Taxonomic notes on the Rhynchosia densiflora group ( Phaseoleae , Fabaceae ) in South Africa and its segregation from Rhynchosia section Arcyphyllum

Rhynchosia sect. Arcyphyllum Torr. & Gray is based on the generic name Arcyphyllum Ell., erected by Elliott (1818) for three species previously placed in Glycine L. by Pursh (1814), viz. Arcyphyllum simplicifolium (Walter) Ell. [=Rhynchosia reniformis (Pursh) DC.], Arcyphyllum difforme Ell. [=Rhynchosia difformis (Ell.) DC.] and Arcyphyllum erectum (Walter) Ell. [=Rhynchosia tomentosa (L.) Hook. & Arn]. However, Elliott (1824) transferred these species back to Glycine without any reference to the name Arcyphyllum. Torrey and Gray (1838), in their treatment of Rhynchosia in North America, recognised the two sections R. sect. (Eu)Rhynchosia, with one species (Rhynchosia caribaea DC.), and Rhynchosia sect. Arcyphyllum (Ell.) Torr. & Gray, with three species [Rhynchosia menispermoidea DC., Rhynchosia tomentosa Torr. & Gray (with five infraspecific variants) and Rhynchosia latifolia Nutt. ex Torr. & Gray]. The distinction between these species was based on growth habit, stem pubescence, stipule shape, leaflet shape and indumentum, flower number, shape of calyx lobes and the degree of connation of the upper lip, as well as fruit shape and pubescence.

[=Rhynchosia difformis (Ell.)DC.] and Arcyphyllum erectum (Walter) Ell.[=Rhynchosia tomentosa (L.) Hook.& Arn].However, Elliott (1824) transferred these species back to Glycine without any reference to the name Arcyphyllum.Torrey and Gray (1838), in their treatment of Rhynchosia in North America, recognised the two sections R. sect.(Eu)Rhynchosia, with one species (Rhynchosia caribaea DC.), and Rhynchosia sect.Arcyphyllum (Ell.)Torr.& Gray, with three species [Rhynchosia menispermoidea DC., Rhynchosia tomentosa Torr.& Gray (with five infraspecific variants) and Rhynchosia latifolia Nutt.ex Torr. & Gray].The distinction between these species was based on growth habit, stem pubescence, stipule shape, leaflet shape and indumentum, flower number, shape of calyx lobes and the degree of connation of the upper lip, as well as fruit shape and pubescence.Endlicher (1840) presented an account of the genus Rhynchosia for America and Australia, listing six sections: sect.Copisma E.Mey., sect.Arcyphyllum Ell.Torr.& Gray, sect.Phyllomatia Wight & Arn., sect.Ptychocentrum Wight & Arn., sect.Pitcheria Nutt.and sect.Rhynchosia, but he did not enumerate the species within these sections.Bentham (1859) discussed ten South American species of Rhynchosia, of which four were newly described, treated in sect.Copisma comprising four species, viz.Rhynchosia phaseoloides (Sw.)DC., Background: Rhynchosia section Arcyphyllum is one of the five sections of Rhynchosia as currently circumscribed.Previous studies in South Africa placed two species of Rhynchosia in this section.Some authors treated the species as a group rather than a section, to avoid phytogeographical confusion because the section is based on the North American generic name Arcyphyllum.Taubert (1894), but he estimated that there were now more than 100 species.In his treatment of the Flora of Tropical Africa, Baker (1871) placed two African species (Rhynchosia debilis Hook.and R. densiflora DC.) in section Arcyphyllum based on their twining habit and the densely clustered flowers.The former species was reduced by Verdcourt (1971) to a subspecies of R. densiflora.In his revision of the genus in South Africa, Baker (1923) retained R. densiflora in this section and added a new species R. connata Baker f.The distinction between these two species was based on the shape of the terminal leaflet (broadly ovate and acuminate in R. densiflora and rhombic-ovate and obtuse in R. connata).A summary of the history of sectional classification of Rhynchosia section Arcyphyllum is presented in Table 1.Baker (1923) further merged sect.Copisma and sect.Orthodanum into sect.Eurhynchosia, now treated as sect.

