Morphology and anatomy of the rhizome and frond in the African species of Polystichum (Pteropsida: Dryopteridaceae)

The generic circumscription of the polystichoid ferns within the Dryopteridaceae and their relationships has been and still is much debated. Although floristic accounts for Polystichum are available for many parts of the world, its morpholo­ gy and evolutionary trends within the genus are still poorly understood. In this study, primarily based on the Polystichum species from Africa and neighbouring islands, the morphology and anatomy of the rhizome and frond are addressed. Several species from other genera within the Dryopteridaceae are used for comparison, to gain a better understanding of generic affinities and evolution in this family.


INTRODUCTION
Polystichum Roth is a cosmopolitan genus comprising 180 to 200 species.Although floristic accounts of the genus exist for many parts of the world, a comprehensive modern taxonomic account for the genus as a whole has never been undertaken.The circumscription and definition of species within the genus are still weak.Reasons for the group's inadequate taxonomic status include the occur rence of common and widespread allopolyploid taxa (D.H. Wagner 1979), sterile FI hybrids, apomictic taxa, substantial phenotypic variation within populations and in most cases, a lack of critical morphological studies.
The morphology and anatomy of Polystichum repre sentatives from the Indian subcontinent was studied by Chandra & Nayar (1982) and those of western North America by D.H. Wagner (1979).Polystichum, Arachniodes Blume, Cyrtomium C.Presl, Dryopteris Adans.and Phanerophlebia C.Presl form a closely related group of genera within the tribe Dryopterideae and are here referred to as the polystichoid ferns.Cyrtomium and/or Phanerophlebia are often included in Polystichum (Tryon & Tryon 1982;Kramer 1990).Yatskievych (1996), however, treated Cyrtomium and Phanerophle bia as distinct genera and showed that both have a clos er affinity with Polystichum than with each other, a hypotheses first proposed by Christensen (1930).The genera Arachniodes, Dryopteris and Polystichum are not always clearly separable, as each include species that display characters generally associated with the other genera.The proposed close affinity of these genera is also supported by the occurrence of a natural hybrid between Dryopteris and Polystichum (W.H. Wagner 1985).Widen et al. (1981) showed that phloroglucinols widespread in Dryopteris also occur in Arachniodes but are rare in Polystichum.Indusia, when present, are peltate in Polystichum and reniform in Arachniodes and This study is primarily based on Polystichum species from Africa, Macaronesia, the Madagascan region and the Marion Island groups, but observations on species from outside of this region are also considered.For com parative purposes several species from other genera within the Dryopterideae were included in the study.The aim is to gain a better understanding of generic affinities within the Dryopterideae and to speculate on possible evolutionary trends within the group.

MATERIALS AND METHODS
Material used in the anatomical study was obtained from the wild and from cultivated plants (Table 1).Voucher specimens are housed at the Compton Herbarium (NBG).Fresh material was fixed in FAA for at least 24 hours.For rhizomes, serial sections were cut by hand, stained with alcian blue for one minute, rinsed in water and tem porarily mounted in glycerine.Camera lucida drawings were prepared at x 7.5, x 15 or x 31.25 magnifications using a Wild 'Heerbrugg' microscope.The stelar struc ture of the rhizome was reconstructed using these sec tions.
For describing the vasculature of the stipe and lamina fresh material was used; freehand serial sections were made at 2.5 or 5 mm intervals along the entire length of the axis, noting where bundles divide or fuse and where pinnae and pinnules originate.
Epidermal features were studied using pinna or pin nule fragments removed from selected specimens, and cleared using household bleach.Once cleared the mate rial was repeatedly rinsed in clean water and sem i permanently mounted in either glycerine or glycerine jelly.Cover slips were sealed with Entellan.

