow deterministic community assembly.

Despite their key role in biogeochemical processes, particularly the methane cycle, archaea are widely underrepresented in molecular surveys because of their lower abundance compared with bacteria and eukaryotes. Here, we use parallel high-resolution small subunit rRNA gene sequencing to explore archaeal diversity in 109 Swedish lakes and correlate archaeal community assembly mechanisms to large-scale latitudinal, climatic (nemoral to arctic) and nutrient (oligotrophic to eutrophic) gradients. Sequencing with universal primers showed the contribution of archaea was on average 0.8% but increased up to 1.5% of the three domains in forest lakes. Archaea-specific sequencing revealed that freshwater archaeal diversity could be partly explained by lake variables associated with nutrient status. Combined with deterministic co-occurrence patterns this finding suggests that ecological drift is overridden by environmental sorting, as well as other deterministic processes such as biogeographic and evolutionary history, leading to lake-specific archaeal biodiversity. Acetoclastic, hydrogenotrophic and methylotrophic methanogens as well as ammonia-oxidizing archaea were frequently detected across the lakes. Archaea-specific sequencing also revealed representatives of Woesearchaeota and other phyla of the DPANN superphylum. This study adds to our understanding of the ecological range of key archaea in freshwaters and links these taxa to hypotheses about processes governing biogeochemical cycles in lakes.

Cryptogams signify key transitions of bacteria and fungi in Arctic sand dune succession.

Primary succession models focus on aboveground vascular plants. However, the prevalence of mosses and lichens, that is cryptogams, suggests they play a role in soil successions. Here, we explore whether effects of cryptogams on belowground microbes can facilitate progressive shifts in sand dune succession. We linked aboveground vegetation, belowground bacterial and fungal communities, and soil chemical properties in six successional stages in Arctic inland sand dunes: bare sand, grass, moss, lichen, ericoid heath and mountain birch forest. Compared with the bare sand and grass stages, microbial biomass and the proportion of fungi increased in the moss stage, and later stage microbial groups appeared despite the absence of their host plants. Microbial communities of the lichen stage resembled the communities in the vascular plant stages. Bacterial communities correlated better with soil chemical variables than with vegetation and vice versa for fungal communities. The correlation of fungi with vegetation increased with vascular vegetation. Distinct bacterial and fungal patterns of biomass, richness and plant-microbe interactions showed that the aboveground vegetation change structured the bacterial and fungal community differently. The asynchrony of aboveground vs belowground changes suggests that cryptogams can drive succession towards vascular plant dominance through microbially mediated facilitation in eroded Arctic soil.

Distinct Anaerobic Bacterial Consumers of Cellobiose-Derived Carbon in Boreal Fens with Different CO2/CH4 Production Ratios.

Northern peatlands in general have high methane (CH4) emissions, but individual peatlands show considerable variation as CH4 sources. Particularly in nutrient-poor peatlands, CH4 production can be low and exceeded by carbon dioxide (CO2) production from unresolved anaerobic processes. To clarify the role anaerobic bacterial degraders play in this variation, we compared consumers of cellobiose-derived carbon in two fens differing in nutrient status and the ratio of CO2 to CH4 produced. After [13C]cellobiose amendment, the mesotrophic fen produced equal amounts of CH4 and CO2 The oligotrophic fen had lower CH4 production but produced 3 to 59 times more CO2 than CH4 RNA stable-isotope probing revealed that in the mesotrophic fen with higher CH4 production, cellobiose-derived carbon was mainly assimilated by various recognized fermenters of Firmicutes and by Proteobacteria The oligotrophic peat with excess CO2 production revealed a wider variety of cellobiose-C consumers, including Firmicutes and Proteobacteria, but also more unconventional degraders, such as Telmatobacter-related Acidobacteria and subphylum 3 of Verrucomicrobia Prominent and potentially fermentative Planctomycetes and Chloroflexi did not appear to process cellobiose-C. Our results show that anaerobic degradation resulting in different levels of CH4 production can involve distinct sets of bacterial degraders. By distinguishing cellobiose degraders from the total community, this study contributes to defining anaerobic bacteria that process cellulose-derived carbon in peat. Several of the identified degraders, particularly fermenters and potential Fe(III) or humic substance reducers in the oligotrophic peat, represent promising candidates for resolving the origin of excess CO2 production in peatlands. IMPORTANCE: Peatlands are major sources of the greenhouse gas methane (CH4), yet in many peatlands, CO2 production from unresolved anaerobic processes exceeds CH4 production. Anaerobic degradation produces the precursors of CH4 production but also represents competing processes. We show that anaerobic degradation leading to high or low CH4 production involved distinct sets of bacteria. Well-known fermenters dominated in a peatland with high CH4 production, while novel and unconventional degraders could be identified in a site where CO2 production greatly exceeds CH4 production. Our results help identify and assign functions to uncharacterized bacteria that promote or inhibit CH4 production and reveal bacteria potentially producing the excess CO2 in acidic peat. This study contributes to understanding the microbiological basis for different levels of CH4 emission from peatlands.

