167 Morphologically, Diplazium can be distinguished from other close related
genera including from Athyrium by characters combination as follow Kato, 1977: 1 stipe bases neither swollen nor bearing pneumatophores; 2 Frond
axes U-shaped with flat base in most species; 3 Acroscopic basal pinnules equal or smaller; lamina margin not cartilaginous, spines absent; 4 Vein free, or
goniopetrid or sagenoid – anstomousing, ending near the margin; 5 Sori dorsal on the vein and linear, either single Asplenoid or double Diplazioid; generally
Asplenoid sori along the acroscopic side of a vein and Diplazioid sori are confined to both sides of the basal acroscopic; both parts of double sori of about equal
length; 6 Scales entire or toothed with consisting of two upturned ends of adjacent marginal cells. This morphological study see Chapter 9 supported the
previous workers Ching 1940; van Alderwerelt van Rosenburgh 1908; Alston 1956; Sledge 1962; Holttum 1940, 1966; Kato 1977, 1995; Edie 1978; Tagawa
Iwatsuki 1988; Andrews 1990; Kramer et al. 1990. in separating Diplazium from Athyrium
.
8.3.4.2. Relationships among species within Diplazium
In the monophyletic of Diplazium, D. porphyrorachis form a clade alone and diverges earliest Clade I, it is the most basally positioned in all studied taxa
Clade II. But the divergency of D. porphyrorachis from the all taxa studied is without well-supported Bootstrap value. Subsequently Clade II diverges into two
main clades without well-supported Boostrap value, viz. Clade II.1. that contains only D. subvirescens TNgP 1012 and Clade II.2. that diverges again into four
main clades, one main well-supported group, Clade A, with Bootstraping value 100 and three main weakly-supported group, Clade B, C, and D. The
relationships inferred among the species included in these cades are discussed below. Before discussing the relationship inferred among the species, the
polytomy occurrence in the topological tree generated is discussed first. In ferns, polytomy relationship patterns among species on the evolutionary
relationship inferred that generated from molecular data are usual. As showed in the Figure 8.1., Clade A2 diverges into six subclades. Subsequently in the
terminal clade, Diplazium from Japan, Eastern Asia and Central America are also
168 forming a polytomy. The polytomy pattern is also showed in the sister clade of D.
riparium that comprised D. hottae, D. xiphophyllum, D. cordifolium, and D.
angutipinna . Dubuisson et al 1998 showed polytomy relationship pattern of the
sister clade of Trichomanes thysanostomum that consisted of T. minutum, T. bipunctatum, T. speciosum,
and D. diaphanum. In the monophyly of the section Lepidoglosa
Elaphoglossaceae, Elaphoglossum asterolepis, E. splendens and E. rufidulum
formed a polytomy Rouhan et al. 2004. Hauk et al 1993 showed the polytomy pattern in the sister clade of Ophioglossum pusillum that included O.
reticulatum, O. vulgatum , and O. richardsiae. The polytomy patterns indicate that
more data, including more taxa and more informative characters, are needed in studying the relationships of the taxa in the future.
D. porphyrorachis. D. porphyrorachis is the sister clade of the main
clade Clade II. Morphologically, D. prophyrorachis is very distict among the West Malesian Diplazium. D. porphyrorachis posses a suite of morphological
characters distinguishing it from other West Malesian species. As presented in Chapter 6 and 9 this species is having diagnostic characters as follow: Lamina
pinnatifid lanceolate apex lobed or toothed, the lower 23–67 deeply pinnatifid into many or numerous, close, spreading, subfalcate, linear oblong, blunt, slightly
crenate, serrate or toothed segments 8-15 mm broad, which have not seldom the sides entire and only the apex serrate; lower segments gradually growing smaller,
the 1-4 lowest free and deflexed. The position of D. prophyrorachis at the basal tree of the rbcL tree is also supported by the topology of the phylogenetic tree
generated from morphological data. In the tree generated from morphological data, D. porphyrorachis is forming a clade that include its closely related species,
viz. D. fuliginosum and D. lomariaceum and also at the basal clade of the monophyly of West Malesian Diplazium.
