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GreatBasinNaturalist54(3),© 1994,pp.256-271 IDENTIFICATION OF PURSHIA SUBINTEGRA (ROSACEAE) FrankW. Reichenhacherl — Abstract. Populations ofPurshia incentralArizonaare intermediatein somecharactersbetween Purshiasubinte- gra, an endangered species, and Purshiastanshuriana, the common cliffrose. These intermediates mayrepresentforms derivedfromahistoryofhybridizationandintrogressionbetweentheputativeparentspecies. Morphologicaldatawere obtained from 216 pressed specimens ofP. stibmtegra, P. stanshuriana, and introgressed forms. Over 50 separate discriminant function analyses (DFA) and principal components analyses (PCA) were nm on numerous combinations of rawand log-transformed data. Thebestvariable suite, providingtheclearest discriminationbetween groups, used log- transformed dataon 15 morphological characters, but DFA post-hoc identifications were 90-100% correct with only 7 characters usingrawdata. DFAdistinguishedfourseparatenodes ofvariation. Twogroupsconsistingof122P. subinte- graand29P.stanslnirianawereeasilydiscriminatedin DFAandweredistinguishedin PCAaswell. Introgressedforms wereconsistently identified in two much less well-defined groups of46and 19 specimens. Introgressed forms are not intermediate between the two supposed parents in some characters, appearingmost similar to P. stanshuriana—in most measuredcharacteristics. Principaldistinguishingcharacteristicsofthefourgr—oupsareasfollows; P.suhintegra usual- lyeglandular, has0-2 leaflobes and short hypanthia-pedicels; P. stanshuriana always abundantly glandular, has 4 leaf lobesandshorthypanthia-pedicels;theintrogressedfonn "Tonto"isusuallyeglandular,has4leaflobesandlonghypan- thia-pedicels;theintrogressedfonn"Verde"isusuallyglandular,has4leaflobesandslightlyshorterhypanthia-pedicels. Keywords: Purshiasubintegra, Purshiastanshuriana,Arizonaclijfrose, clijfrose, endangeredspecies, morphometries, introgression, taxonomy. Purshia suhintegra (Kearney) Henrickson Barbara G. Phillips' 1984 discovery of the (Arizona cliffrose) is protected under federal fourdi locality for P. suhintegra near Horseshoe law as an endangered species (USFWS 1984). Dam along the lowerVerde River. For a federally endangered species like P. At the four locations cited above, Purshia suhintegra it is important, indeedvital, to know suhintegra is restricted to outcrops ofTertiary the taxonomic identity of every individual deposits of limy lacustrine rock formations plant in agiven populationbecause the protec- (Anderson 1986). Soils derived from these tive measures ofthe Endangered Species Act ancient lake basin rocks are characterized by are available to species (including forms that low nitrogen and phosphorus levels, which exhibit characteristics of introgression with limit, or preclude, typically Sonoran Desert other species), but not to theirearly generation species that are common on nearby sites with hybrids. soils derived from igneous and metamorphic Purshia suhintegra is found in four widely rocks (Anderson 1986). Purshia subintegra is a scattered locations from northwestern to species ofthe northern and eastern perimeter southeastern Arizona (Table 1, Fig. 1). The first ofthe Sonoran Desert; all four sites supporting collection was made by Darrow and Benson in the species are atorbelow 1000 m elevation. 1938 (Kearney 1943, Schaack 1987a) near Purshia stanshuriana, the common cliffrose Burro Creek in Mohave County, Arizona. A of the Southwest, is not a Sonoran Desert second population was documented in a col- species and consequently is not sympatric lection by Pinkava, Keil, and Lehto in 1968 with P. suhintegra at three ofthe four P. suhin- (Pinkava et al. 1970) almost 300 km from tegra sites known. In the Verde RiverValley of Burro Creek, near Bylas in Craham County, eastern Yavapai County, Arizona, the highest- Arizona. Anderson (1986) found a third popu- elevation P. suhintegra site, the two species lation on bluffs overlooking the upper Verde occur in close enough proximity that gene River near the town of Cottonwood (referred exchange may occur, at least occasionally. to as the Verde Valley area) and reported on Scattered populations and individuals of 'SouthwesternFieldBiologists,8230EastBroadwayBoulevard,SuiteW8,Tucson,Arizona85710-4002. 256 1994] Identification ofPurshia subintegra 257 Table 1. LocationsoiPurshiaspp.collectionssampledinmultivariatemorphometricanalysis;216specimenscollected formorphologicalanalysisinArizona. Regionand 258 Great Basin Natur.