Revision of Fothergilla (Hamamelidaceae), including resurrection of F. parvifolia and a new species, F. milleri

Abstract Fothergilla is a small genus of deciduous shrubs native to the southeastern United States that depending on circumscription comprises two to four species. Recent treatments recognized only two species in the genus: F. gardenii (tetraploid) and F. major (hexaploid). Until recently, no diploid taxon of Fothergilla was known. However, recent investigations identified a number of diploid populations in Alabama, Florida, Georgia, and South Carolina. A subsequent phylogenomic analysis showed that the diploids segregated into two, well-supported lineages, corresponding to largely allopatric populations. A re-examination of the morphology of diploid plants, in combination with the genetic evidence, has led us to the recognition of two species of diploids in the genus – a resurrected F. parvifolia and a new species (F. milleri W.D. Phillips & J.E. Haynes, sp. nov.) – bringing the total number of recognized species in Fothergilla to four. A revised taxonomic treatment of the genus is provided.


Introduction
Fothergilla L. (Hamamelidaceae, Hamamelidoideae, Fothergilleae) is a small genus of deciduous shrubs native to the southeastern United States that depending on circumscription comprises two to four species (Small 1903;Britton 1905;Small 1913; Harms cultivars (such as 'Mount Airy') represented F. major or F. gardenii. No pentaploids have been identified in nature.
Until recently, no diploid taxon of Fothergilla was known. However, recent sampling and cytometric analysis identified a number of diploid populations in Alabama, Florida, Georgia, and South Carolina (Ranney et al. 2012). This work was followed by phylogenomic analyses, examining the origins of F. gardenii and F. major and their relationship to the diploid populations (Qi et al. 2015). These analyses identified 11 haplotypes of plastid DNA, five of ETS, and 13 of combined plastid-ETS sequences. Of these, no haplotypes were shared between the diploid populations and polyploid taxa (i.e., F. gardenii and F. major). Furthermore, the diploid OTUs segregated into two, well-supported lineages, corresponding to largely allopatric populations. A reexamination of the morphology of diploid plants, in combination with the genetic evidence, has led us here to the recognition of two species of diploids in the genus: a resurrected F. parvifolia and a new species (F. milleri) as described below. A revised taxonomic treatment of the genus is provided.

Methods
Specimens studied in the course of preparing this revision included: (1) 34 accessions of Fothergilla from throughout the southeastern United States, planted and grown in a common garden at the Mountain Horticultural Crops Research and Extension Center in Mills River, North Carolina (Table 1) (Table 1). The Phylogenetic Species Concept (PSC) sensu Nixon and Wheeler (1990), and its method of discovery-Population Aggregation Analysis-was applied to determine if taxa could be recognized. Thirteen binary and three multi-state morphological characters were assessed ( Table 2). These included several novel characters not heretofore explicitly employed in study of the genus, such as the orientation of blades on living plants (e.g., spreading, erect, or drooping) and the ratio of: (1) the width, at the widest point, of the intervening leaf surface between the lowermost secondary vein and the leaf margin (IW), and (2) the length of the midvein interval between the junction of the midvein and lowermost secondary vein and the junction of the midvein and the next-most distal secondary vein on the same side of the leaf (IL; Fig. 1). To determine the IW:IL ratio for the Population Aggregation Analysis, respective measurements were taken from the largest measurable leaf of each of 34 accessions of known ploidy (Table 1). Subsequent to our post-analysis decisions regarding taxon recognition, we took additional IW and IL measurements from 96 loaned herbarium specimens (marked by m in the list of exsiccatae below). We also searched the SERNEC portal (http://sernecportal.org) to identify any additional specimens of the diploid taxa we recognized. This search resulted in six additional specimens, which we added to the list of exsiccatae in the taxonomic treat- ment below (only two of these represented a county not represented in our original loan of specimens). The combined sets of specimens of known ploidy (Table 1) and available to us from the herbaria identified above were the source of the morphological data we provide in our species descriptions.

