Shoot and Floral Development in Calla Palustris (Araceae-Calloideae) Author(S): R

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Shoot and Floral Development in Calla palustris (Araceae-Calloideae) Author(s): R. W. Scribailo and P. B. Tomlinson Reviewed work(s): Source: International Journal of Plant Sciences, Vol. 153, No. 1 (Mar., 1992), pp. 1-13 Published by: The University of Chicago Press Stable URL: http://www.jstor.org/stable/2995699 . Accessed: 12/04/2012 13:42

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SHOOTAND FLORAL DEVELOPMENT IN CALLA PALUSTRIS (ARACEAE-CALLOIDEAE)

R. W. SCRIBAILO'AND P. B. TOMLINSON2 HarvardForest, HarvardUniversity, Petersham,Massachusetts 01366

Callapalustris shows annualproduction of one or two inflorescenceson each ephemeralseasonal shoot. Shoots are sympodial; the terminal inflorescencesare produced in the summer of the year before the mostly perfect flowers open. Overwinteringterminal buds show limited preformationof foliage leaves on renewalshoots; most leaves areneoformed. The spadixproduces a seriesof prominentflower primordia in up to eight irregularorthostichies. Each primordiumdevelops no perianthbut a series of indistinctly trimerousstamen initials in centripetalorder. The gynoeciumoriginates as a single dorsiventrallyasym- metriccollar-like outgrowth that becomesovate and enclosesthe seriesof basalanatropous ovules, showing no evidence of a pseudomonomerouscondition either in development or vasculature.Floral variation within a spadix is considerable,distal flowersare usuallywholly male, basal flowersshow reducedstamen number,but no strictlyfemale flowersare produced.Vascular traces to the floralappendages are derived from an anastomosingsystem that obscuresany uniformpattern. Morphogenetic considerations describe the vascular system better than any referenceto imaginaryevolutionary antecedents. Introduction (1974). Flowers open in the late spring,the fruits Despiteconsiderable floral diversity in Araceae, ripening in late summer of the same year. virtuallyno developmentalstudy of aroidflowers In the classification of the Araceae developed has beencarried out. Diversityof flowersis usu- by Englerand Krause(Engler 1877; Krause 1908), ally described,in evolutionaryterms, as a mod- Calla is included in the subfamily Calloideae to- ificationof a dimerousor trimeroushermaph- gether with three other north temperate genera, roditicflower, even thoughAraceae with perfect Lysichiton, Orontium, and Symplocarpus.Gra- flowershave themselvesconsiderable organiza- yum (1984, 1990) suggests,primarily on the basis tional diversity (Engler1884). Developmental of pollen morphology,that Calla is more isolated studiesof flowersare essentialto a preciseun- within the family and includes it as a monotypic derstandingof morphologythat can revealcom- subfamily,Calloideae, the remainingthree genera mon floralplans of systematicand phyleticsig- being added to other groups. Barabe and Forget nificance(Leins et al. 1989).In this presentstudy (1987) also suggest a position isolated from Ly- we show that the inflorescenceaxis (spadix)of sichiton, Orontium,and Symplocarpuson the ba- Calla initiateslateral floral primordia that lack sis of cladistic analysis, although they still retain any subtendingbracts. All floralappendages are all four genera in the subfamily Calloideae. initiatedwithout reference to anyconsistent sym- Previous accounts of the shoot organizationof metry,in partbecause stamens are "interpolated" Calla go back to Doll (1843) and Wydler (1856) in anirregular centripetal pattern. The gynoecium and have been summarizedby Braun(1859), En- showsno developmentalevidence for a trimerous gler (1877), Krause (1908), and Dudley (1937) condition.The sequenceof appendagescondi- without the addition of significant new data. tions a vascularsystem without obvious regular- Dudley provided additional information on seed ity. and seedling morphology and pollen develop- Callapalustris is a temperateherbaceous rep- ment and clarified the cytology. More recently resentativeof the primarilytropical family Ara- Barabeand Labrecque(1983) describedthe floral ceae,but with a wide circumborealdistribution. vasculatureand concluded that this provided ev- It persistsby meansof creepinghorizontal axes, idence for a pseudomonomerous condition, the whichare either permanently or temporarilysub- gynoecium being interpreted as being made of merged,or at the surfaceof swamps.Leaves are three fused carpels because of the presence of alwaysemergent. The shootunits are ephemeral, three principal carpellary traces. This interpre- lastinglittle more than a year, but populations tation is contradictedby the observations on flo- persist by both regenerativeand proliferative ral vasculatureby Hotta (1971). The concept of branching,in the terminologyof Tomlinson pseudomonery in aroids apparently originated with Engler(1884) and was extensively elaborat- ed upon by Eckardt(1937). Manuscriptreceived June 1991; revisedmunuscript received September1991. Materialand methods 1 Presentaddress: Purdue University, North CentralCam- Populationsof Callapalustris were studied from pus, 1401 South U.S. 421, Westville, Indiana 46391. Black Gum Swamp, Tom Swamp, and Harvard 2 Authorto whom correspondenceand requestsfor reprints Pond, Petersham, and Spectacle Pond, Athol, should be sent. Massachusetts. Since no differences in patterns

