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Plant Systematics and Evolution
ISSN 0378-2697
Volume 299
Number 3
Plant Syst Evol (2013) 299:659-681
DOI 10.1007/s00606-012-0751-0
Comparative morphology of leaf
epidermis in the genus Lithocarpus and
its implication in leaf epidermal feature
evolution in Fagaceae
Min Deng, Qiansheng Li, Shuting Yang,
YanChun Liu & Jin Xu
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ORIGINAL ARTICLE
Comparative morphology of leaf epidermis in the genus
Lithocarpus and its implication in leaf epidermal feature
evolution in Fagaceae
Min Deng •Qiansheng Li •Shuting Yang •
YanChun Liu •Jin Xu
Received: 13 June 2012 / Accepted: 20 December 2012 / Published online: 10 January 2013
ÓSpringer-Verlag Wien 2013
Abstract Leaf epidermal features are considered to be
taxonomically important in Fagaceae. In this study, we
examined and compared leaf epidermal features of 112
specimens, representing 105 species and one variety of
Lithocarpus from China and adjacent areas and Notho-
lithocarpus densiflorus. As a result of the different inter-
pretations of terms in previous studies, trichome
terminology in Lithocarpus and its relatives was re-asses-
sed aiming to reveal the trichome evolutionary patterns in
Fagaceae. Twelve types of trichomes and five types of
trichome bases were detected in Lithocarpus, among which
the broad-based trichome (BBT) is newly reported. Sto-
mata in Lithocarpus are restricted to the cyclocytic type
and their size range is 28.6 ±8.2 lm926.5 ±9.3 lm.
The distribution of epidermal features in Lithocarpus
revealed three distinct morphological groups: glabrous,
BBT, and appressed parallel tufts (APT). The importance
of epidermal features across Fagaceae for taxon delimita-
tion is evaluated. Species of Lithocarpus can be accurately
identified by the presence of APT or flat epidermal cells
combined with non-dark stained subsidiary cells and non-
cutinized trichome bases only, or in addition, fasciculate
trichome bases. The phylogenetic distribution of epidermal
features and their evolutionary trends in Fagaceae is also
discussed.
Keywords Lithocarpus Leaf anatomy Taxonomy
Phylogeny
Introduction
The genus Lithocarpus L. is the second largest genus in the
family Fagaceae, with about 320 Asian species (Govaerts
and Frodin 1998), including 123 species in China (Huang
et al. 1999), and 61 in Borneo (Soepadmo 1972). The
geographical distribution of Lithocarpus spp. extends from
eastern India, southern China, and Japan in the north and
through much of Southeast Asia, including New Guinea
and Malaysia (Cannon and Manos 2001), and it is one of
the dominant tree genera in the evergreen monsoon rain
forests in these regions.
Although the plants in the genus Lithocarpus contribute
significantly to local vegetation, the works conducted on its
taxonomy and systematics are limited. Only two compre-
hensive taxonomical studies have been conducted on
Lithocarpus. Based on reproductive morphology, Barnett
(1944) placed 221 species of Lithocarpus into five sections
and 12 groups. Camus is the most recent author to treat the
genus in its entirety (Govaerts and Frodin 1998). The
subdivision system of Camus (1934–1954) is more com-
plicated; she subdivided 279 species into 14 subgenera,
among which the subgenus Pseudocastanopsis was later
moved to ‘‘fissa group’’ in the genus Castanopsis.This
transfer was accepted by most taxonomists (Barnett 1944;
Forman 1966; Nixon 1997; Huang et al. 1999; Soepadmo
1972), and has been supported by both molecular (Manos
M. Deng S. Yang Y. Liu J. Xu
Shanghai Chenshan Plant Science Research Center,
Chinese Academy of Sciences, Shanghai Chenshan Botanical
Garden, 3888 Chenhua Rd, Shanghai 201602,
People’s Republic of China
Q. Li (&)
School of Ecology, Shanghai Institute of Technology,
Shanghai 201418, People’s Republic of China
e-mail: qianshengli@gmail.com
Q. Li
Shanghai Key Laboratory of Protected Horticultural
Technology, Shanghai 201403, People’s Republic of China
123
Plant Syst Evol (2013) 299:659–681
DOI 10.1007/s00606-012-0751-0
Author's personal copy
and Stanford 2001; Oh and Manos 2008; Chen et al. 2009)
and multiple anatomical studies (Lee 1968; Liu et al.
2009). However, 13 subgenera of Lithocarpus in the work
of Camus (1934–1954) are mostly considered to be sec-
tions rather than subgenera (Cannon and Manos 2001). The
only species distributed in western North America—
L. densiflorus (Nixon 1997) was designated as a new
genus—Notholithocarpus (Manos et al. 2008), the recog-
nition of which is supported by leaf epidermal features
(Jones 1986), pollen morphology and molecular evidence
(Manos et al. 2008). Cannon and Manos (2003) compre-
hensively studied phylogeography of Lithocarpus from SE
Asia. Their results revealed two major clades of cpDNA
haplotypes: one confined to Borneo and the other wide-
spread, but such geographical structures were weak based
on nuclear DNA sequences. The other recent molecular
phylogenetic studies on Lithocarpus were either within a
small region (Borneo) (Cannon and Manos 2000,2001)or
based on a broad sampling to investigate the phylogeny of
Fagaceae (Manos et al. 2001; Oh and Manos 2008).
Leaf epidermal features are informative and valuable for
interpreting relationships at low taxonomical levels in
Fagaceae, and a large number of researchers, for example,
Smiley and Huggins (1981), Jones (1984,1986), Hardin
(1976,1979a,b), Hardin and Johnson (1985), Kvacek and
Walther (1987), Manos (1992), Liu et al. (2009) and
Tschan and Denk (2012) have focused on these charac-
teristics. In Lithocarpus, leaf epidermal features have been
described to varying levels of detail. Jones (1986) com-
prehensively studied leaf epidermal features on a broad
sample of Fagaceae. His work well demonstrated the
potential phylogenetic information in some of these leaf
epidermal features in Fagaceae, but he only sampled 12
species in Lithocarpus. Kvacek and Walther (1987) studied
leaf cuticular characters of megafossils of Fagaceae in
Central Europe and a few extant species. Their study
mentioned ‘‘typical appressed finger-like tufted (to stellate)
hairs only found in Lithocarpus and not met with in other
Fagaceae’’. In more recent work, Zhou and Xia (2012)
studied leaf epidermal features of 52 Chinese Lithocarpus
species. They reported nine trichome types in Lithocarpus,
of which three (fused stellate, appressed laterally attached
(ALA) unicellular and curly thin-walled unicellular tric-
homes) were new to Lithocarpus. They classified the 52
Chinese Lithocarpus species into seven groups based on
trichome types and adaxial epidermal cell wall pattern. All
of the aforementioned studies provided an opportunity to
research and compare more comprehensively the leaf epi-
dermal features of Fagaceae. However, only about 20 % of
the species of Lithocarpus [with an overall total of more
than 300 species world-wide (Huang et al. 1999)] were
surveyed by Zhou and Xia (2012), and except for trichome
types and adaxial epidermal cell wall pattern, the other
epidermal features, such as trichome base, stomata types,
stomatal frequency and size which were thought to be
taxonomically informative were not considered.
Trichome types have been regarded as important in
delimiting species in Lithocarpus (Huang et al. 1999; Zhou
and Xia 2012). Nevertheless, ambiguous descriptions of
hair types and inconsistencies in their naming have led to a
state of confusion in trichome classification. This situation
has serious implications for different studies on Lithocar-
pus and other members of Fagaceae. For example, Zhou
and Xia (2012) considered the ‘‘bulbous’’ type of trichome
equivalent to the capitate or irregularly multiseriate types
as in Jones (1986), but the same structure was regarded as
thin-walled peltate (TWP) by Liu et al. (2009). Further-
more, APT with long rays defined by Jones (1986) was
treated as a subtype of the stellate trichome by Zhou and
Xia (2012). Another example is the TWP trichome detected
in Lithocarpus and Castanopsis by Jones (1986) and Liu
et al. (2009) which was only found in one Lithocarpus
species by Zhou and Xia (2012). A good application using
explicit morphological features to elucidate the phylogeny
should be based on homology and accuracy score of the
characteristic stages. Therefore, such inconsistencies in
trichome terminology increase the difficulty in comparing
the results of the published accounts in Fagaceae, and also
limit the usage of epidermal features for purposes of
identification and systematics.
In the Northern Hemisphere, Fagaceae have rich fossil
records in the Tertiary (Crepet and Daghlian 1980; Jones
1986; Kvacek and Walther 1987; Crepet 1989; Crepet and
Nixon 1989a,b; Uzunova et al. 1997). Due to similarities
of leaf architecture, it is difficult to distinguish leaf fossils
of Lithocarpus,Castanopsis, and Chrysolepis (Jones 1986).
A comparison of epidermal features of Castanopsis,Cas-
tanea, and Chrysolepis reveals the trichome types and
stomatal apparatus as informative in terms of identification
of the three genera (Liu et al. 2009). Appressed parallel
tufts (APT) are believed to be an autapomorphism of
Lithocarpus (Jones 1986; Uzunova et al. 1997). However,
some species in Lithocarpus are without APT (Zhou and
Xia 2012). Whether leaf epidermal features can show that
those Lithocarpus species without APT should be placed in
other evergreen genera in Fagaceae is still unknown, since
no comprehensive studies compared the cuticular features
among those taxa. Therefore, further studies to clarify the
epidermal terminology and a careful comparison of epi-
dermal features in Lithocarpus and other genera in Faga-
ceae were essential to elucidate and understand the patterns
of evolution of epidermal features in Fagaceae. This work
can also facilitate fossil leaf identification of Lithocarpus
and its relatives. The following were the main objectives of
the present study:1. To study the leaf epidermal variability
in Lithocarpus by an extensive survey of species from
660 M. Deng et al.
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available sources and by comparing these with other
morphological features to facilitate grouping of species of
Lithocarpus, elucidate its phylogeny, and the usefulness of
leaf epidermal features to identify extant and fossil leaves
of Lithocarpus and its relatives.2. To compare the different
usage of names for specific trichome types by previous
authors and clarify the terminology used in the present
study;3. To evaluate the evolutionary pattern of important
leaf epidermal characters by using trees obtained from the
most recent molecular analysis.