Rhynchosia.
In two treatments of African Rhynchosia species (Gillett, Polhill & Verdcourt 1971;Verdcourt 2001), R. densiflora was not placed in sect.Arcyphyllum, but treated as a separate group (the R. densiflora group) to avoid phytogeographical confusion because the centre of diversity for sect.Arcyphyllum species is in North America.Based on Torrey and Gray (1838), sections Rhynchosia and Arcyphyllum differ essentially in features of the calyx.In sect.Rhynchosia, the stems are always twining and the calyx is marcescent with subulate segments, of which the lowest one is the longest, while in sect.Arcyphyllum the stems are sometimes erect, or commonly twining or trailing, and the calyx is persistent and foliaceous, with linear or oblong-lanceolate, acuminate segments which are nearly equal.Grear (1978), supported by Fortunato (2000), has concluded that in sect.Arcyphyllum the stems are mostly erect or prostrate and rarely twining.In the South African taxa formerly included in sect.Arcyphyllum, the calyx lobes are foliaceous, linear or linear-lanceolate, and equal or longer than the corolla, but the lowermost lobe is the longest and the stems are climbing as in sect.Rhynchosia.However, the R. densiflora group differs from sect.Rhynchosia in the densely clustered flowers.According to Grear (1978), taxa with trifoliolate leaves in sect.Arcyphyllum tend to have unifoliolate older leaves, but no specimens of the R. densiflora group have this kind of leaf structure (we have also examined some specimens of the East African taxa, i.e.R. densiflora subsp.debilis and subsp.stuhlmannii).For these reasons, we follow Gillett et al. (1971) and Verdcourt (2001) in segregating the R. densiflora group from sect.Arcyphyllum, but refrain from recognising sectional status because preliminary molecular data indicate that Baker's (1923) sectional classification of the South African species of Rhynchosia is not supported (Manyelo 2014).Furthermore, molecular and detailed morphological analyses are currently underway.
The differences are summarised in Table 2. Based on Verdcourt's (1971) circumscription, R. connata falls within the R. densiflora complex in that it has a similar growth form (climbing, procumbent perennial herbs about 0.8 m long), obtuse leaflets and yellow glands on the leaflets.According to Baker (1923), R. connata differs from R. densiflora mainly in the shape of the terminal leaflet (rhombic-ovate in the former and broadly ovate in the latter) as well as stipule shape (ovate in R. connata vs. lanceolate in R. densiflora), and the length of the terminal leaflet petiolule (8 mm -10 mm in R. connata vs. 10 mm -18 mm in R. densiflora).
Examination of numerous herbarium specimens has however revealed that within R. densiflora itself, there is a great deal of morphological variation, and therefore none of these characters can be used for diagnostic purposes in separating these two taxa.It is important to mention that descriptions of R. connata are based on the type specimen only.Verdcourt (1971) observed that the East African plants of the R. densiflora complex exhibit a wide range of morphological variation and as a result concluded that the colour of the glands on the leaves and calyx lobes appears to represent 'a more accurate picture of the evolution of the species' (i.e.R. densiflora).The colour of glands places R. connata firmly in this complex, from which it differs only in the size and the extent of connation of the lobes of the uppermost calyx lip.Therefore, this taxon is here included in R. densiflora as R. densiflora subsp.chrysadenia var.connata (Baker f.) Jaca & Moteetee. As

Materials and methods
Plant material was studied from herbarium specimens housed at Natal Herbarium (NH), Bews Herbarium (NU, formerly called Natal University Herbarium) and the National Herbarium, Pretoria (PRE) (Acronyms from Thiers [2011]).Several field excursions were undertaken by the authors and other groups (including the crew) in search of R. connata.For the floral dissections, flowers were rehydrated in boiling water and mounted in glycerol and illustrations were drawn using a camera lucida attachment.Images of leaf surfaces and anatomical sections were taken using a Zeiss Stereo microscope, 6.3 × micro-lens and a Zeiss compound microscope.For anatomical studies, material from herbarium specimens was treated according to a modification of the method of Feder and O'Brien (1968).Thin sections were made using a 2045 Multicut Rotary Microtome.Staining was performed using the periodic acid Schiff-toluidine blue staining method.For scanning electron microscopy (SEM) studies of the leaf and calyx surface appendages, material was sputter-coated with gold and examined using a Phenon Pro SEM.