I 2 INTERNATIONAL JOURNAL OF PLANT SCIENCES of shoot and floral development were found per year, although the younger spadix in shoots among these collections they have been treated with two spadices may sometimes abort. Phyl- as one. Shoots were sampled at about biweekly lotaxis is distichous throughout,with no change intervals throughout the growing season (May- in the plane of distichy at branching. Although October 1990) and also in the spring of 1991, to no leaves are present in figure3D the distichous a total of about 20 samples of over 150 shoots. phyllotaxis of the shoot is made obvious by the Material was either dissected fresh before fix- arrangementof axillarybuds. In nature,the plane ation or afterfixation in 70%F.A.A. For the study of distichy is horizontalso that plants spreadover of shoot and floral ontogeny fixed spadices were the surface of the substratewith all leaves simi- dehydrated to 95% ethyl alcohol, stained over- larly erected by unequal growth at the base of night in 1% acid fuchsin in 95% alcohol, and each petiole. photographedusing epi-illumination techniques The regenerative renewal shoot is developed (Poslusznyet al. 1980). For scanningelectron mi- by syllepsis (Halle et al. 1978) and is adnate to croscopy(SEM) spadices were dehydrated through the parent axis for about half the associated in- an ethanol series into absolute ethanol, critical- ternode (dotted line in fig. 2). A bicarinatemem- point dried in a SamDri PVT. 3 critical-point branous prophyll is present in the adaxial posi- dryer and coated with gold palladium for 5 min tion (P in figs. 1, 2, 3B). The first foliage leaf on in a Hummer III sputter coater. Spadices were the renewal shoot is inserted on the same side as viewed using an AMR 1000 SEM set at 10 kV at the prophyll, not the opposite side, as would be the Museum of ComparativeZoology of Harvard expected with distichous phyllotaxis (L'o in figs. University. For the study of histology and vas- 1, 2, 3B, 6A). This unusual arrangement also culature at several stages of development, ma- characterizes proliferative vegetative shoots, terial was embedded in paraplast after routine which arise in the axils of each foliage leaf from dehydration in a tertiary butyl alcohol-ethanol dormant buds by prolepsis (PS in fig. 3D). Only series, serially sectioned at 7-10 um in longitu- axillarybuds farthestfrom the shoot tip grow out dinal and transverse planes and stained in saf- and form proliferative shoots with the degree of ranin and alcian green. Serialsections of spadices suppressionof buds increasingmarkedly toward and shoots were analyzedby recordingindividual the shoot tip. The ultimate (Lu) and the unusually sections on a Panasonic GX-4 model AG-1950 placed (Lo) foliage leaf usually lack axillarybuds. multifunction video cassette recorder using a Damage of the leader shoot brings about the re- Javelin Model JE3012 video camera attached to lease of previously dormant buds. a Wild compound microscope. This method of Individual foliage leaves show a regularsym- analysis was complemented by drawings of sec- metry, which influences overall shoot symmetry, tions made with a Wild drawing apparatus.Ad- describedas antidromousby Engler(1877). Each ditional specimens for the study of vasculature leaf has a basal open sheath with overwrapping were preparedby clearingthick sections of older margins, an extended petiole, and a somewhat spadices in 5%alcoholic sodium hydroxide, fol- hastateblade with convolute vernation.The blade lowed by lactic acid, afterwashing in water.These margin of each leaf overwrapsin a direction op- specimens were stained in 1%basic fuchsin, de- posite to that of its sheath;only the overwrapping hydrated, and cleared in xylol. of sheaths is shown in figure 1. Leaves may be designated as right handed or left handed as de- Results terminedby direction of overwrap.Leaves on the same side of the axis alwaysoverwrap in the same SHOOT MORPHOLOGY direction (i.e., leaves in one orthostichy are al- Shoots of Calla are sympodial, each unit end- ways either righthanded or left handed), whereas ing in a terminal spadix (Sx) with an enveloping leaves in the opposite orthostichy always have spathe (Se) (figs. 1, 2, 3C). The regenerativeor contrasted handedness (fig. 1). There is thus a renewal shoot (RS in fig. 3A, B) arises from the consistent contrast in leaf symmetry (antidrom- axil of the penultimate foliage leaf (Lp in figs. 1, ous) except that the renewal shoot can originate 2). The renewal shoot may develop as an ex- in the axil of either a left-handedor a right-hand- tended vegetative axis that either overwinters as ed leaf. a condensed shoot and grows out the following The mouth of the leaf sheath is prolonged as spring(in about 60%of examples) (figs. 1, 2A) or an extended ligule, which encloses all younger immediately proceeds to form a second spadix organs (lig in fig. 3A, B). The prophyll of the on an axis with only three foliage leaves (fig. 2B). regenerativeshoot (P in fig. 1) does not encircle An extended renewal shoot again arises in the the axis at its insertion; symmetry is determined axil of the penultimate foliage leaf of the first distally by its overwrappingmargins (dotted lines renewal shoot (L'p in fig. 2B) and subsequently in fig. 1). Prophyllsof proliferativeshoots encircle overwintersas above. The reproductivepotential the axis completely and overwrapin the manner of a shoot may thereforebe one or two spadices of foliage leaves (Pp in fig. 1). SCRIBAILO & TOMLINSON-DEVELOPMENT IN CALLA 3

