Structural traits of some species of Hydrocotyle (Araliaceae

© 2010 E. SchweizerbartÕ sche Verlagsbuchhandlung, Stuttgart, Germany www.schweizerbart.deDOI: 10.1127/1869-6155/2010/0128-0015 1869-6155/2010/0128-0015 $ 04.50

Received January 16, 2009, in revised form August 3, 2009, accepted August 30, 2009

Plant Div. Evol. Vol. 128/3–4, 329–346E Stuttgart, September 17, 2010

Structural traits of some species of Hydrocotyle (Araliaceae) and their significance for constructing the generic system

By Alexandra I. Konstantinova and Elena Yu. Yembaturova

With 7 figures and 1 table

Abstract

Konstantinova, A.I. & Yembaturova, E.Yu.: Structural traits of some species of Hydrocotyle (Arali-aceae) and their significance for constructing the generic system. — Plant Div. Evol. 128: 329–346. 2010. — ISSN 1869-6155.

The characterization of Hydrocotyle species was traditionally almost entirely based on leaf characters, but these features are quite variable and directly dependent on age and ecological factors. However, we have shown that the fruits of representatives of these species, which display a certain amount of uniformity with regard to their macromorphology are very diverse anatomically. The objective evalu-ation of this kind of polymorphism in fruit anatomy, which is connected, first of all, with the size and shape of the cells constituting the crystalliferous layer in the pericarp, presence or absence of so-called “hydrocyte parechyma” complexes amongst the fruit tissues and other key features for Apiaceae sys-tematics, seems to be extremely important for developing well-grounded concepts of Hydrocotyle generic system.

Keywords: fruit anatomy, generic system, Hydrocotyle, morphology.

Introduction

The taxonomic history of Hydrocotyle L., as well as other representatives of Hydro-cotylaceae Hylander (1945), starts from the works of J. P. Tournefort, who mentioned several species of this genus (H. vulgaris L., H. asiatica L., H. umbellata L.) thus in-troducing its generic name, later validated by C. Linnaeus in 1753 (Shan & Liou 1964). Linnaeus also described H. americana L. and H. chinensis L., and new genera Solandra L. and Centella L. (a little later), which were included in the genus Hydrocotyle by C. Linnaeus filius as H. solandra L.f., H. villosa L.f., H. glabrata L.f. and along with H. virgata L.f., H. linifolia L.f., H. tridentata L.f., H. ranunculoides L.f. and H. erecta L.f., described by him (Richard 1820, Perez-Moreau 1948). Throughout the last centu-ries, various investigators had been discovering new species in this genus; the confu-sion in their synonymics, which arose later on, was initiated by their limited awareness

330 A.I. Konstantinova & E.Yu. Yembaturova, Structural traits of Hydrocotyle

of the studies of both their precursors and contemporaries (Richard 1820). In the 13th edition of “Systema naturae…” of Linnaeus, published in 1791, J.F. Gmelin recorded 17 species of Hydrocotyle, and C.L. Willdenow, following a critical revision, increased the number of species up to 18, though rejecting some of previously described species and including the genus Spanathe Jacq. (described in 1789) in the genus Hydrocotyle, deprived of its generic status — as H. spanathe Willd. C.P. Thunberg, though holding the opinion that Spananthe is an autonomous genus, treated Hydrocotyle as consisting of 21 species (Richard 1820). The number of newly described species continued to grow steadily — thus, A. H. Ruiz & J. Pavon in “Flora Peruviana et Chilensis” (1802) described 9 new Hydrocotyle species (H. triflora Ruiz & Pavon , H. multiflora Ruiz & Pavon, H. globiflora Ruiz & Pavon, H. acutofolia Ruiz & Pavon, H. gracilis Ruiz & Pavon, H. incrassata Ruiz & Pavon et al.), and A. Michaux in “Flora boreali-ameri-cana…” (1803) presented the original description of H. lineata Michx.

In 1820 A. Richard clarified the synonyms and presented rather detailed descriptions of 60 Hydrocotyle species, placed by him in several sections. As he said (Richard 1820), at first, amazed at the huge interspecific distinctions, he had considered advisable to exclude several independent genera from Hydrocotyle, but later, after studying a large number of species thoroughly, he had quit that idea since he had observed how the dis-tinctions had dissolved in each other and had become less and less pronounced. Richard based his sectional divisions mostly on the difference in leaf blade shape and inflores-cence features. According to him, the distinctions in fruit structure of Hydrocotyle spe-cies are not significant enough to be used for setting intergeneric boundaries.

