Bacterial communities vary across populations and tissue type in red mangroves (Rhizophora mangle, Rhizophoraceae) along an expanding front.

Plant-associated microbial communities may be important sources of functional diversity and genetic variation that influence host evolution. Bacteria provide benefits for their hosts, yet in most plant systems we know little about their taxonomic composition or variation across tissues and host range. Red Mangrove (Rhizophora mangle L.) is a vital coastal plant species that is currently expanding poleward and with it, perhaps, its microbiome. We explored variability in bacterial communities across tissues, individuals, and populations. We collected samples from six sample types from 5 to 10 individuals at each of three populations and used 16S rRNA gene (iTag) sequencing to describe their bacterial communities. Core community members and dominant bacterial classes were determined for each sample type. Pairwise PERMANOVA of Bray-Curtis dissimilarity and Indicator Species Analysis revealed significant differences in bacterial communities between sample types and populations. We described the previously unexplored microbiome of the reproductive tissues of R. mangle. Populations and most sample types were associated with distinct communities. Bacterial communities associated with R. mangle are influenced by host geography and sample type. Our study provides a foundation for future work exploring the functional roles of these microbes and their relevance to biogeochemical cycling.

Digital Extended Specimens: Enabling an Extensible Network of Biodiversity Data Records as Integrated Digital Objects on the Internet.

The early twenty-first century has witnessed massive expansions in availability and accessibility of digital data in virtually all domains of the biodiversity sciences. Led by an array of asynchronous digitization activities spanning ecological, environmental, climatological, and biological collections data, these initiatives have resulted in a plethora of mostly disconnected and siloed data, leaving to researchers the tedious and time-consuming manual task of finding and connecting them in usable ways, integrating them into coherent data sets, and making them interoperable. The focus to date has been on elevating analog and physical records to digital replicas in local databases prior to elevating them to ever-growing aggregations of essentially disconnected discipline-specific information. In the present article, we propose a new interconnected network of digital objects on the Internet-the Digital Extended Specimen (DES) network-that transcends existing aggregator technology, augments the DES with third-party data through machine algorithms, and provides a platform for more efficient research and robust interdisciplinary discovery.

Mobilizing the community of biodiversity specimen collectors to effectively detect and document outliers in the Anthropocene.

PREMISE: Biological outliers (observations that fall outside of a previously understood norm, e.g., in phenology or distribution) may indicate early stages of a transformative change that merits immediate attention. Collectors of biodiversity specimens such as plants, fungi, and animals are on the front lines of discovering outliers, yet the role collectors currently play in providing such data is unclear. METHODS: We surveyed 222 collectors of a broad range of taxa, searched 47 training materials, and explored the use of 170 outlier terms in 75 million specimen records to determine the current state of outlier detection and documentation in this community. RESULTS: Collectors reported observing outliers (e.g., about 80% of respondents observed morphological and distributional outliers at least occasionally). However, relatively few specimen records include outlier terms, and imprecision in their use and handling in data records complicates data discovery by stakeholders. This current state appears to be at least partly due to the absence of protocols: only one of the training materials addressed documenting and reporting outliers. CONCLUSIONS: We suggest next steps to mobilize this largely untapped, yet ideally suited, community for early detection of biotic change in the Anthropocene, including community activities for building relevant best practices.

Worldwide Engagement for Digitizing Biocollections (WeDigBio): The Biocollections Community's Citizen-Science Space on the Calendar.

The digitization of biocollections is a critical task with direct implications for the global community who use the data for research and education. Recent innovations to involve citizen scientists in digitization increase awareness of the value of biodiversity specimens; advance science, technology, engineering, and math literacy; and build sustainability for digitization. In support of these activities, we launched the first global citizen-science event focused on the digitization of biodiversity specimens: Worldwide Engagement for Digitizing Biocollections (WeDigBio). During the inaugural 2015 event, 21 sites hosted events where citizen scientists transcribed specimen labels via online platforms (DigiVol, Les Herbonautes, Notes from Nature, the Smithsonian Institution's Transcription Center, and Symbiota). Many citizen scientists also contributed off-site. In total, thousands of citizen scientists around the world completed over 50,000 transcription tasks. Here, we present the process of organizing an international citizen-science event, an analysis of the event's effectiveness, and future directions-content now foundational to the growing WeDigBio event.

The phylogeny and biogeography of Hakea (Proteaceae) reveals the role of biome shifts in a continental plant radiation.