Materials and methods
In this study 105 species, and one variety of Lithocarpus
covering the ten subgenera of Lithocarpus recognized by
Camus (1943–1954), and Notholithocarpus densiflorus
were examined using light microscopy (LM) and scanning
electron microscopy (SEM). All samples were either col-
lected by the authors or were obtained from KUN, IBK and
CSH. The voucher specimens are listed in Table 1. All
relevant slide mounts have been deposited in the herbarium
of Shanghai Chenshan Plant Science Research Center,
Chinese Academy of Sciences, China.
Leaf epidermal materials were prepared from mature
leaves. Laminas were boiled in water for 30 s, and then
macerated overnight in 1:1 (by volume) hydrogen dioxide
solution and glacial acetic acid at 60 °C. Pieces of leaf
epidermis were stained with Safranin–alcohol (50 %) prior
to mounting in glycerin gel. Prepared cuticles were
observed using an Olympus microscope (Model BX 53,
Olympus, Japan).
To check the consistency of epidermal structures, at
least five slides of leaf material were made from different
parts of a single leaf of each studied species. For com-
parison, stomatal frequency (number of stomata per mm
2
)
was calculated.
The material for SEM observation was directly mounted
on stubs without any treatment, and after coating with gold,
the specimens were examined and photographed under an
SEM (Model S-3400N, Hitachi, Japan).
Comparisons of leaf epidermal features across genera in
Fagaceae were based on the present as well as previous
studies (Jones 1986; Manos 1992; Zhou and Wilkinson
1995; Lou and Zhou 2001; Denk 2003; Deng 2007; Liu
et al. 2009; Tschan and Denk 2012) (Table 3). For the large
genera, e.g., Quercus s.l. and Lithocarpus, the leaf tric-
homes of main sections (subgenera) or groups were
selected to represent epidermal feature diversification of
the genera. The dimorphic state of leaf trichome characters
was scored and mapped onto the most recent molecular
phylogeny cladogram of ITS and CRABS CLAW combined
datasets (Oh and Manos 2008) based on the parsimony
method using Mesquite version 2.75 (Maddison and
Maddison 2011).
Results
Stomata were found only on the abaxial surface of the leaf
lamina in Lithocarpus. The stomata and other epidermal
features were consistent within species, and therefore,
represented reliable characters for taxonomic purposes.
Leaf epidermal features, observed through LM and SEM,
are summarized in Table 2which shows that leaf epider-
mal features show large variations between different spe-
cies. More specific interpretations and illustrations of the
microanatomical features are described below:
Adaxial epidermal cells
The adaxial epidermal cells of Lithocarpus as seen under
LM were usually rectangular to polygonal or irregular in
form, with the anticlinal cell walls usually straight
(Fig. 1a–d) to curved (Fig. 1e–i). Six species appeared to
have slightly sinuous to undulate anticlinal cell walls, such
as L. eriobotryoides (Fig. 1i), L.uvariifolius, and L.fordi-
anus. In most species, the anticlinal cell wall thickness
was uniform, but a ridge-like thickening was present in
L. eriobotryoides and L. elizabethiae (Fig. 1l).
Trichomes and trichome bases on the adaxial epidermis
The surface of epidermal cells was flat without special
ornamentation although coated by a thick wax flake in
most species; both LM and SEM detected a few trichome
types (Fig. 1m–p). Some bowl-like, flat, thin and trans-
parent small structures were usually detected by LM
(Fig. 1a, c–e) and SEM (Fig. 1m, p), which were TWP
trichomes (Fig. 1a, c, e, f, m–o), rather fragile and liable
to be lost while preparing the cuticle for observations,
leaving only a basal portion remaining. Unicellular sol-
itary (Fig. 1m–p), fasciculate and stipitate fasciculate
trichomes (Fig. 1p) were found on the abaxial surface in
five species. These three types of trichomes usually have
a large dark stained base with one to two circles of
radially arranged small epidermal cells surrounding them
(Figs. 5,1i).
Abaxial epidermal cells
The morphology of abaxial epidermal cells was diverse; 41
species had a smooth cuticle (Figs. 2,3,4,5a–d, x), while
others possessed special ornamentation which could be
categorized as globular (Fig. 5e, f, l, p) or papilla-like
(Fig. 5i, j, k, m, n, o, q, r, s, u). The globular or papilla-like
Comparative morphology of leaf epidermis 661
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Table 1 List of species, vouchers and collection localities used for the leaf epidermal study
Species Voucher Collection locality Kept place
of voucher
1Lithocarpus amoenus Tsang, W. T. 22822 Huaiji, Guangxi, China IBK
2L. amygdalifolius Lau, S. K. 27622 Hainan, China IBK
3L. amygdalifolius var.
