Cordia, L., 1753, L., 1753

Pätzold, Janin, Feyrer, Benedikt, Saumweber, Johanna, Hilger, Hartmut H. & Gottschling, Marc, 2023, Observations on flower and fruit anatomy in dioecious species of Cordia (Cordiaceae, Boraginales) with evolutionary interpretations, Organisms Diversity & Evolution 23 (1), pp. 77-90 : 80-85

Results

Flower and perianth

Flowers are solitary or composed in small inflorescences comprising up to 15 elements. They are arranged terminal % Fig. 1A–C ), ramiflorous % Fig. 1N ) or cauliflorous. The basic monotelic inflorescence is an acropetalous, ebracteose thyrsoid comprised of monochasial paracladia with few scattered trichomes. The terminal flower is usually not developed, and the early paracladia are less branched than the younger ones % Fig. 1C ). The flowers are shortly pedicellate, subsessile or sessile. In C. cf. grandicalyx , a sweet scent is emitted from the flower, and the cup of the anthetic calyx and corolla is full of nectar.

The flowers of all species under investigation are basically tetracyclic and pentamerous. Only occasionally, they exhibit four or six petals and stamens in C. cf. grandicalyx , or they are trimeric or tetrameric in C. crenata . The calyx is synsepalous and completely fused in bud with no visible lobes % Fig. 1A–C, G ). The botuliform or urceolate calyx reaches its final size before anthesis and opens irregularly in C. cf. grandicalyx and C. sinensis % Fig. 1E, F ; no data for C. crenata ). The surface is glabrous, with a few trichomes accumulating at the apex % Fig. 1A, E–G ). In mature fruit, the calyx is dilated but does not enclose the fruit % Fig. 6L ).

The petals are fused to a terete tube for half of their lengths % Fig. 1H ). The free distal lobes are oblanceolate and involute in C. cf. grandicaly x and C. sinensis % Fig. 1D, N ) and triangular in C. crenata %not shown). The colour of the petals at anthesis is creamy white in C. cf. grandicalyx % Fig. 1C, D, K ), pale yellow in C. sinensis % Fig. 1N ) and creamy in C. crenata %H.H. Hilger , pers. obs.). On the adaxial surface of the corolla and below the free parts of the filaments, white scattered trichomes are found in all species under investigation % Fig. 1K ). In C. cf. grandicalyx , dark red spots develop after lesion damage, and the petals turn completely brownish-red after anthesis % Fig. 1H–J ). The corolla wilts but remains with the young fruit, until it ultimately falls off at fruit maturity.

Androecium

Stamens are developed, irrespectively of the flowers being functionally male or female %or bisexual as supposed for C. crenata ). The filaments are fused to the lower half of the corolla tube in all three species % Fig. 1K ). The anthers consist of two thecae and four microsporangia % Fig. 2D, F ). They reach their final size already in bud, when the filaments are still short and unstretched % Fig. 2L ). The anthers are introrse and dorsifixed and in C. cf. grandicalyx , red and blue spots and lines develop on the abaxial surface % Fig. 1D, K ) near the somewhat swollen connective. These markings may correspond to strongly stained tissues in the anatomical sections of C. crenata % Fig. 2A, D, E, H ) and C. cf. grandicalyx .

The anther walls consist of three layers, namely an epidermis, an endothecium comprising one cell layer and a secretory tapetum of one to few cell layers. Cells of the endothecium can be recognised by anticlinal, fibrillar wall thickenings % Fig. 2B, E ). Intracellular crystals are present along the joint dehiscence lines of two adjacent microsporangia in C. crenata and C. sinensis %visible in phase-contrast microscopy: Fig. 2H, K ) but not in C. cf. grandicalyx . The flowers of C. cf. grandicalyx and C. sinensis are functionally female and therefore the microsporangia contain no microspores. The archespore %which usually develops into microspores and tapetum cells nourishing the microspores) remains undifferentiated in both species % Fig. 2F, G, J ). In C. crenata , pollen grains originate by meiosis of the microspore mother cells and are already developed early during anthesis. The pollen grains are binucleate when mature % Fig. 2C ). The thickened exine represents the outer layer of the pollen grains % Fig. 2B, C ).