At the present time the number of species in Hydrocotyle is about 100 (Willis 1988) or even 130 (Pimenov & Leonov 1993). Moreover, the number of newly described species and varieties of Hydrocotyle (Mathias & Constance 1951, 1975, Constance & Dillon 1990) as well as the varieties acquiring specific status (Webb & Johnson 1982) continues to increase steadily. Although a huge gap in our knowledge of the genus pointed out by M. Mathias (1936) — the absence of full-valued monograph meeting all contemporary standards and embracing the entire genus — is unfortunately still to be filled. A colossal work was needed, nomenclatural aspects of which though were clarified by H. Eichler (Eichler 1987a, b, c) to a considerable extent.

Lately the concepts of placing Hydrocotyle and Trachymene Rudge close to each other within the сore Araliaceae as well as uniting Centella, Micropleura Lag., Platysace Bunge and Mackinlayeae (Plunkett & Lowry 2001, Chandler & Plunkett 2004, Plunkett et al. 2004a) within Apiaceae–Mackinlayoideae Plunkett & Lowry (Plunkett et al. 2004b) have been developed. As we believe, this concept needs to be thoroughly checked as it is at odds with the conclusions of the classical morphology and, in par-ticular, with carpological evidence on many genera Apiaceae-Mackinlayoideae, Apia-ceae-Azorelloideae and Araliaceae s. str. (Konstantinova, unpublished data).

As far as Hydrocotyle is concerned, it is noteworthy that molecular works, much to our regret, have always analyzed a very limited sampling of species of this genus (H. bowlesioides Math. & Const., H. verticillata Thunb., H. vulgaris — Plunkett et al. 2004a; H. bowlesioides, H. modesta Cham. & Schltdl., H. verticillata — Chandler & Plunkett 2004), and this does not give us an impression of Hydrocotyle diversity from the standpoint of recent molecular data.

A.I. Konstantinova & E.Yu. Yembaturova, Structural traits of Hydrocotyle 331

Thus, Hydrocotyle is a large, heterogenous genus, that is poorly defined based on macromorphological, generally vegetative characters alone. It is likely that a thorough work is needed in order to investigate the structure of reproductive sphere in this ge-nus’s species, in particular — not only macromorphological (shape, pubescence), but also anatomical details of pericarp structure. That is why the purpose of our research is to critically evaluate the advisability of such a study for this particular case and reveal the potential taxonomic value of fruit anatomy traits for establishing major groups within Hydrocotyle. Based on these traits, we attempted to line out possible carpologi-cal groups. We did not aim at finally defining these groups and determining their size and number as our limited sampling did not allow us to do that.

Materials and methods

The paper presents results of studying 16 species of the genus Hydrocotyle. The material for the re-search was obtained from the Herbarium of Botanical Institute of Russian Academy of Science (LE) and Herbarium of the Botanical Institute of Academia Sinica, Kunmimg, Yunnan, China (KUN).

The species were sampled the way that allows to, as much as possible, reflect the geographical spread of the genus Hydrocotyle: amongs the species there are cosmopolitan ones (H. bonariensis, H. verticillata), as well as species restricted to the Old World (H. sibthorpioides Lam., H. vulgaris) or to the New World (H. chamaemorus Cham. & Schltdl., Н.mexicana Cham. & Schltdl.). Also, some spe-cies are endemic to certain terrirories (H. poepigii DC. — endemic to Chile, H. hookeri Craib subsp. handelii (H. Wolff) M.F. Watson & M.L. Sheh, H. “dulongensis”, nom. unpubl.1 — to China).

To make the preparations, mainly traditional anatomical procedures were applied: dry fruits were kept in a mixture of 96% ethanol, glycerol and water in equal proportions for several days. To deter-mine the degree of lignification in cell walls as well as to locate tannins and oils corresponding his-tochemical test reactions were conducted (Prozina 1969, O’Brien & McCully, 1981).Transverse sec-tions through the middle part of fruits were examined by means of light and scanning electron microscopes (Hitachi S–450a, CamScan S–2m) at different magnifications.

