University of Vermont ScholarWorks @ UVM College of Agriculture and Life Sciences Faculty Publications College of Agriculture and Life Sciences 1-2-2018 The impact of salinity on mycorrhizal colonization of a rare legume, Galactia smallii, in South Florida pine rocklands Klara Scharnagl Florida International University Vanessa Sanchez Florida International University Eric Von Wettberg Florida International University Follow this and additional works at: https://scholarworks.uvm.edu/calsfac Part of the Community Health Commons, Human Ecology Commons, Nature and Society Relations Commons, Place and Environment Commons, and the Sustainability Commons Recommended Citation Scharnagl K, Sanchez V, von Wettberg E. The impact of salinity on mycorrhizal colonization of a rare legume, Galactia smallii, in South Florida pine rocklands. BMC research notes. 2018 Dec 1;11(1):2. This Article is brought to you for free and open access by the College of Agriculture and Life Sciences at ScholarWorks @ UVM. It has been accepted for inclusion in College of Agriculture and Life Sciences Faculty Publications by an authorized administrator of ScholarWorks @ UVM. For more information, please contact [email protected]. Scharnagl et al. BMC Res Notes (2018) 11:2 https://doi.org/10.1186/s13104-017-3105-8 BMC Research Notes RESEARCH NOTE Open Access The impact of salinity on mycorrhizal colonization of a rare legume, Galactia smallii, in South Florida pine rocklands Klara Scharnagl1,2*, Vanessa Sanchez1 and Eric von Wettberg3,4* Abstract Objectives: The success of restoration plantings depends on the capacity of transplanted individuals or seeds to establish and reproduce. It is increasingly recognized that restoration success depends quite heavily upon biotic interactions and belowground processes. Under stressful abiotic conditions, such as soils salinized by storm surge and sea level rise, symbiotic interactions with soil microbes such as mycorrhizae may be critically important. In this study, we investigate the impact of salinity on percent colonization of roots by arbuscular mycorrhizal fungi, in addition to the impacts of this colonization on plant ftness under saline conditions. Fifty Galactia smallii plants from an ex situ collection were subjected to a salinity treatment for 6 weeks, and 50 plants were untreated. Plants were harvested and assessed for percent colonization by arbuscular mycorrhizal fungi, nodule number, shoot and root dry biomass, and micronutrient content. Results: Colonization by arbuscular mycorrhizae was higher in plants in the salinity treatment than in untreated plants; plants in the salinity treatment were also found to have a lower root:shoot ratio, and higher phosphorus and nitrogen levels. These results support the importance of arbuscular mycorrhizal fungi in restoration eforts of endan- gered plants in fragmented and threatened ecosystems, such as pine rocklands. Keywords: Galactia smallii, Pine rockland, Soil mutualists, AMF (arbuscular mycorrhizal fungi), Saline soils, Sea level rise Introduction Legumes with the capacity to symbiotically obtain Pine rocklands are a dry forest ecosystem unique to nitrogen and phosphorus can be critical to low fertility South Florida, Cuba, and the Bahamas. In South Florida, pine rockland systems. One endangered legume is Galac- pine rocklands are dominated by the trees Pinus elliotti tia smallii, or Small’s milkpea, a procumbent under- and Serenoa repens, yet are host to a large diversity of story legume endemic to the pine rockland ecosystem smaller understory plants, many of which are endemic [4]. Galactia smallii is critically endangered as a conse- to this ecosystem [1]. Pine rocklands once covered a quence of habitat fragmentation [5, 6]. Although endan- large expanse of the southeastern Florida peninsula, yet gered, very little is known about this species. Fairchild today stand at two percent of their original extent and Tropical Botanic Garden has established a program to are highly fragmented [2]. Many fragments are very small replant Galactia smallii in pine rocklands. One potential (under a hectare), and only some of the fragments are factor in restoration is the belowground community [7]. protected [3]. Arbuscular mycorrhizal fungi are known to help plants deal with a variety of environmental stressors [8, 9]. In pine rocklands, where available phosphorus (Pi) readily *Correspondence: [email protected]; [email protected] 2 Plant Biology, Michigan State University, 612 Wilson Rd, East Lansing, MI binds to the calcium carbonate of the limestone, the abil- 48824, USA ity to acquire phosphorus is critical [10]. 