\list [Volume54 subintegra, ". restricted to late Tertiary cal- . . careous, lacustrine deposits ca. 16-21 km north- westofBylas, Graham County, Arizona. Few botanists have adopted Schaack's tax- O V PP--ssutbainnstbeugrra onomy, but most recognize that variation is " IntrogressedForm present and concede that it may have resulted from some form or degree of past hybridization or introgression involving P. subintegra and P. stansburiana. McArthur et al. (1983) reported n = 9 and 2n = 18 for P. subintegra and P. stansburiana, respectively. Phillips et al. (1988) used starch gel electrophoresis to investigate isozymes at 14 loci in three populations ofPurshia stansburiana and four populations ofP. subintegra. The material used in this study was collected by the author and presei^ved in liquid nitrogen from the same plants that provided dried V2 specimens for this study. Phillips et al. (1988) were unable to discern differential patterns of variability useful in identification oftaxonomic Fig. 1. Distribution ofsampling sites, species, and groups; between-groups similarities ranged introgressed forms. See Figure 2 foran e.xpanded viewof VerdeValley. Letters identify sites illustrated in graphs of from 0.925 to 0.992 (Nei [1978] unbiased multivariateanalyses. genetic identity). Fitts et al. (1992) studied Purshia subintegra in Verde Valley, reporting on many impor- relationship ofP. subintegra to P. stanshuriana tant, but heretofore unknown, aspects of the is the subject ofthis study, that o{P. subintegra pollination biology ofthe species. They found to the moiphologically similar P. ericifolia is of that flowers may be pollinated anytime in the interest as well. All other Purshia taxa have first three days after anthesis, the plants are lobed leaves except P. ericifolia. Leaves of P. partially self-compatible, and native and intro- mm ericifolia are about 6 long, simple, acute, duced bees are primaiy pollinators. Reciprocal linear, and eglandular The species is restricted crossing experiments between P. subintegra to limestone outcrops in the Texas Big Bend and what was believed to be P. stansburiana region. Ithas been speculated thatP. subintegra were also conducted by Fitts et al. (1992). As may have evolved from some ancient series of is discussed in the concluding section of this crosses and backcrosses involving P. ericifolia report, plants from which the P. stansburiana and some other Purshia, perhaps P. stansburi- was taken are actually introgressed forms. ana (McArthur et al. 1983). Phylogenetic The purpose of this study was to analyze investigation ofthe whole genus in relation to moiphological charactervariation in species of closely related genera such as Fallugia would Purshia in order to identify the range ofmor- be valuable in interpreting P. subintegra and phological variation in P. subintegra and to shouldbe the logical next step for future work. develop a means of discriminating between Purshia subintegra, an endangered species, Relation toPurshiastansburiana and other non-endangered Purshia taxa with Schaack (1987a, 1987b) suggested that the which P. subintegra is most likely to be con- basionym P. subintegra was based on material fused. This study was undertaken solely to ofhybrid origin, resulting from pasthybridiza- address the need ofnatural resource managers tion of P. stansburiana and a previously to have ameans ofdeterminingwhich individ- unnamed central Arizona Purshia. He conse- uals and populations ofPurshia are protected quently published a new species name, under the Endangered Species Act. The Purshia pinkavae Schaack, to include a very methods used in this studywere carefullycho- pure concept ofwhat had been included in P. sen toobtain this result. 1994] Identification of Purshia subintegra 259 Methods mens were also employed. Table 2 shows the character palette developed for the morpho- A total of216 Purshia plants were sampled metric analysis. The list ofcharacters indicates at 27 widely scattered sites from southeastern that amix ofbinary, categorical, and continuous to northwestern Arizona for measurement and data was used. This was taken into account in analysis of morphometric characters (Fig. 1). subsequent statistical analyses. All measure- Much attention was given to sampling Purshia ments and counts were made under a binocu- in Verde Valley, the only location where it is lar dissecting microscope with a micrometer believed P. subintegra and P. stanshuriana are disk or electronic calipers. Scoringprocedures in close enough proximity that gene exchange are describedin Table 2. might currently be occurring. It was hoped SYSTATversion 