Rhizome
Detailed studies of the rhizome of Polystichum are scant and its morphology remains poorly known.The rhi zome anatomy of Cyrtomium falcatum was studied by Gibson et al. (1984) and found to be basically identical to that of Polystichum.Rhizomes are rarely represented in herbarium collections, hence in modern floristic accounts they are often not mentioned.In many species the rhi zome is bulky and this may explain why most collectors are deterred from collecting them.The Polystichum species studied by Chandra & Nayar (1982) in India, all have short, stout, and erect rhizomes that are mostly unbranched.In African species two distinct rhizome types, namely, erect and decumbent occur.The erect rhi zome type is short and generally unbranched with a large number of fronds crowded in the apical region (Figure 1 A).Up to seven leaf gaps overlap at any given time.This rhizome type is sheathed by a mass of wiry roots, with helically arranged persistent stipe bases several layers deep and, at least in the younger parts, also with paleae, giving greater bulk to the rhizome; the highly dissected stellar structure suggests that it is derived.The decumbent rhizome type is usually long and branched, terete to slightly laterally or dorsoventrally flattened; up to five leaf gaps overlap at a time; the fronds are generally more widely spaced, often exposing the rhizome intemodes (Figure 1B).In both rhizome types paleae form a dense covering especially over the apical region.
Branching of the rhizome takes place through the for mation o f lateral buds at regular or irregular intervals along the main stem.Branching in Polystichum transkeiense W.B.G.Jacobsen is regular.In certain habi tats, P. incongruum J.P.Roux, P. monticola N.C.Anthony & Schelpe and P. pungens (Kaulf.)C.Presl also show regular branching.In P. dracomontanum Schelpe & N.C.Anthony and P. marionense Alston & Schelpe, lat eral buds initiate the formation of long stoloniferous out growths.As a result, large clonal stands are formed by these species.Fronds are initially widely spaced along these stolons but marked primary thickening takes place distally, and the fronds become more closely spaced.Roots are irregularly formed over the entire length of the stolons.Since the thinner stoloniferous branches can rot away more readily, individual rhizomes may become iso lated thus serving as a means of clonal propagation.
Branching and stolon formation is here viewed as an adaptation for plants occurring in environments not always conducive to sexual reproduction.Species with this type of rhizome often occur in more exposed, often dryish habitats that may also be subjected to periodic fires.It is therefore also possible that the clonal habit might be an adaptation to periodic burning.The short, suberect to erect rhizome type is, however, much more common in the genus.In contrast to the decumbent rhi zome type, the erect rhizome of the African species mostly remains unbranched resulting in these species occurring as individuals rather than as clonal clusters.Plants with this rhizome type are also adapted to a wide range of habitat conditions, but tend to be more site-specific.The erect rhizome is generally well seated in the substrate, whereas the decumbent rhizome type mostly grows epigeally, although the stolons of P. dracomon tanum are subterranean.Occasionally the rhizome in P. transkeiense may also be subterranean.
Species in the genera Arachniodes, Cyrtomium, Dryopteris and Phanerophlebia have either erect or decumbent rhizomes.If the cyatheoid ferns are accepted as an ancestor of the dryopteroid ferns, as suggested by Bower (1928); Ching (1940); Copeland (1947); Nayar (1970Nayar ( , 1979) ) and Holttum (1973), then the short erect rhi zome should be viewed as the primitive state and the decumbent rhizome as derived.However, a dennstaedtioid origin as proposed by Holttum (1947) andPichi Sermolli (1977) would suggest the reverse.Hasebe etal.(1995) andPryer et al. (1995) presented strong morphological and molecular evidence that Dennstaedtiaceae branched off the tree below the Dryopteridaceae, with Cyatheaceae considerably below that.The evolutionary transformation of the rhizome habit thus remains open to debate.

Roots
An abundance of wiry adventitious roots occurs irregu larly over the entire surface of the rhizome, often form ing a dense fibrous mass.Roots appear to be long-lived but they are, however, only initiated at or near the grow ing apex of the rhizome.Young roots are whitish to pale brown and mostly simple, whereas older roots are dark brown to black and frequently monopodially branched.Golden brown root hairs form a dense covering a short distance behind the root apex.They do, however, lose their function on the older parts of the roots and become abraded.In the erect rhizome type, roots also provide good anchorage.