As stated by Price 1983 D. prophyrorachis
and its closely related species, D. lomariceum and D. fuliginosum, constitute unusual Malesian Diplazium that agree in the following characters:
rhizome short-erect, with thick wiry black roots; fronds narrowly elliptic, pinnate of subpinnate, apex coadunate, segments numerous, usually dark; scales abundant
on stipe and rachis, narrow, usually dark and shiny; veins free.
169
Figure 8.1. Strict consensus of the 200 equally most-parsimonious trees obtained in maximum parsimony analysis of the rbcL sequence data. Bootstrap values are indicated
above the branches occurring in more than 50 of 100 bootstrap replicates.
170 Clade C. Clade C without well Bootstrap support comprised D.
procumbens 1047, D. asymmetricum 1094, D. subvirescens 1178, D.
asymmetricum 1728, D. procumbens 1281. Due the very low level of Boostrap
value, it is difficult to state the relationship among them. Therefore each species is discussed separately below.
D. asymmetricum. This species in a glance is very similar to D.
procumbens as the two species are having affinity in characters combination:
scales on stipes fallen when mature, dark brown, ovate-lanceolate, margin with blunt teeth; lamina oblong; lobes oblong with blunt or truncate apex; veinlets
forked. Diplazium assymetricum
differs from D. procumbens in the following characters combination: rhizome short, erect; lamina more incised to tripinnate
basiscopic pinnulae or segments and lobes are larger than the acroscopic ones; indusia thicker, margin entire. Therefore it is possible that the two species are
closest related in the phylogenetic tree inferred based on gene rbcL sequence. Diplazium subvirescens.
In the topology of rbcL tree, Diplazium subvirescens
TNgP 1012 constitute the sister clade of Clade II in which this speices is also included in unresolved Clade II.2. TNgP 1178. Cytological
observations on two individuals of this species Chapter 5 showed only triploid apogamous and it is presumed hybrid. Suggestion that this species is originally
hybrids may be support by the evidence from the gene rbcl sequence. Two plants successfully sequenced TNgP 1012 and TNgP 1178 showed differences each
other in 22 nucleotides. This difference number was high enough although morphologically the two collection number are very similar and should be treated
as one species. Morphologically, D. subvirescens is closely related to D. virescens
. As mentioned in the Chapter 9., D. subvirescens has affinity to D. virescens
in the characters combination as follows: rhizome long creeping, black with densely scales on younger part; scales on stipes lineary lanceolate, polish
dark brown with toothed margin; lamina deltoid, deep green, firm herbaceous with veinlets prominent on both surface; sori oblong to linear, medial to supramedial;
indusia thin-membranaceous, laciniate at margin, irregurarly broken at maturity. Two collections number that successfully examined their chromosomes number
TNg 1013 and TNgp 1177 See Chapter 5 showed triploid and the two
171 collentions number are having similar morphological characters with TNgP 1012
and TNgP 1178. Therefore, based on these cytological and rbcL evidence it is suggested that this species are hybrids.
Diplazium procumbens. Diplazium procumbens
distributes on four different clades in rbcL Most Parsimonious tree. Firstly, this species is sister
species of D. pallidum that has subcoriaceous pinnate fronds with lineary lanceolate sharply acuminate toothed pinnae up to 27 pairs less than 3 cm wide
with veins forked 1-3 times and soriferous on upper simple branch, with Bootstrap value 62. Secondly, D. procumbens form a clade with D. dilatatum
TNgP 1011 without supporting Bootstrap value. Thirdly, D. procumbes TNgP 1047; 1281 form a clade with D. asymmetricum and D subvirescens TNgP 1177
without supporting Bootstrap value. The latter, D. procumbens TNgP 1216 form a clade alone and become the sister clade of well resolved of Clade A and
weakly supported of Clade B dan C. D. procumbens
is bipinnate diplazioid fern with rhizome long creeping, very narrow slightly toothed scales, veins copiously forked in the oblong lobes
and sori with thin indusia that break down in the middle. Therefore the relationship between D. pallidum and D. procumbens in the topological tree
generated from gene rbcL is not logical. In the tree generated from morphology, D. pallidum
is closely related to D. prescottianum See Chapter 6 and this relation is more logic.