praecipitiorum
Liang, X. R.63364 Gang-en, Hainan, China IBK
4L. areca Sino-Vietnam Exped. 2388 Fuming, Yunnan, China KUN
5L. attenuatus Chen, H. Y. 6892 Hongkong, China IBK
6L. bacgiangensis Mao, P. Y. 03736 Pinbian, Yunnan, China KUN
7L. balansae Shui Y. M. and Chen W. H. 13676 Lue-chun, Yunnan, China KUN
8L. brachystachyus Hou, K. Z. 73548 Bao-ting, Hainan, China KUN
9L. calolepis Li, Z. J. 1325 Mubian, Guangxi, China IBK
10 L. calophyllus Li, Y. 2127 Ruyuan, Guangdong, China KUN
11 L. calophyllus Chen, S. Q. 13450 Longjing, Guangxi, China IBK
12 L. carolinae Wang, C. W. and Liu, Y. 82574 Pinbian, Yunnan, China KUN
13 L. caudatilimbus Hou, K. Z. 70133 Sanya, Hainan, China KUN
14 L. chifui Gao, X. P. 53679 Ruyuan, Guangdong, China IBK
15 L. chiungchungensis Deng, M. 533 Lingshui, Hainan, China KUN
16 L. chrysocomus Chen, S. Q. 16657 Da-miao-shan, Guangxi, China KUN
17 L. cinereus Zhang, Z. S. 12298 Shangsi, Guangxi, China IBK
18 L. cleistocarpus Yu, P. H. 940 Zhengxiong, Yunnan, China KUN
19 L. confinis Li, H. et al. 790 Yuanjiang, Yunnan, China KUN
20 L. corneus Hu and But 21376 Hongkong, China KUN
21 L. craibianus Shui, Y. M. 0463 Yimeng, Yuexi, Yunnan KUN
22 L. crassifolius Pu-Ge-shan exped. 171 Puge Mount., Vietnam KUN
23 L. cucullatus Teng, L. 7336 Renhua, Guangdong, China KUN
24 L. cyrtocarpus South China Biodiversity Survey Team3263 Shangsi, Guangxi, China IBK
25 L. cyrtocarpus Yen, H. H. Bi-Vitnam Vietnam KUN
26 L. damiaoshanicus Chen, S. Q. 17025 Rongshui, Guangxi, China KUN
27 L. dealbatus Liu, S. E. 19153 Kunming, Yunnan, China KUN
28 L. echinotholus Deng, M. 819 Lingshui, Hainan, China KUN
29 L. elaeagnifolius Chen, S. Q. 11284 Dongfang, Hainan, China KUN
30 L. elizabethiae Chen, W. Y. 6182 Hongkong, China IBK
31 L. elmerrillii Deng, M. 748 Lingshui, Hainan, China KUN
32 L. eriobotryoides N. Guizhou Exped. 183 Kiang-kou, Guizhou, China KUN
33 L. farinulentus Pei, S. J. 59-11284 Mengla, Yunnan, China KUN
34 L. fenestratus Wang, W. C. 10196 Lancang, Yunnan, China KUN
35 L. fenzelianus Liang, X. R 64853 Ding-an, Hainan, China KUN
36 L. floccosus Huang, C. 164285 Fengchuang, Guangdong, China KUN
37 L. fohaiensis Liang, S. F. 59-9365 Mengla, Yunnan, China KUN
38 L. fordianus Kunming working station 57945 Jinghong, Yunnan, China KUN
39 L. glaber Manos, P. S. et al. 1671 Wuyi, Fujian, China KUN
40 L. glaucus Wang, C. 38718 Yuanchun, Guangdong, China IBK
41 L. grandifolius Sun, H et al. 3066 Motou, Tibet, China KUN
42 L. guinieri Gulf Xi-Ke 258 Forest by the beach, Cambodia KUN
43 L. haipinii Wang, C. 37958 Xingyi, Guangdong, China IBK
44 L. haipinii Huang, C. 164372 Fengchuang, Guangdong, China KUN
45 L. hancei Yin, W. Q. 2188 Yuan-Jiang, Yunnan, China KUN
46 L. handelianus Teng, L. 3055 Lingshui, Hainan, China KUN
47 L. harlandii Mao, P. Y. 04303 Yao-shan, Pinbian, Yunnan, China KUN
662 M. Deng et al.
123
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Table 1 continued
Species Voucher Collection locality Kept place
of voucher
48 L. henryi Fu, G. X. and Zhang, Z. S. 1216 Shi-en, Hubei, China KUN
49 L. himalaicus Sun, H et al. 4167 Motou, Tibet, China KUN
50 L. howii Lau, S. K. 28249 Wangning, Hainan, China KUN
51 L. hypoglaucus Qiu, B. Y. 60959 Er-Yuan, Yunnan, China KUN
52 L. irwinii Chen, N. Q. 41798 Lianhua Mount. Hongkong, China KUN
53 L. ithyphyllus Wei, S. F. 121013 Zijin, Guangdong, China KUN
54 L. konishii Saiti, S. 8637 Nan-tou, Taiwan KUN
55 L. laetus Feng, K. M. 5093 Pinbian, Yunnan, China KUN
56 L. laoticus Tao, D. D. 291Tao, D. D. 291 Lue-chun, Yunnan, China KUN
57 L. lepidocarpus Liao, C. C. 1854 Chohsi Hsiang, Yushan National Park,
Taiwan
KUN
58 L. litseifolius Manos, P. S. et al. 1502 Xichou, Yunnan, China KUN
59 L. litseifolius Chen, S. Q. 13474 Longjing, Guangxi, China IBK
60 L. litseifolius Chai, X. T. 58-8652 Xichou, Yunnan, China KUN
61 L. longanoides Wang, C. 40237 Xiangxian, Guangxi, China IBK
62 L. longanoides Chen, N. Q. 41580 Lou-fu-shan, Guangdong, China KUN
63 L. longipedicellatus Deng, M. 503 Lingshui, Hainan, China KUN
64 L. longzhouicus Deng, M. 1000 RongAn, Guangxi, China CSH
65 L. lycoperdon Feng, K. M. 4873 Pinbian, Yunnan, China KUN
66 L. lycoperdon Wang C. W. 82574 Pinbian, Yunnan, China IBK
67 L. macilentus Chen, S. Q. 10270 Canwu, Guangxi, China KUN
68 L. magneinii Mao, P. Y. 04023 Pinbian, Yunnan, China KUN
69 L. mairei Liu, S. E. 16494 Kunming, Yunnan, China KUN
70 L. megalophyllus Feng, K. M. 13242 Ma-li-po, Yunnan, China KUN
71 L. melanochromus Chen, S. Q. 4735 Fangcheng, Guangxi, China KUN
72 L. mianningensis Yin, W. Q. 1363 Tengchong, Yunnan, China KUN
73 L. microspermus Liu, W. X. 531 He-kou, Yunnan, China KUN
74 L. naiadarum Chen, S. Q. 10654 Qiongzhong, Hainan, China KUN
75 L. oblanceolatus Chuang-Jing-Zhi (59)1121 Pinshan, Sichuan, China KUN
76 L. obovatilimbus Hou, K. Z. 74003 Lingshui, Hainan, China IBK
77 L. obscurus Sci. Exped. 1648 Motou, Tibet, China KUN
78 L. oleifolius Chen, D. Z. 679 Da-miao-shan, Guangxi, China KUN
79 L. pachylepis Feng, K. M. 4555 Pinbian, Yunnan, China KUN
80 L. pachyphyllus Wang, C. W. 89985 Longlin, Yunnan, China KUN
81 L. pachyphyllus 780 team 721 Tengchong, Yunnan, China KUN
82 L. paihengii Chen, S. Q. 14240 Da-miao-shan, Guangxi, China KUN
83 L. pakhaensis Mao, P. Y. 04247 Pinbian, Yunnan, China KUN
84 L. paniculatus Wang, C. 43962 Ruyuan, Guangdong, China KUN
85 L. lithocarpaeus Sun, H. et al. 1707 Motou, Tibet, China KUN
86 L. petelotii Liu, W. X. 487 He-kou, Yunnan, China KUN
87 L. propinquus Wang and Liu 82493 Pinbian, Yunnan, China IBK
88 L. pseudoreinwardtii Sino-Russian. 9788 Jinghong, Yunnan, China KUN
89 L. pseudosundaicus Sino-Vietnam Exped. 1385 Qong-shan, Vietnam KUN
90 L. pseudovestitus Teng, L. 2729 Lingshui, Hainan, China KUN
91 L. qinzhouicus Foresty Burea 77 Qingzhou, Guangxi, China IBK
92 L. quercifolius Wei, S. F. 121706 Huiyang, Guangdong, China KUN
93 L. rhabdostachyus Mao, P. Y. 2982 Pinbian, Yunnan, China KUN
94 L. rosthornii Li, G. F. 60925 Nanchuan, Sichuan, China KUN
Comparative morphology of leaf epidermis 663
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ornamentations on epidermal cells were present in 64
species; for example, in L. floccosus (Fig. 6g), L. litseifo-
lius (Fig. 6k), and L. fenestratus (Fig. 6l). In some species,
the thickened areas were all covered by epidermal cells, such
as in L. laoticus (Fig. 6i), L. confinis (Fig. 6j), and L. lepido-
carpus (Fig. 6n). Both flat and globular-papillae thickening
structures can be present on the same species, such as in
L. silvicolarum (Fig. 5n) and in L. skanianus (Fig. 5o).
In most species, the anticlinal walls of the epidermal
cells on the abaxial surface were straight (Figs. 2f, 5b) to
curved (Figs. 2a–d, 5e–s). Eighteen species had undulated
to sinuous anticlinal walls (Figs. 2k, 3b–p).
Stomatal apparatus
All studied species were hypostomatic. The stomata were
confined to small areolar regions of the leaf cuticle, with
each containing ca. 11–29 stomata, and forming rather
dense groups (Fig. 3a). The stomatal size range was
28.6 ±8.2 lm926.5 ±9.3 lm across the species. The
largest stomata were present in L. glaucus (37.6 lm9
35.5 lm) and the smallest (17.9 lm916.0 lm) in
Notholithocarpus densiflorus. The stomatal frequency
ranged from 213 to 574/mm
2
. The lowest and the highest
stomatal frequency were noted in L. grandifolius and
L. uvariifolius, respectively.
The stomata of Lithocarpus species were restricted to
the cyclocytic type. The stomata in N. densiflorus were
mostly anomocytic, but at the center of the areolar region,
stomata larger (25.1 ±2.3 lm920.1 ±3.4 lm) than
others were surrounded by a circle of small epidermal cells
and the stomata were of the cyclocytic type.
The outlines of the pair of guard cells were usually
suborbiculate to broadly elliptical in surface view, with a
length/width (L/W) ratio of 1.1–1.5:1. Guard cells were
often thickened to some degree, and were made up of outer
stomatal ledges or rims. The subsidiary cells were flat. The
stomatal apparatus was easily observable unless shielded
by thick trichome layers. The outlines of the pair of guard
cells of Lithocarpus were usually suborbiculate to broadly
elliptical in surface view, with length/width (L/W) radio of
(1.1)1.2–1.5:1. The stomatal pores, where the guard cells
meet, were almost circular, but truncated in N. densiflorus
(Fig. 5x). While preparing the epidermal samples for LM, a
membranous structure beneath the stomatal pore was
generally noted in 11 flat cuticle species, which was pos-
sibly the remains of the lower anticlinal and/or periclinal
walls of the guard cells, such as in L. corneus,L.eriobot-
ryoides, and L.areca (Fig. 3a, o, p, respectively).
Trichomes and trichome bases on the abaxial epidermis
Trichome types
Twelve types of trichomes were detected in this study.
Detailed explanation of each type is summarized below:
Table 1 continued
Species Voucher Collection locality Kept place
of voucher
95 L. silvicolarum Deng, M. 830 Ma-li-po, Yunnan, China KUN
96 L. skanianus Teng, L. 7382 Renhua, Guangdong, China KUN
97 L. sphaerocarpus Mao, P. Y. 03872 Pinbian, Yunnan, China KUN
98 L. tabularis Mao, P. Y. 4254 Pinbian, Yunnan, China KUN
99 L. taitoensis Manos P. S. et al. 1674 Napo, Guangxi, China KUN
100 L. talangensis Yun, W. Q. 2054 Yuanjiang, Yunnan, China KUN
101 L. talangensis Wu, S. G. 970 Shipin, Yunnan, China KUN
102 L. tenuilimbus Gao, X. P. 50792 Cong-Feng-Pin, Guangdong, China IBK
103 L. touranensis Sino-Vietnam Exped. 1903 Tonkin, Youngfu, Vietnam KUN
104 L. trachycarpus Lin, Z. W. et al. 160 Menghai, Yunnan, China KUN
105 L. truncatus Pei, S. J. 59-10275 Mengla, Yunnan, China KUN
106 L. uvariifolius Zuo, J. L. 22682 Lianxian, Guangdong, China KUN
107 L. variolosus Lianda11632 Chang Mount. Dali, Yunnan KUN
108 L. vestitus Lau, S. K. 27190 Ledong, Hainan, China KUN
109 L. xizangensis Tibet Exped. 74-4411 Motou, Tibet, China KUN
110 L. xylocarpus Sun, H. 84-128 Jingdong, Yunnan, China KUN
111 L. xylocarpus Deng, M. 796 Jingdong, Yunnan, China KUN
112 Notholithocarpus
densiflorus
Bartholonew, B. 1427 SanBenito County, Califonia, USA KUN
664 M. Deng et al.
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Table 2 Leaf epidermal features of Lithocarpus in this study
Scientific name Adaxial Abaxial Morph
group
T type TB Ep Anti
wall
Ep T type TB Anti
wall
Orn ep SA Stomata L 9W(lm) Stoma freq/
mm
2
1L. amoenus twp stb/fb irr str irr apt/f/s/sf/twp aptb/fb/
stb
str-cur pap cyc 23.9–26.2 921.5–23.7 320 APT group
2L. amygdalifolius twp stb irr str irr apt/twp aptb/stb str-cur glo-pap cyc 22.7–24.6 921.1–22.1 320 APT group
3L. amygdalifolius var.