4.0was used to subject the that if hybridization were occurring between data to more than 50 separate discriminant the two taxa, it would be possible to isolate function analyses (DFA) and principal compo- characters useful in discriminating between P. nents analyses (PCA) to identify moqDhologi- subintegra, P. stanshuriana, and the introgressed cal groups and to determine which characters forms. In determining where to sample, it was could be mostconfidently used to separate the important to have some firsthand notion of groups. Numerous combinations ofcharacters where the introgressed forms might be: this were used to group like data (binary, categori- turned out to be more difficult than it might cal, and continuous) and to examine the effects seem given the disparate views of several ofincluding ratios as characters (hypanthium- researchers. Table 1 lists three separate collec- pedicel length/width, sepal length/width, petal tion sites for P. subintegra in Verde Valley length/width) in the data set. Initial analyses (labeled D, E, and F), in addition to several using PCA were run on several combinations locations for introgressed forms and one loca- ofcharacters to identifycharacters responsible tion for P. stanshuriana on mountains over- forwithin-group similarity. looking the valley (Jerome, labeled X). Figure A priori assignments of plants to groups 2 shows Verde Valley and collection sites in required for DFA involved grouping collec- the valley. The author made three separate tion sites in several combinations by morpho- collecting—trips to Verde Valley—1987, 1989, logical, geographical, and ecological criteria. and 1992 each time expanding the scope of Most DFAs were run with the following the samplingeffort to try to obtain a more rep- groupings: (1) 27-group analysis, all 27 collec- resentative sample ofcharactervariation. tion sites coded as separate groups; (2) 3- Morphometric samples from each of the group analysis, 4 P. subintegra sites, 3 P. stans- 216 sampled plants consisted of two to four huriana sites, and all introgressed forms in one 20-40-cm-long branches dried in a standard group; (3) 4-group analysis, 4 P. subintegra herbarium press. Samples were collected in sites, 3 P. stanshuriana sites, and the intro- April 1987, 1989, and 1992 from each of 4 P gressed forms separated into two groups iden- subintegra populations, 3 P. stanshuriana pop- tifiedas "Tonto" and "Verde." ulations, and 20 sites of introgressed forms. Table 1 shows the locations and sample sizes Results of each collection site. A rigorous stratified- random samplingmethodwas employed at the Purshia subintegra can be differentiated P. subintegra and P. stanshuriana sites, and at from P. stanshuriana and introgressed forms least 10 specimens were collected at each site. by leafglandularity and leaflobing. The mean Collections at the introgressed fonn sites were score of leaf glandularity in P. subintegra is made much more subjectively and the sample less than 0.4, and the mean number oflobes/ sizes were much smaller, only one to six speci- leafis 2.5 or less. All others are more glandular mens. or have more leaflobes. A population ofwhat Data on 15 characters judged to be poten- I initially believed to be introgressed forms at tially useful in taxonomic differentiation site a (Fig. 2) in Verde Valley possesses glan- between the 27 groups were obtained from dularity and lobing characteristics ofP. subin- the pressed specimens. Floral characters were tegra and, based on this and the results ofthe heavily relied on, and characters that could be multivariate analyses, should probablybe clas- used in field identification ofunknown speci- sified as P. subintegra. 260 Great Basin Naturalist [Volume54 1994] Identification ofPurshia subintegra 261 Table 2. Characters measured for analysis ofcharacter variation in Purshia stansburiana, introgressed forms, and Purshia subintegra. Descriptions ofthecharactermeasurements used in multivariate analyses and acronyms (in parentheses) usedasvariablelabels inTables 4 and5. Mosthypanthia-pedicel, petal, sepal, pistil, andstamen measurements andcountswerefromthesamefiveflowersfromeachplant. 