Valerie C. Pence

Culture tubes are commonly used for the in vitro growth of plant tissues. Plants grown in culture tubes can vary in phenotype within the tube, with tissues nearer the surface of the medium being more hyperhydric than those nearer the cap. This is most common in unvented tubes and with medium gelled with gellan gum rather than agar. This study examined whether a gradient in humidity or water availability exists within the culture tube that might explain differences in growth related to distance from the medium. To measure humidity, microsensors were placed at three levels within tubes capped with solid or vented caps with gel medium and repeated with agar medium. Readings were compared with ambient levels outside of the tube for six weeks, under conditions of 16 hrs light:8 hrs dark each day. Humidity in the unvented tubes was mostly saturated. In contrast, humidity in the vented tubes showed a strong vertical gradient. In both vented and unvented tubes, humidity near the medium was higher than near the cap. The experiments with agar and gel gave similar results. To examine water availability, uptake into cellulose or cotton plugs was measured. Water uptake was significantly greater from gel than from agar medium, and only agar showed a significant gradient in the plugs through that time period, regardless of venting. Both the humidity gradients and water uptake differences suggest that one or both may account for differences in exposure of the tissues to water and could affect phenotype depending on the sensitivity of the species and distance of the tissue from the medium.

The conservation of threatened exceptional plants, which cannot be conserved by seed banking, requires in vitro technologies for many of the approaches needed for their long-term ex situ conservation. This study evaluated the current in vitro plant literature, as represented in Web of Science, to determine its taxonomic overlap with the families and genera of the 775 species currently listed as exceptional. Web of Science was searched using the terms micropropagation, somatic embryogenesis, zygotic embryo, and cryopreservation, and the target genera and families were identified in the more than 19,000 articles evaluated. There were five families with significant overlap between the in vitro literature and exceptional species: Fabaceae, Asteraceae, Orchidaceae, Arecaceae, and Rutaceae. However, there was less overlap at the level of genus, with Citrus, Coffea, and Quercus having the most articles. Significant gaps were also found, with 14 exceptional families and half of the exceptional genera having no representation in the Web of Science search results. The 20 exceptional species with the most articles were all economically important species, and these had 343 threatened congeners that could be prioritized for research. A highly important group of exceptional plants that was significantly under-represented in the literature was tropical woody species, which form the backbone of the diversity of the world’s threatened rainforests. Overall, there are areas of strength upon which to build future work, but significant gaps where research should be prioritized for effectively conserving exceptional plants.

Worldwide, oak species are threatened with extinction due to habitat loss, pathogens, and changing fire regimes. Ex situ conservation through tissue culture may protect the remaining genetic diversity of Quercus dumosa, or the coastal sage scrub oak, from further loss. We designed three basal salt formulations based on the mineral composition of shoot tips and first leaves from mature Q. dumosa and explored carbohydrate source, stress-mitigating compounds, and plant growth regulator concentrations to develop a method of cultivating many Q. dumosa culture lines in vitro. All three novel basal salt formulations led to decreased necrosis compared with commercial basal salt formulas WPM, MS, and DKW. Substitution of 30 g L⁻¹ sucrose with glucose and adding 250 mg L⁻¹ ascorbic acid, 5.2 mg L⁻¹ SNP sodium nitroprusside, and 103 mg L⁻¹ y-aminobutyric acid improved culture health overall. In an experiment involving 115 culture lines, 0.66 mg L⁻¹ 6-benzylaminopurine produced the highest average shoots per explant, but 0.33 mg L⁻¹ produced the greatest proportion of shoots 2 cm or greater. Incubation for 24 h in 20 mg L⁻¹ indole-3-butyric acid led to the most rooting. These methods show promise for the ex situ conservation of many genotypes of endangered Q. dumosa.

Introductions are a critical tool in the recovery of many imperiled species, yet adequate evaluation and development of best practices has lagged. Importantly, long‐term post‐introduction data are typically lacking, as well as suitable comparisons to wild populations to provide a baseline against which to assess performance. Here, we report on three experimental introductions of Crotalaria avonensis (Fabaceae), a federally endangered perennial herb that is narrowly endemic to scrub of the Lake Wales Ridge in peninsular Florida, U.S.A. We synthesize 10 years of post‐introduction monitoring at both the introduced and a nearby, protected wild population to (1) develop best practices for conservation, and (2) evaluate the success of the introduction. First, our study identified best practices that included using transplants propagated from stem cuttings, as well as several factors that may increase seed germination such as habitat choice, seed burial, and litter addition. Second, during the 10 years following the introduction, population density in the introduced population was higher than in a nearby protected, wild population, and a comparison of vital rates revealed that this result was due to relatively high clonal and seedling recruitment rates in the introduced population. Furthermore, the source population, which occurred on unprotected lands, precipitously declined during this time period, further highlighting the importance of safeguarding plants from that population. We report that a new, growing population of C. avonensis has been established to date, with important implications for the species' conservation as well as how introductions are evaluated.