Fronds
All the Polystichum species within the study area are evergreen; the fronds remain alive for at least two annual cycles.The fronds are persistent and are shed through decay or mechanical damage only.A type of frond re-ori entation as described by Nooden & W.H. Wagner (1997), occurs in P. wilsonii H.Christ.In this high altitude species, which frequently grows in exposed sites, a large percentage of the fronds may collapse during the cold winter months.The plants are, however, never entirely deciduous: Frond length varies considerably among species and in P. incongruum it can attain a length of 1.8 m.There is a strong correlation between the rhizome type and the number of fronds borne by a plant.Species with an erect rhizome always bear more fronds than those with a decumbent rhizome.In P. transvaalense N.C.Anthony, up to 27 fronds are borne by each plant, whereas in species with a decumbent rhizome, the number of fronds rarely exceeds seven.Fronds in the Polystichum species studied are monomorphic, with no evident differentiation between the sterile and fertile fronds.The fronds are dif ferentiated into a distinct stipe and lamina.Vernation of the fronds is initially circinate, but as the stipe and lami na elongate, the juvenile fronds become hook-shaped in species such as P. dracomontanum, P. luctuosum (Kunze) T.Moore, P. transvaalense and P. wilsonii.Monomorphic fronds are most common in the Dryopterideae, but in Dryopteris, e.g.D. dracomontanum Schelpe & N.C.An thony.they are dimorphic.

Stipe
The use of stipe characters in fern taxonomy has been studied by Lin & De Vol (1977, 1978).They clearly illu strate that stipe characters, especially anatomy, are more useful at the generic and family level than at the species level.The stipe in all the studied species of Polystichum is firm.In species with erect rhizomes the stipes grow directly upwards, whereas in species with a decumbent rhizome, they are initially strongly curved upwards.Stipe length and diameter are variable but may be up to 930 mm long in P. incongruum and up to 10 mm diam. in P. volkensii (Hieron.)C.Chr.In most species the stipe base is variously rounded adaxially and abaxially, but distally they all become shallowly to deeply sulcate adaxially.In most taxa the basal part of the stipe is castaneous (brownish) to ebeneous (black) and often lus trous (nitid), becoming paler distally in dried material.In live plants, however, the distal part of the stipe generally rem ains green.The stipe bases of the African Polystichum species appear not to be modified into dis tinct trophopods as described by W.H. Wagner & Johnson (1983).Trophopods have been reported for all the Phanerophlebia species with the exception of P. macrosora (Baker) Underw.(Yatskievych 1996).In most species the dorsolateral aerophore line is conspicuous throughout the length of the stipe, generally being some what paler in colour than the surrounding tissue.In P. luctuosum the aerophore line is often green, thus con trasting strongly with the generally castaneous to ebeneous stipe.The stipe is always clothed with paleae, the density of which shows considerable variation amongst the species.

Lamina
Most Polystichum species in the study area have 2pinnate pinnatifid to 3-pinnate laminae, with 1-pinnate laminae recorded in P. falcinellum (Sw.)C.Presl, P. kalambatitrense Tardieu, P. macleae (Baker) Diels and P. maevaranense Tardieu only.P. macleae has laminae ranging from 1-pinnate to 2-pinnate.Lamina length varies considerably between species with the longest laminae, up to 925 mm, having been recorded in P. volkensii (Hieron.)C.Chr.Lamina outline also shows variatk)n between species, but within a species the degree of variation is fairly restricted.
Pinnae and pinnules are borne subopposite to alter nate on the rachis and secondary rachis.Proximally the pinnae and pinnules are usually short-stalked but distally they become sessile and eventually adnate.They are mostly herbaceous in texture, but in P. dracomontanum and P. marionense, both of which grow in harsh condi tions, the pinnules are usually coriaceous.The lamina is always discolorous with the adaxial surface darker in colour.Adaxially the pinnules are pale to dark green in colour, but in P. dracomontanum and P. incongruum at Hogsback in the Eastern Cape, exposed and older fronds generally turn bronze.
Polystichum is characterised by a largely acroscopic pinna development (Figure 1C, D), but basiscopically developed laminae are often present in P. vestitum (G.Forst.)C.Presl and P. whiteleggei Watts.The degree to which the proximal acroscopic pinnules are developed shows considerable inter-and infraspecific variation.Interspecific variation is exemplified especially in P. drepanum (Sw.)C.Presl, P. incongruum and P. macleae.
In Arachniodes and Dryopteris the laminae are most ly basiscopically developed (Figure IE, F).In Arach niodes and in Cyrtomium and Phanerophlebia, where most species have 1-pinnate laminae, the pinnae are also acroscopically developed and often conspicuously auricled (Figure 1G, H, M).
In Polystichum the pinnae are mostly oblong-acumi nate to narrowly oblong-acuminate in outline, but in some species they may be ovate, narrowly triangular, or in P. marionense often deltoid.In some species the proxi mal pinnae are reduced towards the base of the frond, and often also strongly conduplicate and deflexed.In most species the proximal pinnae are usually widely spaced with no or little overlap with the more distal ones.Towards the lamina apex, however, the pinnae frequent ly become imbricate.The number of stalked pinnae per lamina ranges from 12 to 15 in P. maevaranense, but up to 45 in P. setiferum (Forssk.)T.Moore ex Woyn.Pinna length in most species falls within the 100 to 200 mm range, but in P. marionense the pinnae may reach a maxi mum length of only 36 mm.whereas in P. drepanum the proximal pinnae may be up to 450 mm long.
Pinnules are always inaequilateral, asymmetric and variously ovate, trullate or rhomboid in outline, with an acroscopic auricle.The base is mostly truncate acroscopi cally whilst basiscopically it is mostly narrowly to broad ly cuneate.Margins may be lobate.serrate, doubly ser rate or dentate with the teeth being obtuse, pungent or aristate.Pichi Sermolli (1972), in describing P. kilimanjaricum Pic.Serm., emphasized the number and direction of the pinnule awns, but we found them to be extremely variable and of no taxonomic value.In P. marionense the pinnule margins of plants growing in exposed conditions are often revolute.The number of short-stalked pinnules per pinna ranges from five in P. marionense to 27 in P. zambesiacum Schelpe.Pinnules are mostly small, but in P. drepanum the proximal acroscopic pinnule can be up to 83 x 15 mm.Indumenta occur on both the adaxial and abaxial lamina surfaces of most species with the abaxial surface generally more densely set.