praecipitiorum
twp stb irr str irr apt/twp aptb/stb str-cur pap cyc 26.6-29.0 920.9–25.5 363 APT group
4L. areca twp stb poly-irr str-cur irr bbt/f/s/sf btb/fb sin None cyc 22.4–28.0 920.3–24.6 491 BBT group
5L. attenuatus twp stb irr str irr apt/twp aptb/stb str-cur None cyc 23.4–23.9 922.0–22.2 341 APT group
6L. bacgiangensis twp stb irr str-cur irr ala/apt/twp aptb/stb str-cur None/glo cyc 25.8–24.1 920.2–23.7 491 APT group
7L. balansae twp stb irr str irr apt/twp aptb/stb str-cur None cyc 20.6–24.8 919.4–23.0 469 APT group
8L. brachystachyus twp stb irr str irr apt/twp aptb/stb str-cur None cyc 19.6–24.0 917.5–20.0 512 APT group
9L. calolepis NA stp irr str irr apt/twp aptb/stb str pap-oa cyc 22.7–23.4 917.9–22.5 533 APT group
10 L. calophyllus twp stb irr str irr apt/su/twp aptb/stb str-cur pap cyc 21.5–21.5 917.6–20.4 469 APT group
11 L. calophyllus twp stb irr str-cur irr apt/su/twp aptb/stb str-cur oa cyc 30.8–32.3 930.1–31.5 341 APT group
12 L. carolinae twp stb irr str irr apt/twp aptb/fb/
stb
str-cur pap cyc 28.0–32.3 923.8–28.9 277 APT group
13 L. caudatilimbus twp stb irr str irr apt/twp aptb/stb str-cur None cyc 19.9–21.3 916.0–19.5 448 APT group
14 L. chifui twp stb irr str irr apt/twp aptb/stb str-cur glo-pap-
oa
cyc 25.2–28.2 924.2–26.7 405 APT group
15 L. chiungchungensis NA stb irr str-cur irr apt/twp aptb/stb str-cur glo-pap cyc 23.9–29.2 920.7–22.3 299 APT group
16 L. chrysocomus twp stb irr str irr apt/twp aptb/stb str-cur pap cyc 23.8–27.7 923.0–26.7 320 APT group
17 L. cinereus twp stb irr str irr apt/twp aptb/stb str-cur None-
glo-pap
cyc 26.4–32.0 922.4–29.0 384 APT group
18 L. cleistocarpus twp stb irr str irr apt/twp aptb/stb str-cur glo-pap cyc 24.0–25.7 923.3–24.2 341 APT group
19 L. confinis twp stb irr str rou apt/twp aptb/stb cur pap/oa cyc 24.8–24.4 918.6–21.7 427 APT group
20 L. corneus twp stb poly-irr str irr bbt btb und-
sin
None cyc 28.0–36.5 926.7–35.5 363 BBT group
21 L. craibianus twp stb irr str irr apt/twp aptb/stb str-cur pap cyc 24.9–34.0 920.1–28.5 256 APT group
22 L. crassifolius twp stb irr str irr apt/twp aptb/stb str flat cyc 28.0–30.2 925.0–28.4 427 APT group
23 L. cucullatus twp stb irr str irr apt/s/f/twp aptb/
ctb/stb
str-cur rou-pap cyc 21.6–25.9 921.6–23.0 405 APT group
24 L. cyrtocarpus f/s/sf fb rec-
poly-
irr
str irr bbt/f/s/sf/st/
uc
btb/fb cur-sin None cyc 24.3–28.8 921.6–23.6 363 BBT group
25 L. cyrtocarpus f/s/sf fb poly-irr str irr bbt/f/s/sf/st/
uc
btb/fb sin None cyc 20.3–25.0 918.5–20.1 320 BBT group
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Table 2 continued
Scientific name Adaxial Abaxial Morph
group
T type TB Ep Anti
wall
Ep T type TB Anti
wall
Orn ep SA Stomata L 9W(lm) Stoma freq/
mm
2
26 L. damiaoshanicus twp stb irr str iso-
irr
apt/twp aptb/stb str-cur oa cyc 19.3–25.3 919.1–22.1 235 APT group
27 L. dealbatus f/s/
twp
stb/fb irr str irr apt/f/s/twp aptb/fb/
stb
str-cur pap cyc 24.4–25.5 920.0–24.2 320 APT group
28 L. densiflorus NA ctb/
stb
irr str-cur irr mu/su/twp ctb/stb str-cur None an/
cyc
17.9–27.1 916.0–22.3 427 APT group
29 L. echinotholus twp stb/
ctb
irr str irr apt/twp aptb/stb str-cur pap cyc 24.4–26.9 921.2–23.8 384 APT group
30 L. elaeagnifolius twp stb irr str-cur irr apt/twp stb/stb str-cur glo-pap cyc 26.1–30.2 922.2–25.7 341 APT group
31 L. elizabethae NA stb irr cur-
und
irr apt/twp aptb/stb str-cur glo-pap cyc 23.9–26.2 921.5–23.7 235 APT group
32 L. elmerrillii twp stb irr str irr apt/twp aptb/stb str-cur None cyc 19.6–24.9 919.3–22.2 320 APT group
33 L. eriobotryoides bbt btb poly-irr und-
sin
irr bbt/f/ro/s/sf/
twp/uc
btb/fb/
stb
und-
sin
None cyc 26.1–29.8 925.6–26.4 320 BBT group
34 L. farinulentus twp stb irr str irr apt/twp aptb/stb str None cyc 24.1–25.2 917.2–21.4 405 APT group
35 L. fenestratus twp stb irr str irr apt/twp aptb/stb str-cur pap cyc 21.0–26.1 919.3–21.7 405 APT group
36 L. fenzelianus twp stb irr str irr apt/twp aptb/stb str-cur pap cyc 19.2–21.8 919.1–21.1 341 APT group
37 L. floccosus twp stb irr cur-
sin
irr apt/twp aptb/stb str-cur pap cyc 25.5–27.1 921.0–24.1 386 APT group
38 L. fohaiensis NA stb irr str-cur irr twp stb cur-sin None cyc 24.7–28.1 919.0–27.2 433 Glabrous
group
39 L. fordianus bbt btb poly-irr sin irr bbt/f/s/sf/uc btb/fb/
stb
sin None cyc 19.6–27.0 919.0–19.9 469 BBT group
40 L. glaber NA ctb irr str irr apt/bu/twp aptb/
ctb/stb
str-cur pap cyc 23.7–27.7 920.7–27.4 320 APT group
41 L. glaucus NA stb poly-irr und-
sin
irr twp stb str-cur None cyc 36.4–37.6 931.4–35.5 235 Glabrous
group
42 L. grandifolius NA stb rec-poly cur-str irr twp stb sin None cyc 26.8–27.2 924.9–26.6 213 Glabrous
group
43 L. guiuieri twp stb irr str irr twp/apt/twp aptb/stb str-cur None cyc 25.5–26.8 924.8–26.4 277 APT group
44 L. haipinii bbt btb poly-irr str irr bbt/f/s/sf/twp btb/fb cur-sin None cyc 28.9–33.8 928.6–33.1 363 BBT group
45 L. haipinii bbt btb poly-irr str irr bbt/s/f/sf btb/fb cur-sin None cyc 29.9–32.2 924.6–24.9 405 BBT group
46 L. hancei absent absent irr str rou-
irr
twp/su stb cur None cyc 24.3–30.9 921.1–26.4 469 Glabrous
group
47 L. handelianus f/s/sf/
twp
stb/fb irr str irr apt/f/s/twp aptb/fb/
stb
cur pap cyc 23.7–27.7 920.8–27.5 341 APT group
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Table 2 continued
Scientific name Adaxial Abaxial Morph
group
T type TB Ep Anti
wall
Ep T type TB Anti
wall
Orn ep SA Stomata L 9W(lm) Stoma freq/
mm
2
48 L. harlandii twp stb irr str irr twp stb rou-
sin
None cyc 23.7–27.7 920.7–27.4 491 Glabrous
group
49 L. henryi twp stb irr str iso apt/twp stb rou oa cyc 19.2–21.8 919.1–21.1 299 APT group
50 L. himalaicus twp stb irr str irr twp stb str-cur None cyc 28.2–28.8 924.6–27.2 256 Glabrous
group
51 L. howii f/sf stb poly-irr cur-str irr bbt/f/st/twp btb/ctb/
fb
cur-sin None cyc 20.2–25.4 916.8–22.9 427 BBT group
52 L. hypoglaucus twp stb irr str irr apt/twp aptb/stb str-cur glo cyc 23.7–27.7 920.8–27.5 320 APT group
53 L. irwinii twp stb irr str irr apt/twp aptb/stb cur pap cyc 25.5–27.1 921.0–24.1 320 APT group
54 L. ithyphyllus NA stb irr str irr twp stb str-cur None cyc 25.5–30.9 924.7–27.7 341 Glabrous
group
55 L. konishii bbt stb poly-irr und irr bbt/uc btb/stb sin None cyc 25.3–28.4 921.5–24.3 491 BBT group
56 L. laetus twp stb irr str irr apt/twp aptb/stb str-cur pap cyc 21.6–25.9 921.6–23.0 469 APT group
57 L. laoticus twp stb irr str irr apt/twp aptb/stb str-cur pap cyc 26.4–29.8 922.8–28.2 341 APT group
58 L. lepidocarpus twp stb irr str rou apt/twp aptb/stb rou oa cyc 26.1–27.7 923.8–25.7 452 APT group
59 L. litseifolius twp stb irr str irr apt/twp aptb/stb str-cur pap cyc 25.6–26.8 922.4–26.6 427 APT group
60 L. litseifolius twp stb irr str irr apt/twp aptb/stb cur pap cyc 27.3–30.2 925.3–27.5 275 APT group
61 L. litseifolius twp stb irr str irr apt/twp aptb/stb str-cur pap-oa cyc 21.5–21.5 917.6–20.4 299 APT group
62 L. longanoides twp stb irr str-cur irr-
rou
apt/twp aptb/stb str-cur glo or oa cyc 25.1–28.7 923.5–22.8 401 APT group
63 L. longanoides twp stb irr str-cur irr-
iso
apt/twp aptb/stb str-cur None/oa cyc 21.7–26.2 921.6–26.0 384 APT group
64 L. longipedicellatus twp stb irr str irr apt/twp aptb/stb str-cur None cyc 22.7–23.7 917.6–20.7 384 APT group
65 L. longzhouicus f/sf stb poly-irr und-
sin
irr bbt/twp/uc btb/stb cur-sin None cyc 36.4–25.0 931.4–20.1 299 BBT group
66 L. lycoperdon twp stb irr str irr apt/twp aptb/stb str-cur oa cyc 26.1–27.7 923.8–25.7 341 APT group
67 L. lycoperdon twp stb irr str irr apt/twp aptb/stb str-cur pap cyc 27.9–27.4 922.7–24.4 277 APT group
68 L. macilentus NA stb irr str irr apt/ala/twp stb str-cur glo-pap cyc 22.7–23.4 921.1–23.2 414 APT group
69 L. magneinii twp stb irr str irr apt/twp aptb/stb str-cur None cyc 21.8–25.8 921.1–25.5 363 APT group
70 L. mairei NA stb irr str irr apt/twp aptb/stb str glo-pap cyc 23.7–29.4 921.7–22.5 299 APT group
71 L. megalophyllus twp stb irr str-
und
irr twp stb cur-sin flat cyc 29.1–33.9 928.8–30.8 350 Glabrous
group
72 L. melanochromus NA ctb irr str iso apt/twp aptb/stb rou oa cyc 21.8–25.8 921.1–25.5 363 APT group
73 L. mianningensis NA stb irr str irr apt/twp aptb/stb str-cur pap cyc 27.3–32.5 923.9–26.1 384 APT group
74 L. microspermus twp stb irr str irr apt/twp aptb/stb str-cur glo cyc 19.8–23.0 919.6–20.6 371 APT group
Comparative morphology of leaf epidermis 667
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Table 2 continued
Scientific name Adaxial Abaxial Morph
group
T type TB Ep Anti
wall
Ep T type TB Anti
wall
Orn ep SA Stomata L 9W(lm) Stoma freq/
mm
2
75 L. naiadarum twp stb irr str irr-
rou
twp stb str-cur flat cyc 23.7–29.4 921.7–22.5 501 Glabrous
group
76 L. oblanceolatus twp stb irr str irr twp stb str flat cyc 29.1–32.7 926.1–31.8 471 Glabrous
group
77 L. obovatilimbus twp stb irr str irr-
iso
apt/twp aptb/stb str-cur pap-oa cyc 26.6–29.0 920.9–25.5 320 APT group
