Leaf Epicuticular WAX Morphology and Trichomes Presence on The Sudanese Acacias

Abstract

This is the first study of the epicuticular wax on the leaves of Sudan Acacias. It was carried out in Birkbeck College, University of London. Morphology of leaf epicuticular wax and trichomes presence were examined on twelve Sudanese Acacia species from Khartoum and Erkowit (Eastern Sudan). Comparison of the scanning electron micrographs showed that, the epicuticular wax structure was similar on both the adaxial and abaxial surfaces among the studied species.Two main types of wax were distinguished: upright flakes and smooth flakes. Both with dense or sparce irregular particles. The variation in the wax deposits between the studied species is due to specific responses to environmental elements. The distribution of trichomes was found to be of three different types: single row on the margin; a narrow row on the margin; or trichomes distributed all over the surface which was found only in A. albida (new name = Faidherbia albida) and that suggests an extreme adaptation to water loss.

Full text

IJBRITISH-47 ISSN 2349-9419 www.ijbritish.com
80 | I J B R I T I S H
Volume 1 2014 Issue 3 SEP-OCT IJBRITISH
Leaf Epicuticular WAX Morphology and Trichomes
Presence on The Sudanese Acacias
Maha A.Y. Kordofani¹, Martin Ingrouille² and Abdul Rahman H. Baleela,³
¹Department of Botany, Faculty of Science, University of Khartoum, Khartoum, Sudan
²Biology Dedpartment, Birkbeck College, University of London, UK.
³ University of ALBaha, ALBaha, Saudi Arabia.
Abstract:
This is the first study of the epicuticular wax on the leaves of Sudan Acacias. It was carried out in Birkbeck
College, University of London. Morphology of leaf epicuticular wax and trichomes presence were examined
on twelve Sudanese Acacia species from Khartoum and Erkowit (Eastern Sudan). Comparison of the
scanning electron micrographs showed that, the epicuticular wax structure was similar on both the adaxial
and abaxial surfaces among the studied species.Two main types of wax were distinguished: upright flakes
and smooth flakes. Both with dense or sparce irregular particles. The variation in the wax deposits between
the studied species is due to specific responses to environmental elements. The distribution of trichomes was
found to be of three different types: single row on the margin; a narrow row on the margin; or trichomes
distributed all over the surface which was found only in A. albida (new name = Faidherbia albida) and that
suggests an extreme adaptation to water loss.
Key words:
Acacia; epicuticular wax; trichomes; leaf.