Rachis
Chromatically the rachis does not differ significantly from the stipe, being green throughout, but in older fronds it may become dark brown proximally.The aerophore line that extends from the stipe is generally visible with the naked eye and may be paler or darker in colour than the surrounding tissue.The rachis is mostly straight throughout its length but distally it may become slightly flexuose in some species.Adaxially the rachis forms a V-shaped sulcus along its entire length.In P. luc tuosum the sulcus is shallow and not very prominent.Holttum (1959) stressed the importance of the external shape of the rachis in defining related groups: in the Dryopteris-group of ferns, to which Polystichum be longs, the rachis has a median sulcus that opens adaxial ly to admit the sulcus of the secondary rachises; the raised edges of the secondary rachis sulcus join the sides of the costa sulcus with the edge of the pinnule-lamina decurrent on the secondary rachis as a lateral wing.
Our observations on live Polystichum.Cyrtomium and Dryopteris material show that the rachis sulcus does not always open to admit the secondary' rachis sulcus, as the pinna stalk is often terete, especially in the lower part of the lamina (Figure II.J, K).In Arachniodes, however, the sulci of the rachis and that of the lower order axes are confluent (Figure 1L).Paleae similar to those occurring on the stipe extend to the rachis.Palea density on the rachis is, however, determined by the species and to a lesser degree also by the prevailing environmental con ditions.

Bulbils
Bulbils are here defined as buds or outgrowths capa ble of developing into an independent plant.Although external stimuli may contribute to the formation of bul bils, this ability largely appears to be fixed genetically.In the polystichoid ferns the ability to produce gemmae appears to be restricted to Polystichum, Dryopteris and Phanerophlebia juglandifolia (Willd.)J.Sm.Within Polystichum a diverse group of species is capable of pro ducing bulbils.They have been recorded in sections Lasiopolystichum Daigobo, Metapolystichum Tagawa, Macropolystichum Daigobo, Cyrtomiopsis Tagawa, Stenopolystichum Daigobo and Haplopolystichum Tagawa emend.Daigobo.Bulbils in Polystichum are borne either at the retuse apex of a terminal pinna, at the apex of an extended glabrous rachis or, as in the case of the species within the study area, adaxially on the rachis near the frond apex in or near the pinna 'axils'.One to three bul bils per frond appear to be the norm, but in P. pauciaculeatum Bonap. up to five bulbils are borne by a frond.Paleated bulbils may also occur near the pinna apices in P. tsaratananense Tardieu and occasionally on the stipe of P. setiferum (Moore 1855).African and Madagascan species bearing gemmae include P. crinulosum, P. kilimanjaricum, P. maevaranense, P. magnificum F.Ballard, P. pauciaculeatum, P. tsaratananense and P. volkensii and total 27% of the species within the defined area.Within the Madagascan region 62% of the species are bulbiliferous compared with the 64% of species occur ring in the West Indies (Mickel 1997).
Most taxa which produce bulbils seem to occur in moist or tropical conditions where this form of vegeta tive reproduction may contribute to more rapid colonisa tion in areas of fierce competition.Bulbil formation in the ferns, its distribution and ecological implications, requires further study.The fact that bulbil formation occurs in clearly unrelated taxa suggests that it originat ed independently in these groups.