Methane-cycling microbial communities and methane emission in natural and restored peatlands.

We addressed how restoration of forestry-drained peatlands affects CH(4)-cycling microbes. Despite similar community compositions, the abundance of methanogens and methanotrophs was lower in restored than in natural sites and correlated with CH(4) emission. Poor establishment of methanogens may thus explain low CH(4) emissions on restored peatlands even 10 to 12 years after restoration.

Host's genetic background determines the outcome of reciprocal faecal transplantation on life-history traits and microbiome composition.

BACKGROUND: Microbes play a role in their host's fundamental ecological, chemical, and physiological processes. Host life-history traits from defence to growth are therefore determined not only by the abiotic environment and genotype but also by microbiota composition. However, the relative importance and interactive effects of these factors may vary between organisms. Such connections remain particularly elusive in Lepidoptera, which have been argued to lack a permanent microbiome and have microbiota primarily determined by their diet and environment. We tested the microbiome specificity and its influence on life-history traits of two colour genotypes of the wood tiger moth (Arctia plantaginis) that differ in several traits, including growth. All individuals were grown in the laboratory for several generations with standardized conditions. We analyzed the bacterial community of the genotypes before and after a reciprocal frass (i.e., larval faeces) transplantation and followed growth rate, pupal mass, and the production of defensive secretion. RESULTS: After transplantation, the fast-growing genotype grew significantly slower compared to the controls, but the slow-growing genotype did not change its growth rate. The frass transplant also increased the volume of defensive secretions in the fast-growing genotype but did not affect pupal mass. Overall, the fast-growing genotype appeared more susceptible to the transplantation than the slow-growing genotype. Microbiome differences between the genotypes strongly suggest genotype-based selective filtering of bacteria from the diet and environment. A novel cluster of insect-associated Erysipelotrichaceae was exclusive to the fast-growing genotype, and specific Enterococcaceae were characteristic to the slow-growing genotype. These Enterococcaceae became more prevalent in the fast-growing genotype after the transplant, which suggests that a slower growth rate is potentially related to their presence. CONCLUSIONS: We show that reciprocal frass transplantation can reverse some genotype-specific life-history traits in a lepidopteran host. The results indicate that genotype-specific selective filtering can fine-tune the bacterial community at specific life stages and tissues like the larval frass, even against a background of a highly variable community with stochastic assembly. Altogether, our findings suggest that the host's genotype can influence its susceptibility to being colonized by microbiota, impacting key life-history traits.

Detection of methanogenic Archaea in peat: comparison of PCR primers targeting the mcrA gene.

Methanogens (domain Archaea) have a unique role in the carbon cycle as producers of the greenhouse gas methane (CH(4)). Methyl-coenzyme M reductase (MCR) is a vital enzyme in CH(4) production, and the mcrA gene coding for a subunit of MCR has been employed as a specific marker for the detection and differentiation of methanogen communities. A critical step in assessing environmental mcrA diversity is the selection of PCR primers. The objective of this study was to compare the diversity coverage of three published mcrA primer sets MCR, ME and ML (also known as MCR and Luton-mcrA) and their ability to discern methanogen communities in a drained peatland. The primers were applied to DNA extracts from unfertilised and ash-fertilised peat from two different depths. Amplified mcrA communities were cloned and sequenced, and the sequences were divided into operational taxonomic units (OTUs) by restriction fragment length polymorphism (RFLP) and sequence analysis. All primers recovered characteristic OTUs associated with the peat depths and treatments and confirmed a previous observation of low methanogen diversity. The minor differences in OTU ranges of the primers did not greatly affect the observed community composition. However, as the proportions of several OTUs varied strongly, the primers provided different quantitative representations of mcrA communities. We concluded that the ML and MCR primers had better amplification ranges than the ME set, but the use of MCR with peat samples was problematic due to poor amplification. Consequently, the ML primers were best suited for mcrA analysis of peatland methanogen communities.