IJBRITISH-47 ISSN 2349-9419 www.ijbritish.com
81 | I J B R I T I S H
Volume 1 2014 Issue 3 SEP-OCT IJBRITISH
INTRODUCTION
The use of characters of the leaf surface as an aid in
taxonomical studies was discussed by Stace (1984). Within
the Mimosaceae the studies have focused on the anatomy of
the leaf surface. Cutler (1969) studied the leaf of A. albida
and reported that the adaxial and abaxial surfaces are the
same, hairs are unicellular, elongated, with a few septa and
stomata are mostly paracytic. Pettigrew and Watson (1973)
studied the leaf surface of some of the Australian Acacias
and reported the presence of paracytic and cyclocytic
stomata. Equally the hairs were found to vary from smooth
to granular, short to long and in some species hairs may take
the form of epidermal prickles and trichomes all over the
phyllodes. Bleckmann et al (1980) reported that the cuticular
structure and thickness in leaflets of Prosopis velutina
Woot. varies with the environment, and in addition to that
they reported the presence of trichomes, finger like
projections and irregular plates
The epicuticular wax, which covers the surface of higher
plants, appears as crystallized bodies in a great diversity of
form and thickness. Over the previous three decades,
detailed studies on wax structure have been carried by a
number of investigators. Amelunxen et al (1967) presented a
system of classification: granules; rods and filaments; plates
and scales; layers and crusts; aggregate and liquid or
viscous coatings.
Jeffree et al (1975) showed that the form in which the wax is
deposited on the leaf surface is very largely dependent upon
its chemical constitution since it can be dissolved and
recrystallized from solution without changing its form.
Barthlott and Wollenweber (1981) recognized seven types
of wax deposits in about 5000 species grouped into either
continuous (flat sheets, smooth or sculptured) or
discontinuous (flat plates, particles, rodlets, upright flakes:
(random / regular), and ribbons or filaments. According to
Baker (1982) all plants carry an amorphous wax film upon
which may be superimposed crystalline structures in the
form of plates, tubes ribbons rods and filaments and
dendrites. Lyshede (1982) working on the xerophytic plants,
observed that the wax deposited around the stomata was
found to be thicker than on the surface. Christopher&
Reinhard (2010) found that wax coating on plant surfaces
considered to be non uniform in composition, where the
percentage of primary alcohol approximately equal between
intracuticular and epicuticular layers.
Studies focused on wax deposits in the Mimosaceae include
Bocher (1975) who noticed the presence of small flakes on
the surface of the leaves of Prosopis sp. whereas around the
stomata a thick wax coating was deposited. Baker (1982)
reported the presence of plates on Acacia leaves. Canning
(1978) studied the wax structure in the leaves of some
Australian Acacias and reported that species fall into four
classes: those with wax tubes, those with wax in angular
plates, stellate wax in a basal position and no wax present. .
Duarte & Wolf (2005) repoted that the epidermal cells were
coated with filament of epicuticular wax and there are
unicellular,non glandular trichomes in Acacia podalyriifolia
in Australia.
Since Acacias are xerophytes, and xerophytes were found to
adapt themselves to the environment by changing their wax
structure (Lyshede, 1982), the study of the relationship
between the morphology of the wax and its distribution in
the Sudan Acacias and the environment was thought to be
useful.
II. Material and Methods
10 leaflets from mature leaves of twelve Acacia species (A.
asak (Forssk.) Willd. ,A. etbaica Schweinf. , A. raddiana
Savi, A. tortilis (Forssk.) Hayne , A. sieberiana DC., A.
Albida Delile (new= Faidherbia albida Delile) , A.
gerrardii Benth., A. mellifera (Vahl) Benth. , A. seyal, A.
nilotica (L.) Willd. ex Delile , A. oerfota (Forssk.)
Schweinf. and A. ehrenbergiana Hayne) were collected
from Erkowit and Khartoum in the Sudan. They were
preserved in herbarium sheets. Different methods were tried
for the preparation of the leaflets, but Barthlott’s and
Wollenweber’s (1981) method was used because in all other
methods the wax was found to be dissolved. The material
was transferred to stubs with double adhesive tape and
coated with gold using the E.M. Scope sputter coater and

IJBRITISH-47 ISSN 2349-9419 www.ijbritish.com
81 | I J B R I T I S H
Volume 1 2014 Issue 3 SEP-OCT IJBRITISH
examined using the JEOL SM- 35-CF scanning electron
microscope in Birkbeck College, University of London.
Both abaxial and adaxial surfaces of the leaflets were
studied. The study was repeated using three different leaves
from each of the three different plants for each species from
each location. Photomicrographs were taken for each species
using 18 kv. and different magnifications (x 440 for wax, X
4400 for trichomes The terminology used is after Barthlott
and Wollenweber (1981). To help in understanding the
terminology used the characters are shown in Plates (1&2).
III. Results
A comparison of the abaxial and adaxial surfaces of leaves
of the twelve Acacia species showed no differences between
them. All the following descriptions are from the adaxial
surfaces.
No variation was observed between the three replicates of
each species from each site studied nor within the species
between the different sites.
A.asak and A.etbaica showed similarity in most characters.
A .raddiana, A. gerrardii, A. mellifera and, .A.sieberiana are
also similar to each other.
Wax morphology and trichome distribution were found to be
different between the studied taxa. Two main types of wax
on the leaflets were found: up right flakes (with dense or
sparse irregular particles) and smooth flakes (with dense or
sparse irregular particles).
The distribution of trichomes was found to be of three
different types: a narrow row on the margin which was
found in the leaves of A. asak, A. raddiana, and A.
ehrenbergiana; single row on the margin which was found
in all the other species; or trichomes distributed all over the
surface which was found in A. albida.
The wax morphology for each species is described and
presented with an accompanying photomicrograph in Plate
(3):1-12.
Plate(1):Terminology used for wax morphology.
a = Up-right flakes. b = Smooth flakes
c = Dense irregular particles. d = Sparse irregular particle
Plate (2): Distribution of Trichomes
a- Single row of Trichome on the margin e.g. A. mellifera
leaf.
b- Narrow row of Trichome on the margin e.g. A. asak leaf.