Venation
Venation patterns in the genera of the Dryopterideae are diverse.This is the principle character commonly used to segregate Cyrtomium and Phanerophlebia from Polystichum.In Polystichum the veins of all the species in the study area are always free and anadromous (Figure 1C.D), but in the circumboreal P. braunii (Spenn.)Fee. at least some of the veins are catadromous (Kramer 1987).Most veins run into the teeth where they always terminate near the lamina margin.In P. volkensii the veins show a slight enlargement towards their apices.
In the fertile pinnules of 2-pinnate species of Polystichum, the veins mostly terminate in the soral receptacle midway between the costa and the margin.This feature is also found in several Arachniodes and Dryopteris species.Variations, however, do occur with the fertile vein often extending for a short distance beyond the sorus.However, in 1-pinnate Polystichum and free-veined Phanerophlebia species, the venation of the fertile pinnae shows no or little differentiation from that of the sterile pinnae, in that the veins bearing sori are not conspicuously shortened, thus extending well beyond the sorus (Figure 1H).Also in most Dryopteris species the sori-bearing veins are not shortened and extend to the margin (Figure IF).Cyrtomium and some Phanerophlebia species have reticulate venation (Figure 1G).Studies showed that reticulation in these genera have different origins (Mitsuta 1977).In Cyrtomium the reticulations have either a costal or a discal origin, whereas in Phanero phlebia they are exclusively marginal (W.H. Wagner 1979).The areolae formed by the reticulations in Cyrtomium all have one or two (often three) free excur rent veinlets on which the sori are borne.In Phanero phlebia the areolae have no included veinlets.

Rhizome: stelar structure
The stelar structure of the Polystichum rhizome can best be classified as a dictyostele as defined by Schmid (1982).By this definition, two or more perforations (leaf, root or branch gaps) overlap along the vascular cylinder.In those Polystichum species with the erect rhizome type, the number of vascular bundles or meristeles (leaf, root and branch traces excluded) may number as many as eight (Figure 2A).In the decumbent rhizome type, how ever, the number is typically lower at four or five (Figure 2B), but in P. monticola up to seven vascular bundles may occur.These bundles are variable in size and shape and are situated approximately in a medial ring.
The rhizome of Cyrtomium falcatum is a radially symmetrical dictyostele but here only three traces vascularise each frond; the lowest one diverging into the abaxial region of the frond axis and the other two traces into the adaxial region of the frond axis (Gibson et al. 1984).
The rhizome branch trace in P. transkeiense is a strand that originates laterally from one of the larger vas cular bundles of the main axis.This trace is dorsoventral and haplostelic as defined by Schmid (1982).The first frond borne by this rhizome branch is not associated with a leaf gap.However, when the second and third fronds develop, non-overlapping leaf gaps are formed.At this stage the stele is still dorsoventral.A true dictyostele is established later.
Root traces branch at irregular distances off the dorsal or lateral surfaces of the main axis vascular bundles and are not associated with root gaps.Chandra & Nayar (1982) claim that they originate from the base of the leaf trace as well, but we were unable to confirm this.The stele in the roots of most species is circular in cross sec tion, whereas in P. luctuosum it is often conspicuously elliptic with the protoxylem centres forming the distal poles.In all the Polystichum species studied the roots were found to be consistently diarch and exarch (Figure 2C, D).In Cyrtomium caryotideum var.micropterum, a taxon often included in Polystichum (Kramer 1990).the roots were found to be tnarch.