The frequency of evolutionary biome shifts during diversification has important implications for our ability to explain geographic patterns of plant diversity. Recent studies present several examples of biome shifts, but whether frequencies of biome shifts closely reflect geographic proximity or environmental similarity of biomes remains poorly known. We explore this question by using phylogenomic methods to estimate the phylogeny of Hakea, a diverse Australian genus occupying a wide range of biomes. Model-based estimation of ancestral regions indicates that Hakea began diversifying in the Mediterranean biome of southern Australia in the Middle Eocene-Early Oligocene, and dispersed repeatedly into other biomes across the continent. We infer around 47 shifts between biomes. Frequencies of shifts between pairs of biomes are usually similar to those expected from their geographic connectedness or climatic similarity, but in some cases are substantially higher or lower than expected, perhaps reflecting how readily key physiological traits can be modified to adapt lineages to new environments. The history of frequent biome-shifting is reflected in the structure of present-day assemblages, which tend to be more phylogenetically diverse than null-model expectations. The case of Hakea demonstrates that the radiation of large plant clades across wide geographic areas need not be constrained by dispersal limitation or conserved adaptations to particular environments.

Paraphyly changes understanding of timing and tempo of diversification in subtribe Hakeinae (Proteaceae), a giant Australian plant radiation.

PREMISE OF THE STUDY: Subtribe Hakeinae (526 spp.) represents a large Australian plant radiation central to our understanding of that flora's evolution and ecology. It contains Grevillea-the third largest plant genus in Australia and a group inferred to have among the highest diversification rates in the angiosperms. However, we lack a robust phylogenetic framework for understanding subtribe Hakeinae and recognize that Grevillea lacks an unambiguous synapomorphy supporting its monophyly. METHODS: We used four plastid and one nuclear DNA region from a taxonomically even sampling of a third of the species to infer a time-calibrated phylogeny of Hakeinae and absolute diversification rates of major clades. We developed the R package addTaxa to add unsampled taxa to the tree for diversification rate inference. KEY RESULTS: Grevillea is paraphyletic with respect to Hakea and Finschia. Under most parameter combinations, Hakea contains the major clade with the highest diversification rate in Hakeinae, rather than Grevillea. The crown age of the Grevillea+Hakea+Finschia crown group is about double that of prior estimates. CONCLUSIONS: We demonstrate that the paraphyly of Grevillea considerably enlarges the number of Australian descendants from its most recent common ancestor but has also misled investigators who considered a single operational taxonomic unit as adequate to represent the genus for inferences of diversification rate and timing. Our time-calibrated phylogeny can form the basis of future evolutionary, comparative ecology, and biogeography studies involving this large Australian plant radiation, as well as nomenclatural changes.

Digitization workflows for flat sheets and packets of plants, algae, and fungi.

Effective workflows are essential components in the digitization of biodiversity specimen collections. To date, no comprehensive, community-vetted workflows have been published for digitizing flat sheets and packets of plants, algae, and fungi, even though latest estimates suggest that only 33% of herbarium specimens have been digitally transcribed, 54% of herbaria use a specimen database, and 24% are imaging specimens. In 2012, iDigBio, the U.S. National Science Foundation's (NSF) coordinating center and national resource for the digitization of public, nonfederal U.S. collections, launched several working groups to address this deficiency. Here, we report the development of 14 workflow modules with 7-36 tasks each. These workflows represent the combined work of approximately 35 curators, directors, and collections managers representing more than 30 herbaria, including 15 NSF-supported plant-related Thematic Collections Networks and collaboratives. The workflows are provided for download as Portable Document Format (PDF) and Microsoft Word files. Customization of these workflows for specific institutional implementation is encouraged.

Finding our way through phenotypes.

Despite a large and multifaceted effort to understand the vast landscape of phenotypic data, their current form inhibits productive data analysis. The lack of a community-wide, consensus-based, human- and machine-interpretable language for describing phenotypes and their genomic and environmental contexts is perhaps the most pressing scientific bottleneck to integration across many key fields in biology, including genomics, systems biology, development, medicine, evolution, ecology, and systematics. Here we survey the current phenomics landscape, including data resources and handling, and the progress that has been made to accurately capture relevant data descriptions for phenotypes. We present an example of the kind of integration across domains that computable phenotypes would enable, and we call upon the broader biology community, publishers, and relevant funding agencies to support efforts to surmount today's data barriers and facilitate analytical reproducibility.

Five task clusters that enable efficient and effective digitization of biological collections.

This paper describes and illustrates five major clusters of related tasks (herein referred to as task clusters) that are common to efficient and effective practices in the digitization of biological specimen data and media. Examples of these clusters come from the observation of diverse digitization processes. The staff of iDigBio (The U.S. National Science Foundation's National Resource for Advancing Digitization of Biological Collections) visited active biological and paleontological collections digitization programs for the purpose of documenting and assessing current digitization practices and tools. These observations identified five task clusters that comprise the digitization process leading up to data publication: (1) pre-digitization curation and staging, (2) specimen image capture, (3) specimen image processing, (4) electronic data capture, and (5) georeferencing locality descriptions. While not all institutions are completing each of these task clusters for each specimen, these clusters describe a composite picture of digitization of biological and paleontological specimens across the programs that were observed. We describe these clusters, three workflow patterns that dominate the implemention of these clusters, and offer a set of workflow recommendations for digitization programs.