1. Leafpubescence. (LEFPUB) The adaxial surface were measured under a binocular dissecting ofPurshialeavesisdenselytomentose,thoughthe microscope usingamicrometerdisk orelectronic midvein is often bare. The abaxial surface ranges calipers. The length/width ratio was also entered fi-om completely glabrous to completelyobscured asacharactervariable(HYPRAT). bylongarachnoid hairs. Twenty leaves from each ofthe 216 plant specimens were scored on an 8-10. Sepal dimensions. (SEPLGTH, SEPWDTH) All index ofleafpubescence density. Only the dorsal sepals (usually 5) from five flowers from each of (abaxial) surface was scored. The scale ranged the 216 plants were dissected and measured from 1 (completelytonearlycompletelyglabrous) under a microscope using a micrometer disk or to5(denselypubescent). electronic calipers. Maximum (basal) width and length ofthe sepals were recorded. The length/ 2. Leafglands. (LEAFGLAN) Ten leaves from each width ratio was also entered as a character vari- ofthe 216 plants were examined and scored for able(SEPRAT). presence or absence ofimpressed-punctate glands. 11-13. Petal dimensions. (PETLGTH, PETWDTH) Afl petals (usually 5) from five flowers from each of 3. Hypanthium-pedicel glands. (HYPGLAN) Five the 216 plants were dissected and measured hypanthia (with pedicels) from each plant were under a microscope using a micrometer disk or examined and scored for presence or absence of electroniccalipers. Maximumwidthandlengthof stipitateglands. the petals were recorded. The length/width ratio wasalsoenteredasacharactervariable(PETRAT). 4. Leaflobes. (LOBES) The number oflobes on each of20leaves from each ofthe216 plants was 14. Pistilnumber. (PSLSFLR) Flowersnormallycon- counted. Theleaftipwas notcounted. Lobes var- tained 2-A pistils. Aborted pistils were easilydis- ied in distinctness, i.e., from much longer than tinguished by their small size (<1.25 mm long) widetomerebumpsontheedgeoftheleaf Even andbrowntodarkbrowncolor.Viablepistilswere the most minorlobes were scored. Figure 3 illus- pale yellow with silvery-white achene hairs and trates variation inleaflobingamongP. subintegra, were nearly always longer than 1.25 mm. The the "Tonto" and "Verde" introgressed forms, and total numberofpistils per flowerwas counted on P. stansburiana, and provides an example oflobe each ofthe 216specimens, as wellas the number scoring. ofviableandabortedpistils. 5-7. Hypanthium-pedicel dimensions. (HYPLGTH, 15. Number of stamens. (STMNS) Stamens were HYPWDTH) Length and maximum width offive countedinfiveflowersfromeachof216plants. hypanthia-pedicels from each ofthe 216 plants Table 3 lists mean values and standard Principal Components Analysis errors obtained for each of the 15 characters Rotated factor scores derived from three included in the analysis for each of the four PCAs are listed by character in Table 4 and identified groups of Purshia spp. Figure 4 are graphed in Figure 5. The first three factor illustrates the distribution ofvariation in sepal axes together account for 73-87% ofvariance and hypanthium-pedicel dimensions. Note that sepal length and width are highly and in the data. Horizontal relationships on the positively correlated among groups, while FACTOR(2)/FACTOR(l) graphs for each hypanthium-pedicel width and length are not. analysis (x-axis, Figs. 5A-5C) are primarily P. subintegra plants have shorter, narrower based on leaf lobing, while vertical relation- sepals, whileP. stansburiana havelonger, wider ships are based on glandularity. The horizontal sepals; introgressed forms are intermediate. relationship is again based on glandularity in "Tonto" forms have very long, wide hypanthia- the FACTOR(3)/FACTOR(2) graphs (y-axis, pedicels, while "Verde" forms have slightly Figs. 5A-5C), but the vertical relationship (z- shorter, butmuch narrower, hypanthia-pedicels; axis. Figs. 5A-5C) is mostly influenced by neither ofthe two introgressed forms is inter- hypanthium-pedicellength. mediate in hypanthium-pedicel dimensions PCAs illustrate similarities of the three between the supposed parent species, P. groups ofPurshia spp. to each other, butgraphs subintegra andP. stansburiana. must be interpreted carefully. It appears that 262 Great Basin Naturalist [Volume54 P. subintegra n<imiini cm 1 0001 1 1 201 2 101001022 X.75 "Verde" ^/Vi 44444 3 6 4 2 4 4 4 Fig. 3. Variation in leaflobing and size in Piirshia subintegra, "Verde" and "Tonto" introgressed forms, and Piirshia stansburiana. AiTaysof20leaves scoredforpubescence, glandularity, andlobingfromtypicalspecimens ineachgroup. Numberofleaflobesscoredforeachleafandmeanscoreforeachplantareshown. P. stansburiana groups (X-Z) are lost among this reason, DFAs were firstrun on datamatri- "Verde" introgressed forms (b, c, d, e, f, g, h, i, ces in which groups were assigned according k, 1, m), butacareful examination ofthe distri- to the 27 collection sites for each plant, rather bution of points along FACTOR(3) (z-axis) than a preconceived notion of the