Frond: stelar structure o f the axes
The stelar structure of the stipe base is characterised by four to five (rarely seven) vascular bundles arranged in a U-shape towards the adaxial surface.Khullar & Gupta (1979) reported up to nine vascular bundles in P. biaristatum (Blume) T.Moore, whereas smaller species like P. obliquum (D.Don) T.Moore merely have two.In Polystichum the arc is a broken line with the larger vas cular bundles located in a dorsolateral position at either end and the smaller ones between (Figure 2E).The smaller vascular bundles are the first to branch from the lateral ends of two adjacent rhizome meristeles and the larger ones are the last to branch from the rhizome meris teles.A reduction in the number of vascular bundles usu ally takes place along the length of the stipe as some of the smaller bundles merge.An increase often occurs as a bundle may also split to fuse again after a short distance.The xylem strand in the larger meristeles is curved and hooked adaxially, the so-called hippocampus shape when viewed in cross section (Lin & De Vol 1977) (Figure 2F).Xylem bundles in the smaller meristeles are circular to broadly elliptic in cross section.In Dryopteris we observed a consistently larger number of vascular bun dles (up to 12) in the stipe.
Xylem tissue in the vascular bundles is composed of helical and scalariform tracheids with thin plates of xylem parenchyma irregularly dispersed between them.Protoxylem of the smaller and larger vascular bundles has an endarch arrangement.The phloem, which forms a sheath around the xylem, is more prominent in the outer dorsolateral and ventral sides of the xylem.The pericycle consists of large parenchymatous cells two to four cell layers deep.The endodermis is characterised by inner and radial walls that are secondarily thickened and in which numerous simple and branched pits of variable sizes are scattered.Pit apertures are always elliptic in outline.
In all the species of the Dryopterideae studied the arrangement of the vascular tissue in the rachis is essen tially the same as that of the stipe.In the rachis, howev er, there is a further reduction in the number of vascular bundles towards the lamina apex.Also here the smaller bundles initially merge with one another at irregular intervals and often also divide as in the stipe.The final small bundle eventually merges with one of the larger dorsolateral bundles some distance from the lamina apex.The two dorsolateral meristeles eventually fuse and continue as a single vascular bundle to the lamina apex (Figure 2J).
The vascular tissue serving each pinna, branches as a single dorsolateral trace from one of the two larger vas cular bundles.This trace soon divides and the two (rarely three) bundles that are formed run parallel to one anoth er for most of the pinna length before they finally merge near the pinna apex.This condition holds true for 1-pinnate species and those with more compound laminae suggesting the costa of 1-pinnate species to be analogous with the secondary rachis of species with more com pound laminae.Costules and veins branch from one of the two pinna meristeles or the terminal bundle of the pinna apex and remain single-stranded.

Rhizome: non-vascular tissue
In most of the species investigated the non-vascular tissue of the rhizome is distinctly differentiated into parenchym atous ground tissue and sclerenchym a. Histologically the cortex and pith are identical, consist ing of small isodiametric parenchyma cells.Towards the epidermis, however, the cells become compressed with the anticlinal walls somewhat sinuate.A narrow sheath of sclerenchyma several cell layers deep is situated beneath the epidermis.In the outer cortical layer this sclerenchyma usually extends to the root trace.In P. transkeiense a small sclerotic cap is formed on the outer periphery of the root trace only, whereas in P. dra comontanum and P. incongruum a sclerotic sheath often forms around the entire root trace.Chandra & Nayar (1982) also found sclerenchyma bundles associated with the departing leaf trace bundles.Small to large groups of partially to moderately thick-walled sclerenchymatous cells occur at random throughout the ground tissue of most species and often extend to the stipe bases.Their abundance appears to be determined by age and the envi ronment with more cell aggregates present in older parts of the rhizome as well as in plants occurring in exposed habitats.In older rhizomes they appear to be concentrat ed near the larger meristeles.Considering the abundance and distribution of sclereid clusters in the ground tissue of the rhizome, Chandra & Nayar (1982) suggested some evolutionary trends and species relationships within Polystichum.D.H. Wagner (1979) also found the pres ence of sclereid clusters in the rhizome of P. munitum (Kaulf.)C.Presl diagnostic in separating it from P. imbricans (D.C.Eaton) D.H.Wagner where they are mostly absent.However, because of their variability, we consid er them to be of no taxonomic value for the species we studied.The deposition of phenolic substances in the sclerenchymatic clusters in especially the older parts of the rhizome may act as a preservative against decay, thus promoting longevity.
The cortex of Polystichum roots is composed of a dense inner sclerenchymatic sheath and parenchymatous outer cortex.In old roots the outer non-sclerotic cortical layer decomposes.Sclerenchyma cells opposite the pro toxylem poles are generally not as strongly lignified as the rest of the sheath (Figure 2D).Schneider (1996) terms these non-sclerotic cells opposite the protoxylem poles passage cells, a feature that characterises the Davallia type of root.