Results and discussion
Population Aggregation Analysis revealed four distinct aggregate profiles, each corresponding to one of the major lineages identified by Qi et al. (2015) ( Table 3; Fig. 2). In contrast to the tetraploids and hexaploids, which bear leaves mostly spreading (profiles 3 and 4), diploids bear leaves either distinctly drooping (profile 1) or erect (profile 2) (Fig. 3). Diploids with drooping leaves (profile 1) also exhibit green, ovate laminas with marginal dentation beginning at the middle of the blade, as well as obtuse seed apices, in contrast to diploids bearing erect leaves (profile 2), which exhibit blue-green or gray-green, obovate laminas with marginal dentation beginning only at the top third of the blade and seed apices that are acute. A re-examination of type material revealed that a name already exists for the drooping-leaved diploids representing profile 1: F. parvifolia Kearney (holotype: Kearney s.n., NY-02514026; Fig. 4). This name was originally published in Small (1903), but lumped beneath F. gardenii by subsequent authors (Ernst 1963;Radford et al. 1968;Weaver 1969;Meyer 1997 andWeakley 2015). The mean IW:IL ratio for F. parvifolia accessions from the common garden at the Mountain Horticultural Crops Research and Extension Center (Table 1; known ploidy) is 0.86 (s.d. = 0.25, n = 4) and 0.96 (s.d. = 0.22, n = 7) when including additional herbarium specimens. There are no prior names applicable to plants referred to profile 2, which we here recognize as representing a new species, described as F. milleri below. The mean IW:IL ratio for F. milleri accessions from the common garden at the Mountain Horticultural Crops Research and Extension Center (Table 1; known ploidy) is 0.26 (s.d. = 0.13, n = 4) and 0.29 (s.d. = 0.10, n = 14) when including additional herbarium specimens.
Fothergilla gardenii is the appropriate name for the tetraploids representing profile 3. With the removal of the diploid components previously lumped under that name, our taxon concept of F. gardenii is necessarily narrower than that of recent authors such as Weaver (1969) and Meyer (1997). The mean IW:IL ratio for F. gardenii accessions from the common garden at the Mountain Horticultural Crops Research and Extension Center (Table 1; known ploidy) is 0.29 (s.d. = 0.09, n = 10) and 0.33 (s.d. = 0.09, n = 49) when including additional herbarium specimens.  Fothergilla major is the appropriate name for the hexaploids referred to profile 4. The average IW:IL ratio for F. major accessions from the common garden at the Mountain Horticultural Crops Research and Extension Center (Table 1; known ploidy) is   (Table 3). For accession details, including localities, see Table 1. 0.99 (s.d. = 0.37, n = 16) and 0.96 (s.d. = 0.32, n = 60) when including additional herbarium specimens. Our taxon concept of F. major is consistent with that of Weaver (1969) and Meyer (1997). However, it should be noted that although the leaf bases of F. major have often been generally described as cordate or rounded (if sometimes asymmetrically so), the shape is actually a combination of a short cuneate section adjoining the petiole that broadens out laterally into the more general cordate or rounded shape. We here use the terms V-cordate and V-rounded to describe this type of base (the "V" representing the cuneate section) and consider it structurally distinct from the neatly cordate leaf bases of F. parvifolia (Fig. 1). Based on these results, Fothergilla is here recognized as a genus of four species endemic to the southeastern United States: F. gardenii (4x), F. major (6x), F. milleri (2x), and F. parvifolia (2x). An updated taxonomic treatment follows. Description. Shrubs, rhizomatous, perennial, to 8 m tall; clump forming, usually multi-stemmed. Bark smooth, gray to reddish-brown. Stems stellate-pubescent when young, sparsely pubescent to glabrate when mature. Vegetative buds naked, densely stellate-pubescent. Leaves deciduous, simple, alternate, petiolate; stipules lanceolate to ovate; blades pinnately-veined, ovate, obovate, or oblong, bases oblique to symmetric, rounded, truncate, V-cordate or cordate, margins crenate to serrate from at or below middle to apex, or only near apex, apices acute to obtuse, surfaces sparsely to densely stellate-pubescent or glabrous, abaxially glaucous or not. Inflorescence terminal, spikes, erect, appearing with or before leaves. Flowers: mostly perfect, proximal often staminate; calyx lobes 5-7, connate, forming shallow hypanthium; apetalous; stamens 10-32, adnate to hypanthium, filaments white, anthers yellow, basifixed, 2-loculed; gynoecium, adnate to hypanthium, 2-carpellate, connate below, divergent near apex into separate styles, semi-inferior. Fruit: capsules, in groups of 3 or more, loculicidal, gray to brown, densely stellate-pubescent throughout with long, simple trichomes mixed in predominantly on and above persistent hypanthium; remnant style beaks conspicuous, abscising with maturity. Seeds: 2 per capsule, glossy, hard, nearly white, mottled, or solid redbrown to brown, ellipsoid to slightly ovoid, round to slightly flattened near apex in crosssection, apex round, obtuse, or acute to acuminate, when acuminate often recurved. Notes. Fothergilla seeds vary in color and shape, but the surface texture is consistent throughout. The seed surface is smooth and glossy with a conspicuous hilum scar at the base. In F. major, seeds are variable in color: completely white, mottled, or brown. In F. gardenii, seeds are mottled white-brown, with white conspicuously appearing around the margins of the seeds. In both diploid taxa, seeds are consistently red-brown to brown. In F. gardenii and F. major, it appears that color is affected by age and storage, as older herbaria specimens have nearly completely white seeds.
Phenology. Flowering beginning late Mar; fruiting by late Apr through Jul. Distribution and habitat. This species can be found along the Atlantic coastal plains of North Carolina, South Carolina, and Georgia (Fig. 5). It occurs in pocosins, savannas, and ecotones. It can be found in both sandy and peaty soils from mesic to wet conditions. It has been found in association with Acer rubrum L., Amelanchier obovalis Notes. Fothergilla gardenii was apparently cultivated in England as early as 1765, grown at Kew Gardens by 1789, and English and French plant nurseries were offering seeds for sale by the early 1800s (Weaver 1971).
The species appears well adapted to periodic fires and is shade intolerant. Populations found in recently burned sites appear more abundant and robust, while populations in sites that have not been burned are generally outcompeted by other plants.