Results

One of the primary methodological principles of our study was the strict necessity of obtaining comparable results. Therefore, even though our study has revealed signifi-cant diversity in morphological and anatomical features of fruit structure in the inves-tigated species, we still start from a general overview, which can be and should be given. The difference between Hydrocotyle species is presented in Table 1.

Fruit morphology

The fruit consists of two similar mericarps, markedly compressed from both sides.Mericarp outline is usually lanceolate from the back edge (oval in H. medicagi

noides Turcz.) and semi-circular (H. aconitifolia A. Rich., H. hookeri Craib subsp.

1 The species was incorrectly described by H. Li in “Flora of Dulongjia Region (NW Yunnan)” in 1993.


Tabl

e 1.

Bas

ic fe

atur

es o

f fru

it st

ruct

ure

in th

e sp

ecie

s of t

he g

enus

Hyd

roco

tyle

.

Spec

ies

Tann

ifero

us c

ells

Hyd

rocy

te

pare

nchy

ma

Cri

stal

lifer

ous c

ells

Secr

etor

y ri

b ca

nals

End

ocar

pVe

ntra

l vas

cula

r bu

ndle

s1

23

45

67

H. a

coni

tifol

ia

A.R

ich.

In 1

or 2

sube

pide

rmal

la

yers

, in

grou

ps o

r si

ngly

Mis

sing

With

mar

kedl

y ho

rses

hoe

thic

kene

d (w

ithou

t lig

nific

atio

n)

cell

wal

ls. F

orm

a

cont

inuo

us b

elt.

Min

ute,

com

pres

sed,

no

ticea

ble

only

in so

me

ribs (

mor

e of

ten

in th

e do

rsal

and

late

ral o

nes)

Thin

, 2-la

yere

dM

issi

ng

H. b

onar

iens

is

Lam

.In

1 o

r 2 in

ner l

ayer

s of

mes

ocar

p, fo

rm a

co

ntin

uous

bel

t

Dev

elop

ed

in ri

bsSi

mila

r to

“hyd

rocy

te

pare

nchy

ma”

cel

lsM

inut

e, c

ompr

esse

d,

notic

eabl

e on

ly in

som

e rib

s (m

ore

ofte

n in

the

dors

al a

nd la

tera

l one

s)

Thic

k, o

f 5–6

fibe

r la

yers

Mis

sing

H. b

owle

sioi

des

Mat

h. &

Con

st.

In 1

or 2

sube

pide

rmal

la

yers

, as a

con

tinuo

us

belt

– th

roug

hout

the

entir

e m

esoc

arp

th

ickn

ess

Mis

sing

Slig

htly

hor

sesh

oeth

icke

ned

wal

ls; a

s a

cont

inuo

us b

elt

Not

seen

at m

atur

ity

2–3

laye

red;

fibe

rs

in th

e ou

ter l

ayer

s ar

e ve

ry la

rge

in

diam

eter

Mis

sing

H. c

ham

aem

orus

C

ham

. & S

chltd

l.In

1 o

r 2 su

bepi

derm

al

laye

rs, i

n gr

oups

or

sing

ly

Form

an

en

tire

shea

thSi

mila

r to

“hyd

rocy

te

pare

nchy

ma”

cel

lsN

ot se

en a

t mat

urity

Th

in, 2

-laye

red

Mis

sing

H. d

ulon

gens

isIn

the

epid

erm

al la

yer,

form

a c

ontin

uous

bel

t, so

met

imes

inte

rrup

ted

near

the

ribs

Mis

sing

Min

ute,

with

ho

rses

hoe

wal

l th

icke

ning

Not

seen

at m

atur

ity2-

laye

red

Mis

sing

.

H. d

isse

cta

H

ook.

f.Su

bepi

derm

al la

yer;

as

a co

ntin

uous

bel

tM

issi

ngW

ith m

ore

or le

ss

even

ly th

icke

ned

wal

lsN

ot se

en a

t mat

urity

Thic

k, o

ut o

f 4 to

6

fiber

laye

rsM

issi

ng


H. h

ooke

ri C

raib

su

bsp.

chi

nens

is

(Dun

n ex

. R. H

. Sh

an &

S. L

. Lio

u)

M. F

Wat

son

&

M. L

. She

h

2 to

4 su

bepi

derm

al

laye

rs –

alm

ost e

ntire

m

esoc

arp

Mis

sing

With

mor

e or

less

ev

enly

thic

kene

d

wal

ls; f

orm

a

cont

inuo

us b

elt

Not

seen

at m

atur

ity2-

laye

red

Mis

sing

H. h

ooke

ri su

bsp.

