Ludo A. H. Muller

Genomic Resources Notes accepted 1 April 2015 – 31 May 2015

This article documents the public availability of transcriptomic resources for (i) the stellate sturgeon Acipenser stellatus, (ii) the flowering plant Campanula gentilis and (iii) two endemic Iberian fish, Squalius carolitertii and Squalius torgalensis.

Soil aggregates as massively concurrent evolutionary incubators

Soil aggregation, a key component of soil structure, has mostly been examined from the perspective of soil management and the mediation of ecosystem processes such as soil carbon storage. However, soil aggregation is also a major factor to consider in terms of the fine-scale organization of the soil microbiome. For example, the physico-chemical conditions inside of aggregates usually differ from the conditions prevalent in the bulk soil and aggregates therefore increase the spatial heterogeneity of the soil. In addition, aggregates can provide a refuge for microbes against predation since their interior is not accessible to many predators. Soil aggregates are thus clearly important for microbial community ecology in soils (for example, Vos et al., 2013; Rillig et al., 2016) and for microbially driven biogeochemistry, and soil microbial ecologists are increasingly appreciating these aspects of soil aggregation. Soil aggregates have, however, so far been neglected when it comes to evolutionary considerations (Crawford et al., 2005) and we here propose that the process of soil aggregation should be considered as an important driver of evolution in the soil microbial community. There are several features that make soil aggregates specifically interesting, and perhaps even unique, in terms of a setting for microbial evolution (Table 1). Soil aggregation is a continuous and dynamic process in which the formation and disintegration of individual microand macroaggregates are separated in time by periods of relative stability. Each individual soil aggregate may provide a unique environmental compartmentalization of the soil microbial community that is, to a large extent, isolated from its surroundings and that can be thought of as an ‘incubator’ for microbial evolutionary change. Because of their isolation, different aggregates can be regarded as ‘concurrent incubators’ that allow enclosed microbial communities to pursue their own independent evolutionary trajectories during their lifetime (‘incubation period’). The huge number of aggregates that exists at any moment in time validates their conceptualization as ‘massively concurrent incubators’ for microbial evolutionary change (Figure 1). Upon disintegration of soil aggregates (‘incubation cycle ends’), formerly enclosed microbial communities are released and allowed to interact with the microbial community of the soil at large. This combination of features (isolation, large number and relative stability) sets soil aggregates apart from other microbial habitats that may also provide temporary isolation of microbial communities, such as the animal intestinal tract and other parts of the animal body (see also Cordero and Datta, 2016), leaves, roots, and many aquatic habitats (Table 1). However, these habitats do not provide the same combination of extent of isolation, duration of isolation and number of concurrent ‘incubators’ as soil aggregates do. We discuss these specific characteristics of soil aggregates next, before describing how evolutionary change in aggregates can occur and explaining how this system can be tackled empirically.