dormant renewal shoots was 5.4 (SD = 1.2) although the range was from four to eight leaves. pp ~~~~~~~~~~La On shoots producing two spadices the first renewal shoot always terminated in a spadix after the production of three foliage leaves. The sub- sequentlyproduced renewal shoot thus tended to have only four or five leaves compared with a maximum of eight found in shoots having only \ f A - # a single spadix. The two or three outermostleaves W ~~~~~~~~Lu present in terminal shoots at the time of over- winteringwere alwaysreduced, although they still B (r'-''', p- g W - ~~~~~~...... ,k -'-'::':--::--... It S P possessed the morphology of foliage leaves. The series of parts illustratedin figures 1-3 can only be exposed by dissection of the overwintering renewal shoots. These observations indicate that the majority of leaves are formed in the season of growth, as the renewal shoot extends. Floral expansion on the preformed spadices occurs in early springwith firstflowering occurring in May, although floweringcan continue into late June as the second spadix present on some plants ex- pands. Each previous renewal shoot dies in late summer up to the Fig.1 Calla palustris.Diagrammatic transverse section of region of short intemodes, several floweringshoot with renewalshoot in axil of its penultimate intemodes below the point of last year'sflowering foliageleaf (Lp).Foliage leaves, hatchedexcept L'o; prophylls (i.e., below the node of originof the renewalshoot; (P) of regenerativeshoot (Pp) of proliferativeshoot, black; fig. 3C) so that any population is representedby spathe (Se), stippled; spadix (Sx), heavily stippled; La, an- plantsthat are a maximum of 2 yr old. Senescence tipenultimatefoliage leaf of floweringshoot; Lu, ultimateleaf of floweringshoot; Lo, adaxial foliage leaf of renewalshoot of leaves in the fall months yields rhizomes of not alternatewith prophylland interruptingnormal distichy. the appearanceshown in figure 3C and D where no leaves except those reduced leaves of the terminal shoot are present on the axis in late fall and spring, and older parts of rhizome segments SHOOT PHENOLOGYAND DEMOGRAPHY have collapsed and decayed. The shoot system may be described as "poly- Proliferativeshoots diverge at 900 to the parent phyllous," since there is no fixed number of fo- axis (PS in fig. 3D), growing through the back of liage leaves on each renewalshoot; values ranged the sheath of the subtendingleaf. Because of the from 16 to 30 (x = 23.5, SD = 3.9; N = 31). annual necrosis of basal parts of the shoot each Seasonality of growth is reflected in the long in- daughter proliferative shoot becomes autono- temodes produced during early summer exten- mous from the parent axis within one year of sion and the short intemodes duringlate summer production.The clonal population thereforecon- and fall when dormancy is induced. This series sists of shoots that migrate some 10-50 cm per of short intemodes correspondsto the region of annum, depending upon shoot vigor. The "pe- sympodial branching,which begins in the middle rennial" nature of the plant is thus determined of August and is associated with production of by shoot units that are intrinsicallyannual in their spadices that will flower the following May as life cycle. Despite the common basic plan of shoot mapped in figures 1 and 2. In figure 3C and D organization there is considerable plasticity in the region of short intemodes is to the right. In shoot size dependingon the number of parts pro- figure3C growthwas terminatedwith the spadix duced per year, the amount of intemodal exten- (Sx) now representedby the scar where the in- sion and the number of proliferativeshoots pro- fructescence has abscised. The shoot shown in duced. figure 3D did not flower in the previous year. Many proliferativeshoots do not flower for sev- SPADIX PHENOLOGY eral years until some critical size is attained. Spadices are initiated in the July or August The precise degree of preformation of foliage previous to the year that they will flower. De- leaves on the renewalshoot within the winter bud velopment occurs rapidly such that the majority varies considerablyand is dependent on whether of first spadices on shoots have flowers at the a shoot producesone or two spadices.Of 21 shoots point of early gynoecial closure (fig. 4F) by the examined in the months of September and Oc- time dormancy sets in. Development of first spa- tober the mean number of preformed leaves on dices is well synchronized such that they are at 4 INTERNATIONAL JOURNAL OF PLANT SCIENCES similar stagesof development at the time of overwintering.It is only on the second spadixof shoots producing two spadices that the early develop- Lp mental stage of flowers can be found in the fall months. In these cases spadices were often just showing the beginningof gynoecial initiation (fig. 4D, E) when they went dormant. La Lu ,1 Despite the advanced stage of development of these first spadices on shoots in the fall months, Lu~~~~~~' Se spadices were only one-half to one-third the size of spadices at anthesis the following spring, in- dicatingthat extensive expansion of spadices and Lp flowersoccurs in the springmonths prior to flow- up ering. This is illustratedwell if one compares fig- p ~~~~Se ure 5A (a spadix at time of dormancy)with figure 5E (flowers close to the time of anthesis). Both figuresare the same approximate magnification. La L~~~~~u

