Recent development and implementation of crop cryopreservation protocols has increased the capacity to maintain recalcitrant seeded germplasm collections via cryopreserved in vitro material. To preserve the greatest possible plant genetic resources globally for future food security and breeding programs, it is essential to integrate in situ and ex situ conservation methods into a cohesive conservation plan. In vitro storage using tissue culture and cryopreservation techniques offers promising complementary tools that can be used to promote this approach. These techniques can be employed for crops difficult or impossible to maintain in seed banks for long-term conservation. This includes woody perennial plants, recalcitrant seed crops or crops with no seeds at all and vegetatively or clonally propagated crops where seeds are not true-to-type. Many of the world's most important crops for food, nutrition and livelihoods, are vegetatively propagated or have recalcitrant seeds. This review will look at ex situ conservation, namely field repositories and in vitro storage for some of these economically important crops, focusing on conservation strategies for avocado. To date, cultivar-specific multiplication protocols have been established for maintaining multiple avocado cultivars in tissue culture. Cryopreservation of avocado somatic embryos and somatic embryogenesis have been successful. In addition, a shoot-tip cryopreservation protocol has been developed for cryo-storage and regeneration of true-to-type clonal avocado plants.

Recent developments in the cryopreservation space has increased the trend in germplasm collections established through cryopreserved in vitro material. Cryopreservation of recalcitrant seeds through embryos and embryonic axes, is not uncommon. Tropical and sub-tropical plants are not acclimated to the cold season, therefore have no in-built natural resilience to the cold. Also, larger seeds from trees, such as avocado (Persea americana Mill.), mango (Mangifera indica) and durian (Durio zibethinus L.) are sensitive to desiccation, chilling and freezing stress, making them unsuitable for seed banking or cryopreservation. Alternatively, as seeds do not carry the same genetic make-up as the mother plant, especially in the context of woody rainforest species of which the cross-pollination is dominant; seed conservation does not serve the purpose of germplasm preservation. Other plant material and methods are needed for these plants to be successfully stored in liquid nitrogen (LN). One such method commonly used is shoot-tip cryopreservation which ensures the clonal fidelity of germplasm. There are many problems when using shoot tips of tropical recalcitrant-seeded species. These include: 1) the toxic effects of cryoprotective agents towards structural integrity; 2) optimum developmental stage for success and 3) oxidative stress associated with excision injury leading to necrosis triggering cell death and hindering regeneration for the shoot tips in culture. A pre-requisite for any cryopreservation system is the availability of an established tissue culture regeneration platform. This review will outline conservation strategies for avocado with special emphasis on attempts and improvements made in the cryopreservation space for storing this horticulturally important crop ‘avocado’ at ultra-low temperatures.

Slow rates of molecular evolution at low taxonomic levels hamper studies of relationships among species and subsequent biogeographic and evolutionary analyses. An example is the genus Brahea, which is among the most poorly understood lineages of American palms and is characterized by a wide variety of growth forms and intermediate morphological features. We generated c. 400 kb of genome-scale data from all three genomes for the 11 currently described Brahea spp. to infer phylogenetic relationships, reconstruct ancestral growth form, estimate ancestral geographical ranges and test for niche differences among closely related species with geographical overlap. Relationships receive strong support and conform to previous subgeneric assignments, except for placement of the dwarf species B. moorei in Brahea subgenus Erythea. Our robust phylogenetic hypothesis reveals trends in growth form, including an overall increase in height in the B. armata clade and independent evolution of dwarf forms from taller ancestors in the B. pimo and B. dulcis clades. Ancestral range estimation reveals roles of dispersal (e.g. B. edulis on Guadalupe Island) and putative divergence within geographical regions in some cases (e.g. in the B. armata clade) but is equivocal in others (e.g. in the B. pimo clade). We find evidence of niche non-equivalency among species in the B. armata clade in north-western Mexico and some evidence of niche non-equivalency between B. berlandieri and B. dulcis, the former of which is synonymized under B. dulcis. Our findings have implications for the complex biogeographic history in Central America and Mexico, suggesting that divergence within regions and dispersal are the predominant processes of species diversification. Future studies should include population-level sampling across the genus, along with morphological and ecological information, to assess distinctness among species and, particularly, levels of gene flow, in an integrative fashion.