Time-calibrated phylogeny of the woody Australian genus Hakea (Proteaceae) supports multiple origins of insect-pollination among bird-pollinated ancestors.

PREMISE OF THE STUDY: A past study based on morphological data alone showed that the means by which plants of the Australian genus Hakea reduce florivory is related to the evolution of bird pollination. For example, bird pollination was shown to have arisen only in insect-pollinated lineages that already produced greater amounts of floral cyanide, a feature that reduces florivory. We examine a central conclusion of that study, and a common assumption in the literature, that bird pollination arose in insect-pollinated lineages, rather than the reverse. METHODS: We combined morphological and DNA data to infer the phylogeny and age of the Australian genus Hakea, using 9.2 kilobases of plastid and nuclear DNA and 46 morphological characters from a taxonomically even sampling of 55 of the 149 species. KEY RESULTS: Hakea is rooted confidently in a position that has not been suggested before. The phylogeny implies that bird pollination is primitive in Hakea and that multiple shifts to insect pollination have occurred. The unexpectedly young age of Hakea (a crown age of ca. 10 Ma) makes it coincident with its primary bird pollinators (honeyeaters) throughout its history. CONCLUSIONS: Our study demonstrates that Hakea is an exception to the more commonly described shift from insect to bird pollination. However, we note that only one previous phylogenetic study involved Australian plants and their honeyeater pollinators and that our finding might prove to be more common on that continent.

Contrasted patterns of hyperdiversification in Mediterranean hotspots.

Dating the Tree of Life has now become central to relating patterns of biodiversity to key processes in Earth history such as plate tectonics and climate change. Regions with a Mediterranean climate have long been noted for their exceptional species richness and high endemism. How and when these biota assembled can only be answered with a good understanding of the sequence of divergence times for each of their components. A critical aspect of dating by using molecular sequence divergence is the incorporation of multiple suitable age constraints. Here, we show that only rigorous phylogenetic analysis of fossil taxa can lead to solid calibration and, in turn, stable age estimates, regardless of which of 3 relaxed clock-dating methods is used. We find that Proteaceae, a model plant group for the Mediterranean hotspots of the Southern Hemisphere with a very rich pollen fossil record, diversified under higher rates in the Cape Floristic Region and Southwest Australia than in any other area of their total distribution. Our results highlight key differences between Mediterranean hotspots and indicate that Southwest Australian biota are the most phylogenetically diverse but include numerous lineages with low diversification rates.

Origin, adaptive radiation and diversification of the Hawaiian lobeliads (Asterales: Campanulaceae).

The endemic Hawaiian lobeliads are exceptionally species rich and exhibit striking diversity in habitat, growth form, pollination biology and seed dispersal, but their origins and pattern of diversification remain shrouded in mystery. Up to five independent colonizations have been proposed based on morphological differences among extant taxa. We present a molecular phylogeny showing that the Hawaiian lobeliads are the product of one immigration event; that they are the largest plant clade on any single oceanic island or archipelago; that their ancestor arrived roughly 13 Myr ago; and that this ancestor was most likely woody, wind-dispersed, bird-pollinated, and adapted to open habitats at mid-elevations. Invasion of closed tropical forests is associated with evolution of fleshy fruits. Limited dispersal of such fruits in wet-forest understoreys appears to have accelerated speciation and led to a series of parallel adaptive radiations in Cyanea, with most species restricted to single islands. Consistency of Cyanea diversity across all tall islands except Hawai ;i suggests that diversification of Cyanea saturates in less than 1.5 Myr. Lobeliad diversity appears to reflect a hierarchical adaptive radiation in habitat, then elevation and flower-tube length, and provides important insights into the pattern and tempo of diversification in a species-rich clade of tropical plants.

A smaller Macadamia from a more vagile tribe: inference of phylogenetic relationships, divergence times, and diaspore evolution in Macadamia and relatives (tribe Macadamieae; Proteaceae).