FLORAL DIFFERENTIATION Fig.2 Calla palustris. Diagram of shoot constructionin region of flowering.A, Spadix with renewalshoot in axil of The inflorescence axis extends above the pri- Lp. B, Spadix with first-orderrenewal shoot terminatingin a mordium of the spathe as an elongated ellipse. second spadix after producingonly four leaves (P + L'o + L 'p + L'u);second-order vegetative renewal shoot (including Floral primordia are initiated as prominent out- P + L"o)from axil of L'p. Hatchingand labelingas in fig. 1. growths of the surface in such a way that an ir- regular, somewhat angular mound is produced (fig. 4A). There is no subtending bract that pre- imal regions of the spadix are describedlater un- cedes floral initiation. Primordia are initiated in der gender variation. acropetal order, and the whole of the axis is ul- No perianth is initiated. The first appendages timately consumed so that the last-formed pri- are stamen primordiathat arise at the periphery mordium is pseudoterminal. The most obvious of the floral primordium, which becomes flat- pattern is the series of oblique, irregularortho- tened. Stamen primordia originate as protuber- stichies, each with about five or six primordia; ances that are initially dome shaped, soon be- parastichiesare equally irregularso that it is not coming dorsiventral and flattened and clear if the series is spiral or whorled (fig. 4B). subsequentlybecoming bilobed. There is no ob- Primordia may or may not at first be in contact vious circumferentialsequence to the initiation with each other early in development; there is of stamens except for a strong tendency for the considerable variability between inflorescences. first-formedstamen to be in an abaxial position Irregularcontact laterestablishes the angularout- reminiscent of a subtending bract (A in fig. 4A). line of the primordia,which is accentuatedby the An indistinct trimery may be recognized, which initiationof floralappendages. The irregularshape is largely a consequence of the irregularlyhex- of the primordiais correlatedwith the subsequent agonal shape that the primordium adopts when irregularityin sequence of appearance of floral appendagesare first evident. The abaxial stamen parts, so that a uniform acropetal and radially often develops precociously and remains larger symmetricalarrangement of parts, e.g., stamens, than later-initiated stamens until late in devel- is not possible. An ultimate extension of this is opment, particularlyin upper flowers of the spa- the development of "twin" primordia,which re- dix. Later initiated stamens appear in approxi- semble two primordiafused laterallyand always mated centripetal order (fig. 4F, G; stamen in the horizontal plane. primordianext to gynoecium), but it is clear that Description of development of flowersinitially new primordia can be inserted tangentially be- deals with those in the central region of the spa- tween existing primordia. As organogenesispro- dix, which are bisexual; those in distal and prox- gresses the increasing contact between adjacent

--4 Fig.3 Calla palustris.Shoot construction.Bar = 300 ,im, A. Bars = 2 cm, B-D. A, Epi-illuminationphotograph of partly dissected shoot apex; renewal shoot (RS) with enveloping prophyll, ultimate leaf of shoot (Lu) with inflated sheathingbase enclosingspathe and spadix (cf. fig. 2A). B, Laterstage of a shoot similarto A with developingrenewal shoot (RS) and flowering shoot of previous sympodial unit. Bicarinateprophyll (P) and its immediatelysucceeding leaf (L'o) insertedon the same side of the axis. Se, spathe;Lu, ultimate foliage leaf. C, Sympodialunit with leaves removed, scar of spadix (Sx) of previous unit. D, Vegetative shoot of same age arisingfrom a previous terminal bud; short intemodes to right indicate position of shoot at time of previous year's dormancy.PS, proliferativeshoots from previouslydormant axillarybuds. PS