ha

ndel

i (H

. Wol

ff)

M. F

. Wat

son

&

M. L

. She

h

In 1

to 3

sube

pide

rmal

la

yers

; diff

usel

y th

roug

hout

the

entir

e m

esoc

arp

Mis

sing

With

mar

kedl

y ho

rses

hoe

thic

kene

d

cell

wal

ls; f

orm

a

cont

inuo

us la

yer

Not

seen

at m

atur

ity2-

laye

red

Mis

sing

. Ta

nnife

rous

cel

ls

in th

e co

mm

issu

ral

zone

H. m

edic

agin

oide

s Tu

rcz.

Exoc

arp

and

su

bepi

derm

al la

yer;

fo

rm a

bel

t whi

ch

brea

ks o

nly

near

the

win

gs (d

orsa

l and

in

term

edia

te ri

bs)

Mis

sing

With

hor

sesh

oe

thic

kene

d ce

ll w

alls

Dis

cern

ible

onl

y in

the

dors

al ri

b2-

laye

red

On

the

basi

s of

vent

ral v

ascu

lar

bund

les a

sc

lere

nchy

mal

st

rand

(the

col

umn

whi

ch d

oes n

ot

segr

egat

e fr

om th

e m

eric

arpi

a tis

sues

) is

form

ed

H. m

exic

ana

C

ham

. & S

chltd

l.M

issi

ngFo

rm a

n

entir

e sh

eath

With

ext

rem

ely

th

icke

ned

(hor

sesh

oe

shap

ed) c

ell w

alls

; fo

rm a

con

tinuo

us la

yer

Min

ute,

com

pres

sed,

di

scer

nibl

e on

ly in

som

e rib

s

2-la

yere

dM

issi

ng

H. n

epal

ensi

s H

ook.

In th

e su

bepi

derm

al

laye

r; fo

rm a

di

scon

tinuo

us b

elt

Mis

sing

With

mor

e or

less

ev

enly

thic

kene

d w

alls

; fo

rm a

con

tinuo

us b

elt

Not

seen

at m

atur

ity2-

laye

red

Mis

sing

. Cav

ity in

th

e co

mm

issu

ral

zone

H. p

oepp

igii

DC

.In

the

sube

pide

rmal

la

yer;

form

alm

ost

unin

terr

upte

d be

lt

Form

an

en

tire

shea

thW

ith h

orse

shoe

th

icke

ned

cell

wal

ls;

form

a c

ontin

uous

bel

t

Con

spic

uous

in a

ll rib

s2-

laye

red

Mis

sing


H. s

ibth

orpi

oide

s La

m.

In th

e su

bepi

derm

al

laye

r; fo

rm a

lmos

t un

inte

rrup

ted

belt

Mis

sing

With

mor

e or

less

ev

enly

thic

kene

d w

alls

; fo

rm a

con

tinuo

us b

elt

Not

seen

at m

atur

ity2-

laye

red;

add

ition

al

grou

ps o

f fibe

rs o

f m

esoc

arpi

al o

rigin

oc

cur n

ear t

he d

orsa

l rib

and

com

mis

sura

Mis

sing

H. u

mbe

llata

L.

In 2

–3 in

ner l

ayer

s of

mes

ocar

p; fo

rm a

co

ntin

uous

bel

t

Dev

elop

ed

in ri

bsSi

mila

r to

“hyd

rocy

te

pare

nchy

ma”

cel

lsN

otic

eabl

e on

ly in

so

me

ribs (

mor

e of

ten

in

the

inte

rmed

iate

one

s)

3–4-

laye

red

Mis

sing

H. v

ertic

illat

a Th

unb.