78 L. obscurus NA stb irr str irr twp stb str flat cyc 27.3–29.7 920.0–23.8 437 Glabrous
group
79 L. oleifolius NA stb irr str-cur irr ala/apt/twp aptb/stb str-cur None-
glo-pap
cyc 25.2–25.9 923.7–24.1 435 APT group
80 L. pachylepis f/s/sf fb/stb poly-irr str-cur irr bbt/s/f/sf btb/fb und-
sin
None cyc 19.3–25.3 919.3–23.5 348 BBT group
81 L. pachyphyllus twp stb irr str irr apt/twp aptb/stb str-cur None cyc 22.7–25.1 920.0–23.9 469 APT group
82 L. pachyphyllus twp stb irr str irr apt/twp aptb/stb str-cur None cyc 19.8–25.5 919.6–21.1 529 APT group
83 L. paihengii NA stb irr str irr apt/twp aptb/stb str-cur pap cyc 24.7–28.1 919.0–27.2 320 APT group
84 L. pakhaensis NA stb irr str irr apt/twp aptb/stb str-cur None cyc 22.5–25.6 921.3–23.4 457 APT group
85 L. paniculatus NA ctb irr str irr apt/twp aptb/stb str-cur pap cyc 23.9–28.6 918.2–23.8 427 APT group
86 L. lithocarpaeus twp stb irr str irr apt/twp aptb/stb str-cur pap cyc 27.9–33.4 920.8–24.4 258 APT group
87 L. petelotii NA stb/
ctb
irr str irr apt/s/f/twp fb/stb cur-sin None cyc 27.0–27.7 923.5–27.6 363 APT group
88 L. propinquus twp stb irr str irr apt/twp aptb/stb str-cur None/glo cyc 21.7–25.2 918.6–20.8 299 APT group
89 L. pseudoreinwardtii twp stb irr str irr apt/twp aptb/stb cur-sin None cyc 22.9–22.6 916.7–20.7 497 APT group
90 L. pseudosundaicus twp stb irr str-cur irr apt/s/twp fb/stb str-cur pap cyc 22.9–26.8 921.6–24.7 341 APT group
91 L. pseudovestitus twp stb irr str irr apt/twp aptb/stb str-cur None cyc 21.7–23.3 920.3–21.2 427 APT group
92 L. qinzhouicus twp stb irr str irr apt/twp aptb/stb str-cur pap cyc 28.8–29.7 924.8–27.0 341 APT group
93 L. quercifolius twp stb poly-irr str-
und
irr bbt/uc btb/stb sin None cyc 25.4–28.8 921.7–24.5 256 BBT group
94 L. rhabdostachyus f/twp fb/stb irr str irr apt/f/s/sf/twp aptb/fb/
stb
str-cur None cyc 25.3–26.7 923.9–26.4 299 APT group
95 L. rosthornii NA stb/
ctb
irr str-cur irr apt/f/s/sf/twp aptb/stb str-cur glo-pap cyc 24.9–25.6 920.2–24.5 363 APT group
96 L. silvicolarum twp stb irr str-cur irr apt/s/twp aptb/
btb/fb
str-cur pap cyc 21.4–24.3 919.5–24.0 410 APT group
97 L. skanianus NA stb irr str-
und
irr apt/f/s/sf/twp aptb/fb/
stb
str-cur pap cyc 29.1–35.7 927.3–28.8 235 APT group
98 L. sphaerocarpus twp stb irr str irr apt/twp aptb/stb str-cur pap cyc 22.7–24.1 921.4–23.3 405 APT group
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Table 2 continued
Scientific name Adaxial Abaxial Morph
group
T type TB Ep Anti
wall
Ep T type TB Anti
wall
Orn ep SA Stomata L 9W(lm) Stoma freq/
mm
2
99 L. tabularis twp stb irr str irr apt/bu/twp aptb/stb str-cur pap cyc 25.6–26.8 922.4–26.6 256 APT group
100 L. taitoensis twp stb irr str irr apt/twp aptb/stb str flat cyc 25.2–31.5 923.0–30.1 299 APT group
101 L. talangensis twp stb/fb irr str irr apt/twp aptb/fb/
stb
str-cur pap cyc 19.3–25.3 919.1–22.1 320 APT group
102 L. talangensis twp stb/
ctb
irr str irr apt/twp aptb/fb/
stb
str-cur pap cyc 23.0–26.7 923.0–26.7 384 APT group
103 L. tenuilimbus s/f/
twp
stb/fb irr str irr apt/twp aptb/fb/
stb
str-cur flat-pap cyc 22.3–27.4 922.0–22.2 491 APT group
104 L. touranensis NA stb/
ctb
irr str irr twp stb str None cyc 24.8–27.4 922.6–27.1 341 Glabrous
group
105 L. trachycarpus twp stb irr str irr apt/twp aptb/stb cur pap cyc 24.4–26.7 923.3–25.2 448 APT group
106 L. truncatus twp stb irr str irr apt/twp aptb/stb str-cur None cyc 24.0–24.6 915.8–24.0 256 APT group
107 L. uvariifolius s/fs/
bbt
fb/stb poly-irr und-
sin
irr bbt/f/s/sf btb/fs/
stb
sin None cyc 22.7–29.9 921.9–27.6 576 BBT group
108 L. variolosus twp stb irr str irr apt/twp aptb/stb str-cur pap cyc 24.3–28.8 921.6–23.6 341 APT group
109 L. vestitus twp stb irr str irr apt/twp aptb/stb str flat cyc 22.2–23.1 920.0–22.3 384 APT group
110 L. xizangensis f/twp stb/fb irr str iso apt/s/f/twp aptb/fb/
stb
cur oa cyc 24.3–28.8 921.6–23.6 320 APT group
111 L. xylocarpus twp stb irr cur to
str
irr apt/twp aptb/stb str-cur glo cyc 24.9–26.4 915.9–21.4 320 APT group
112 L. xylocarpus twp stb irr cur to
str
irr apt/twp aptb/stb str-cur glo cyc 24.4–28.2 915.9–24.6 256 APT group
T type trichome types, ala appressed laterally attached unicellular, apt appressed parallel tufts, bbt broad based trichomes, bu branched uniseriate, ffasciculate, mu multiradiate, ro rosulate,
ssolitary unicellular, sf stipitate fasciculate, su simple uniseriate, st stellate trichomes, twp thin-walled peltate, uc unicellular conical trichome, NA not available, TB trichome base type, aptb
appressed parallel tuft base, btb broad trichome base, ctb compound trichome base, fb fasciculate trichome base, stb simple trichome base, Ep Shape of epidermal cells, irr irregular, rec
rectangle, poly polygonal, iso isodiametric, Anti Wall Anticline wall shape, str straight, cur curved, und undulate, sin sinus, rou rounded, Orn ep Ornentmental on epidermal cells, pap papilate
thickening, glo global thickening, oa overall thickening, none flat no special ornentments, TB Trichome base, stb simple trichome base, ctb compound trichome base, aptb appressed parallel
tufts base, btb broad trichome base, SA stomata aperture, cyc cyclocytic, an anomocytic, Stoma Size stomata size, Stoma freq stomata frequency, Morph group morphological group
Comparative morphology of leaf epidermis 669
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Thin-walled peltate trichomes Jones (1986) recorded this
trichome type as an intermediate type consisting of thin-
walled cells which possess a unicelled to 2–3-celled stalk
and an irregularly shaped stellate to discoidal cap com-
posed of many randomly oriented cells (Fig. 5v). This type
of trichome is generally present in all species of Litho-
carpus. The shape of these trichomes is diverse among
various species. The stalk cell(s) is usually isodiametric
and stained darker than epidermal cells, which indicates a
glandular trichome type. The simplest TWP trichome has
only a transparent skirt-like rim on the stalk cell, such as in
L.balansae (Figs. 5b, 6a) and L. oleifolius (Fig. 5s). The
large TWP trichomes were rather fragile (L. calophyllus
Fig. 6q, L. craibianus Fig. 6t). In most cases, while pre-
paring the cuticle for LM, most of the peltate trichome was
usually lost, with only a round simple trichome base
remaining, or with only the unicelled stalk left (Fig. 6l, o,
u, v). The trichome base was usually large, with a diameter
ranging from ca. 9.3 to 17.0 lm, and the basal portion
remaining without any obvious stain.
Broad base trichomes (BBTs) This type of trichome is
usually composed of two parts: the basal broad cell portion
or foot cell (Hill 1983), and an upper, long or short, barrel-
shaped structure. This trichome type was present in 11
species; for example, L. areca (Fig. 3h, p) L. quercifolius
(Fig. 3c), L. konishii (Fig. 3f), L. howii (Figs. 3k, 4j, k),
L. uvariifolius (Figs. 3g, 4b), L. fordianus (Figs. 3i, 4c, d),
and L. corneus (Fig. 4m, n). The trichome itself is usually
almost transparent. This trichome type is very similar to
simple uniseriate (SU), but is easily distinguishable due to
its rather prominent convex basal broad cells. The diameter
of the basal broad cell portion is large, usually
17.3–25.6 lm. The convex broad basal portion also can be
Fig. 1 Characteristics of adaxial epidermal cells. a–lLM. a–kbar
50 lm; aLithocarpus chrysocomus, showing a small TWP and
its simple trichome base; bL. pseudoreinwardtii;showingSU;
cL. tenuilimbus; showing F and TWP; dL. variolosus;eL. amoenus;
fL. cucullatus;gL. obscurus;hL. pachylepis, the arrow indicates SU;
iL. eriobotryoides;jL. uvariifolius, showing fasiculate trichome
base; kNotholithocarpus densiflorus, showing compound trichome
base; lL. elizabethiae,bar 20 lm; m–pSEM, showing TWP. m
L. fenestratus,bar 50 lm; nL. hancei,bar 10 lm; oL. harlandii,bar
20 lm; pL. hypoglaucus,bar 50 lm, showing SF
670 M. Deng et al.
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found in TWP, but is different from its closed terminal
structure, in contrast to the flat, free structure in TWP.
Under SEM, BBT can be easily distinguished from TWP
and SU.
Appressed laterally attached unicellular This trichome
type is similar to the solitary unicellular trichome, except
that they are attached laterally, and are generally thin-
walled and difficult to detect. Usually, the distance from
the trichome base to the end of the hair is extremely short,
much shorter than in other trichomes. The trichome base is
simple-celled, but the basal portion is smaller and darker
stained than in TWP and BBT. This trichome type was
detected only in L. macilentus (Fig. 5q, r) and L. oleifolius
(Fig. 5s) although the typical trichome bases were gener-
ally found in several other species bearing APT, indicating
its possible existence in those species as well.