classifica- shows that, primarily on the basis ofhypanthi- tion ofeach plant. A series of27-group analy- um-pedicel length, P. stansburiana are more ses were run (Figs. 6A, 6D, 6G) using the 15-, similarto each other. P. subintegra and P. stans- 7-, and 4-character sets. This clearly showed buriana groups (A-F and X-Z, respectively) that the data naturally fall into at least three or are usually closest to each other. Introgressed four groups, depending on how many charac- forms (a-r) are more loosely grouped together ters are used. ornearP. subintegra orP. stansburiana. Results ofthe 27-group DFAs were used to reclassify each plant into one ofthree or four Discriminant Function Analysis groups. Bylas, Burro Creek, Horseshoe Lake, Univariate and multivariate F-tests con- and Verde Valley (sites D, E, F) plants were ducted as part of DFA indicated that 14 of 15 classified as P. subintegra. Plants collected characters used were significant at or below from Jerome, Skull Valley, and Sonoita were the .001 level of significance. Sepal length/ placed in P. stansburiana. All otherplants were width ratio was the only character that did not placed in "Verde" or "Tonto" introgressed produce a high F-number This was true ofall forms, or were combined together in a single character combinations and a priori grouping group of introgressed forms. Figure 6 illus- assumptions. trates results ofnine DFA analyses with three Two-dimensional graphical illustrations of character suites, each analyzed on two addi- canonical factor scores often used to show tional apriori grouping assumptions. groupings derived from DFA are partially Character suites containing as few as two dependent on initial group assignments. For characters (e.g., hypanthium-pedicel length 1994] Identification ofPurshia subintegra 263 Table 3. Characteristics of27groups ofsampled Purshia spp. Mean values and standard errors (in parentheses) are listedfor15measuredcharactersof27samplesitesand216specimens. 264 Great Basin Naturalist [Volume54 17 B 15 p.etanaburiana 13 11 (jrg.'- .IntrogressedForm IntrogressedForm 9 7 Btanaburiana 4.5 MEAN NO. LOBES/LEAF MEAN HYPANTHIUM WIDTH (mm) 6.5 Fig. 4A-C. Plots ofkey characters used in multivariate analyses. Mean values for each ofthe 27 collection sites. Ellipses include .05 and .90 confidence intervals about bivariate data means foreach ofthe twoPiirshia spp. and 5.5 two introgressedforms. SeeTable 1 forlistofcodesiden- tifying each collection site and Table 2 for list ofcharac- IntrogressedForm ters. Only twoellipses arevisible in Figure4AbecauseP. stanshuriana sitesX,Y,andZscoredexactly 1.0forallleaf glandularity samples; thus there was no standard errorto measureforthatgroup. 3.5 2.5 2.0 2.5 3.0 3.5 4.0 4.5 5.0 5.5 MEAN SEPAL WIDTH (mm) "Verde plants have slightly shorter hypan- Discussion " thia-pedicels (mean 9.2 mm) and glandular- punctate leaves. Generally, "Tonto" plants are Multivariate analysis of character variation found in Gila County, Arizona, on lakebed clearly indicates that the four Purshia subinte- deposits around Roosevelt Lake in Tonto gra populations sampled exhibit a coherent Creek basin. Note that Camp Verde, site n, is syndrome of characters. Although there is included in the "Tonto " introgressed form some variation between populations, the tax- despite its location in Verde Valley in close onomy developed by Schaack (1987b) is not proximity to the "Verde" introgressed form. supported by the analyses. Specimens claimed This assignment resulted from inspection of to be of hybrid origin by Schaack (1987a, previous DFAs and PCAs. Pinal Creek "Tonto" 1987b) from among the type material collected plants (site p) are frequently given the highest at Burro Creek should be considered to repre- probability ofbelonging to the "Verde" intro- sent P. subintegra. It may be speculated that gressed form despite their location far away leaf lobing and leaf and hypanthium-pedicel from any other "Verde" collection site (Fig. 2). glandularity exhibited by these specimens Plants at these collection sites illustrate the arose from alimited exchange ofgenetic infor- high degree of variability in phenotypic mation with P. stansburiana during a period of expression ofintrogressedforms. briefproximityin the ranges ofthe two species. 1994] Identification ofPurshia subintegra 265 Table 4. Principal components analysis ofPurshia spp. showingfactoraxis loadings resultingfrom analysisoftlie fiill set ofmeasured characters and two abbreviated sets. Scores from the first three factor axes are given for each characterset. 15-characterset