Corrigendum: LocoGSE, a sequence-based genome size estimator for plants.

[This corrects the article DOI: 10.3389/fpls.2024.1328966.].

Deciphering microbial communities of three Savoyard raw milk cheeses along ripening and regarding the cheese process.

Different Savoyard cheeses are granted with PDO (Protected Designation or Origin) and PGI (Protected Geographical Indication) which guarantees consumers compliance with strict specifications. The use of raw milk is known to be crucial for specific flavor development. To unravel the factors influencing microbial ecosystems across cheese making steps, according to the seasonality (winter and summer) and the mode of production (farmhouse and dairy factory ones), gene targeting on bacteria and fungus was used to have a full picture of 3 cheese making technologies, from the raw milk to the end of the ripening. Our results revealed that Savoyard raw milks are a plenteous source of biodiversity together with the brines used during the process, that may support the development of specific features for each cheese. It was shown that rinds and curds have very contrasted ecosystem diversity, composition, and evolution. Ripening stage was selective for some bacterial species, whereas fungus were mainly ubiquitous in dairy samples. All ripening stages are impacted by the type of cheese technologies, with a higher impact on bacterial communities, except for fungal rind communities, for which the technology is the more discriminant. The specific microorganism's abundance for each technology allow to see a real bar-code, with more or less differences regarding bacterial or fungal communities. Bacterial structuration is shaped mainly by matrices, differently regarding technologies while the influence of technology is higher for fungi. Production types showed 10 differential bacterial species, farmhouses showed more ripening taxa, while dairy factory products showing more lactic acid bacteria. Meanwhile, seasonality looks to be a minor element for the comprehension of both microbial ecosystems, but the uniqueness of each dairy plant is a key explicative feature, more for bacteria than for fungus communities.

LocoGSE, a sequence-based genome size estimator for plants.

Extensive research has focused on exploring the range of genome sizes in eukaryotes, with a particular emphasis on land plants, where significant variability has been observed. Accurate estimation of genome size is essential for various research purposes, but existing sequence-based methods have limitations, particularly for low-coverage datasets. In this study, we introduce LocoGSE, a novel genome size estimator designed specifically for low-coverage datasets generated by genome skimming approaches. LocoGSE relies on mapping the reads on single copy consensus proteins without the need for a reference genome assembly. We calibrated LocoGSE using 430 low-coverage Angiosperm genome skimming datasets and compared its performance against other estimators. Our results demonstrate that LocoGSE accurately predicts monoploid genome size even at very low depth of coverage (<1X) and on highly heterozygous samples. Additionally, LocoGSE provides stable estimates across individuals with varying ploidy levels. LocoGSE fills a gap in sequence-based plant genome size estimation by offering a user-friendly and reliable tool that does not rely on high coverage or reference assemblies. We anticipate that LocoGSE will facilitate plant genome size analysis and contribute to evolutionary and ecological studies in the field. Furthermore, at the cost of an initial calibration, LocoGSE can be used in other lineages.

Multiple genomic solutions for local adaptation in two closely related species (sheep and goats) facing the same climatic constraints.

The question of how local adaptation takes place remains a fundamental question in evolutionary biology. The variation of allele frequencies in genes under selection over environmental gradients remains mainly theoretical and its empirical assessment would help understanding how adaptation happens over environmental clines. To bring new insights to this issue we set up a broad framework which aimed to compare the adaptive trajectories over environmental clines in two domesticated mammal species co-distributed in diversified landscapes. We sequenced the genomes of 160 sheep and 161 goats extensively managed along environmental gradients, including temperature, rainfall, seasonality and altitude, to identify genes and biological processes shaping local adaptation. Allele frequencies at putatively adaptive loci were rarely found to vary gradually along environmental gradients, but rather displayed a discontinuous shift at the extremities of environmental clines. Of the 430 candidate adaptive genes identified, only 6 were orthologous between sheep and goats and those responded differently to environmental pressures, suggesting different putative mechanisms involved in local adaptation in these two closely related species. Interestingly, the genomes of the 2 species were impacted differently by the environment, genes related to signatures of selection were most related to altitude, slope and rainfall seasonality for sheep, and summer temperature and spring rainfall for goats. The diversity of candidate adaptive pathways may result from a high number of biological functions involved in the adaptations to multiple eco-climatic gradients, and a differential role of climatic drivers on the two species, despite their co-distribution along the same environmental gradients. This study describes empirical examples of clinal variation in putatively adaptive alleles with different patterns in allele frequency distributions over continuous environmental gradients, thus showing the diversity of genetic responses in adaptive landscapes and opening new horizons for understanding genomics of adaptation in mammalian species and beyond.