D S 6 INTERNATIONAL JOURNAL OF PLANT SCIENCES floral primordia appears to constrain both the the development of the gynoecium is i-ndicated shape of floralprimordia and sites of stamen ini- by the numbered sequence in figure 4D and E. tiation and expansion (fig. 4C-G). The tendency At no time during initiation of the gynoecium is for stamens to be initiated and to expand at lo- there evidence of the trimerous development of cations of available space between adjacentfloral structuresinterpretable as separate carpels. De- primordia is visible in figure 4C in the flower velopment of the adaxial part of the gynoecium immediately below the arrowhead. lags considerablybehind the abaxialportion until Typical numbers of stamens are nine to 12 per later in ontogeny. The shape of the developing flower, but this is difficult to establish precisely gynoecium varies considerablyand is dependent from surfacemorphology because of the absence upon space available for its enlargement as de- of a limiting perianth and the close packing of termined by continued stamen initiation and ex- flowers.Stamen number per floweris more easily pansion. The gynoecium tends to be somewhat determinedfrom sections, since the xylem of sta- angular according to the onrginalshape of the men connective tracesis orientedtoward the cen- floralprimordium. The centerof the primordium ter of the flower. Barabe and Labrecquereport within the torus remainsflat as the futureplacenta 1O-1 2 stamens per floweron this basis. However, (fig. 6B). as reportedbelow, stamen numberper floweralso Further floral development involves enlarge- varies accordingto position in the inflorescence. ment of the receptacle, so that appendages are Sections show evidence of abortedstamens; these somewhat separated,together with continued up- are always inner and presumed late-differentiat- growth of the gynoecium wall (fig. 4F). As the ing stamens. gynoecium continues to develop, it becomes Further stamen development involves broad- widely flask shaped, and the opening becomes ening of the future connective as the filament progressivelynarrower (figs. 4F, G; 6B, C), ob- becomes slightly bilobed (arrows in fig. 4F, G). scuringthe developmentof the basal ovules which The connective itself becomes bilobed as the sep- can only be revealed by further dissection. Gy- arate microsporanglaare formed. As develop- noecium closure is completed by its distal exten- ment continues, the two lobes of each anther be- sion to form an abbreviatedstyle (fig. 5E, arrow). come delimited (fig. 5C) and are arrangedin a The short hollow canal of the style opens directly divergentmanner. The two halves of each stamen onto the stigma which has no visible papillate remain appressed along their inner margins (as- structure(figs. 5E, 6C). terisks in fig. 5GCE) until anthesis when rapid Ovule primordiaare initiated on the flat, floor- filament elongation produces an evident bilobed like receptacleas closureofthe gynoeciumoccurs. stamenwith the four microsporangiaexternally The ovary has free-centralplacentation. A num- visible.At laterstages of floraldevelopment, as ber of placentallobes are initiated, corresponding the boundariesof individualflowers coalesce, it to the number of ovules that will appear(e.g., six becomesincreasingly difficult to distinguishthe in fig. 5C). The number of ovules present varied flowerof originfor outer stamens. In this respect, widely. The first indication of ovule initiation is it is of somehelp that the planeof dorsiventrality meristematic upgrowth of the flattened interior of the stamensremains somewhat tangential to floor of the ovary (fig. 6B). As development con- the circle circumscribedby the ovary of each tinues, placentallobes become distinguishableas flower. Apart from late interpolationof stamens, distinct protuberances(o in fig. 6C). The ovules furtherirregularity results because stamens may develop as outgrowthsfrom the top of these lobes not complete development of functional micro- with the inner and outerinteguments successively sporangiaor are aborted, as mentioned above. enclosing the nucellus from the proximal end of Gynoecium initiation is marked by the torus- the primordium (Io, Ii, in fig. 6D). like extension of a mound of tissue, immediately within the innermost series of stamen primordia. GENDER VARIATION Initiation of the gynoecium always begins abax- In all spadices examined the distal four to eight ially with gradualuplifting of the ovary wall in a flowers are always male. All male flowers ex- bidirectional fashion until a complete ring pri- amined showed initiation of stamens in the usual mordium is formed. For a brief period the torus fashion but no subsequent gynoecial initiation. is U-shaped(1 in fig. 4E). A progressiveseries in Male flowers typically displayed a central flat-