Frond: non-vascular tissue
In cross section the stipe base is usually circular, tetragonal or transversely broadly ovate in outline.In the upper part of the stipe, however, the adaxial surface is usually variously sulcate, as is the rachis.In cross section the epidermal cells are small and isodiametric in outline and their walls generally densely lignified, similar to the underlying band of sclerenchyma.In young material this layer is interrupted dorsolaterally by the aerophore lines studied by Davies (1991).In younger fronds the lateral aerophore lines are composed of parenchymatous cells with large intercellular spaces and are usually not very rich in phenolic substances.With age, however, this tis sue also becomes sclerified.Stomata occurring on the stipe and rachis are confined to the aerophore line and may be raised but are mostly flush with the epidermis.A tanniferous sclerenchyma layer is deposited on the inner tangential and radial walls of the cell layer adjoining the endodermis.The density of this layer is determined by age with that in older fronds being more prominent.
The lamina anatomy of species with 1-pinnate fronds differs slightly from those species with more compound laminae.In cross section the pinnules are flat with a shal low V-shaped sulcus adaxially along the costa that has slightly raised margins.The costa and veins that are situ ated equidistant between the adaxial and abaxial surfaces are enveloped by a narrow layer of parenchymatous tis sue, a bundle sheath, that stretches from the adaxial to the abaxial epidermis.This tissue becomes lignified as the frond ages.In P. macleae, a 1-pinnate species, the pinna structure is similar except that the costa is signifi cantly enlarged abaxially.This enlarged costa is com posed of parenchymatous mesophyll in which the two vascular strands that run parallel to each other occur.

Epidermal cells
Epidermal cells can provide many characters of taxo nomic value (Stace 1984), such as cell size, shape, ori entation and anticlinal wall undulation.Cell size, howev er, may be influenced by ploidy level and environmental factors (Metcalfe & Chalk 1950).
Epidermal cell size in Polystichum has been studied by Chandra (1977), who concluded that Polystichum has a closer affinity with Cyrtomium than it has with Arachniodes and Dryopteris.Epidermal cells in all the species of this investigation are elongated parallel to the veins and are irregular in shape.Cell size of the adaxial and abaxial epidermis differ, those of the abaxial epi dermis usually being larger.The anticlinal walls of the epidermal cells are generally sinuous to deeply lobed (Figure 2H), but in P. crinulosum (Desv.)J.P.Roux the cells walls are almost straight (Figure 2G).Epidermal cells occurring abaxially along the veins are generally narrow and their anticlinal walls are not as deeply sinu ate as those of the intercostal cells.Cell size varies con siderably between species, with the smallest cells in P. ammifolium and the largest in P. wilsonii (Table 2).
Stomata in Polystichum are confined to the abaxial surface of the lamina and are positioned in the same plane as the epidermal cells, oriented with their longitudinal axis parallel to the lateral veins (Figure 21).Anomocytic, eupolocytic, copolocytic and staurocytic stomata have been reported for Polystichum (Van Cotthem 1970;Chandra 1977;Mehra & Soni 1983).In the species dealt with here, we found eupolocytic stomata to be the most common type, followed by the anomocytic state.A few copolocytic stomata were noted in P. aculeatum (L.) Roth.P. dracomontanum.P. falcinellum.P. incongruum and P. kalambatitrense.Guard cell length is fairly vari able: the smallest stomata were recorded in P. crinulosum and the largest in P. falcinellum (Table 3).In P. setiferum stoma initials often abort during early development.Stoma size appears to be related to ploidy level (Table 3).