Fothergilla major
Phenology. Flowering beginning late Mar; fruiting late May through mid-Jul. Distribution and habitat. This species can be found in the mountains and Piedmont of Alabama,Arkansas,Georgia,North Carolina,South Carolina,and Tennessee (Fig. 5). Habitat in which this species is found is variable, from rocky/xeric to peaty/ mesic, but generally upland deciduous forest (typically oak-dominated) with mountain and Piedmont-associated species. However, in the Uwharrie National Forest, Montgomery County, North Carolina, F. major can be found growing in seepage community with Coastal Plain-affinity species. In this habitat, where burning is frequent, this species may be <1 m in height. However, height variance is apparently not simply a factor of fire frequency. Fothergilla major varies considerably in habit, ranging from colonial shrubs less than 2 m tall, typically found growing on sandy dampish sites in the southwestern part of the range and lower elevations (for example populations from Marshall Co., Alabama and Blount Co., Alabama) to larger tree forms growing to 6 m, with offsets but no rhizomes, typically found in more upland mountain dry forest sites with igneous geology found to the North and East (for example populations in Scott Co., Tennessee). Commonly associated upland species include Acer rubrum, Aesculus sylvatica W. Bartram, Alnus serrulata, Aronia arbutifolia ( Conservation. Fothergilla major is considered Vulnerable rangewide, ranked by Na-tureServe as follows: G3; Alabama (S2), Arkansas (S1), Georgia (S1), North Carolina Diagnosis. Fothergilla milleri is morphologically most similar to F. gardenii, but differs from the latter by leaves held erect (vs. spreading in F. gardenii), blades bluegreen or gray-green (vs. green in F. gardenii), petioles 1/3-1/2 the length of the IL (vs. ¾ the length of the IL or longer in F. gardenii), and seed apices acute to acuminate (vs. rounded or obtuse in F. gardenii).
Phenology. Flowering Mar-May; fruiting May-Sep. Distribution and habitat. This species range is restricted to Georgia, South Carolina, and Alabama (Fig. 5). Because there are few herbarium records for this species, little is known about its exact distribution and environmental restrictions. According to the few notes available on herbarium specimens, it occurs in seepages and margins of bogs, bay swamps, and watercourses.
Conservation. The conservation status of this species needs to be assessed. It is presently known from only eight counties, and would appear to have an imperilment status at least as severe as that of F. major. Consequently, only skeletal collections data are provided below.