With a range of programs focused on local plant and insect conservation, the Cincinnati Zoo & Botanical Garden works with partners and our communities to restore landscapes and thriving ecosystems for wildlife and people. We used organizational identity theory (OIT) to evaluate the current strategies and practices of five programs and determine opportunities for adaptation to better achieve our organizational conservation goals. Case studies ranged from habitat restoration of wetlands and gardens to community engagement to encourage individual gardeners and the reintroduction of endangered plants and insects. We present program characteristics and how collaborative partnerships facilitate opportunities for zoos to lead the conservation of local flora and fauna. The OIT framework reveals components critical to strategy implementation and best practices relevant to other zoos, aquariums, and botanical gardens when evaluating their collaborative conservation initiatives.

Premise: Of the approximately 430 species of oaks (Quercus spp.) that have been assessed, 31% are threatened with extinction and in need of safeguarding. However, oak seeds cannot be seed banked, and thus rely on alternative strategies such as in vitro culture for ex situ conservation. One challenge to this approach is low culture initiation rates. Our objective was to identify factors that may improve the establishment of shoot cultures in vitro using new growth collected from mature trees.Methods: Shoot cuttings were harvested from individuals of five different oak species (Q. alba, Q. bicolor, Q. macrocarpa, Q. muehlenbergii, and Q. palustris). Shoots were cultured onto medium with or without 50 µM silver thiosulfate (STS), a known inhibitor of the stress hormone ethylene. Cultures were grown for one month, at which point shoots were assessed for survival.Results: Shoot survival was significantly greater in shoots cultured on medium containing STS compared to the control group, with the overall survival rate increasing from 65% to 73%.Discussion: Increasing the survival rate of newly established cultures is important in ensuring that material collected from endangered species has the best chance for survival, which is critical for successful ex situ conservation.

Premise: The effective ex situ conservation of exceptional plants, whether in living collections or cryo‐collections, requires more resources than the conservation of other species. Because of their expertise with rare plants, botanical gardens are well positioned to lead this effort, but a well‐developed strategy requires a clear understanding of the resources needed.Methods: Grant funding was obtained from the Institute of Museum and Library Services to support a three‐year project on cryobanking, and to provide smaller grants to 10 other botanical gardens for one‐year projects on either (1) seed behavior studies or (2) the development of protocols for in vitro propagation or cryopreservation.Results: Nine of the partner gardens worked on 19 species (one was unable to continue due to the COVID‐19 pandemic), while the larger project focused on 14 species. A point system was developed for tasks accomplished, and the average costs per point of the larger and smaller projects were similar. Labor accounted for half the costs. Projects focused on species in the Asteraceae and Orchidaceae had lower costs per point than other species.Discussion: Both large and small projects can contribute to a strategy for exceptional plant conservation for similar costs. Prioritizing species with lower costs could help advance the field while allowing time for work on more difficult species to develop.

Cryopreservation is increasingly important as a conservation tool, particularly for threatened exceptional species. The goal of this study was to investigate the current knowledge of plant cryopreservation through a search of the literature in Web of Science and align that with the 775 species currently identified on the Working List of Exceptional Plants. While there is a good foundation in plant cryopreservation research, particularly with economically important species, there are significant gaps in research on families that contain the largest numbers of currently known exceptional species, including the Dipterocarpaceae, Rhizophoraceae, and Pittosporaceae. Even families well represented in both in the literature and on the List of Exceptional Plants had much less overlap at the level of genus. Tropical trees, a significant portion of exceptional species, were not as well represented in the literature as herbaceous species. Over 70% of all articles dealt with in vitro cryopreservation, with much less emphasis on other methods (seed, embryo, dormant bud, and pollen) that will be more cost-effective for species where they can be applied. While the research on plant cryopreservation to date provides a strong foundation and is being utilized effectively for conserving the diversity of a number of economically important species, this study revealed significant gaps that can help prioritize future research to more effectively conserve the diversity of threatened exceptional species.