Microform-related community patterns of methane-cycling microbes in boreal Sphagnum bogs are site specific.

Vegetation and water table are important regulators of methane emission in peatlands. Microform variation encompasses these factors in small-scale topographic gradients of dry hummocks, intermediate lawns and wet hollows. We examined methane production and oxidization among microforms in four boreal bogs that showed more variation of vegetation within a bog with microform than between the bogs. Potential methane production was low and differed among bogs but not consistently with microform. Methane oxidation followed water table position with microform, showing higher rates closer to surface in lawns and hollows than in hummocks. Methanogen community, analysed by mcrA terminal restriction fragment length polymorphism and dominated by Methanoregulaceae or 'Methanoflorentaceae', varied strongly with bog. The extent of microform-related variation of methanogens depended on the bog. Methanotrophs identified as Methylocystis spp. in pmoA denaturing gradient gel electrophoresis similarly showed effect of bog, and microform patterns were stronger within individual bogs. Our results suggest that methane-cycling microbes in boreal Sphagnum bogs with seemingly uniform environmental conditions may show strong site-dependent variation. The bog-intrinsic factor may be related to carbon availability but contrary to expectations appears to be unrelated to current surface vegetation, calling attention to the origin of carbon substrates for microbes in bogs.

Microbial ecology in a future climate: effects of temperature and moisture on microbial communities of two boreal fens.

Impacts of warming with open-top chambers on microbial communities in wet conditions and in conditions resulting from moderate water-level drawdown (WLD) were studied across 0-50 cm depth in northern and southern boreal sedge fens. Warming alone decreased microbial biomass especially in the northern fen. Impact of warming on microbial PLFA and fungal ITS composition was more obvious in the northern fen and linked to moisture regime and sample depth. Fungal-specific PLFA increased in the surface peat in the drier regime and decreased in layers below 10 cm in the wet regime after warming. OTUs representing Tomentella and Lactarius were observed in drier regime and Mortierella in wet regime after warming in the northern fen. The ectomycorrhizal fungi responded only to WLD. Interestingly, warming together with WLD decreased archaeal 16S rRNA copy numbers in general, and fungal ITS copy numbers in the northern fen. Expectedly, many results indicated that microbial response on warming may be linked to the moisture regime. Results indicated that microbial community in the northern fen representing Arctic soils would be more sensitive to environmental changes. The response to future climate change clearly may vary even within a habitat type, exemplified here by boreal sedge fen.

Spatial patterns of microbial diversity and activity in an aged creosote-contaminated site.

Restoration of polluted sites via in situ bioremediation relies heavily on the indigenous microbes and their activities. Spatial heterogeneity of microbial populations, contaminants and soil chemical parameters on such sites is a major hurdle in optimizing and implementing an appropriate bioremediation regime. We performed a grid-based sampling of an aged creosote-contaminated site followed by geostatistical modelling to illustrate the spatial patterns of microbial diversity and activity and to relate these patterns to the distribution of pollutants. Spatial distribution of bacterial groups unveiled patterns of niche differentiation regulated by patchy distribution of pollutants and an east-to-west pH gradient at the studied site. Proteobacteria clearly dominated in the hot spots of creosote pollution, whereas the abundance of Actinobacteria, TM7 and Planctomycetes was considerably reduced from the hot spots. The pH preferences of proteobacterial groups dominating in pollution could be recognized by examining the order and family-level responses. Acidobacterial classes came across as generalists in hydrocarbon pollution whose spatial distribution seemed to be regulated solely by the pH gradient. Although the community evenness decreased in the heavily polluted zones, basal respiration and fluorescein diacetate hydrolysis rates were higher, indicating the adaptation of specific indigenous microbial populations to hydrocarbon pollution. Combining the information from the kriged maps of microbial and soil chemistry data provided a comprehensive understanding of the long-term impacts of creosote pollution on the subsurface microbial communities. This study also highlighted the prospect of interpreting taxa-specific spatial patterns and applying them as indicators or proxies for monitoring polluted sites.

