Taxonomic significance of the abaxial lemma surface in southern African members of Helictotrichon ( Poaceae )

Results: Based on whether the lemma surface is smooth, papillate, scaberulous, scabrid or a combination of these, five groups of species are distinguished. All species, except Helictotrichon barbatum which has a smooth lemma surface, have small prickles (scaberulous). Helictotrichon leoninum, Helictotrichon quinquesetum and Helictotrichon rogerellisii have only small prickles, whilst Helictotrichon dodii, Helictotrichon hirtulum, Helictotrichon namaquense and Helictotrichon roggeveldense have, in addition, medium to large prickles (scabrid). Helictotrichon capense, Helictotrichon longifolium, Helictotrichon longum and Helictotrichon turgidulum have a combination of small prickles and papillae whilst Helictotrichon galpinii and Helictotrichon natalense have a combination of all three. A key to the groups and photos of the different types of surfaces are provided.


Introduction
Helictotrichon Besser ex Schult.& Schult.f.(Mashau, Fish & Van Wyk 2010b) [excluding Amphibromus Nees, Avenula (Dumort.)Dumort., Helictochloa Romero Zarco and Tricholemma (Röser) Röser] (Romero-Zarco 2011; Röser et al. 2009) is a genus of C 3 grasses belonging to the subfamily Pooideae and tribe Poeae (Soreng, Davis & Voionmaa 2007) in the Poaceae.The genus, with about 40 species worldwide (Gibbs Russell et al. 1990;Mabberley 2008), is most diverse in the temperate regions of the northern hemisphere, especially Europe, from where it extends southwards through the East African mountains (with about five species) to southern Africa, with 14 species in the Flora of southern Africa (FSA) region and one species in Madagascar.Biogeographically the link between the African and Mediterranean members of the group is via Frank White's Afromontane Archipelago-like Regional Centre of Endemism (Van Wyk & Smith 2001).Schweickerdt (1937) revised Helictotrichon in South Africa and recognised 12 species, one of which was only known from the type collection, and a number of other species only from a very few herbarium specimens.The present availability of more herbarium collections has necessitated a revision of the group for the FSA region, this having already resulted in the description of two new species (Mashau, Fish & Van Wyk 2010a).
Southern African members of Helictotrichon have an inflorescence which is a narrow, open, contracted to spike-like panicle with solitary and pedicelled spikelets.Spikelets are 7 mm -30 mm long (excluding awns), laterally compressed and disarticulate above the glumes and between the 2-6 florets.Glumes are usually shorter than the spikelet, equal or unequal, hyaline or subhyaline, and the apex is acute, acuminate or shortly awned.Lemmas are firmer than the glumes, being firmly membranous to leathery, and they have scarious or hyaline apices.The lemma is rounded dorsally, 5-11-nerved, the apex acute or acuminate and 2-lobed (rarely 4-lobed).The awn, which is longer than the body of the lemma, is geniculate and twisted, or only slightly so, and the callus is hairy.The palea is shorter than the lemma, 2-keeled and ciliate.
As many species of Helictotrichon are difficult to identify using only macromorphology, the primary objective of this study was to investigate the usefulness of the micromorphology of the lemma surface for differentiating amongst the southern African members of Helictotrichon.The elucidation of new features to facilitate identification will contribute towards knowledge of the southern African flora in general and infrageneric diversity of Helictotrichon in particular.Moreover, most species of Helictotrichon are relatively valuable grazing grasses as they remain green until late in winter.Reliable species identification will therefore benefit many end users of plant names, including ecologists, pasture scientists and farmers.

Materials
The abaxial lemma surface features were studied in 14 species of Helictotrichon (Table 1) from the FSA region.Lemmas were dissected from herbarium specimens housed in the National Herbarium (PRE), Pretoria (Table 1).

Procedures
Dry lemmas (24 samples) were mounted on aluminium stubs using double-sided adhesive tape, sputter-coated with gold and viewed under a Jeol J840 scanning electron microscope (SEM).The entire lemma, as well as enlargements of the middle-dorsal and distal (below the awn insertion) areas were photographed and compared.Terminology used to describe the surface features follows Ellis (1979).