Fig.4 Calla palustrisSEM (A, B, F, G) and epi-illumination(C-F) photographsof developing spadix. Bars = 100 pm. A, B, Spadixfrom side (A) and above (B) to show acropetalinception of irregularflower primordia. A, Primordiumof firststamen. C, P. E, Successivestages of inceptionof floralorgans; numbers in D and E show successive stagesin the asymmetricinception of the gynoecium. F, G, Later stages of stamen development, with irregularinterpolation of central stamens after gynoecial development;arrows show bifurcationand enlargementof connective. 4,1X,*' Z}4 8 INTERNATIONAL JOURNAL OF PLANT SCIENCES tened region in place of the site of gynoecial ini- In development (fig. 7A-C), traces to the ear- tiation (arrowin fig. 5A, D). Late in development liest-formed stamens are first recognized when furtherstamens may be initiated in this region. the stamen primordiumis only about 30,m high In addition to the presenceof distal male flow- (fig. 7A). The first-formedstamen traces are usu- ers in Calla there is a strongtendency for increas- ally the most deep-seated but can become inter- ing femaleness toward the base of most spadices. connected by extensive anastomoses before the Increasingfemaleness is expressed through both placental plexus is conspicuous (fig. 7C). Differ- decreased stamen number and increased gynoe- entiation of carpel traces within extending carpel cial size and number of ovules (fig. 5B). Distal walls occurs relatively late (cf. figs. 6B, 7B). Car- female flowers typically had two to three ovules, pel traces at this and subsequent stages may ap- while basal flowers more often had six to eight pear basally discontinuous (e.g., fig. 7C) because ovules. The extent of the gradient in femaleness the placental plexus with which they unite dif- can be quite extreme (fig. 5B). Also compare the ferentiates last in terms of recognizablydiscrete size of gynoecia in figure 5A with that in figure vascular strands. With the differentiationof the 5B whereboth figuresare the same magnification. procambial strands of this plexus ovular traces Although perfect flowers below the male flowers are also evident and carpel traces become clearly often had smaller gynoecia than proximal flow- continuous with it and, indirectly, with the axial ers, no flower with a visibly aborted gynoecium system of the spadix via furtherperipheral anas- containing aborted or no ovules was observed. tomoses (fig. 7D). The system is without numer- Perfect flowers are protogynous, as noted by ical regularitybecause of extensive anastomosing Kunth (1892). and variation from base to apex of the spadix. Carpel traces are at a maximum of 12 to 13 at FLORALVASCULATURE the middle height of the mature gynoecium but The spadix includes a series of axial vascular are fewer in the smaller, distal flowers (cf. range bundles within a lacunose parenchyma(fig. 6E); of flower size in fig. 5B). peripheralbundles are somewhat more crowded. Anastomosing between bundles is extensive, not Discussion only in association with flowersbut also between Accurate descriptions of shoot organizationin axial bundles. Figure 7D representsthe main fea- Calla palustrisfollow a long historical precedent. tures of vasculaturewithout showing all bundles The first description of shoots of C. palustriswas of one flower. A conspicuous feature of mature that of Doll (1843), who noted the anomalous flowers is a plexus of anastomosing strands im- position of the first foliage leaf on the same side mediatelybelow the placenta(fig. 7D, p.p.). From as the prophyllon each branchaxis. Wydler(1856) this plexus single traces to the ovules (o.t.) are observed and illustrated the leaf rolling charac- derived directly.The carpelwall includes bundles teristics, symmetry, and the same anomaly in that also anastomose, convergingapically and to phyllotaxis. He noted that the latteranomaly was a lesser extent basally. The carpel bundles at ma- absent from proliferative shoots, a point in dis- turity are continuous with the margin of the pla- agreementwith all subsequentauthors, including cental plexus. Stamens each have a single trace ourselves. (s.t.) that is derived either directly from the axial Braun(1859) repeatedmuch of Wydler's(1856) system (when its origin may be quite deep-seated, descriptionand added details of shoot phenology e.g., fig. 7A), or indirectly from a branch of the including the observations that the history of placentalcomplex. Tracesto stamens on the same shoots could be traced "for many years." This radius sometimes diverge from a common strand observation is contrary to our own that shoots (s.t. in fig. 7C). We saw no direct evidence of are a maximum of 2 yr old and may representa stamen traces to differentflowers divergingfrom variable effect of habitat on shoot survivorship. a common source; however, the extensive anas- Braun primarily discussed the occasional pres- tomosing of strandswithin the spadix before trac- ence of spadices possessing two or three spathes. es to specificorgans can be recognizedmeans that Engler (1877) and Wamstorf (1883) in turn re- indirect connection must occur. peated Braun's description and added details of