Tribe Macadamieae (91 spp., 16 genera; Proteaceae) is widespread across the southern hemisphere on all major fragments of Gondwana except New Zealand and India. Macadamia is cultivated outside its natural range as a "nut" crop (notably in Hawaii, where it is the principal orchard crop). We sampled seven DNA regions and 53 morphological characters from the tribe to infer its phylogeny and address the common assumption that the distribution of the extant diversity of the tribe arose by the rafting of ancestors on Gondwanan fragments. Macadamia proves to be paraphyletic with respect to the African genus Brabejum, the South American genus Panopsis, and the Australian species Orites megacarpus. We erect two new generic names, Nothorites and Lasjia, to produce monophyly at that rank. The earliest disjunctions in the tribe are inferred to be the result of long-distance dispersal out of Australia (with one possible exception), rather than vicariance. Evolution of tardy fruit dehiscence is correlated with these dispersals, and the onset of the Antarctic Circumpolar Current (ACC) precedes them. We suggest that the ancestors of extant diversity arrived on their respective continents via the ACC, and we recognize that this is a mechanism precluded, rather than facilitated, by Gondwana's terrestrial continuity.

Are any primroses (Primula) primitively monomorphic?

Primula (c. 430 species) and relatives (Primulaceae) are paradigmatic to our understanding of distyly. However, the common co-occurrence of distyly and monomorphy in closely related groups within the family has made the interpretation of its evolution difficult.Here, we infer a chloroplast DNA (cpDNA) phylogeny for 207 accessions, including 51% of the species and 95% of the sections of Primula with monomorphic populations, using Bayesian methods. With this tree, we infer the distribution of ancestral states on critical nodes using parsimony and likelihood methods. The inferred cpDNA phylogeny is consistent with prior estimates. The most recent common ancestor (MRCA) of Primula is resolved as distylous using both methods of inference. However, whether the distyly in Primula, Hottonia, and Vitaliana arose once or three independent times is not clear. We conclude that monomorphism in descendants of the MRCA of Primula is derived from distyly in all cases. Thus, scenarios for the evolution of distyly that rely on the persistence of primitive monomorphy (such as in Primula section Sphondylia) require re-evaluation.

Buzz-pollinated Dodecatheon originated from within the heterostylous Primula subgenus Auriculastrum (Primulaceae): a seven-region cpDNA phylogeny and its implications for floral evolution.

We sequenced seven cpDNA regions from 70 spp. in Dodecatheon, Primula subgenus Auriculastrum, and outgroups, reconstructed their cpDNA phylogeny with maximum parsimony, and determined branch support with bootstrap frequencies and Bayesian posterior probabilities. Strongly supported conclusions include the (1) paraphyly of Primula subgenus Auriculastrum with respect to a monophyletic Dodecatheon, (2) sister relationship between the North American Dodecatheon and the Californian P. suffrutescens, (3) novel basal split in Dodecatheon to produce one clade with rugose and one clade with smooth anther connectives, (4) monophyly of all sections of Primula subgenus Auriculastrum, and (5) exclusion of the enigmatic Primula section Amethystina from the similar Primula subgenus Auriculastrum. These results support the origin of the monomorphic, buzz-pollinated flower of Dodecatheon from the heterostylous flower of Primula. We marshal evidence to support the novel hypothesis that the solanoid flower of Dodecatheon represents the fixation of recessive alleles at the heterostyly linkage group (pin phenotype). Of the remaining traits associated with their solanoid flowers, we recognize at least six likely to have arisen with the origin of Dodecatheon, one that preceded it (flower coloration, a transfer exaptation in Dodecatheon), and one that followed it (rugose anther connectives, an adaptation to buzz pollination).

Historical biogeography and the origin of stomatal distributions in Banksia and Dryandra (Proteaceae) based on their cpDNA phylogeny.

Banksia and Dryandra have undergone extensive speciation and adaptive radiation, especially in Australia's isolated Southwest Botanical Province. We derive a phylogeny for these groups based on cpDNA sequences and use it to reconstruct their historical biogeography and evolution of leaf traits thought to be adapted to drought and/or nutrient poverty. Slowly evolving regions (trnL intron, trnL/trnF spacer) are used to resolve large-scale relationships; faster evolving regions (rp116 intron, psbA/trnH and trnT/trnL spacers) are used to resolve relationships among closely related species. Banksia is paraphyletic with respect to Dryandra. The lineage underwent a basal split into two clades (here named /Cryptostomata and /Phanerostomata), and four infrageneric taxa supported by morphological cladistic analyses (series Spicigerae, Abietinae, Tetragonae, and Banksia) are not monophyletic. Dispersal-vicariance analysis resolves a southwestern Australian origin for the lineage, with two later expansions to the east followed by vicariance events. Stomatal crypts arose with the /Cryptostomata, which is characterized by tough, long-lived leaves and common in southwestern Australia. Sequestering of stomata also arose multiple times in /Phanerostomata, which is characterized by softer, short-lived leaves and common in moister coastal areas, via inrolling of the margins of narrow leaves and restricting stomata to shallow pits. The hypothesis that sclerophylly preadapted the plants to xeromorphy is supported in the case of shallow stomatal pits and deep stomatal crypts, but not narrow, needle-like leaves.