In 2

–3 in

ner l

ayer

s of

mes

ocar

p; fo

rm a

co

ntin

uous

bel

t

Dev

elop

ed in

th

e do

rsal

and

la

tera

l rib

s;

som

etim

es

also

in th

e in

term

edia

te

ones

With

mor

e or

less

ev

enly

thic

kene

d

wal

ls

Not

icea

ble

only

in

som

e rib

s (m

ostly

in

the

dors

al a

nd th

e in

term

edia

te o

nes)

3-la

yere

dM

issi

ng

H. v

ulga

ris L

.In

the

sube

pide

rmal

la

yer;

form

an

entir

e

belt

whi

ch in

terr

upts

ne

ar ri

bs

Dev

elop

ed

in ri

bsSi

mila

r to

“hyd

rocy

te

pare

nchy

ma”

cel

lsN

otic

eabl

e on

ly in

so

me

ribs (

mos

tly in

th

e do

rsal

one

)

2–3-

laye

red

Mis

sing


Fig. 1. External fruit morphology of Hydrocotyle species. — a, H. vulgaris. — b, H. bowlesioides. — c, H. umbellata. — d, H. mexicana. — e, H. dissecta. — f, H. bonariensis. — g, H. verticillata. — h, H. medicaginoides. — i, H .poeppigii. — j, H. sibthorpioides. — k, H. aconitifolia. — l, H. chamaemorus.


Fig. 2. External fruit morphology of Hydrocotyle species. — a, H. hookeri subsp. handelii. — b, H. nepalensis. — c, H. “dulongensis”, nom. unpubl.. — d, H. hookeri subsp. chinensis.

chinensis (Dunn ex R.H. Shan & S.L Liou) M.F. Watson & M.L. Sheh, H. nepalensis Hook., H. umbellata, H. vulgaris, H. sibthorpioides) or slightly ovate (H. dissecta Hook. f., H. “dulongensis”, H. poeppigii) from the side view (Fig.1, 2). The mericarp surface is smooth or tuberculate; the knobs appear because large tanniferous cells of mesocarp origin are located subepidermally. Fruit surface sculpturing may be elabo-rated due to peculiar protuberances of exocarp cells, grouped together — trichomes (H. bowlesioides, Fig. 6 c, d and H. hookeri subsp. handelii, Fig. 6a), however, the majority of species examined appeared to be lacking those projections (Fig. 6b).

Calyx teeth (sepals) are inconspicuous. The number of ribs appeared to be five and they are usually poorly defined, fili-

form. On the contrary, in H. medicaginoides, the dorsal and the intermediate ribs of each mericarp are large, wing-shaped, and in H. bowlesioides, H. dissecta, H. mexicana, H. sibthorpioides they are found to form a scarious edging.

The stylopodium is oblique toward the comissure, low-conical (H. chamaemorus, H. hookeri subsp. chinensis, H. hookeri subsp. handelii, H. poeppigii) or almost flat (H. bowlesioides, H. “dulongensis”, H. medicaginoides).

The stylodium is either short and upright (H. aconitifolia, H. nepalensis, H. verticillata, H. vulgaris) or long (H.chamaemorus, H. mexicana, H. poeppigii), with a capitate (H. umbellata, H. bonariensis, H. sibthorpiodes) or a clavate (H. poeppigii, H. medicaginoides, H. verticillata) stigma.


Fruit anatomy

Outline of the transverse section (Fig. 3, 4). If cut in the middle, the mericarp in the cross section is oval (H. dissecta, H. “dulongensis”, H. bowlesioides, H. nepalensis), almost round (H. chamaemorus, H. poeppigii), shaped as an elongated triangle with the widest side between the marginal ribs (H. umbellata, H. bonariensis, H. vulgaris), or almost diamond-shaped (H. hookeri subsp. handelii, H. medicaginoides).

The exocarp is single-layered, consisting of cells slightly elongated tangentially. The exocarp cells in H. medicaginoides and H. mexicana are square or radially elon-gated. The outer cell walls, commonly slightly thickened, are covered with a thin cu-ticle layer. The exocarp cells can serve as reservoirs for tannins, like in H. medicaginoides (Fig. 3d), but more often mesocarp cells appear to be tanniferous (Fig.3a, b, e-h; 4a-h; 5b). In fruits of H. bowlesioides and H. hookeri subsp. handelii some trichome-forming exocarp cells (gathered in groups) have secondarily thickened walls and actu-ally appear to be “hydrocyte parenchyma” cells.