A 2-million-year-old ecosystem in Greenland uncovered by environmental DNA.

Late Pliocene and Early Pleistocene epochs 3.6 to 0.8 million years ago1 had climates resembling those forecasted under future warming2. Palaeoclimatic records show strong polar amplification with mean annual temperatures of 11-19 °C above contemporary values3,4. The biological communities inhabiting the Arctic during this time remain poorly known because fossils are rare5. Here we report an ancient environmental DNA6 (eDNA) record describing the rich plant and animal assemblages of the Kap København Formation in North Greenland, dated to around two million years ago. The record shows an open boreal forest ecosystem with mixed vegetation of poplar, birch and thuja trees, as well as a variety of Arctic and boreal shrubs and herbs, many of which had not previously been detected at the site from macrofossil and pollen records. The DNA record confirms the presence of hare and mitochondrial DNA from animals including mastodons, reindeer, rodents and geese, all ancestral to their present-day and late Pleistocene relatives. The presence of marine species including horseshoe crab and green algae support a warmer climate than today. The reconstructed ecosystem has no modern analogue. The survival of such ancient eDNA probably relates to its binding to mineral surfaces. Our findings open new areas of genetic research, demonstrating that it is possible to track the ecology and evolution of biological communities from two million years ago using ancient eDNA.

High resolution ancient sedimentary DNA shows that alpine plant diversity is associated with human land use and climate change.

The European Alps are highly rich in species, but their future may be threatened by ongoing changes in human land use and climate. Here, we reconstructed vegetation, temperature, human impact and livestock over the past ~12,000 years from Lake Sulsseewli, based on sedimentary ancient plant and mammal DNA, pollen, spores, chironomids, and microcharcoal. We assembled a highly-complete local DNA reference library (PhyloAlps, 3923 plant taxa), and used this to obtain an exceptionally rich sedaDNA record of 366 plant taxa. Vegetation mainly responded to climate during the early Holocene, while human activity had an additional influence on vegetation from 6 ka onwards. Land-use shifted from episodic grazing during the Neolithic and Bronze Age to agropastoralism in the Middle Ages. Associated human deforestation allowed the coexistence of plant species typically found at different elevational belts, leading to levels of plant richness that characterise the current high diversity of this region. Our findings indicate a positive association between low intensity agropastoral activities and precipitation with the maintenance of the unique subalpine and alpine plant diversity of the European Alps.

Postglacial species arrival and diversity buildup of northern ecosystems took millennia.

What drives ecosystem buildup, diversity, and stability? We assess species arrival and ecosystem changes across 16 millennia by combining regional-scale plant sedimentary ancient DNA from Fennoscandia with near-complete DNA and trait databases. We show that postglacial arrival time varies within and between plant growth forms. Further, arrival times were mainly predicted by adaptation to temperature, disturbance, and light. Major break points in ecological trait diversity were seen between 13.9 and 10.8 calibrated thousand years before the present (cal ka BP), as well as break point in functional diversity at 12.0 cal ka BP, shifting from a state of ecosystem buildup to a state where most habitat types and biotic ecosystem components were in place. Trait and functional diversity stabilized around 8 cal ka BP, after which both remained stable, although changes in climate took place and species inflow continued. Our ecosystem reconstruction indicates a millennial-scale time phase of formation to reach stable and resilient levels of diversity and functioning.

The generality of cryptic dietary niche differences in diverse large-herbivore assemblages.