Simple uniseriate This type of trichome is composed of a
single column of at least two or more thin-walled struc-
tures, apparently cells. This trichome type is not common in
Lithocarpus and was present only in L. calophyllus (Fig. 5w),
L. hancei (Fig. 2u) and. N. densiflorus (Figs. 5x, 6x).
Branched uniseriate Branched uniseriate (BU) trichomes
were detected in two species of Lithocarpus only. This type
of trichome is similar to TWP in cellular composition, but
in shape is usually elongated and branched at least once.
Fig. 2 Characteristics of abaxial epidermis of glabrous group.
a–lLM, bar 50 lm; aLithocarpus glaucus;bL. fohaiensis;
cL. obscurus;dL. oblanceolatus;eL. naiadarum;fL. touranensis;
gL. ithyphyllus;hL. megalophyllus;iL. harlandii;jL. himalaicus;
kL. grandifolius;lL. megalophyllus;m–vSEM, bar 20 lm.
mL. himalaicus;nL. fohaiensis;oL. harlandii;pL. touranensis;
qL. grandifolius;r–sL. ithyphyllus;tL. obscurus;u–vL. hancei;
uarrow indicates SU; vshowing stomatal wax plug
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These trichomes, irregular in shape, produce a large num-
ber of oddly shaped forms, such as in L. tabularis (Fig. 6s)
and L. glaber (Fig. 5t).
Rosulate This trichome type consists of unicellular, open,
thin-walled elements, arranged in small bushy tufts. The
trichome base is simple in Lithocarpus. The rosulate (Ro)
trichome is not common in species of Lithocarpus and was
detected only in one species, L. eriobotryoides (Fig. 4g).
Appressed parallel tufts APT trichomes were the most
common in Lithocarpus spp.; 85 species were found to bear
APT trichomes. The trichome consists of (1)2–8(12) thick-
walled, unicellular elements that are nearly coplanar and
approximately parallel to the leaf surface as well as to each
other (Figs. 5a–s, 6a–n). This trichome’s basal portion is
usually convex and raised above the epidermal cell to form
a linear band along the subsidiary cells of the stomata,
which can partly (Figs. 5d, f, h, i, n; 6a–j) or mostly
(Figs. 5c, e, p; 6k–o) cover the stomata. The convex APT
trichome bases in extreme types were surrounding the
stomata’s subsidiary cells. The stomata were shielded by the
upper APT rays, such as in L. lithocarpaeus (Fig. 5m). In
another extreme type, the APT trichome bases were fused
together in a round fashion with the trichome rays fanning out
to form a ‘‘stellate’’-like structure, such as in L. pseud-
oreinwardtii (Fig. 6c), L. vestitus (Fig. 6f) and L. paihengii
(Fig. 6o). A similar trichome type was also reported as fused
stellate or stellate by Jones (1984,1986) and Zhou and Xia
(2012). However, the stellate trichome formed by a cluster of
APTs discovered in the present study can be easily distin-
guished from the typical stellate trichome by its convex base
without dark staining compared to the flat, and dark-stained
‘‘flower-like’’ compound trichome base found in typical
stellate and fused stellate trichomes (St).
Stellate trichomes Stellate trichomes usually consisted of
(3)4–12 non-glandular, unicellular, and generally thick-
Fig. 3 Characteristics of abaxial epidermis of BBT group (LM).
aLithocarpus areca,bar 100 lm; b–lbar 50 lm; bL. longzhouicus;
cL. quercifolius;dL. pachylepis;eL. cyrtocarpus;fL. konishii;
gL. uvariifolius;hL. areca;iL. fordianus;jL. haipinii;kL. howii;
lL. eriobotryoides;m–pbar 20 lm. mL. corneus;nL. cyrtocarpus;
oL. eriobotryoides;pL. areca
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walled elements that radiate from a common point of
attachment in a parallel or nearly parallel fashion to the leaf
surface. Of all the studied species of Lithocarpus, this tri-
chome type was present only in two species, L. howii
(Figs. 3k, 4i) and L. cyrtocarpus (Fig. 4h). In this trichome,
the base is compound and one side of the epidermal cell
wall adjacent to the base cells was dark stained.
Unicellular conical trichome (UC) This trichome type
also was uncommon and in the present study was detected
only in five species: L. quercifolius (Fig. 3c), L. konishii
(Fig. 3f), L. fordianus (Figs. 3i, 4c), L. cyrtocarpus (Fig. 4i)
and L. longzhouicus. It is not a typical conical trichome as
defined by Jones (1986) that was short and thick walled. The
wall of this trichome was thin and the whole trichome was
stained as revealed by the present study, indicating that it
might be a glandular type. The trichome bases were simple
and round and without cutinized cell wall (Fig. 3c, f, i).
Solitary unicellular trichome This solitary unicellular
trichome (S) was generally thick-walled, fairly long and straight
and dark-stained (Figs. 3a, h, i, 4a). It is a basic element to form
the fasciculate trichome and the stipitate fasciculate trichome.
This trichome type was present in 20 species including
L. uvariifolius (Fig. 4a), L. areca (Fig. 3a), L. fordianus
(Fig. 3i), L. rhabdostachyus (Fig. 5g), L. silvicolarum (Fig. 5n),
L. skanianus (Figs. 5o, 6p) and L. xizangensis (Fig. 6p).
Fasciculate trichomes Fasciculate trichomes (F) were
composed of 2–6(8) solitary structures that were joined
together at the base. This type of trichome existed in
19 species of Lithocarpus including L. haipinii (Fig. 3j);
L. eriobotryoides (Fig. 3l); L. uvariifolius (Fig. 4a); L. areca
(Fig. 3a) and L. xizangensis (Fig. 6p).
Stipitate fasciculate trichomes Stipitate fasciculate tric-
homes (SF) are very similar to fasciculate trichomes,
Fig. 4 Characteristics of abaxial epidermis of BBT group (SEM).
a,bLithocarpus uvariifolius.abar 100 lm; bbar 20 lm, showing
BBT; c, d L. fordianus;cbar 20 lm, showing BBT and UC; dbar
10 lm, the high magnification of BBT; eL. haipinii,bar 20 lm;
f,gL. eriobotryoides,bar 20 lm; fshowing TWP; gshowing Ro;
hL. cyrtocarpus, showing St and BBT, bar 50 lm; i–kL. howii.
ishowing UC, bar 20 lm; jshowing St, bar 50 lm; kshowing BBT,
bar 20 lm; lL. longzhouicus, showing BBT, bar 10 lm; m,n
L. corneus.mshowing BBT, bar 20 lm; nas M, BBT under high
magnification bar 10 lm; o,pL. konishii;oshowing BBT, bar
20 lm; phigh magnification of BBT on the same slides of O, bar
10 lm
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Fig. 5 Characteristics of abaxial epidermis of APT group and
Lithocarpus densiflorus (LM). a,c–n,q,s,t,xbar 50 lm; b,o,r,
u,wbar 20 lm; a,bL. balansae;bhigh magnification of A, arrow
indicates TWP; cL. taitoensis;dL. pseudoreinwardtii;eL. micro-
spermus;f.L. mianningensis;gL. rhabdostachyus;hL. petelotii;
iL. echinotholus;jL. carolinae;kL. rosthornii, arrow indicates
TWP: lL. elaeagnifolius;mL. lithocarpaeus;nL. silvicolarum;
oL. skanianus;pL. paihengii, the showing APT trichome bases;
q,rL. macilentus;rthe arrow indicates ALA and its dark stained
simple trichome base; sL. oleifolius, the arrow indicates ALA;
tL. glaber, showing BU; uL. pseudosundaica, the arrow indicates
TWP; vL. chrysocomus, showing the TWP with long stalk cells;
wL. calophyllus, showing SU. xNotholithocarpus densiflorus,
showing central large cyclocytic stomata and other anomocytic
stomata
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except that in the former trichome type, only the lower
part of the trichome cells was fused (Figs. 3l, 5p).
Thirteen species of Lithocarpus were found to have this
trichome and the species possessing SF, usually have F
and S trichomes as well. The trichome bases of the S, F
and SF were similar and coexisted frequently; they may
represent the same trichome morphological stage in
Lithocarpus.
Fig. 6 Characteristics of abaxial epidermis of APT group (SEM)
and Notholithocarpus densiflorus,bar 20 lm. aL. balansae;
bL. petelotii;cL. pseudoreinwardtii;dL. guinieri;eL. silvicolarum;
fL. vestitus;gL. floccosus;hL. glaber;iL. laoticus;jL. confinis;
kL. litseifolius;lL. fenestratus;mL. pseudovestitus;nL. lepido-
carpus;oL. paihengii;pL. skanianus;qL. calophyllus, showing
TWP; rL. lepidocarpus, showing TWP; sL. tabularis, showing dense
BU; tL. craibianus, showing well-developed TWP; uL. amoenus,
showing TWP; vL. handelianus, showing TWP; w,xNotholithocar-
pus densiflorus. X. the high magnification of X on the same slide,
showing SU and crystalline wax flake
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Table 3 Comparison of leaf trichome types of various genera within Fagaceae
Trichome types Taxa
Lithocarpus Notholithocarpus
densiflorus
Chrysolepis Castanopsis Castanea Quercus s.l. Fagus Trigonobalanus
s.l.
BBT Glabrous APT subg
Cyclobalanopsis
sect.
Quercus
s.s.
sect.
Protobalanus
sect.
Lobatae
sect.
Cerris
Simple solitary ?*(1) 0 ?*(1) 0 ?
8
(1) ?
2, 8
(1) ?
2, 8
(1) ?
5, 7
(1) ?
2
(1) ?
2, 3
(1) ?
1, 2
(1) ?
2,
9
(1)
?
6
(1) ?
2
(1)
Unicellar conical ?*(1) 0 0 0 0 0 ?
2
(1) 0 0 0 0 0 ?
6
(1) ?
2
(1)
Appressed
laterally
attached
00 ?*(1) 0 0 0 0 ?
7
(1) 0 0 ?
2
(1) 0 0 0
Stellate ?*(1) 0 0 0 ?
2, 8
(1) 0 ?
2, 8
(1) ?
5, 7
(1) ?
2
(1) ?
2, 3
(1) ?
1, 2
(1) ?
2,
9
(1)
00
Fused stellate 0 0 0 0 0 0 0 ?
5, 7
(1) ?
2
(1) 0 0 ?
2,
4
(1)
00
Fasciculate ?*(1) 0 ?*(1) 0 0 ?