Vegetative morphology
Taxa of the R. densiflora group are characterised by mostly twining, erect or prostrate stems and unifoliolate or trifoliolate leaves, although the South African taxa are exclusively trifoliolate.Plants are vigorous climbing, procumbent or ascending perennial herbs, 0.2 m -8.0 m long.Stems are slender, firm-herbaceous, shortly pubescent to densely grey-pilose and glandular (Figure 1).Leaflet shape ranges from elliptic-ovate, rhomboidal to rhomboidal-ovate or almost round.In R. densiflora subsp.chrysadenia and R. densiflora subsp.connata, the terminal leaflets are ellipticovate, rhombic-ovate, obtuse to subacute or apiculate, whereas the lateral leaflets are acute or apiculate, oblique, rounded to cuneate at the base.The vestiture is finely scaly pubescent on the midrib and other veins to softly pubescent on both sides of the leaflets, and both surfaces of the leaflets are covered with small orange glands although these are more abundant on the upper surface (Figure 2).SEM micrographs and cross sections of the leaf revealed that these glands are bulky-capitate and are located in depressions of the epidermis (Figure 3).Cross sections of the petiole of R. densiflora subsp.chrysadenia revealed that the petiole has a somewhat irregular shape with orbicular epidermal cells and a ring of five isolated bundles (Figure 3b).

Reproductive morphology
The R. densiflora group is characterised by flowers mostly in dense sessile, subsessile or pedunculate racemes.The inflorescences are axillary racemes and comprise many flowers, a character that is considered ancestral in Rhynchosia (Grear 1978).The inflorescence varies from 10 mm to 130 mm long or more (Grear 1978).The extent of fusion of the upper calyx lobes is a useful diagnostic character between typical R. densiflora subsp.chrysadenia and R. densiflora subsp.chrysadenia var.connata; in the former, the upper calyx lobes are connate up to halfway, whereas in the latter the upper calyx lobes are connate to more than halfway.The standard petal is glabrous and eglandular.The lamina of the wing petal is generally oblong, without surface sculpturing, and is glabrous.There seems to be some variation in keel petal length and width in R. densiflora subsp.chrysadenia, and in var.connata the keel is slightly narrower (8 mm × 4 mm) than in var.chrysadenia (10mm -12mm × 3 mm -4 mm).Climbing, procumbent or ascending perennial herb 0.   list: 428 (1989).Rhynchosia chrysadenia Taub. in Pflanzenw.Ost-Afr.C: 222 (1895); Bak.f., Legum. Trop. Afr.: 470 (1929).

Distribution and ecology:
Rhynchosia densiflora subsp.chrysadenia var.chrysadenia is widespread from northeast and northwest South Africa,

Diagnostic characters:
Rhynchosia densiflora subsp.chrysadenia var.connata is similar to R. densiflora subsp.chrysadenia var.chrysadenia in growth form, from which it can be distinguished by its generally much shorter 14 mm -33 mm long inflorescence, (opposed to 26-105[-130] mm) and peduncles that are 3 mm -7 mm long (vs. 5 mm -15 mm long).The lobes of the upper calyx lip are connate to more than half the length or sometimes almost connate to the apex, whereas in R. densiflora subsp.chrysadenia they are connate to the middle or below the middle.Baker (1923) mentioned that this species is an ally of R. stuhlmannii Harms (now R. densiflora subsp.stuhlmannii [Harms] Verdc.), which occurs mainly in tropical Africa.

Distribution and ecology:
Rhynchosia densiflora subsp.chrysadenia var.connata is known only from the type locality in Camperdown, KwaZulu-Natal, South Africa (Figure 5), in grasslands at altitudes of about 770 m above the sea level (a.s.l).Several attempts were made to locate live specimens in the area, all of which were futile.