Cytological observations on eight individuals of D. procumbens from five localities in Java See Chapter 5., Praptosuwiryo Darnaedi 2004, 2005 showed
triploid apagamous. DNA rcbL sequence showed that nucleotides differences within D. procumbens are high enough, viz. from 1 – 10. As mentioned above,
whereas, interspecific nucleotides differences in Diplazium are 4 – 37 Table 8.4..
It is indicate that D. procumbens may be polyphyletic and originally hybirids. In Malesia, D. procumbens is distributed in Malay Peninsula Holttum 1940,
Sumatra present study and Jawa Praptosuwiryo 1999. The position of D. procumbens and D. subvirescens in two or more
different clades may be correlated with the hybrid occurrence in these species. Funk 1985 stated that if hybridization has occurred among the species of a taxon
172 under cladistic analysis the results are varied but always present additional
difficulties. Hybridization results in incongruent intersecting data that obscure the underlying hierarchy.
Clade B. Clade B consisted of D. donianum, D. tomentosum, D.dilatatum and D. simplicivenium without Boostrap support. It is difficult to make a
statement about the relationship among these species due to the very low level of the Bootstrap value, but between D. simplicivenium and D. dilatatum,
D. simplicivenium
may has affinity to D. dilatatum. The two species
morphologically very similar see Chapter 9. Whereas the relationship between D. donianum
and D. tomentosum is difficult to explain and not logic. The two species are morphologically very different See Chapter 9. The affinity of D.
donianum shoud be with D. halimunese not included in rbcL analysis, See
Chapter 9, and D. maoenense Ching. Moore et al 2002 revealed that D. donianum
and D. maoenense are closely related species. D. donianum is different from D. maoenense in the following characters: D. maonense has lobed terminal
pinna wuth free lobes in lower part, pinnae crenate at margin and serrate near apex whereas D. donianum has unlobed terminal pinna, pinnae entire at margin and
slightly serrate near apex. Clade A. Clade A with well supported Bootstrap values level 100
included 30 species in which species from Japan D. chinense, D. rhachidosorus, D. mesosorum, D. cavalerianum, D. sibiricum, D. squamigerum
and D. okudairaekami
, Eastern Asia D. wichurae Central America D. lonchophyllum nested. This Clade diverged into two clade, viz A1 that only consisted D.
megasegmentum and Clade A2 that include 29 species. Subsequently A2 diverged
into five branches that are polytomy: 1 A2.1 had one species only D. poiense; 2 well supported clade of A2.2. 65 that comprised D. bantamense, D.
subserratum, D. speciosum, D. lobbianum , and D. silvaticum; 3 Clade A2.3. that
include two individual of D. umbrosum with strong Bootstrap support; 4 Clade A2.4 that comprised D. accedens, D. polypodioides and D. subpolypodioides
without Bootstrap value support; 5 Clade A2.5. that included D. pallidum, D. procumbens
TNgP 1163, D. esculentum and species from Japan, Eastern Asia and Central America; and Clade A2.6. Subsequently, Clade A2.6. diverges into two
173 monophyletic group with well-supported Bootstrap value, ‘sorzogonense’ Group
Clade A2.6.1 and ‘riparium’ Group Clade A2.6.2. The first group with well- supported Bootstrap value 96 includes two species, D. sorzogonense and D.