Mesocarp. One or several subepidermal layers usually are located where large tannin-containing cells are concentrated. In some species these cells make a continu-ous belt girdling the entire perimeter of mericarp cross section and breaking only where the ribs are located (H. dissecta, H. medicaginoides, H. sibthorpioides — Fig. 3d, 4a, g); in the other ones they are sporadic (H. chamaemorus, H. poeppigii — Fig. 4e, f) or are completely missing (H. mexicana — Fig. 3c). In fruits of H. “dulongensis”, H. hookeri subsp. chinensis, H. hookeri subsp. handelii and H .nepalensis, the entire me-socarp zone consists of such tanniferous cells (Fig. 3e-h). In a number of species the major part of the mesocarp consists of roundish cells with slightly thickened lignified walls, forming thick strands under the rib vascular bundles (H. vulgaris, H. umbellata, H. bonariensis — Fig. 3a, b; 4b, c). These cells can constitute a continuous belt around the endocarp (H. mexicana, H. chamaemorus, H. poeppigii — Fig. 3c, 4e, f). Their walls possess many large pore canals (Fig. 7f). The researchers have interpreted such cell complexes differently: “tissu aerifer” (Briquet 1899), “pleenchyma” (Tamamshy-an & Vinogradova 1969), “aerofores” (Kozo-Polyanski 1916), “hydrocyte parenchy-ma” (Klimochkina 1950). This tissue appears at relatively late stages of fruit develop-ment between a bundle and crystalliferous layer and while the fruit continues to develop, it causes the bundle to move a significant distance away (Pervukhina 1950). Apparently it serves to lighten the entire fruit construction and, being a sort of a float, holds the hydrochorous fruit at water surface.

In the species whose pericarp lacks such a tissue (Fig. 3d-h, 4a, d, g, h), the most of the mesocarp commonly comprises parenchyma deformed in the course of fruit devel-opment (H. hookeri subsp. chinensis, H. dissecta, H. “dulongensis”, H. bowlesioides, H. hookeri subsp. handelii, H. nepalensis, H. sibthorpioides, H. medicaginoides).

The innermost mesocarp layer is represented by crystalliferous cells in all studied Hydrocotyle species; some researchers treated this layer as a part of the endocarp (Kli-mochkina 1950) or mesocarp (Tseng 1967). As we believe, in this case such terms as “exocarp”, “mesocarp” or “endocarp” could be used only in conditional (topographical) sense, because Apiaceae fruits develop from the inferior ovary, constituted by elements of different (hypanthial and carpellary) origin which congenially become inseparable


Fig. 3. Schemes of cross section through the middle of mericarps: — a, H. verticillata. — b, H. umbellata. — c, H. mexicana. — d, H. medicaginoides. — e, H. hookeri subsp. chinensis. — f, H. hookeri subsp. handelii. — g, H. nepalensis. — h, “H. dulongensis”. — Excp = Exocarp, Endcp = endocarp, Tc = tanniferous cells, Hp = “hydrocyte parenchyma”.


Fig. 4. Schemes of cross section through the middle of mericarps. — a, H. dissecta. — b, H. bonariensis. — c, H. vulgaris. — d, H. bowlesioides. — e, H. chamaemorus. — f, H. poeppigii. — g, H. sibthorpioides. — h, H. aconitifolia.


Fig. 5. Fragments of cross section through the middle of mericarps between ribs in Hydrocotyle spe-cies. — а, H. mexicana, — b, H. vulgaris. — Excp = Exocarp, Mscp = mesocarp, Endcp = endocarp, Tc = tanniferous cells, Hp = “hydrocyte parenchyma”, Crc = crystalliferous cells.

(Shibakina 1984). We believe the above mentioned layer belongs to the mesocarp since its cells in a number of representatives show striking similarity to those of mesocarp “hydrocyte parenchyma” (Tikhomirov & Konstantinova 1995). Radial and inner tangential walls in crystalliferous cells are often thicker than the outer (tangen-tial) ones (Fig. 5a, 7c) — that is why in cross section these cells look as if they have horseshoe wall thickening (H. hookeri subsp. handelii, H. mexicana, H. poeppigii — Fig. 3c, f, 4f, 7a, c); or crystalliferous cells just are strikingly similar to “hydrocyte parenchyma” cells (H. bonariensis, H. chamaemorus, H. umbellata, H. vulgaris — Fig. 5b; 7d, e).