Gynoecium

A normally developed gynoecium is present in C. cf. grandicalyx % Fig. 3A, B, D, E ) and C. sinensis % Fig. 3C, F ), whereas it is only rudimentary in the functionally male flowers of C. crenata % Fig. 3J, K ). The gynoecium is superior, bicarpellate and coenocarpous-syncarpous in all species, but a style is only developed in functionally female or bisexual flowers. The shape of the gynoecium is obovoid % C. cf. grandicalyx : Fig. 1J ) or ovoid % C. sinensis : Fig. 5A ). Internally, the synascidiate zone is rather short, and the symplicate region comprises the main part of the gynoecium. The symplicate region corresponds to the ventral slit that is orientated perpendicularly to the coalescence plane of the two carpels % Figs. 3E, F, J ; 4N ; 6F ). It continues distally to the asymplicate zone with the first splitting of the style. The gynoecium is glabrous throughout its entire development.

Stomata in the epidermis of the gynoecium are occasionally found in all three species % Fig. 5 ). They vary in size, morphology and arrangement with adjacent cells. In C. cf. grandicalyx and C. sinensis , they are found scattered from the base of the gynoecium % Fig. 5E, J ) through the base of the style and sometimes even on the proximal style. Stomata are more frequent in the distal than the proximal region of the gynoecium of C. sinensis % Fig. 5A, B ). In C. crenata , stomata are found from the base through the equator of the gynoecium. Distal stomata are smaller and appear somewhat vestigial % Fig. 5G ) in comparison to the proximal ones % Fig. 5F ). The precise function of the stomata remains elusive. An inconspicuous, collar-like disc is present in C. crenata % Fig. 5H ) and C. cf. grandicalyx , but is lacking in C. sinensis . No stomata are found on this organ.

In its most distal part, the style separates into four branches and terminates into four papillate stigmata % Figs. 1C, D, H ; 3A–C ; 5A). Each branch has the length of approximately one-third of the mature gynoecium. The stylar branches are rather narrow in bud and with maturation, the lobes expand and become oblanceolate to obovate in shape of the outline. They are folded in bud and subsequently stretched at anthesis. The transmitting tissue originates right above the level of the funiculi and proceeds through the distal carpels and terminates within the stigmata % Fig. 3C, D ).

Each carpel is supplied by a usually prominent dorsal bundle % Figs. 3J , 4N–Q , 6B–D ), whereas lateral and ventral bundles are more or less distinctive. The dorsal bundles continue into the stylar vascular bundles % Fig. 4O, P ). The primary %or true) septa arise from the fused boundaries of the two involute carpels, each bearing two ovules. The architecture of the functional gynoecium is determined by the presence of different secondary septa, namely four basal septa, two false septa and an apical septum. Only the ventral bundles supply the %true) septa % Fig. 4N ). The false septa % Fig. 3E–G, J ) continue to the apical septum, and this structure is developed at a 90° angle to the carpels’ coalescence plane. In the male flowers of C. crenata , the apical septum is absent due to the rudimentary development of the carpels’ apices % Fig. 3K ), but false septa are nevertheless present % Fig. 3J ).

Each primary loculament is twice two-parted by the true and the false septa, resulting in four single ovules each located in its own locule % Figs. 3E, F, J , 4N ). In the regions of the funiculi and of the ventral bundles, crystals are found in the cells % Fig. 3G ). Each ovule comprises one multi-layered integument with a distinctive micropyle and a nucellus enclosing the embryo sac. In C. cf. grandicalyx , the embryo sac is stalked % Fig. 3H ). Ovules in functionally male flowers of C. crenata lack an embryo sac % Fig. 3J, K ).

Fruit and seed development

At early ontogenetic states, the corolla tube is ripped % Fig. 1J ) and finally falls off the fruit. At maturity, the calyx becomes slightly larger in C. cf. grandicalyx but never encloses the fruit. The remains of the style persist in fruit % Fig. 6L, M ). The mature fruit is spherical to ellipsoid in shape. The colour is greenish to yellowish white in C. cf. grandicalyx % Fig. 6M ) and orange in C. crenata and C. sinensis .