Effects of mountain formation and uplift on biological diversity

The formation and uplift of mountain ranges constitute major geological phenomena that can have long-lasting effects on the evolutionary diversification of living organisms. They provide opportunities for adaptive evolution through an increase of spatial heterogeneity of the landscape, including elevation, and the generation of a wide variety of ecologically diverse biotopes, and affect the migration of organisms and the distribution of species since mountain ranges can act as both biological corridors and ecological barriers. Hence, it should come as no surprise that mountain ranges host a large proportion of the biological diversity on earth (Barthlott et al., 2007; Jenkins et al., 2013). The reviews of Wen et al. (2014) and Luebert and Weigend (2014) included in this Research Topic present accounts of plant diversification processes in two major mountain regions of the world: the Andes and the Qinghai-Tibetan Plateau. Both studies show that plant diversifications have occurred in relatively recent geological times, primarily since the Middle Miocene, and therefore followed the late uplift phases of the high mountain ranges of both the Andes and the Qinghai-Tibetan region during the last 15–20 million years (Garzione et al., 2008; Wang et al., 2008). It is thus likely that the formations of those mountain ranges are at least partially responsible for the observed diversification processes, as proposed in other, more recent studies (e.g., Favre et al., 2015; Sanchez-Baracaldo and Thomas, 2014). Rapid diversification processes are documented for the Paramo clade of the plant genus Hypericum in the northern Andes (Nurk et al., 2013). The high diversity of this group originated recently (2.3–5.6 mya) and its diversification rate is well above the background diversification of Hypericum. This pattern of diversification is also seen in other endemic plant groups of the Paramo flora, and the region has the highest average diversification rate among all biodiversity hotspots in the world (Madrinan et al., 2013). Given the similar ages of these lineages (Luebert and Weigend, 2014) and of the high-elevation environments of the Paramo (Mora et al., 2010), these diversifications may have been triggered by the formation of high-mountain habitats in the northern Andes. Increased speciation rates would have occurred along with mountain uplift and habitat diversification, as observed in other Andean groups such as hummingbirds (Chaves et al., 2011) and butterflies (Despland, 2014), confirming the ideas initially proposed by Simpson (1975). Similar diversifications as those observed in the Paramo ecosystem have also occurred in plants of the Qinghai-Tibetan Plateau (Wen et al., 2014), but more studies are necessary to gain insights into any large-scale pattern (Favre et al., 2015). Mutke et al. (2014) report the distribution patterns of four Andean plant groups to reflect habitat heterogeneity rather than uplift history or barrier effects of mountain ranges, supporting, at least partially, the hypothesis that direct drivers of plant diversification in both the Qinghai-Tibetan Plateau and the Andes include plant-pollinator interactions, local adaptation to diverse environmental conditions and polyploidization (Luebert and Weigend, 2014; Wen et al., 2014). Although not reported in this Research Topic, the occurrence of polyploidization has been shown for the European-centered plant genus Campanula. Polyploid species of this genus are concentrated in the Campanula rotundifolia-complex, a mountain clade of Pliocene origin (Mansion et al., 2012). The significance of plant-pollinator interactions for the isolation of plant populations and plant diversification in mountain ranges, on the other hand, has been shown for three Penstemon species by Kramer et al. (2010). The different studies reported in this Research Topic clearly illustrate the potential effects of mountain uplift and formation on species diversification, at least in two major mountain regions of the world. A synthesis of biological diversification on mountains is, however, still far from being achieved and the potentially high complexity of the involved history, geography and biological processes encourages further research (Hoorn et al., 2013; Favre et al., 2015; Luebert and Weigend, 2014; Wen et al., 2014). Nevertheless, we hope that the collection of papers in this Research Topic will be of interest to scientists and will stimulate development of new studies and syntheses. We sincerely thank the authors and the reviewers for their efforts and contributions that made this Research Topic possible.

Analysis of nuclear microsatellites reveals limited differentiation between Colchic and Hyrcanian populations of the wind-pollinated relict tree Zelkova carpinifolia (Ulmaceae)

UNLABELLED • PREMISE OF THE STUDY The Caucasus represents one of the worlds biodiversity hotspots and includes the climatic refugia Hyrcan on the southern coast of the Caspian Sea and Colchis on the eastern coast of the Black Sea, where different species survived during the Quaternary climatic oscillations. We evaluated the genetic diversity of the relict tree Zelkova carpinifolia shared between the two refugia and distributed throughout the Caucasus and adjacent areas.• METHODS Specimens were collected from 30 geographical sites in Azerbaijan, Georgia, Iran, and Turkey and screened for variability at eight nuclear microsatellite loci. The genetic diversity among and within populations was assessed using a set of statistical measures.• KEY RESULTS We detected 379 different genotypes from a total of 495 individuals with varying degrees of clonal reproduction at the different sites. Low to intermediate levels of genetic diversity were observed at all sites, and strong differentiation between sampling sites was absent. In addition, we observed no clear genetic differentiation between the Colchis and Hyrcan. Bayesian clustering of the genotypes revealed three populations with high levels of admixture between the sampling sites.• CONCLUSIONS The lack of strong genetic structure of studied populations of Z. carpinifolia contrasts with a previous study based on chloroplast markers and suggests that long-distance pollen dispersal is an important factor of gene flow among populations of Z. carpinifolia. The present study does not reveal any particular site with particularly isolated genotypes that would deserve more attention for conservation purposes than others, although some sites should be considered for further investigation.

Evidence for nonallopatric speciation among closely related sympatric Heliotropium species in the Atacama Desert

The genetic structure of populations of closely related, sympatric species may hold the signature of the geographical mode of the speciation process. In fully allopatric speciation, it is expected that genetic differentiation between species is homogeneously distributed across the genome. In nonallopatric speciation, the genomes may remain undifferentiated to a large extent. In this article, we analyzed the genetic structure of five sympatric species from the plant genus Heliotropium in the Atacama Desert. We used amplified fragment length polymorphisms (AFLPs) to characterize the genetic structure of these species and evaluate their genetic differentiation as well as the number of loci subject to positive selection using divergence outlier analysis (DOA). The five species form distinguishable groups in the genetic space, with zones of overlap, indicating that they are possibly not completely isolated. Among-species differentiation accounts for 35% of the total genetic differentiation (FST = 0.35), and FST between species pairs is positively correlated with phylogenetic distance. DOA suggests that few loci are subject to positive selection, which is in line with a scenario of nonallopatric speciation. These results support the idea that sympatric species of Heliotropium sect. Cochranea are under an ongoing speciation process, characterized by a fluctuation of population ranges in response to pulses of arid and humid periods during Quaternary times.