Ecological niche differences are necessary for stable species coexistence but are often difficult to discern. Models of dietary niche differentiation in large mammalian herbivores invoke the quality, quantity, and spatiotemporal distribution of plant tissues and growth forms but are agnostic toward food plant species identity. Empirical support for these models is variable, suggesting that additional mechanisms of resource partitioning may be important in sustaining large-herbivore diversity in African savannas. We used DNA metabarcoding to conduct a taxonomically explicit analysis of large-herbivore diets across southeastern Africa, analyzing ∼4,000 fecal samples of 30 species from 10 sites in seven countries over 6 y. We detected 893 food plant taxa from 124 families, but just two families-grasses and legumes-accounted for the majority of herbivore diets. Nonetheless, herbivore species almost invariably partitioned food plant taxa; diet composition differed significantly in 97% of pairwise comparisons between sympatric species, and dissimilarity was pronounced even between the strictest grazers (grass eaters), strictest browsers (nongrass eaters), and closest relatives at each site. Niche differentiation was weakest in an ecosystem recovering from catastrophic defaunation, indicating that food plant partitioning is driven by species interactions, and was stronger at low rainfall, as expected if interspecific competition is a predominant driver. Diets differed more between browsers than grazers, which predictably shaped community organization: Grazer-dominated trophic networks had higher nestedness and lower modularity. That dietary differentiation is structured along taxonomic lines complements prior work on how herbivores partition plant parts and patches and suggests that common mechanisms govern herbivore coexistence and community assembly in savannas.

Interspecific coprophagia by wild red foxes: DNA metabarcoding reveals a potentially widespread form of commensalism among animals.

Vertebrate animals are known to consume other species' faeces, yet the role of such coprophagy in species dynamics remains unknown, not least due to the methodological challenges of documenting it. In a large-scale metabarcoding study of red fox and pine marten scats, we document a high occurrence of domestic dog DNA in red fox scats and investigate if it can be attributed to interspecific coprophagia. We tested whether experimental artifacts or other sources of DNA could account for dog DNA, regressed dog occurrence in the diet of fox against that of the fox's main prey, short-tailed field voles, and consider whether predation or scavenging could explain the presence of dog DNA. Additionally, we determined the calorific value of dog faeces through calorimetric explosion. The high occurrence of dog DNA in the diet of fox, the timing of its increase, and the negative relationship between dog and the fox's main prey, point to dog faeces as the source of DNA in fox scats. Dog faeces being highly calorific, we found that foxes, but not pine martens, regularly exploit them, seemingly as an alternative resource to fluctuating prey. Scattered accounts from the literature may suggest that interspecific coprophagia is a potentially frequent and widespread form of interaction among vertebrates. However, further work should address its prevalence in other systems and the implications for ecological communities. Tools such as metabarcoding offer a way forward.

Tempo and drivers of plant diversification in the European mountain system.

There is still limited consensus on the evolutionary history of species-rich temperate alpine floras due to a lack of comparable and high-quality phylogenetic data covering multiple plant lineages. Here we reconstructed when and how European alpine plant lineages diversified, i.e., the tempo and drivers of speciation events. We performed full-plastome phylogenomics and used multi-clade comparative models applied to six representative angiosperm lineages that have diversified in European mountains (212 sampled species, 251 ingroup species total). Diversification rates remained surprisingly steady for most clades, even during the Pleistocene, with speciation events being mostly driven by geographic divergence and bedrock shifts. Interestingly, we inferred asymmetrical historical migration rates from siliceous to calcareous bedrocks, and from higher to lower elevations, likely due to repeated shrinkage and expansion of high elevation habitats during the Pleistocene. This may have buffered climate-related extinctions, but prevented speciation along elevation gradients as often documented for tropical alpine floras.

ORTHOSKIM: In silico sequence capture from genomic and transcriptomic libraries for phylogenomic and barcoding applications.