batuayauense , meanwhile the second group with Bootstrap value 51 includes
D. cordifolium, D. angustipinna, D. xiphophyllum, D. hotta and D riparium. The
relationships among species within these clades are dicussed below. Clade A.2.6.2 =‘riparium Group’. This study showed that the closely
related species D. angustipinna and D. cordifolium; D. xiphophyllum and D. hottae
inferred from phylogenetic analysis using morphology are supported by gene rbcL sequence data. The monophyletic of ‘riparium Group’ is characterized
by the entire scales and simple pinnate frond with terminal distinct terminal pinnae, terminal pinnae conform to lateral pinnae. In the monophyletic of
‘riparium Group’ D. riparium is at basal clade and diverge earliest. Of all species within ‘riparium group’, but D.riparium, are dryland species. D. riparium grows
on both riparian and dryland areas. In this group D. cordifolium and D angustipinna
are the most closely related and become the sister group of D. xiphophyllum
and D. hottae. D. cordifolium and D. angustipinna have similar morphological characters in glossy light brown lanceolate scales on stipes and
copiously anastomosing veins up to 13-12 way of margin. In the topology of rbcL tree, the relationship of D. xiphophyllum and D. hottae are not resolved.
Tagawa 1972 stated that D. hottae whic is occurring on Malay Peninsula and in Northern Sumatra is allied to D. subintegrum Holtt.
Affinity between D. dilatatum and D. simplicivenium. D. dilatatum and D. simplicivenium
form a clade without supporting Bootstrap value Clade B2. The closest related of the two species in rbcL tree is supported by the similarity in
morphological appearance such as the lineary lanceolate of black-margined toothed scales, the tuft of gigantic bipinnate fronds, and oblong subtriangular
pinnulae. However, in the morphological tree Sees Chapter 6 these species are separated into two different terminal clades, but still in the same middle clade.
Relationships among non West Malesian Species. Clade A2.5.2 without high Bootstraping value, include D. rachidosorus, D. mesosorum, D.
cavalerianum, D. squamigerum, D. wichurae, D. okudairaekami, D.
174 lonchophyllum
and D. esculentum. As revealed also by Sano et al 2000, in the rbcL
tree in this study, D. wichurae from eastern Asia and D. lonchophyllum from Central America consistently form a clade in the rbcL trees and nested among
species from Japan D. chinense, D. rachidosorus, D. mesosorum, D. cavalerianum, D. squamigerum, D. okudairaekam
and West Malesian species. It is indicate that more detailed molecular phylogenetic studies that include
species from wider areas should be for phylogeographic studies of Diplazium. Phylogenetic relationships of D. dilatatum group and the others groups of
Kato 1977. The member of D. dilataum group are spread into many different terminal clades and also positioned at the subbasal clades. Whereas D.
mesosorum dan D. javanicum group are also mixed in the clade in which the
member of D. dilatatum group forming a calde. It is indicate that Diplazium groups established by Kato 1977 was not monophyletic. Therefore the all of
West Malesian Diplazium can not be referred to the Kato’s Diplazium groups. Phylogenetic relationships of the section Anisogonium and Eudiplazium
van Alderwerelt van Rosenburgh 1908. The rbcL tree is congruent with the phylogenetic tree generated from morphological data in drawing the polyphyletic
of the section Anisogonium and Eudiplazium. Both the member of the section Anisogonium
and Eudiplazium are mixed, the species of Anisogonium nested in the many places in which the member of Eudiplazium forming terminal clades.
This study represents the first attempt to explore the phylogeny of West Malesian Diplazium based on gene rbcL sequences. These results showed that
some clades generating from this sequence are congruence with the clades of phylogenetic tree generated from morphological data. However this result is
preliminary because this study examined only 29 species from West Malesia. Molecular phylogenetic study that includes all species of Diplazium from West
Malesia and also use more molecular marker would give the more robust or well resolved phylogenetic hyphothesis tree.
175
8.4. Conclusions