Ethical considerations
The study did not involve the collection of any fresh samples as only previously collected dried herbarium material was used.

Results
The lemma provides some of the most useful diagnostic features to differentiate amongst the species of Helictotrichon in the FSA region.In some species the micromorphology of the abaxial lemma surface is variable from the base to the point of awn insertion, but the particular pattern of variability appears to be constant for a species.Taxonomically useful lemma characters as seen with the SEM are compared in Table 2.The main character states are whether the lemma surface is smooth, papillate, scaberulous (prickles small) or scabrid (prickles medium or large) (Figures 1a-h and 2a-f).Prickles are described as small if they are shorter than 42 μm, and as medium or large if they are longer than 56 μm.Based on lemma surface micromorphology the species can be classified into five groups.These are keyed out in Box 1.

Group 1: Lemma surface smooth
Helictotrichon barbatum.A smooth lemma is the most obvious character distinguishing H. barbatum from all other southern African members of Helictotrichon (Figure 1a).

Group 3: Lemma surface papillate, scaberulous and scabrid
Helictotrichon galpinii and Helictotrichon natalense are the only two species which have a combination of all the features that may be present on the lemma surface, namely papillae, small prickles and medium or large prickles (Figure 1d; 2b).

Discussion
Although Schweickerdt (1937) included the lemma surface characters for each species description in his key, he only used it to separate some species that are closely related and then usually only as one of the many distinguishing characters used.In southern Africa, Chippindall (1955) closely followed Schweickerdt's key, whilst Stapf (1899) mentioned the lemma surface in the species descriptions but did not use it at all in the key.Descriptive terms such as granulate, finely or coarsely granular, scaberulous and scabrid are used by the various authors, but there is no doubt that they all refer to the prickles as seen under the SEM and referred to as such in the present contribution.
In addition to all these characters, the SEM shows that the lemma surface of H. dodii has small (scaberulous) as well as medium to large prickles (scabrid The SEM shows that the lemma surface of these two species has a combination of small (scaberulous) and medium to large prickles (scabrid).
Helictotrichon galpinii is easily distinguished by the broad glumes that are as long as the spikelet and the apparently scabrid lemma surface which is easily seen under a light microscope.However, the SEM images show the lemma surface of H. galpinii matching that of H. natalense as both have a combination of all three lemma surface types, namely papillate, small and medium to large prickles, present.Helictotrichon natalense occurs at lower altitudes (1900 m) than H. galpinii and both are summer rainfall species.
Helictotrichon rogerellisii is similar to H. longifolium in that both species have setaceous leaves and loosely flowered spikelets.However, they differ in their distribution (geographical range) as H. rogerellisii has only been found in the Bredasdorp District, Western Cape (Agulhas Plain Subcentre, Cape Floristic Region), in fynbos, mainly in shallow, humic soils between limestone outcrops on coastal plain.Helictotrichon longifolium is found along the    Centre), especially in grasslands, and mainly on moist and rocky mountain slopes.These two species also differ in a number of other characters (Mashau et al. 2010a) and under the SEM the lemma surface of H. longifolium has papillae and small prickles (scaberulous) (Group 2) whereas H. rogerellisii has small prickles only (Group 5).
Using macromorphological characters, H. leoninum is the only southern African species with the rachilla internodes glabrous -this character state makes the species distinctly different from all others.The lemma surface of H. leoninum, H. quinquesetum and H. rogerellisii are covered only with small prickles (scaberulous; Group 5).All three species occur in the winter rainfall area in and around Cape Town, to as far east as Bredasdorp.
Although the lemma surface in the FSA species of Helictotrichon is not sufficiently variable to distinguish between all species, it is still a very useful character for identification.It also allows for species that are possibly closely related to be grouped together.The use of the SEM enhances this character especially to distinguish whether the lemma surface is smooth, papillate or has small prickles mixed with larger prickles -details not seen or not easily identified under the light microscope.

TABLE 1 :
List of species and specimens (all in the National Herbarium, Pretoria) examined using scanning electron microscopy.

TABLE 2 :
Comparison of the abaxial lemma surface micromorphology in southern Africa species of Helictotrichon.