CONCLUSIONS
This study illustrates our poor understanding of the genus.Polystichum, like other polystichoid ferns, is characterised by a set of features, most of which are also shared by other members within the assemblage, rather than a set of unique characters (Table 4).The genus can, however, be readily distinguished by an adaxially sulcate rachis with four to five (rarely seven) vascular bundles arranged in a U-shape, axes that are adaxially shallowly sulcate.the sulcus of the secondary axes confluent or not with that of the main axis, 1-pinnate to 3-pinnate fronds with (mostly) acroscopically developed pinnae and pin nules, anadromous free venation, indumentum composed of paleae (rarely also clavate glandular hairs), sori occur ring dorsally on the veins or at a vein apex, and a peltate indusium (secondarily lost in a number of species).
In Polystichum the short, erect rhizome type is the most common, occurring in a wide range of taxa of which many are not considered related and is also wide spread in the other polystichoid genera.Species belong ing to this group are mostly forest dwelling and occur as individuals.Species in the second rhizome type can reproduce vegetatively and have the ability to grow in a wider range of habitats.This clone-forming group can consequently be found growing in forests, forest margins or open habitats.The sharing of resources by clone-form ing plants (Hutchings 1997) may compensate for the fewer fronds produced by each rhizome branch.Chandra & Nayar (1982) and D.H. Wagner (1979) con sider the presence and distribution of sclerenchvmatic cell aggregates in the cortex and pith of the rhizome of taxo nomic value.Our observations, however, suggest their presence and number is influenced by age and habitat and we consequently consider them of no taxonomic value.
Root morphology and anatomy also appear to have little taxonomic value at the species or genus level.In Polystichum both the Dennstaedtia type and Davallia type of cortex occur (Schneider 1996).These root types are widespread within the Pteropsida occurring in sever al groups generally not considered related.The latter type, characterised by the presence of passage cells opposite the protoxylem poles, we observed in P. luctuosum only.All species had diarch roots apart from Cyrtomium caryotideum var.micropterum, where triarch roots were observed, a feature that requires further study.
Frond morphology in African Polystichum, as in most polystichoid ferns, is of the monomorphic type.Only in Dryopteris, however, do several species have dimorphic fronds.This derived feature is widespread and within the Pteropsida must have arisen independently on numerous occasions.The morphology and anatomy of the axes in polystichoid ferns also exhibit some important evolu tionary trends.Holttum (1959) considered the external shape of the rachis and the way in which the shape is modified when a secondary rachis is attached to it, of sig nificant evolutionary and taxonomic importance.He used this feature in separating fern groups with a similar habit and sori.Our observations in Polystichum, howev er, show that this feature exhibits intra-and interspecific variation in the pinnae stalks often being terete rather than sulcate.The pinna sulcus is thus not confluent with that of the rachis.The large number of small vascular bundles in the stipe and rachis of Dryopteris taxa we studied appears to be unique within the Dryopterideae, suggesting it to be of taxonomic value, but the evolu tionary significance of this is not understood.The sec ondary rachis being homologous with the costa in 1-pinnate species of polystichoid ferns suggests frond simpli fication to be derived (Mitsuta 1977;Yatskievych 1989Yatskievych , 1996)).Within the polystichoid ferns as defined earlier, only certain Polystichum and Dryopteris species have the ability of producing bulbils on the laminae.In these gen era one or more bulbils are mostly borne on the rachis behind the apical pinna.In the West Indies, however, most of the Polystichum species bear bulbils, either at a retuse apex of the terminal pinna or at the apex of an extended rachis.The ability to produce bulbils is here viewed as a derived feature.The distinct way in which the bulbils are borne suggest that this feature has also arisen independently on more than one occasion in Polystichum.
Ontogenetic studies show that monopodially branch ed free veins are ancestral and that reticulate venation within the polystichoid ferns is derived (Mitsuta 1977).It is widely accepted that reticulate veins are derived inde pendently in Cyrtomium and Phanerophlebia.Yatskievych (1996) is also of the opinion that within Phanero phlebia it arose independently on two occasions.Sorusbearing veins terminating near the lamina margin are here considered ancestral, whereas modified veins termi nating in the sorus or extending for a short distance beyond the sorus are considered derived.
The epidermal cell anticlinal walls show considerable variation in the degree to which they are undulated and are here not considered to be of any taxonomic value.Polystichoid ferns are all hypostomatic, and the stomata are mostly of the anomocytic type.Our observations confirm that within closely related species ploidy level is reflected in the guard cell length as P. pungens, an octoploid (pers.obs.) has larger stomata than P. incongruum which is tetraploid (Table 3).
Judging by the information provided, more detailed studies on a wider range of species are required if a bet ter understanding of Polystichum, and its affinity with the other genera in the Dryopterideae is to be formed.Further studies may reject or support some of the specu lative evolutionary trends proposed here.The view of Yatskievych (1989) who suggested Polystichum to be polyphyletic, is supported here.