IJBRITISH-47 ISSN 2349-9419 www.ijbritish.com
82 | I J B R I T I S H
Volume 1 2014 Issue 3 SEP-OCT IJBRITISH
c- Distribution of Trichomes all over the surface of A. albida
leaf
Plate (3): Wax description of the studied Taxa ( X. 440)
1- A. asak: upright flakes, star- like discrete and with sparce
irregular particles. (Trichome forms a narrow row on the
margin).
2- A. etbacia: Upright flakes, star- like forming a net – like
structure with sparce irregular particles. (Trichomes are
found in a single row on the margin).
3- A. raddiana: Smooth flakes, continuous with sparce
irregular particles. (Trichomes form a narrow row on the
margin).
4- A. tortilis: Upright flakes, star-like discrete, with dense
irregular particles. (Trichomes form a single row on the
margin).
5- A. sieberiana: Smooth flakes, continuous with dense
irregular particles. (Trichomes form a single row on the
margin) .
6- A. albida: Upright flakes, star-like with spaces, with
sparce irregular particles. (Trichomes distributed all over
the surface).

IJBRITISH-47 ISSN 2349-9419 www.ijbritish.com
83 | I J B R I T I S H
Volume 1 2014 Issue 3 SEP-OCT IJBRITISH
7- A. gerrardii: Smooth flakes, continuous with sparce
irregular particles. (Trichomes are found a single row on
the margin) .
8- A.mellifera: Smooth flakes, continuous with sparce
irregular particles. (Trichomes are found a single row on
the margin) .
9- A .seyal: Smooth flakes, continuous with sparce irregular
particles. (Trichomes form a single row on the margin).
10- A. nilotica: Smooth flakes, continuous with dense
irregular particles. (Trichomes form a single row on the
margin).
11- A nubica: Smooth flakes, continuous with dense
irregular particles. (Trichomes form a single row on the
margin).
12- A. ehrenbergiana: Upright flakes, star-like with spaces,
with sparce irregular particles. (Trichomes form narrow row
on the margin).