Fourteen polymorphic microsatellite markers for the threatened Arnica montana (Asteraceae).

Premise of the study: Microsatellite markers were developed to investigate population genetic structure in the threatened species Arnica montana. Methods and Results: Fourteen microsatellite markers with di-, tetra-, and hexanucleotide repeat motifs were developed for A. montana using 454 pyrosequencing without and with library-enrichment methods, resulting in 56,545 sequence reads and 14,467 sequence reads, respectively. All loci showed a high level of polymorphism, with allele numbers ranging from four to 11 in five individuals from five populations (25 samples) and an expected heterozygosity ranging from 0.192 to 0.648 across the loci. Conclusions: This set of microsatellite markers is the first one described for A. montana and will facilitate conservation genetic applications as well as the understanding of phylogeographic patterns in this species.

Development of Nuclear Microsatellites for the Arcto-Tertiary Tree Zelkova carpinifolia (Ulmaceae) Using 454 Pyrosequencing

Premise of the study: The current study aimed at developing nuclear microsatellite markers for the relict tree species Zelkova carpinifolia, which is threatened in its natural range in the South Caucasus. Methods and Results: Pyrosequencing of an enriched microsatellite library on the Roche FLX platform using the 454 Titanium kit produced 86,058 sequence reads, most of which contained short tandem repeats. Eighty microsatellite loci identified using the software package QDD version 1 were selected and tested for proper PCR amplification. Of these, 13 allowed proper amplification and were shown to be polymorphic among a sample of 25 Z. carpinifolia specimens from various geographic origins. Conclusions: The set of microsatellite markers will be suitable for the assessment of genetic diversity in Z. carpinifolia. They will allow for an examination of phylogeographic patterns as well as of population structure and gene flow within this species.

Phylogeography and population genetics of the riparian relict tree Pterocarya fraxinifolia (Juglandaceae) in the South Caucasus

We aimed to (i) assess the extant genetic diversity of the riparian relict tree Pterocarya fraxinifolia across its current distribution range in the South Caucasus, including the past refugial areas Colchis and Hyrcan, and (ii) test if a separation of these areas is reflected in its phylogeographic history. Genetic diversity of natural populations was examined using nuclear microsatellite and plastid DNA markers. Spatial genetic structure was evaluated using Bayesian clustering methods and the reconstruction of plastid DNA networks. Divergence times of Colchic and Hyrcanian populations were estimated via divergence dating using a relaxed molecular clock. Allelic richness, private allelic richness, and expected heterozygosity were significantly higher in Hyrcan than in Colchis and the Greater Caucasus, and significant genetic differentiation was revealed between the two groups. Whereas only two plastid haplotypes were detected for the Colchic and Caucasian populations, the Hyrcanian populations displayed 11 different haplotypes. Significant isolation by distance was detected in Hyrcan. The most recent common ancestor of all P. fraxinifolia haplotypes was dated to a time well before a suggested glaciation period in the Caucasus during the late Pliocene (5.98 Ma [11.3–2.48 Ma HPD]). The widespread Colchic haplotype that also occurs along the southern slope of the Greater Caucasus and reaches south-eastern Azerbaijan has appeared more recently (0.24 Ma [1.41–0 Ma HPD]). This diversification pattern of Colchic haplotypes from ancient Hyrcanian haplotypes suggests a colonization of the region from south-east to north-west that predates the last glacial maximum (LGM). Natural populations of P. fraxinifolia show low-to-intermediate levels of genetic diversity and a significant decrease of diversity from Hyrcan to Colchis. However, the genetic differentiation between Colchic-Caucasian and Hyrcanian populations for nuclear markers suggests that independent gene pools existed in both areas at least since the LGM. Particular attention to conservation seems justified for the more diverse Hyrcanian populations.

Development of Microsatellite Markers for Crepis mollis (Asteraceae)

Premise of the study: Polymorphic microsatellite markers were developed for the threatened species Crepis mollis (Asteraceae) to investigate population and conservation genetics. Methods and Results: Illumina sequencing was conducted on pooled genomic DNA from 10 individuals of two populations. Ten polymorphic and 10 monomorphic microsatellite loci with di-, tri-, tetra-, penta-, and hexanucleotide repeat motifs were developed and characterized in C. mollis. In the polymorphic markers, up to 17 alleles per locus were detected with an observed and expected heterozygosity ranging from 0.120 to 0.780 and 0.102 to 0.834, respectively. Furthermore, the polymorphic markers were tested for cross-amplification in three congeneric species (C. biennis, C. foetida, and C. sancta) and amplified in up to three loci. Conclusions: The markers developed in this study are the first microsatellites tested on C. mollis and will be useful for performing population and conservation genetic studies in this threatened species.