--4 Fig.5 Calla palustris. SEM photographsof late stages of floral development. Bars = 500 ,m, A, B, E. Bars = 100 ,um,C, D. A, Entirespadix from the side with fully formed flowers;arrows indicate failure of gynoecium to develop, producingmale flowers.B, Spadix with strong female tendency, the basal flowerswith precociousgynoecia and few stamens;arrows indicate incompleteclosure of gynoecialtorus. C, Flowerwith top of gynoeciumremoved to show basal placentae(arrow), each of which will producean ovule. D, Spadixfrom above to show apical wholly male flowers.E, Matureflowers in overwinteringcondition; arrow,stigma; asterisks, septum between pairs of anther sacs in individual stamens (connective obscured,cf. fig. 4G). 4~~~~~~~~

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n~~~ f4~~~~~~~~~~~~~~~~~~~~~~~~O

IL 10 INTERNATIONAL JOURNAL OF PLANT SCIENCES furtherirregularities including a second spadix in mens may make contact with carpel traces only the axil of the ultimate foliage leaf. indirectlyvia bundles of the placentalplexus. The We have never observed the production of a diagram by Barabe and Labrecque(their fig. 8) second spadix in the position discussed above. seems purely imaginary, although it is the basis Rather, our observations indicate that when sec- for their recognitionof a pseudomonomerousgy- ond spadices are producedthey occur with a high noecium. Hence, the tricarpellateancestral model degree of regularity,as illustrated in figure 2B. is used to generate supporting data in a purely None of the earlier authors discuss the presence circular way. In their cladistic analysis Barabe of spadices in this position. and Forget(1987) conclude that Ca/la should not Our observations confirm on the basis of de- be included in the Calloideae but suggest no al- velopmental evidence the absence of a perianth ternative placement. A comparison between it in Calla,as reportedby all earlierauthors. Engler and the othermembers ofthe Calloideaeof Krause (1884) mentions that no perianth was visible at (1908) may not be appropriate.One could there- any stage of development but does not indicate fore choose other outgroupsto createan alternate how young were the flowers he examined. De- typologybecause numerous other membersof the velopmentalinformation also establishesthe lim- family have unilocular ovaries. The implication ited radial symmetry in floral organization,since that a trimerous gynoecium is a necessary pre- at no stage can a precise trimerous condition be cursorto all such types seems unfounded.Eckardt recognized,either in the sequence of stamen ini- (1937) used the criterion of three lobes to the tiation or in the differentiationof the gynoecium. stylarapex as evidence of pseudomonomery.Our At most there can be an irregularhexagonal out- figures4D-G and 5A, B show that the closure of line to the flower, seemingly the result of close- the stylarcanal is irregulareven though figure5E packing as the separatefloral primordiaimpinge (arrow)might be uniquelyselected to show a three- on each other.Bilateral symmetry may be claimed lobed condition. by the initiation of the first stamen in an abaxial In addition, Barabe and Labrecque(1983) in- position and the U-shapedtorus of the young gy- terpret the outer stamens of Ca/la as modified noecium. tepals, largely on the basis again of comparison On the basis of floral vasculatureand compar- with putative relatives, especially Orontium,that ison with Orontiumand Lysichiton,Barabe and has six stamens. No developmental evidence for Labrecque(1983) suggestthat the flower is trim- this interpretationis forthcoming, since stamens erous in its organization and that six of the 12 are all initiated in a like manner and the absence stamens of Callacorrespond to the six tepals of of precise symmetry does not allow any recog- Orontium.Furthermore, they suggestthat the gy- nition of successive whorls of floral parts on the noecium is pseudomonomerous and is derived basis of position. from three carpels, primarily because they iden- The desire to see structuresin terms of ante- tify three principalcarpel traces amid the 10 bun- cedent evolutionary "types"that provide the ba- dles that run in the carpel wall. These principal sis for an interpretative"Bauplan" is also reflect- bundles were recognized because they were said ed in the work of Eyde et al. (1967) on floral each to share a common origin with one of the vasculaturein aroids. Certainflowers are referred stamen traces and were the sole suppliers of the to as structurally"aberrant"" and are avoided in single circumferentialbundle that becomes the vascular analysis by choosing only those at the supply to the series of basal ovules. Additional center of a group of flowers from the mid-level bundles of the carpel wall are derivatives of the of a spadix or the center of a group of unisexual principalbundles; they are describedas "second- flowers. Such rejectedflowers, of course, are only ary bundles" and are not indicated in their in- aberrantin relation to some hypothetical ances- terpretativediagram (fig. 8 of Barabe and Labr- tral type to which all are expected to conform. It ecque 1983). It was suggested that the principal should be emphasized that aroid flowers do not bundles of the gynoecium correspondto the dor- function independently, since the spadix is the sal bundle of three separate carpels. Two of the unit in reproduction.Some appreciation should derivative bundles are then consideredto be ven- be shown for meristic variation within a spadix, tral. However, we have shown that in ontogeny especially in those aroids that possess some per- there is no evidence for a trimerous condition. fect flowers, since deviations from a hypothetical Furthermore,our observation of floral vascu- hermaphroditicground plan may be the raw ma- lature differs considerably from that reportedby terial upon which natural selection could work. Barabeand Labrecque(1983) but agrees in prin- The developmentalvariation within and between ciple with the brief description by Hotta (1971). aroid flowers needs to be surveyed more com- We find no evidence for a consistent number of pletely. primarycarpel traces;the placental plexus is not A morphogeneticinterpretation of the vascular represented by a single girdling trace, and sta- system of the Calla flower seems a realistic al- 9"'t~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~I