4 Plant and Soil Science, University of Vermont, Burlington, VT 05403, USA Full list of author information is available at the end of the article © The Author(s) 2017. This article is distributed under the terms of the Creative Commons Attribution 4.0 International License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/ publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated. Scharnagl et al. BMC Res Notes (2018) 11:2 Page 2 of 5 We were interested in the role that AMF play in the like a pulse of saline stress rather than a constant saline recovery of rare native legumes in South Florida [11]. stress. Furthermore, we were interested in whether symbionts After 6 weeks the plants were harvested. Teir roots such as AMF help their host plants combat salinity stress were washed with tap water, soil samples were collected [12]. A common threat to pine rocklands in South Flor- from each pot, and roots and shoots were separated for ida is sea level rise due to global climate change [13]. It weighing and identifcation of associated soil microbial is predicted that sea level may rise as much as a meter symbionts. Once root tips collected for analysis, both in the coming century [14]. Tis would engulf the many roots and shoots were dried overnight in an 80 °C drying parts of South Florida that are only a meter above mean oven, and weighed. sea level and increase salinization of soils throughout the Twenty root tips, 1–3 cm in length, were randomly col- peninsula [14, 15]. A shadehouse study was conducted to lected from each individual G. smallii plant’s roots and investigate the efects of the addition of a saline solution placed in 10% KOH and left overnight. Root tips were on the AMF relationship of the G. smallii plants. washed three times with tap water, bleached in chlorine for 1–2 min, and washed three times with tap water. Root Main text tips were acidifed with 1% HCl, and placed in 0.05% Materials and methods Trypan blue solution overnight. Finally, root tips were In 2009, nearly 100,000 Galactia smallii individuals were removed from Trypan blue, rinsed with tap water, and observed near the Homestead Airforce Base [6, 16]. Since observed under a compound microscope [18]. development is planned for the base, conservation eforts Percent of colonized roots for each plant was calculated are underway to relocate some of these individuals. As a as: part of this efort, in January 2012, 500 adult plants from % of colonized roots =[# of roots observed with AMF/total the Homestead Airforce Base were collected and moved # of roots observed]×100. into ex situ conservation at Fairchild Garden. Tese plants are being maintained in an open-air rare plant Root colonization includes the observation of arbus- nursery for seed collection. During the 2012 collection cules, vesicles, hyphae or hyphal coils within root cells. efort, plants were dug up with as much of their original Spores and propagules observed around the root tips soil as possible, which was then mixed with standard pot- were not considered observable evidence of colonization. ting soil (with sterilized composted bark, coconut coir We also counted nodules to assess the association with and perlite) and placed in pots (1 L volume). Te root rhizobial bacteria. systems had 3 months to adjust to potting before the Soil samples were weighed, rinsed with tap water and experiment began. One hundred plants from this collec- sieved through 1 mm, 250 µm and 35 µm sieves in order tion were used. to capture spores. 0.5 mL of spore solution was trans- Tese 100 plants were placed in a shade house on ferred to flter paper on a slide, and observed under raised benches, receiving periodic watering from sprin- 100× magnifcation. Spores were counted, and identi- klers connected to a groundwater source. We selected fed to genus level using morphological characteristics this sample size to retain the other translocated plants [19, 20]. In order to determine number of spores per to establish a conservation nursery. For 6 weeks, these gram soil, simple stoichiometry was used to balance the 100 plants received an addition (2 mL per plant) of P/N- number of spores per 0.5 mL solution to the total vol- limited Hoagland’s solution [17] each week. Fifty of the ume of spore solution per sample (50 mL) and the initial plants received a salt treatment: 4–5 mL of 100 ppm weight of the soil sample (ranging between 12 and 24 g saline solution using stock sodium chloride and dou- wet weight). We compared percent of roots that were ble deionized water. Tis salinity treatment mimics the colonized, spore count per gram of soil, number of fun- seasonal rise in salinity that can occur in Florida coastal gal genera and diversity index between treatments with a pine rockland patches due to salt water intrusion into two-sample t-test in PROC GLM in SAS 9.3 [21]. the aquifer at the end of the dry season (April–June), or Plant dry biomass was used as a proxy for plant ftness.