In all species, the fruit is a drupe %or deriving from it, see description below for C. crenata ) preserving the basic architecture of the ovary. Pericarp development is complex, and the course of the dorsal and lateral bundles may help to homologise the different tissues in its various expressions. In postanthetic flowers of C. crenata % Figs. 4D–F , 6C, D, F ), the bundles are notably distinctive and embedded in a tissue, whose cells lignify early during fruit maturation corresponding to %an inner portion of) the mesocarp. However, the prospective stony endocarp is still parenchymatic at this ontogenetic stage and only supplied by the ventral bundles % Fig. 6C, D ). The outer mesocarp is parenchymatic as well and comprises a minor portion of the maturing fruit % Fig. 6F ). The mature endocarp is ellipsoid with a smooth surface % Fig. 6K ).

The principal fruit anatomy of C. sinensis is similar to C. crenata . However, the dorsal and lateral bundles are not embedded in lignified tissue but in strongly stained parenchymatic tissue % Figs. 3C, F , 6B ). A comparable strongly stained tissue is also found in C. cf. grandicalyx % Fig. 4K–M ), but dorsal and lateral bundles are only weakly recognisable in this species. In C. sinensis , the strongly stained tissue is sharply delimited from the remaining pericarp % Figs. 4Q , 6B ), but rather scattered within the remaining parenchymatic portions of the pericarp in C. cf. grandicalyx % Figs. 3D, E , 4K–M , 6A, E ). The further the fruit develops, this tissue becomes more intensely stained. In C. sinensis , the tissue consists of intact cells % Fig. 4A–C ) whereas in C. cf. grandicalyx , cellular borders cannot be inferred leading to a syncytial appearance % Fig. 4G–J ). In mature fruits of C. cf. grandicalyx , the portion of the mesocarp directly abutting on the endocarp is flashily pink in stained cross sections % Fig. 6A, E ).

All four ovules in functionally female flower have the same size at anthesis, but seeds develop in reduced number, indicated by fruits with one % Fig. 6B, D, F, J ), two % Fig. 6A, C, E, H ) or three seeds %or at least developed locules: Fig. 6G ). Mature seeds consist primarily of the embryo, whereas the endosperm is sparse % Fig. 6G, J ). The differential growth of seeds results from the asymmetry of the fruit, particularly of the endocarp. The locules with the abortive ovules are left as small, collapsed slits in the endocarp % Fig. 6G, J ).

Dehiscence lines of the endocarp are predetermined already early in development % Fig. 4N , 6A–D, F ). Each locule is opened by two such lines, one of which is structurally connected with a dorsal bundle. The mature embryo consists of reniform, plicate cotyledons and the radicula. Two types of cotyledons could be distinguished: In C. cf. grandicalyx , they have a notably long petiole, and the shoot apex is barely above the substrate % Fig. 6N ) whereas in C. crenata , the hypocotyl is very long, raising the two shortly petiolate cotyledons well above the substrate % Fig. 6O ).

Figures

Fig.1 Flower and perianth of C. cf. grandicalyx %blue frames) and C. sinensis %pink frames; A–C, L–N: field images; D–K: stack shot images). A, B Inflorescence in bud. C inflorescence at anthesis. D Anthetic flower. E Opening calyx. F Calyx, longitudinally opened. G Bud, calyx partly removed to show the corolla still closed. H Postanthetic corolla, androecium %both turning brown) and gynoecium. J Postanthetic corolla with disrupting corolla tube. K Opened corolla tube with stamen insertion and anthers showing red lines and spots %exemplarily indicated by an arrow). L Shrubby habitus. M, N Inflorescence %abbreviations: a, anther; b, bud; c, calyx; co, corolla; f, filament; fl, flower; g, gynoecium; t, trichomes)

◂Fig.6 Gynoecial development, fruit and seedling of C. crenata %yellow frames), C. cf. grandicalyx %blue frames) and C. sinensis %pink frames; A–F light microscopy, G–K stereo microscopy of endocarp, mesocarp removed; L–O field images; TS in horizontal orientation). A, B TS of anthetic flower %note two to three abortive ovules and strongly stained, peripheral tissue). C, D TS of anthetic flower %note two to three abortive ovules and lignifying portions of prospective mesocarp). E Young fruit %note developing endocarp and flashily pink portions of the mesocarp).F TS of postanthetic flower %note three abortive ovules and lignifying portions of prospective mesocarp). G TS of endocarp, with three developed embryos removed %note scanty endosperm).H Endocarp. J TS of endocarp. K Endocarp. L Immature fruits. M Mature fruits. N Seedlings %note short hypocotyl and long petioles of cotyledons). O Seedlings %note long hypocotyl and short petioles of cotyledons; image taken from cultivated plant, accession number 2012–0005, in the Botanical Garden Munich) %LS, longisection; TS, transverse section; ao, abortive ovule; cot, cotyledon; db, dorsal bundle; c, calyx; ec, endocarp; ens, endosperm; ex, exocarp; fr, fruit; h, hypocotyl; int, integument; lb, lateral bundle; mc, mesocarp; o, ovule; pet, petiolus; sty, style; ut, peripheral tissue; vs, ventral slit)