IJBRITISH-47 ISSN 2349-9419 www.ijbritish.com
84 | I J B R I T I S H
Volume 1 2014 Issue 3 SEP-OCT IJBRITISH
Discussion
The present finding of equal wax deposition on the abaxial
and adaxial surfaces conform to Cutler's (1969) and
Bleckmann et al. (1980)) findings on Prosopis velutina and
Acacia greggii and A. albida respectively. Bleckmann et al.
(1980) explain this fact as a result of the normal leaf
development to which the abaxial surface is initially
exposed to sunlight, as the leaf expands to maturity the
adaxial surface is then exposed.
The variation in the wax deposits between the studied
species regarded as specific responses to environmental
elements. Environmental factors such as the intensity and
quantity of light, relative humidity and U.V. radiation are
important factors in the development of both epicuticular
wax and cuticle (Bleckmann et al., 1980).
Correlation between surface wax morphology and
reflectivity in the U.V. region of the spectrum has been
suggested by Clarke& lister (1975). They assumed that this
could enable plants to tolerate extreme light intensities in
habitat altitudes. However, Davis (1978) study of leaves of
dwarf orange varieties grown outdoors produced little
relationship between U.V. light and cuticle patterns.
Up—right flakes occurred in 50% of the species studied.
Up-right flakes (=rough surface) are known to reflect and
scatter high light and heat intensities. This is in agreement
with Harlley's (1982) finding on Eucalyptus and Cutler
(l982) on one study of Aloes of South Africa where
roughness of the leaf-surface was found to be correlated
with habitat. In arid conditions the plants are characterized
by having an up-right flakes whereas smooth flakes are
found in plants which face wet conditions or a dry habitat
sometimes during the year. The results reported here are in
agreement with these hypotheses For example A. tortilies
which is always found in arid conditions, has got up-right
flakes whereas A. raddiana which favours wet habitats
(deep rooting in khor bottoms) is characterized by having
smooth flakes, A. asak and A. etbica which were collected
from Erkowit are characterized by having upright flakes,
possibly a response to high intensity of light in the high
altitude in Erkowit.
Other functions suggested for the presence of wax in the
plants beside the normally acceptable protective function are
self cleaning in dusty places and water loss prevention. It
has been suggested also that wax discourages predation
particularly by insects (Harlley, 1982). Also epicuticular
wax could cause an apparent resistance to herbivores
(Dragan et al., 2008).
The functions of trichomes in the plants have been studied
by many authors including Shields (1950) and Metcalfe &
Chalk (1979). According to Woolley (1964), trichome
affects transpiration by influencing the water diffusion
boundary layer and decrease air movement at the leaf
surface. Lyshede (1977) suggested that trichomes are used
by the plants to absorb water from the environment. In
addition to that trichomes can have an effect on the
temperature regulation of many plants by the reduction in
absorption of radiant energy or by energy dissipation
(Ehleringer et al, 1976). Fahn (1986) described the presence
of trichomes in xerophytes as an adaptation associated with
arid condition. Further more trichomes act as a physical
obstacle aphids feeding(Bahlmann,2003).
The presence of trichomes in this study (of marginal
distribution in all the species of Acacia studied except in A.
albida (where they occur all over the surface) may be
attributed to the fact that they cause turbulence which helps
to reduce water loss. In the case of A. albida the distribution
of trichomes suggests an extreme adaptation to water loss.
A. albida is the only species in full leaf in the dry season.
References
[1]Amelunxen, F., Morgenroth, K. and Picksak,
T. ( 1967 ). Untersuchungen and der epidermis
demstereoscan- Elektronenmikroskep. Z.
Pflanzenphysiol. 57:79-95.
[2] Bahlmann, L., Govender, P. and Botha A.M.
(2003). Leaf Epicuticular Wax Ultrastructure and
Trichome Presence on Russian Wheet
Aphids(Duraphis noxia) Resistant and Susceptible
Leaves. African Entomology 11(1):59-64.
[3] Baker, E.A. (1982 ), Chemistry and
Morphology of Plant Epicuticular Waxes. In The
Plant Cuticle. Cutler, D.F., Alvin, K.L. & Price,