AbstractBackground and AimsSlow rates of molecular evolution at low taxonomic levels hamper studies of relationships among species, and subsequent biogeographic and evolutionary analyses. An example is the genusBrahea, which is among the most poorly understood lineages of American palms and is characterized by a wide variety of growth forms and intermediate morphological features.MethodsWe generated approximately 400 kb of genome-scale data from all three genomes for the 11 currently described species ofBraheato infer phylogenetic relationships, reconstruct ancestral growth form, estimate ancestral geographic ranges, and test for niche equivalency among closely related species with geographic overlap.Key ResultsRelationships receive strong support, and conform to previous subgeneric assignments, except for placement of the dwarf speciesB. mooreiwithin subgenusErythea.Our robust phylogenetic hypothesis reveals trends in growth form including an overall increase in height in theB. armataclade, and independent evolution of dwarf forms from taller ancestors in theB. pimoandB. dulcisclades. Ancestral range estimation reveals roles of dispersal (e.g.B. edulison Guadalupe Island) and sympatric speciation in some cases (e.g. in theB. armataclade), but is equivocal in others (e.g. in theB. pimo clade). We find evidence of niche non-equivalency among species within theB. armataclade in northwestern Mexico, and some evidence of niche non-equivalency betweenB. berlandieriandB. dulcis, the former of which is synonymized underB. dulcis.ConclusionsOur findings have implications for the complex biogeographic history in Central America and Mexico, suggesting that sympatric speciation and dispersal are the predominant processes of species diversification. Future studies should include population-level sampling across the genus, along with morphological and ecological information, to assess distinctness among species and, particularly, levels of gene flow, in an integrative fashion.

PREMISE OF THE STUDY: The antimicrobial properties and toxicity of Euphorbia plant latex should make it a hostile environment to microbes. However, when specimens from Euphorbia spp. were propagated in tissue culture, microbial growth was observed routinely, raising the question whether the latex of this diverse plant genus can be a niche for polymicrobial communities.METHODS: Latex from a phylogenetically diverse set of Euphorbia species was collected and genomic microbial DNA extracted. Deep sequencing of bar-coded amplicons from taxonomically informative gene fragments was used to measure bacterial and fungal species richness, evenness, and composition.KEY RESULTS: Euphorbia latex was found to contain unexpectedly complex bacterial (mean: 44.0 species per sample; 9 plants analyzed) and fungal (mean: 20.9 species per sample; 22 plants analyzed) communities using culture-independent methods. Many of the identified taxa are known plant endophytes, but have not been previously found in latex.CONCLUSIONS: Our results suggest that Euphorbia plant latex, a putatively hostile antimicrobial environment, unexpectedly supports diverse bacterial and fungal communities. The ecological roles of these microorganisms and potential interactions with their host plants are unknown and warrant further research.

Despite progress based on multilocus, phylogenetic studies of the palms (order Arecales, family Arecaceae), uncertainty remains in resolution/support among major clades and for the placement of the palms among the commelinid monocots. Palms and related commelinids represent a classic case of substitution rate heterogeneity that has not been investigated in the genomic era. To address questions of relationships, support and rate variation among palms and commelinid relatives, 39 plastomes representing the palms and related family Dasypogonaceae were generated via genome skimming and integrated within a monocot-wide matrix for phylogenetic and molecular evolutionary analyses. Support was strong for 'deep' relationships among the commelinid orders, among the five palm subfamilies, and among tribes of the subfamily Coryphoideae. Additionally, there was extreme heterogeneity in the plastid substitution rates across the commelinid orders indicated by model based analyses, with c. 22 rate shifts, and significant departure from a global clock. To date, this study represents the most comprehensively sampled matrix of plastomes assembled for monocot angiosperms, providing genome-scale support for phylogenetic relationships of monocot angiosperms, and lays the phylogenetic groundwork for comparative analyses of the drivers and correlates of such drastic differences in substitution rates across a diverse and significant clade.