Sometimes horseshoe thickenings of cell walls are very poorly defined (H. bowlesioides), sometimes, on the contrary, they are well pronounced, but not lignified (H. aconitifolia). The crystalliferous layer can form a continuous belt surrounding the en-docarp following the perimeter of the mericarp cross section (H. bowlesioides, H. “du-longensis”, H. hookeri subsp. chinensis, H. hookeri subsp. handelii, H. mexicana, H. nepalensis, H. poeppigii, H. sibthorpioides), or can be broken near the commissure or the dorsal rib (H. bonariensis, H. umbellata, H. vulgaris, H. medicaginoides, H. verticillata). Compared to other pericarp cells, the crystalliferous cells could be very large


Fig. 6. Fruit surface. — a, H. hookeri subsp. handelii, papillose surface (trichome-forming exocarp cells). — b, H. mexicana surface sculpturing. — c, d, H. bowlesioides, papillose surface (trichome-forming exocarp cells).

and constitute approximately one half of the pericarp thickness, as it takes place in fruits of H. aconitifolia (Fig. 4h).

As a rule, rib bundles in mature fruits are obliterated and represented only by a few xylem vessels. Secretory canals appeared to be minute and compressed by surrounding parenchyma; in some ribs they could not be determined.

The endocarp is even in thickness, thin in most fruits, more often 2-layered, very rarely thick — then consisting of up to 6 fiber layers. In the former case the fibers are usually oriented in mutually perpendicular directions (H. medicaginoides, H. mexicana, H. bowlesioides, H. chamaemorus — Fig. 5a), in the latter — primarily tangen-tially oriented (H. bonariensis, H. dissecta, H. vulgaris). Near the commissure and the dorsal ribs the fibers of all endocarp layers become oriented lengthwise. The species in which the pericarp lacks “hydrocyte parenchyma”, additional groups of sclerenchymal fibers of mesocarpial origin, adherent to endocarp, can be found (H. sibthorpioides — Fig. 4g).

Commissure. Generally, elements of ventral vascular bundles do not occur in the commissural area (Fig. 7b) — which is where parenchymal tissue of large thin-walled cells, well defined in mature fruits, is located (H. vulgaris, H. bonariensis — Fig. 4b,


Fig. 7. Fragments of cross section through the middle of mericarps. — a, H. poeppigii, innermost mesocarp crystalliferous cell layer in the pericarp. — b, H. poeppigii, commissural area of the pericarp with no ventrals. — c, H. mexicana, innermost crystalliferous cell layer, horseshoe wall thickening in crystalliferous cells. — d, H. bonariensis, medial cross section in the rib zone. — e, H. umbellata, cross section in the rib zone: note the endocarp, cells of the innermost crystalliferous layer and “hydrocyte parenchyma”. — f, H. umbellata, pores in “hydrocyte parenchyma” cell walls. — Mscp = mesocarp, Endcp = endocarp, Hp = “hydrocyte parenchyma”, Crc = crystalliferous cells, Сm = com-missure.

c). Later on this tissue can collapse and be replaced by hollow space (H. hookeri subsp. chinensis, H. “dulongensis”, H. nepalensis — Fig. 3e-h). Rarely a thick sclerenchymal strand appears on the basis of existing ventral vascular strands (H. medicaginoides — Fig. 3d).


Seed coat. As in all Apiaceae, possessing unitegmal seeds, it is represented by ex-otesta — thin structureless lamina. The boundaries between its markedly deformed cells usually cannot be identified.

Discussion

Richard (1820) made intrageneric divisions, based mostly upon the characters of the vegetative sphere (primarily leaf shape) and noted how difficult it was to identify spe-cies on the basis of fruit traits. This is not surprising, since the fruits of Hydrocotyle are very small and can hardly be determined by macromorphological features. In addition, it is not always possible to assess the peculiarities of fruit structure in different species, because, even though flowers and fruits often coexist within an inflorescence for a long period of time, the plant does not stay in the flowering and fruiting stage all the time. That is why to date the identification of Hydrocotyle species is traditionally based upon the leaf characters (Shan 1936, Constance & Dillon 1990). However, the use of leaf blade shape as the key feature often does not seem advisable even for identification purposes due to its marked variability depending on the season and habitual properties of a particular individual, probably conditioned by ecological and age-specific fac-tors.

Without denying the evident significance of leaf blade morphology features and other traits of the vegetative sphere in different Hydrocotyle species, the authors wish to specifically emphasize the importance of carpological characters. Taking these traits into consideration is equally (if not more) important for building up the concept of intrageneric system.