2, 8
(1) 0 ?
5, 7
(1) ?
2
(1) ?
2, 3
(1) ?
1, 2
(1) ?
2,
9
(1)
0?
2
(1)
Stipitate
fasciculate
?*(1) 0 ?*(1) 0 0 ?
8
(1) 0 ?
5, 7
(1) ?
2
(1) 0 ?
1, 2
(1) ?
2,
4
(1)
00
Appressed
parallel tuft
00 ?*(1) 0 0 0 0 0 0 0 0 0 0 0
Multiradiate 0 0 0 ?
2
(1) 0 0 0 ?
5, 7
(1) 0 ?
3
(1) ?
2
(1) 0 0 0
Thick walled
peltate
00 00 ?
2, 8
(1) 0 0 0 0 0 0 0 0 0
Thin-walled
peltate
?*(1) ?*(1) ?*(1) 0 0 ?
2, 8
(1) ?
2, 8
(1) 0 0 0 0 0 0 0
Broad base
trichome
?*(1) 0 0 0 0 0 0 0 0 0 0 0 0 0
Rosulate ?*(1) 0 0 0 0 0 0 ?
7
(1) 0 ?
3
(1) ?
2
(1) ?
2
(1) 0 0
Capitate 0 0 0 0 0 0 ?
2
(1) 0 ?
2, 9
(1) ?
2
(1) ?
1, 2
(1) ?
2, 4,
9
(1)
00
Simple uniseriate ?*(1) ?*(1) ?*(1) ?
2,
*(1) 0 0 ?
2, 8
(1) ?
2, 5, 7
(1) ?
2, 9
(1) ?
2, 3, 9
(1) ?
1, 2
(1) ?
2,
9
(1)
?
6
(1) ?
2, 5
(1)
Branched
uniseriate
00 ?*(1) 0 0 0 0 0 ?
2
(1) ?
2
(1) 0 ?
2
(1) 0 ?
2
(1)
Glandular peltate 0 0 0 0 0 0 0 0 0 0 0 0 0 ?
2, 5
(1)
Papillae-global
thickening
00 ?*(1) 0 0 0 0 ?
2, 5, 7
(1) 0 0 0 0 0 ?
2
(1)
Jelly fish-like 0 0 0 0 0 0 0 ?
5, 7
(1) 0 0 0 0 0 0
19 trichome types are mapped onto the phylogenetic cladogram (Oh and Manos 2008) with character states are presented in the circles (Fig. 7)
Data sources: * current study,
1
Hardin (a, b),
2
Jones (1986),
3
(Manos 1992),
4
Zhou and Wilkinson (1995),
5
Lou and Zhou (2001),
6
Denk (2003),
7
Deng (2007),
8
Liu et al. (2009),
9
Tschan and Denk (2012)
Stage 0, absent; 1, present
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Trichome bases Trichome bases were scattered on the
abaxial epidermis in all the species. Based on their mor-
phology, they can be placed under four different categories:
(1) Simple trichome base In this category, the cell wall of
the basal portion was usually more or less cutinized and
stained darker than the epidermal cells. The epidermal cells
around the trichome base were unmodified (Fig. 2a–e, g–l);
AL, TWP, BBT, SU, BU, Ro and UC have this category of
trichome base. Usually, the basal portion is small (diam. ca.
9.3–14.4 lm) and thickly cutinized in AL, SU, and Ro, but
large (diam. ca. 15.3–25.6 lm) and less cutinized in TWP
and BBT; (2) Compound trichome base Compound tri-
chome based cells were surrounded by 1–2 layers of thick
cutinized (dark stained) small-sized epidermal cells,
which gave the appearance of a ‘‘flower-like’’ structure
(Fig. 1e, k). Only St and multiradiate trichomes had this
type of trichome base; (3) Fasciculate trichome base The
trichome bases in this category were round with dark
stained walls and 1–2(3) rows of epidermal cells sur-
rounding the trichome bases; these epidermal cells are
smaller than the normal epidermal cells (Figs. 1j, 3e, h, l;
5g, h, p); SU, F and SF have this type of trichome base;
(4) APT trichome base This trichome base type was dis-
tributed adjacent to the subsidiary cells. The basal portion
was small convex, swollen and not obviously cutinized.
These trichome bases usually surrounded 2–16(18) sto-
mata. In some extreme types, the convex basal portion
formed a ring surrounding the stomata (Fig. 5m). Only
APT was observed to possess this type of trichome base in
the present study.
Wax flake
In all examined species of Lithcarpus, both the adaxial and
abaxial cuticles were covered with a thin to thick wax
flake. It is easy to detect the stomata and non-glandular
type trichomes (including APT, SU, S, F and SF) through
the wax flake by SEM, but SEM could not fully reveal the
key features of glandular and intermediate types since they
were soft and covered by wax flake, unless assisted by LM.
The wax flake was smooth (Figs. 2n–q, 4b–p, 5a–l) or
composed of irregular particles (Fig. 2r, s, u) in Lithocar-
pus, but the crystalline wax flake was only found in
N. densiflorus (Fig. 6w, x).
Evolutionary pattern of leaf epidermal features
in Fagaceae
The epidermal characters of each investigated species have
been summarized in Table 2. Nineteen leaf trichome types
of Fagaceae genera based on the present and previous
studies were compared (Table 3), and mapped on to the
molecular phylogeny cladogram of Oh and Manos (2008)
(Fig. 7). The results show solitary trichomes are a plesio-
morphism in Fagaceae; four characteristics were autapo-
morphic: APT, BBT to APT and BBT group respectively in
Lithocarpus, TWP to Chrysolepis, glandular peltate to
Trigonobalanus, Jellyfish-like to a species in Quercus
subg. Cyclobalanopsis. One characteristic was synapo-
morphic that the presence of multiradiate trichomes sup-
ports the clade N. densiflorus ?Quercus s.l.
Fig. 7 Character state mapping
of the 19 types of trichomes
among genera of Fagaceae
(Table 3, 1–19). Phylogram was
based on CRABS CLAW
sequences (Oh and Manos
2008). Solid circles are
autapomorphy or
synapomorphy, gray circles are
homoplasy. Characteristic
number is above the circle;
character stage is given in the
circle
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Discussion
Comparison of trichome types reported
from Lithocarpus in previous studies and the refinement
of terminology
Although the epidermal morphology is diverse among the
various Lithocarpus species, the characters shown are
reliable diagnostic features for identification purposes and
are consistent; nevertheless, the terminologies applied in
previous anatomical investigations cause great confusion
for purposes of comparison.
Trichome morphology, having rich variations, has been
regarded as an important taxonomical characteristic in
Fagaceae. Jones (1986) recorded 13 trichome types in
Lithocarpus in which multiradiate trichomes were only
found in L. densiflorus (now Notholithocarpus densiflorus).
The rest of the trichome forms were detected in the present
study as well. Jones (1986) and Zhou and Xia (2012)
recorded the presence of papillae and fused stellate tric-
homes in Lithocarpus. Based on a previous study, the
papillae were composed of a cutinized thickening of the
cuticle above the epidermal cells; therefore, this structure is
ornamentation on epidermal cells rather than a trichome
type (Deng 2007). Similar to Jones (1986) and Zhou and
Xia (2012), in the present study, many fused stellate-like
trichomes were detected in Lithocarpus under SEM
(Fig. 6c, o). A comparison of the sample slides by LM
revealed that these stellate-like trichomes were actually
composed of clustered coplanar APT with their hair-like
rays closely set together in the lower part (Fig. 6o). Their
trichome base is consistent with the typical APT with a
swollen, non-dark stained cell wall (Fig. 5p), which is not
similar to the typical stellate trichomes detected in Quercus
s.l. Therefore, based on the present investigation, we still
recognize this stellate-like trichome with typical APT tri-
chome based as a modified APT.
Zhou and Xia (2012) recently reported ‘‘curly thin-
walled unicellular trichomes’’ in L. macilentus and bulbous
trichomes in L. handelianus and L. amoenus. However, in
the present study, based on LM observation of the same
species, L. macilentus, the ‘‘curly thin-walled trichome’’,
was almost transparent, suggesting that this trichome is a
glandular type. The elongate membranous structures were
connected to each other above the dark stained simple
trichome base, the same as in the large TWP trichome
detected in Castanopsis by Liu et al. (2009) and Jones
(1986) and should be attributed to the TWP trichome.
Observations on the epidermis of L. handelianus and
L. amoenus under LM and SEM in the present study did not
detect any bulbous trichomes with ‘‘uniseriate stalks with a
single markedly enlarged terminal cell’’ as reported by
Zhou and Xia (2012). Instead, a careful comparison of the
figures with those in Zhou and Xia (2012), showed that the
‘‘bulbous trichome’’ recorded by these authors was actually
a small TWP that was composed of a unicellular stalk with
an upper small skirt or flat discoidal cap (L. amoenus,
Fig. 6u; L. handelianus, Fig. 6v). This trichome was
equivalent to the ‘‘collapsed bulbous trichomes’’ in Cas-
tanea reported by Hardin and Johnson (1985). However, it
is easy to distinguish the elongated stalk cell and capitate
tip of bulbous trichomes, from a transparent, small free rim
membranous structure above the stalk in small TWP. This
type of small peltate trichome was not only detected in all
the species of Lithocarpus, but also in Castanopsis and
Castanea. However, the trichome types of species in BBT
and glabrous groups show great differences between the
current study and that of Zhou and Xia (2012), although on
the same species (L. hancei,L. harlandii,L. iteaphyllus,
L. corneus,L. konishii,L. naiadarum,L. quercifolius). Our
results show that TWP and SU were generally found in the
two groups, further more, species from BBT group have a
variety of trichome types, e.g., F, SF, St and BBT as well.
However, these obvious trichome types were not detected
by Zhou and Xia (2012). As a result, we believe that their
grouping and cladistic approaches based on trichome types
of Chinese Lithocarpus are problemetic as well.
Systematic and phylogenetic implications
The epidermal features show some degree of similarity as
well as differences among the genera of Fagaceae and
species of Lithocarpus; therefore, they can offer some clues
for the division of Lithocarpus.