Low-coverage whole genome shotgun sequencing (or genome skimming) has emerged as a cost-effective method for acquiring genomic data in nonmodel organisms. This method provides sequence information on chloroplast genome (cpDNA), mitochondrial genome (mtDNA) and nuclear ribosomal regions (rDNA), which are over-represented within cells. However, numerous bioinformatic challenges remain to accurately and rapidly obtain such data in organisms with complex genomic structures and rearrangements, in particular for mtDNA in plants or for cpDNA in some plant families. Here we introduce the pipeline ORTHOSKIM, which performs in silico capture of targeted sequences from genomic and transcriptomic libraries without assembling whole organelle genomes. ORTHOSKIM proceeds in three steps: (i) global sequence assembly, (ii) mapping against reference sequences and (iii) target sequence extraction; importantly it also includes a range of quality control tests. Different modes are implemented to capture both coding and noncoding regions of cpDNA, mtDNA and rDNA sequences, along with predefined nuclear sequences (e.g., ultraconserved elements) or collections of single-copy orthologue genes. Moreover, aligned DNA matrices are produced for phylogenetic reconstructions, by performing multiple alignments of the captured sequences. While ORTHOSKIM is suitable for any eukaryote, a case study is presented here, using 114 genome-skimming libraries and four RNA sequencing libraries obtained for two plant families, Primulaceae and Ericaceae, the latter being a well-known problematic family for cpDNA assemblies. ORTHOSKIM recovered with high success rates cpDNA, mtDNA and rDNA sequences, well suited to accurately infer evolutionary relationships within these families. ORTHOSKIM is released under a GPL-3 licence and is available at: https://github.com/cpouchon/ORTHOSKIM.

Late Quaternary dynamics of Arctic biota from ancient environmental genomics.

During the last glacial-interglacial cycle, Arctic biotas experienced substantial climatic changes, yet the nature, extent and rate of their responses are not fully understood1-8. Here we report a large-scale environmental DNA metagenomic study of ancient plant and mammal communities, analysing 535 permafrost and lake sediment samples from across the Arctic spanning the past 50,000 years. Furthermore, we present 1,541 contemporary plant genome assemblies that were generated as reference sequences. Our study provides several insights into the long-term dynamics of the Arctic biota at the circumpolar and regional scales. Our key findings include: (1) a relatively homogeneous steppe-tundra flora dominated the Arctic during the Last Glacial Maximum, followed by regional divergence of vegetation during the Holocene epoch; (2) certain grazing animals consistently co-occurred in space and time; (3) humans appear to have been a minor factor in driving animal distributions; (4) higher effective precipitation, as well as an increase in the proportion of wetland plants, show negative effects on animal diversity; (5) the persistence of the steppe-tundra vegetation in northern Siberia enabled the late survival of several now-extinct megafauna species, including the woolly mammoth until 3.9 ± 0.2 thousand years ago (ka) and the woolly rhinoceros until 9.8 ± 0.2 ka; and (6) phylogenetic analysis of mammoth environmental DNA reveals a previously unsampled mitochondrial lineage. Our findings highlight the power of ancient environmental metagenomics analyses to advance understanding of population histories and long-term ecological dynamics.

Altitudinal Zonation of Green Algae Biodiversity in the French Alps.

Mountain environments are marked by an altitudinal zonation of habitat types. They are home to a multitude of terrestrial green algae, who have to cope with abiotic conditions specific to high elevation, e.g., high UV irradiance, alternating desiccation, rain and snow precipitations, extreme diurnal variations in temperature and chronic scarceness of nutrients. Even though photosynthetic green algae are primary producers colonizing open areas and potential markers of climate change, their overall biodiversity in the Alps has been poorly studied so far, in particular in soil, where algae have been shown to be key components of microbial communities. Here, we investigated whether the spatial distribution of green algae followed the altitudinal zonation of the Alps, based on the assumption that algae settle in their preferred habitats under the pressure of parameters correlated with elevation. We did so by focusing on selected representative elevational gradients at distant locations in the French Alps, where soil samples were collected at different depths. Soil was considered as either a potential natural habitat or temporary reservoir of algae. We showed that algal DNA represented a relatively low proportion of the overall eukaryotic diversity as measured by a universal Eukaryote marker. We designed two novel green algae metabarcoding markers to amplify the Chlorophyta phylum and its Chlorophyceae class, respectively. Using our newly developed markers, we showed that elevation was a strong correlate of species and genus level distribution. Altitudinal zonation was thus determined for about fifty species, with proposed accessions in reference databases. In particular, Planophila laetevirens and Bracteococcus ruber related species as well as the snow alga Sanguina genus were only found in soil starting at 2,000 m above sea level. Analysis of environmental and bioclimatic factors highlighted the importance of pH and nitrogen/carbon ratios in the vertical distribution in soil. Capacity to grow heterotrophically may determine the Trebouxiophyceae over Chlorophyceae ratio. The intensity of freezing events (freezing degree days), proved also determinant in Chlorophyceae distribution. Guidelines are discussed for future, more robust and precise analyses of environmental algal DNA in mountain ecosystems and address green algae species distribution and dynamics in response to environmental changes.