As a result of our study of 16 Hydrocotyle species significant polymorphism of fruit anatomy within the genus was revealed. The studied representatives can be grouped according to the greatest similarity in fruit structure:1) species with “hydrocyte parenchyma” complexes in the mesocarp and crystallifer-

ous cells similar to those of this zone (H. bonariensis, H. chamaemorus, H. umbellata,, H. vulgaris);

2) species with “hydrocyte parenchyma” complexes in the mesocarp and crystallifer-ous cells strikingly different from those of these zone, with horseshoe-shaped (H. mexicana , H. poepigii) or relatively evenly thickened (H. verticillata) cell walls;

3) species lacking “hydrocyte parenchyma”, with a few tanniferous cells in outer pericarp layers, forming a broken belt around the main massif of the mesocarp parenchyma (H. aconitifolia, H. bowlesioides, H. dissecta, H. medicaginoides, H. sibthorpioides).

4) species lacking “hydrocyte parenchyma”, but with a great number of tanniferous cells, forming an unbroken or discontinuous belt around the endocarp or adjacent compressed layers of mesocarp parenchyma (H. nepalensis, H. hookeri subsp. handelii, H. dulongenis).Trying to evaluate the correlation between these groups, determined on the basis of

fruit anatomy traits, and the geographic distribution of species they consist of, the fol-lowing can be noted: if Group 1 and Group 3 include both cosmopolitan species and


those from the Old and the New World, Group 2 is composed of the New World spe-cies (with horseshoe-shaped cell walls) and Group 4, on the contrary — of those from the Old World. It is possible, however, that if the number of species sampled grew, these tendencies would not be so obvious.

Besides, some species investigated are characterized by particularly interesting car-pological features, such as:• the formation of wings on the basis of the dorsal and intermediate ribs (H. medi

caginoides).• the presence of extremely large crystalliferous cells with unevenly thickened (with

no lignification) walls; these cells constitute a considerable part of pericarp thick-ness (H. aconitifolia);

• the presence of special groups of lignified fibers of mesocarpic origin, adhering to the crystalliferous layer from the outside (H. sibthorpioides).According to the data obtained, an important conclusion can be drawn stating that

fruit structure features in Hydrocotyle representatives are of great interest for taxono-mists studying this genus; therefore, a detailed carpological investigation of all species is needed.

Conclusions

Fruit structure traits, so meaningful for compiling the taxonomic system of Apiaceae in general, have been neglected in the construction of the Hydrocotyle generic system. The present research was undertaken to fill (at least partly) the gap in our knowledge of the carpology of both Araliaceae at large and the genus Hydrocotyle in particular. It was revealed that by the nature of micromorphological (anatomical) features of fruit structure, various Hydrocotyle species demonstrated a wide range of interesting pecu-liar traits connected with the size and shape of cells of the pericarp crystalliferous layer, presence or absence of special “hydrocyte parenchyma”, location of tannin-con-taining cells, etc. On the basis of these traits we so far were able to reveal only 4 pro-visory carpological groups within Hydrocotyle. Generally, we highly evaluated the taxonomic potential of fruit anatomy traits in Hydrocotyle for furnishing the generic system hoping that this research will stimulate a series of carpological studies dedi-cated specifically to the genus Hydrocotyle. We also hope that the investigation of carpological polymorphism in Hydrocotyle, stated in the present paper will help to elucidate its phylogenetic relationships with other representatives of Araliaceae, on the one hand, and Apiaceae subfamily Mackinlayoideae, on the other hand.

Acknowledgements

The authors would like to express their sincere gratitude to the late Prof. Vadim N. Tikhomirov (Department of Higher Plants, Moscow State University) who initiated and profoundly supervised this research at its early stages. We wholeheartedly thank Prof. Alexander P. Melikian (Department of Higher Plants, Moscow State University) — we could not have done this work without his active participation and valuable remarks. Special thanks to Dr. Tatyana V. Lavrova for the material provided for research.


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Addresses of the authors: Dr. Alexandra Konstantinova, Department of Higher Plants, Biological Faculty, M.V.Lomonosov

Moscow State University, 119992, Moscow, Russian Federation; e-mail: [email protected]; Dr. Elena Yu. Yembaturova Russian State Agrarian University — K.A. Timiryazev MSKHA, Depart-ment of Botany, ul. Timiryazevskaya, 49 127550 Moscow, Russian Federation; e-mail: [email protected]

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Structural traits of some species of Hydrocotyle (Araliaceae