Archaeal rRNA diversity and methane production in deep boreal peat.

Northern peatlands play a major role in the global carbon cycle as sinks for CO(2) and as sources of CH(4). These diverse ecosystems develop through accumulation of partially decomposed plant material as peat. With increasing depth, peat becomes more and more recalcitrant due to its longer exposure to decomposing processes. Compared with surface peat, deeper peat sediments remain microbiologically poorly described. We detected active archaeal communities even in the deep bottom layers (-220/-280 cm) of two Finnish fen-type peatlands by 16S rRNA-based terminal restriction fragment length polymorphism analysis. In the sediments of the northern study site, all detected archaea were methanogens with Rice Cluster II (RC-II) and Methanosaetaceae as major groups. In southern peatland, Crenarchaeota of a rare unidentified cluster were present together with mainly RC-II methanogens. RNA profiles showed a larger archaeal diversity than DNA-based community profiles, suggesting that small but active populations were better visualized with rRNA. In addition, potential methane production measurements indicated methanogenic activity throughout the vertical peat profiles.

Seasonality of rDNA- and rRNA-derived archaeal communities and methanogenic potential in a boreal mire.

Methane (CH(4)) emissions from boreal wetlands show considerable seasonal variation, including small winter emissions. We addressed the seasonality of CH(4)-producing microbes by comparing archaeal communities and the rates and temperature response of CH(4) production in a boreal fen at three key phases of growing season and in winter. Archaeal community analysis by terminal restriction fragment length polymorphism and cloning of 16S ribosomal DNA and reverse-transcribed RNA revealed slight community shifts with season. The main archaeal groups remained the same throughout the year and were Methanosarcinaceae, Rice cluster II and Methanomicrobiales-associated Fen cluster. These methanogens and the crenarchaeal groups 1.1c and 1.3 were detected from DNA and RNA, but the family Methanosaetaceae was detected only from RNA. Differences between DNA- and RNA-based results suggested higher stability of DNA-derived communities and better representation of the active CH(4) producers in RNA. Methane production potential, measured as formation of CH(4) in anoxic laboratory incubations, showed prominent seasonality. The potential was strikingly highest in winter, possibly due to accumulation of methanogenic substrates, and maximal CH(4) production was observed at ca. 30 degrees C. Archaeal community size, determined by quantitative PCR, remained similar from winter to summer. Low production potential in late summer after a water level draw-down suggested diminished activity due to oxygen exposure. Our results indicated that archaeal community composition and size in the boreal fen varied only slightly despite the large fluctuations of methanogenic potential. Detection of mRNA of the methanogenic mcrA gene confirmed activity of methanogens in winter, accounting for previously reported winter CH(4) emissions.

Methanogen communities and Bacteria along an ecohydrological gradient in a northern raised bog complex.

Mires forming an ecohydrological gradient from nutrient-rich, groundwater-fed mesotrophic and oligotrophic fens to a nutrient-poor ombrotrophic bog were studied by comparing potential methane (CH(4)) production and methanogenic microbial communities. Methane production was measured from different depths of anoxic peat and methanogen communities were detected by detailed restriction fragment length polymorphism (RFLP) analysis of clone libraries, sequencing and phylogenetic analysis. Potential CH(4) production changed along the ecohydrological gradient with the fens displaying much higher production than the ombrotrophic bog. Methanogen diversity also decreased along the gradient. The two fens had very similar diversity of methanogenic methyl-coenzyme M reductase gene (mcrA), but in the upper layer of the bog the methanogen diversity was strikingly lower, and only one type of mcrA sequence was retrieved. It was related to the Fen cluster, a group of novel methanogenic sequences found earlier in Finnish mires. Bacterial 16S rDNA sequences from the fens fell into at least nine phyla, but only four phyla were retrieved from the bog. The most common bacterial groups were Deltaproteobacteria, Verrucomicrobia and Acidobacteria.