I 10

Fig.6 Calla palustris.Spadix anatomy. Bars = 100 Am,A-D. Bar = 1 mm, E. A, Transversesection of shoot in region of renewalshoot; Sx, peduncleof spadix (below spathe,which does not appearat this level);Lu, ultimatefoliage leaf of sympodial unit; P, prophyll;L'o, first foliage leaf of renewal shoot inserted on same side as P. B, Transversesection of spadix with developing flower;arrow, floor of ovary as future placental region;A, anther; G, carpel walls. C, Later stages with flower in longitudinalsection, the gynoeciumclosed; o, young ovules. D, Flowerin transversesection, placentaewith developing ovules, each with outer (Jo)and inner (Ii) integumentsdeveloping; GI, gynoecium wall; arrow,filament of stamen with a single vascular bundle. E, Longitudinalsection of immature spadix to show lacunose constructionand terminal portion with wholly male flowers(arrow). 12 INTERNATIONAL JOURNAL OF PLANT SCIENCES ternative to evolutionary hypotheses. Anasto- moses between inflorescencetraces are partof the axial vascular system before floraldifferentiation begins. The first visible traces to the flower are those of the first stamens, each of which becomes S I a morphogeneticpole that becomes connectedby eQ a0 o 0 0 0 procambialdevelopment to the axial system, by 0 dedifferentiation of intervening parenchyma. Carpeltraces within the gynoeciumoriginate later and may appearto be discontinuousbasally. They may correspondto the bundles of the "demand" type in the interpretationof monocotyledonous branch vascular attachment (Zimmermann and f~~~~~~~~~~~~~ Tomlinson 1972). Carpel traces connect either --~~~~~~~~~~~~~- with each other within the gynoecium or with the a,b.~ ~ ab developing placental complex below the flower, which comes to serve as the dominant morphogenetic pole in later stages of floral development since this region retainsits meristematiccapacity longest (cf. fig. 6B, E). Traces to stamens may also make the connection and so give the im- pression that a stamen and carpel bundle have a common origin. Developmentally this is not pos- Fig.7 Callapalustris. Diagrams of floralvasculature drawn sible; one cannot invoke a vascular system dom- from severaladjacent sections superimposed on one structural outline; transversesections in A and B, R L.S in C and D. inated by carpel traces that clearly appearlate in Bars = 200 gtm;a.b., axial bundle;f floral primordium;g, floralontogeny. Anastomosingbetween morpho- gynoecium;g.t., carpel trace within gynoecium; o.t., ovule genetic poles accounts for the lack of regularity trace;p.p., placentalcomplex; s, stamen;s.t., stamentrace. A, in the system because the floweris not very sym- Whole spadix, first stamen bundlesjust differentiated;meri- metrical and the gynoecium is never obviously stematic region of floralprimordia, cross-hatched. B, Young flowerwith gynoeciumalmost closed; carpeltraces present in trimerous. Consequently we are forced to reject gynoecium wall. C, Older flower to show extensive anasto- the notion of pseudomonomery in Calla since mosing of stamentraces, still independentof placentalplexus. there is no structuralor developmental evidence D, Fully developed flower, with well-developed placental for its existence. plexus, continuous via anastomoses with carpel and stamen Our own observations on vascular anatomy traces. make the point that despite the discreteness of the floral primordium, the vascular supply to a Acknowledgments single flower does not have unitary organization. This researchwas supportedby a Natural Sci- Appendages of a single flower are irrigated in- ences and EngineeringResearch Council of Can- directly from several different axial bundles of ada postdoctoral fellowship to R. W. Scribailo. the spadix because of the anastomosingnature of We thank Ellen Moriartyfor preparingfigures 1 the vascular supply. and 2.

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