IJBRITISH-47 ISSN 2349-9419 www.ijbritish.com
85 | I J B R I T I S H
Volume 1 2014 Issue 3 SEP-OCT IJBRITISH
C.E. ( eds). 139-165. Academic Press, London ,
New York.
[4] Barthlott , W. & Wollenweber , E. (1981 ).
Zur Feistrukture, Chemie und Taxonomischen
Signifikanz Epicuticularer Wachse und Ahnicher
Sekrete . Trop. Subtrop. Pflanzenwelt. 32: 35-
97.
[5] Bleckmann, C.A., Huall, H.M. and Hoshaw.
R.W. (1980 ). Cuticular Ultrastructure of
Prosopis velutina and Acacia greggii leaflets.
Botanical Gazette, 141(1): 1-8.
[6] Bocher, T.W. (1975), Structure of the
Multimodal Photosynthetic Thorns in Prosopis
kuntzei Harms. Det Kongelige Danske
Videnskabernes Selskab. Biologiske Skrifter,
20(8):1-43.
[7] Canning, E.M. (1978). Studies of the Waxes
on Acacia Leaves and Phyllodes in Australia.
Kew Bulletin, Fasc. 1.
[8] Christopher, B., Reinhard, J. (2010).
Composition Differences Between Epicuticular
and Intracuticular Wax Substructure: How Do
Plants Seal Their Epidermal Surfaces. J.
Experimental Botany, Vol.62,Issue 3, pp. 841-853.
[9] Clark,J.B. and Lister,G.R. (1975).
Photosynthetic Action Spectra of Trees. II. The
Relationship of Cuticle Structure to the Vissible
and ultraviolet Properties of Needles From Four
Coniferous Species. Plant Physiology,55:407-413.
[10] Cutler, D.F. (1969). The Vegetative
Anatomy of Acacia albida Del.Kew Bulletin,
2312: 203- 208.
[11] Cutler,D.F.(1982). Cuticular Sculpturing
and Habitat in Certain Aloe Species (Liliaceae)
From South Africa. In: Cutler,D.F., Alvin,K.L.
and Price,C.E., eds.The Plant Cuticle. Linnean
Society Symposium Series 10. London: Academic
Press, 425-444.
[12] Davis, G.(1978). Effect of Light Quality and
Irradiance on Development of Citrus Mitis Leaf
Cuticles. Bot. Gaz., 139(4):390-392.
[13] Dragan, A.Z, Nevenla, V. and Stanislav,T.
(2008).Epicuticular Wax Content in the Leaves of
Cabbage (Brassica oleracea L. var. capitata) as a
Mechanical Barrier Against Three Insect Pests.
Acta Agriculturae Slovenica, 91:2, 361-370.
[14] Durate, M.R. and Wolf, S. (2005).
Anatomical Characters of the Phyllode and Stem
of Acacia podalyriifolia A.Cunn.ex G.Don
(Fabaceae). Rev. Bras. Farmacogn. Vol.15 no.1
Joao Pessoa.
[15] Ehleringer, J.R. and Mooney, H.A. (1978).
Leaf Hairs:Effects on Physiological Activity and
Adaptive Value to a Desert Shrub. Oecologia,
37:183-200.
[16] Fahn, A. (1986). Structural and Functional
Properties of Trichomes Of Xeromorphic Leaves.
Annals of Botany, 57:631- 637.
[17] Harlley, R. (1982). Do waxes on Juvenile
Eucalyptus Leaves Provide Protection from
Grazing Insects. Australian Journal of Ecological,
7: 347-352.
[18] Jeffree, C.E., Baker , E.A. and Holloway,
P.T. (1975). Ultrastructure and Recrystallization
of Plant Epicuticular Wazes. New Phytologist, 75:
539-549.
[19] Lyshede, O.B .(1977), Structure and
Function of Trichomes in Spartocytisus filipes.
Botaniska Notiser, 129: 395-404.
[20] Lyshede, O.B. (1982) . Structure of the
Outer Eepidermal Wall in Xerophytes. In The

IJBRITISH-47 ISSN 2349-9419 www.ijbritish.com
86 | I J B R I T I S H
Volume 1 2014 Issue 3 SEP-OCT IJBRITISH
Plant Cuticle.Cuyler, D.F. Alvin , K.L & Price,
C.E (eds). 87-97.Academic Press. London.
[21] Metcalfe, C.R. and Chalk , L. (1979) .
Anatomy of the dicotyledons, 2nd edn. Vol. I.
Systematic anatomy of leaf and stem, with a brief
history of the subject. Clarendon Press:
Oxford. 276 pp.
[22] Pettigrew, C.J. and Watson, L.. 1973. On the
Identification of Sterile Acacia’s and the
Feasibility of Establishing and Automatic key-
generating System. Aust. J. Bot., 21: 141-150.
[23] Shields, L.M. (1950). Leaf Xeromorphy as
Related to Physiological and Structural
Influences. Bot. Rev., 16: 399-447.
[24] Stace, C.A. (1984). The Taxonomic
Importance of The Leafe Surface. In Current
Concepts in Plant Taxonomy, Heywood, V.H. and
Moore, D.M. (eds), 67-94. Academic Press,
London.
[25] Woolley, J.T. (1964). Water Relations of
Soyabean Leaf Hairs. Agron. J. 56: 569-571.