Fig. 2 Androecium of C. crenata %yellow frames), C. cf. grandicalyx %blue frame) and C. sinensis %pink frames; A–K light microscopy; H, K polarised light; L scanning electron microscopy). A LS of functionally male flower %note strongly stained tissue adjacent to connective). B, C Binuclear pollen. D, E TS of functionally male flower %note strongly stained tissue adjacent to the connective). F TS of functionally female flower %note the undifferentiated archespore). G LS of functionally female flower %note strongly stained tissue adjacent to the connective and undifferentiated archespore). H TS of functionally male flower %note crystals at dehiscence line of microsporangia). J, K Secantial section of young, functionally female flower %note deposited crystals). L bud, with calyx and corolla removed %note short filaments) %LS, longisection; TS, transverse section; a, anther; arc, archespore; c, calyx; cn, cell nuclei; co, corolla; con, connective and adjacent tissue; cr, crystals; f, filament; g, gynoecium; ms, microsporangium; p, pollen; ta, tapetum; th, theca)

◂Fig.3 Gynoecium of C. crenata %yellow frames), C. cf. grandicalyx %blue frames) and C. sinensis %pink frames; A, B stack shot images; C–K light microscopy; G polarised light; TS in horizontal orientation). A, B Anthetic female flower, calyx and corolla partly removed. B LS of gynoecium. C LS of functionally female flower %note strongly stained peripheral tissue of corolla, anther and gynoecium). D LS of gynoecium. E, F TS of functionally female flower %note strongly stained, peripheral tissue). G TS of functionally female flower %note crystal deposition). H LS of ovule %note stalked embryo sac). J TS of functionally male flower with non-functional ovules. K LS of functionally male flower %style lacking, original position indicated by an asterisk) %LS, longisection; TS, transverse section; a,anther; bs, basal septum; c, calyx; car, carpel; co, corolla; db, dorsal bundles; es, embryo sac; fs, false septum; lb, lateral bundles; o, ovule; stg, stigma; sty, style; t, trichomes; tt, transmission tissue; ut, peripheral, strongly stained tissue; vb, ventral bundles; vs, ventral slit)

Fig.5 Stomata of C. crenata %yellow frames), C. cf. grandicalyx %blue frames) and C. sinensis %pink frames; A–K, H scanning electron microscopy; D, L light microscopy). A Gynoecium with accumulation of stomata towards the style. B distal part of the gynoecium %arrows indicating stomata). C, D Single stomata. E Stomata preferentially occurring at the proximal part of the gynoecium %arrows indicating stomata). F Stoma from the proximal region of the gynoecium. G Stoma from the distal region of the gynoecium. H Lacerate, collar-like disc %ovary removed). J Proximal part of the gynoecium %arrow indicating stoma). K–M Stomata of varying shapes %co, corolla; dis, disc; ec, ectocarp; mc, mesocarp)

◂Fig. 4 Gynoecium of C. crenata %yellow frames), C. cf. grandicalyx %blue frames) and C. sinensis %pink frames; A–R light microscopy; TS in horizontal orientation). A Secantial section. B, C TS %note cellular organisation). D Secantial section. E, F TS %note cellular organisation). G Secantial section of young gynoecium showing cellular organisation. H–J TS %note lacking cellular organisation, localisation in K–M). K–M LS at different levels from outside to inside of the same specimen %note strongly stained peripheral tissue; asterisks indicate tissue illustrated in H–J). N TS %note dehiscence lines of the prospective endocarp). O, P TS showing transmission tissue and dorsal bundles at top of style. Q vascularisation at base of gynoecium %note strongly stained peripheral tissue).R Vascularisation at base of flower %LS, longisection; TS, transverse section; db, dorsal bundle; dl, dehiscent line; ep, epidermis; lb, lateral bundle; tt, transmission tissue; ut, peripheral tissue; vb, ventral bundle; vs, ventral slit)