Geology and climate influence rhizobiome composition of the phenotypically diverse tropical tree Tabebuia heterophylla

same species, growing in contrasting conditions may respond to each situation with pheno- types that increase their fitness [1, 2]. Climate and geology influence the plant’s phenotype both directly and indirectly, including through adaptations achieved by altering the physiology and/or morphology of the plant [7], increasing its survival rates and geographic range [1]. The effect of environmental conditions on plant phenotypic plasticity has been thoroughly studied, although there are other factors modulating the plant phenotype, such as microorganisms associated to plant roots, which just recently gathered scientific attention. The rhizosphere–a thin layer of soil surrounding plant roots–is influenced by both the root and its associated microbiota (e.g. bacteria and fungi) [8–10], which is recruited by the plant [10, 11]. However, bacteria colonizing the rhizosphere vary due to abiotic factors, including climatic conditions and nutrient availability [9–11]. These abiotic factors contribute to bacte- rial growth rates, competition (e.g. antibiotic production), and metabolism. The effects of cli- mate and geology on the rhizobiota, and their roles in modulating the phenotypic plasticity of the plant, deserve further attention. With this in mind, we hypothesized that environmental interactions could modify rhizosphere microbial composition and their ecological functions which in turn have an impact in the plant’s phenotype. Our underlying hypotheses are that: 1) climate and 2) geology will independently alter the rhizobiota community composition.

Several groups of microbes alter the plant’s response to the environment such as mycor- rhizal fungi, nitrogen-fixing bacteria, and plant growth-promoting bacteria [3, 12–15]. These groups can be beneficial or detrimental to the plant, as they modify rhizosphere processes, which in turn regulate nutrient cycling, water uptake and the response to environmental stress [3, 16, 17]. Therefore, the rhizosphere microbiome has the potential to indirectly increase plant’s fitness by enhancing plant nutrient uptake and altering the plant phenotyp [3]. Our goal was to both characterize the microbial diversity in the rhizosphere of T. heterophylla, a morphologically variable tree, while identifying influential conditions that may be shaping the rhizosphere microbiota and perhaps their influence on phenotype. Tabebuia heterophylla, or Roble Blanco, is an excellent study subject to test our hypotheses. This plant grows throughout Puerto Rico in a broad altitudinal and geographical range. Exposed to contrasting climates and soils of different parent material, this plant exhibits phe- notypic variability in morphological, physiological, reproductive, and growth traits [18]. Nota- bly varying morphological traits such as number and size of leaflets, and size of petioles, are commonly found through the different populations of T. heterophylla in the island. To address our hypotheses, we characterized the rhizosphere microbiome of T. heterophylla under varying geologies and climatic conditions (i.e. temperature and precipitation), to give

insight into the extant of the microbial diversity and to further understand the relative role of biotic and abiotic factors in plant phenotypic plasticity. Materials and methods Sampling design, sample collection, and storage Tabebuia heterophylla is a tree native to the Caribbean archipelago and is amply distributed in the Island of Puerto Rico. We identified 6 populations of T. heterophylla developing on soils derived from contrasting parent materials and climatic conditions. For each of the three parent materials (volcanic, serpentine, and limestone/karst formations–henceforth designated as karst) [19], we identified two sites experiencing different climatic conditions (a combination of temperature and precipitation) (Fig 1, Table 1). Site 1 (S1 Fig) was located in a high altitude volcanic area in Cayey (Bosque de Carite), 18˚ 06’ 28.9’’ N 66˚ 04’ 20.7’’ W; Site 2 (S2) was located in a low altitude area in Guayama 18˚ 01’ 24.4’’ N 66˚ 07’ 44’’ W. Serpentine sites corresponded to sites 3 and 4, Site 3 (S3) was in the PLOS ONE Rhizobiota of the tropical tree Tabebuia heterophylla PLOS ONE | https://doi.org/10.1371/journal.pone.0231083 April 7, 2020 2 / 16 (project title “Microbial community contributions to stress adaptation and plasticity in the tropical tree Tabebuia heterophylla”, to FGV). Additional support was provided by PR-INBRE (NIH/NIGMS- award number P20 GM103475) (to FGV), a CREST Grant (NSF-HRD 0734826) to the University of Puerto Rico-Rı´o Piedras Campus (to ESV), and a NIMHD- RCMI grant U54 MD007600 from RCMI. The

funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. Competing interests: The authors have declared that no competing interests exist.