Systematic status of Notholithocarpus densiflorus
N. densiflorus shared several epidermal features with the
genus Quercus s.l. Multiradiate trichomes, although not
detected in Lithocarpus in this study, but previously
recorded by Jones (1986), the small size and distinct sub-
prolate shape of the pollen gain (Manos et al. 2008), the
scattered typical compound trichome bases and the crys-
talline wax flake on the epidermis were all found in
N. densiflorus and species of Quercus s.l., but not in
Lithocarpus. The similarities of epidermal features sup-
ported the closer relationship of N. densiflorus to Quercus
rather than to Lithocarpus which is confirmed by molecular
phylogenetic approaches that support the close relationship
of N. densiflorus to Quercus,Castanea, and Castanopsis
(Oh and Manos 2008). Remarkably, N. densiflorus has
amphitypical stomata. This feature is uncommon in Faga-
ceae. It also supports the findings of Manos et al. (2008)
that N. densiflorus represents a separate genus in Fagaceae.
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Intergeneric phylogeny in Fagaceae
Castanopsis,Chrysolepis and Castanea share a distinct
cupule feature and were once placed together in Castanea
s.l. (Oersted 1871; Prantl 1894). The sister group rela-
tionship between Castanopsis and Castanea was supported
by wood anatomy (Lee 1968), pollen sculpture (Wang and
Chang 1991) and molecular phylogeny (Manos et al. 2001;
Oh and Manos 2008). Unexpectedly, Oh and Manos (2008)
showed that Chrysolepis and Lithocarpus (excl. L. densi-
florus) form an independent clade. Based on epidermal
features, flat subsidiary cells were shared by the two gen-
era. The trichome types in Chrysolepis are limited, except
for the unique thick-walled peltate as an autapomorphism,
SU and St are shared in most genera in Fagaceae. Leaf
epidermal features offered little information on the rela-
tionship of the two genera.
In the taxonomic system of Camus (1934–1954), Cas-
tanopsis ‘‘fissa-group’’ was mistakenly treated as the sub-
genus Pseudocastanopsis under the genus Lithocarpus.
With two notable reproductive features, the dichasium-
cupule, and the inter-valve zones in the very young cupule
that distinguish it from Lithocarpus, most taxonomists
recognize ‘‘fissa-group’’ as a distinct taxon of Castanopsis
(Barnett 1944; Forman 1966; Nixon 1997; Huang et al.
1999). Comparison of epidermal features reveals the
presence of solitary, fasciculate and fused fasciculate tric-
homes in some species of Lithocarpus, ‘‘fissa group’’ and
some Castanopsis species. But these species in Lithocarpus
either have speckled TWP trichomes and/or broad-based
trichomes or distinct APT, in contrast to extremely well-
developed TWP with a clear center in ‘‘fissa group’’ and
the species of Castanopsis. The subsidiary cell in ‘‘fissa
group’’ and other Castanopsis sepcies are thickened, but
flat in Lithocarpus (Liu et al. 2009). The trichome types
indicate that ‘‘fissa group’’ is closer to other Castanopsis
species rather than species in Lithocarpus.
In a recent taxonomical revision, Castanopsis long-
zhouica was transferred to Lithocarpus based on its unv-
alved cupule features (Chen et al. 2009). The epidermal
features of this species, sinuous anticlinal epidermal cell
walls, and F, BBT, and TWP the same as in the BBT group
in Lithocarpus, support its placement in Lithocarpus.
Therefore, the leaf epidermal features can usefully delimit
the species of Lithocarpus from those of Castanopsis.
Infrageneric phylogeny of Lithocarpus based on epidermal
features
The variation of epidermal features in Lithocarpus is quite
interesting. Three morphologically distinct groups were
revealed: (1) BBT group The species in this group have no
APT. The flat cuticle, sinuate anticlinal epidermal cells,
BBT, UC, SU, Fa, SF and membranous relic of lower
periclinal and anticlinal wall of the guard cells were
shared among the species (including L. howii,L. corneus,
L. haipinii,L. konishii,L. fordianus,L. uvariifolius,
L. quercifolius,L. areca and L. longzhouicus). On the other
hand, all of these species possess a more or less serrated
leaf margin, and corrugated cotyledon. The species of this
group were regarded as ‘‘subgenus Cryptostylis’’ by Camus
(1934–1954), or ‘‘group of L. corneus’’ (Barnett 1944). The
highly consistent reproductive and foliar features suggest
that this group is a natural clade in Lithocarpus; (2)
Glabrous group This group has only TWP trichomes, with
large trichome bases ranging from 15.5 to 25.5 lm. Their
epidermal cells were flat and the anticlinal wall of the
epidermis was mostly straight-curved to round; (3) APT
group This group has the broadest species composition and
epidermal variations. Almost all kinds of epidermal fea-
tures detected in Lithcarpus were present in this group,
except for the UC and sinuous anticlinal wall of the epi-
dermal cells. The cupule and acorn morphology varied
from the cupule totally enclosing or half enclosing the
acorn, to enclosing the acorn only at the base. These
extremely diverse features, of both vegetative and repro-
ductive morphology, indicate that this group possesses
well-developed and different strategies for adapting to the
highly heterogeneous environments that prevail in SE Asia.
Cannon and Manos (2003) sampled the representative
geographic ranges of Lithocarpus to study the phylogeog-
raphy of the genus. Although their cladistic analysis did not
resolve the phylogeny in Lithocarpus, their study revealed
two major cpDNA clades with one clade restricted to
Borneo and another clade widespread. However, less is
known about epidermal features of the Lithocarpus species
distributed in Borneo. Cannon and Manos (2000) studied
the leaf epidermal features of four endemic Bornean
Lithocarpus from section Synaedrys. Their results showed
the four Bornean Lithocarpus species in section Synaedrys
all possess typical APT and TWP, which are consistent
with the trichome types in APT group in our study. Whe-
ther the Bornean species with cpDNA endemism also have
unique leaf epidermal features is still unknown and should
be studied in the future. Based on nuclear CRABS CLAW
sequences ML tree of Fagaceae, three clades were present
in Lithocarpus:(L. balansae ?L. laoticus ?grandifo-
lius)?[(L. corneus ?L. pachylepis)?(L. dealbatus,
L. xylocarpus, etc.)] (Oh and Manos 2008). This cluster is
partly supported by epidermal features, that BBT
(L. corneus and L. pachylepis) and APT (e.g., L. dealbatus,
L. xylocarpus and L. silvicolarum) were clustered in dif-
ferent clades. The glabrous group species L. grandifolius,
APT group species L. laoticus and L. balansae, formed an
independent clade. The tree topology changed when ITS
sequences were added to the analysis (Oh and Manos 2008).
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Neither CRABS CLAW nor its combination with ITS sequence
can offer a robust resolution to Lithocarpus clades.
Leaf epidermal features have close correlations to
environmental factors (Haworth and McElwain 2008). The
variations of stomatal aperture, trichome morphology and
wax flake on the cuticle may be of ecological significance.
As a result, epidermal features are not only controlled by
their genetic bases but are also shaped by environmental
factors. Whether the three distinct epidermal groups have a
genetic base or a convergent evolutionary pattern to adapt
to environmental factors needs to be further explored with
finer phylogenetic resolution and analysis using climate
variations to reveal patterns of homoplasy.
Comparing the stomatal frequency of evergreen Cas-
tanopsis (229–516/mm
2
), Chrysolepis (416/mm
2
), Trigo-
nobalanus s.l. (320/mm
2
) (Liu et al. 2009) and Quercus
(115–405/mm
2
) (Lou and Zhou 2001; Deng 2007) with that
of deciduous Fagus (315–677/mm
2
) (He et al. 2007),
Quercus (405–682/mm
2
) (Zhou and Wilkinson 1995) and
Castanea (476–726/mm
2
) (Liu et al. 2009), the deciduous
species mostly have higher stomatal frequency to facilitate
high CO
2
assimilation.
Paleobotanical implications
Leaf cuticle features are useful in identifying fossil leaves
of Fagaceae. The sinuous anticlinal wall of epidermal cells
is widely detected in Fagaceae, especially in deciduous
taxa, such as Fagus,Quercus and Castanea (Jones 1986;
Liu et al. 2009), but this characteristic is also present in
some other evergreen taxa belonging to Lithocarpus,
Quercus and Castanopsis (Lou and Zhou 2001; Liu et al.
2009; Zhou and Xia 2012). Therefore, it is a homoplastic
feature, but can be applied at lower taxonomic levels for
identification purposes.
The trichome and trichome base types on the abaxial
surface were special and almost consistent, making them
important features in paleobotanical studies. APT in
Lithocarpus (Jones 1986; Uzunova et al. 1997), TWP
trichomes in Chrysolepis, glandular peltate trichomes in
Trigonobalanus, multiradiate trichomes in Quercus s.l. and
Notholithocarpus densiflorus are autapomorphic to these
genera and useful in identifying leaf fossils. For the species
in Lithocarpus without APT, their trichome bases are
restricted to broad trichome base and fasciculate trichome
base. Although the broad trichome bases were also found in
some species of Castanopsis and Quercus, they were
diversified and always combined with small simple tri-
chome base with dark stained cell wall and/or compound
trichome base. These trichome base features can be easily
distinguished using cuticle samples and offer a good
diagnostic feature to accurately distinguish the species of
Lithocarpus from those of Castanopsis and Quercus.
Stomatal features, including stomatal type, stomatal
size, and stomatal frequency can also assist in identifying
fossil leaves of Fagaceae. Cyclocytic stomata with flat
subsidiary cells were present in Lithocarpus and Quercus.
In Castanopsis, the subsidiary cells were thickened
according to Liu et al. (2009), but the range of stomatal
size in Lithocarpus (17.9–37.6 lm916.0–35.5 lm) is
different from that of Quercus subg. Cyclobalanopsis
(10.2–20.4 lm95.1–12.3 lm) (Lou and Zhou 2001;
Deng 2007). The stomatal size in Lithocarpus
(27.6 ±8.2 lm926.5 ±9.1 lm) is larger than that in
Castanopsis (21.3 ±4.6 lm918.7 ±5.5 lm), although
their stomatal frequency was similar (213–574/mm
2
in Litho-
carpus and 229–516/mm
2
in Castanopsis)(Liuetal.2009).
Acknowledgments We thank Mr. Allen Coombes of the Herbarium
and Botanic Garden of the University of Puebla and Professor Arshad
Ali of University of Florida for their kindest help in correcting the
language of the manuscript. Gratitude is expressed to the curators of
KUN, IBK, CSH and SWFC for providing specimens. This work was
supported by grants from the National Natural Science Foundation of
China (31100154 and 31270267); Shanghai Municipal Natural Sci-
ence Foundation (11ZR1435500); the Shanghai Municipal Adminis-
tration of Forestation and City Appearances (F112419), and the
Innovation Program of Shanghai Municipal Education Commission
(12YZ157).
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