Connecting high-throughput biodiversity inventories: Opportunities for a site-based genomic framework for global integration and synthesis.

High-throughput sequencing (HTS) is increasingly being used for the characterization and monitoring of biodiversity. If applied in a structured way, across broad geographical scales, it offers the potential for a much deeper understanding of global biodiversity through the integration of massive quantities of molecular inventory data generated independently at local, regional and global scales. The universality, reliability and efficiency of HTS data can potentially facilitate the seamless linking of data among species assemblages from different sites, at different hierarchical levels of diversity, for any taxonomic group and regardless of prior taxonomic knowledge. However, collective international efforts are required to optimally exploit the potential of site-based HTS data for global integration and synthesis, efforts that at present are limited to the microbial domain. To contribute to the development of an analogous strategy for the nonmicrobial terrestrial domain, an international symposium entitled "Next Generation Biodiversity Monitoring" was held in November 2019 in Nicosia (Cyprus). The symposium brought together evolutionary geneticists, ecologists and biodiversity scientists involved in diverse regional and global initiatives using HTS as a core tool for biodiversity assessment. In this review, we summarize the consensus that emerged from the 3-day symposium. We converged on the opinion that an effective terrestrial Genomic Observatories network for global biodiversity integration and synthesis should be spatially led and strategically united under the umbrella of the metabarcoding approach. Subsequently, we outline an HTS-based strategy to collectively build an integrative framework for site-based biodiversity data generation.

Chloroplast genome assembly of Handroanthus impetiginosus: comparative analysis and molecular evolution in Bignoniaceae.

MAIN CONCLUSION: Bignoniaceae species have conserved chloroplast structure, with hotspots of nucleotide diversity. Several genes are under positive selection, and can be targets for evolutionary studies. Bignoniaceae is one of the most species-rich family of woody plants in Neotropical seasonally dry forests. Here we report the assembly of Handroanthus impetiginosus chloroplast genome and evolutionary comparative analyses of ten Bignoniaceae species representing the genera for which whole-genome chloroplast sequences were available. The chloroplast genome of H. impetiginosus is 159,462 bp in size and has a similar structure compared to the other nine species. The total number of genes was slightly variable amongst the Bignoniaceae, ranging from 124 in H. impetiginosus to 144 in Anemopaegma acutifolium. The inverted repeat (IR) size was variable, ranging from 24,657 bp (Tecomaria capensis) to 40,481 bp (A. acutifolium), due to the contraction and retraction at its boundaries. However, gene boundaries were very similar among the ten species. We found 98 forward and palindromic dispersed repeats, and 85 simple sequence repeats (SSRs). In general, chloroplast sequences were highly conserved, with few nucleotide diversity hotspots in the genes accD, clpP, rpoA, ycf1, ycf2. The phylogenetic analysis based on 77 coding genes was highly consistent with Angiosperm Phylogeny Group (APG) IV. Our results also indicate that most genes are under negative selection or neutral evolution. We found no evidence of branch-site selection, implying that H. impetiginosus is not evolving faster than the other species analyzed, notwithstanding we found site positive selection signal in several genes. These genes can provide targets for evolutionary studies in Bignoniaceae and Lamiales species.