Marsilea:

Habit and Habitat of Marsilea:

Marsilea is commonly known as “pepper wort” or “water fern”
(although it is a fern but hardly resembles a true fern). It is
represented by about 53 species which are cosmopolitan in
distribution but abundantly found in tropical countries like Africa and
Australia. About 9 species have been reported from India.

Either the species are hydrophytic or amphibious i.e., they grow

rooted in mud or marshes and shallow pools or are completely
submerged or partially or entirely out of water in wet habitats. M.
hirsuta is an Australian xerophytic species. M. hirsuta and M.
quadrifolia are two most common Indian species usually found
growing in marshy places, wet soil or near muddy margins of ponds
and are commonly found in U.P., Punjab, Bihar, Delhi etc.

External Features of Marsilea:

The mature sporophyte is an herbaceous plant. Its underground
rhizome spreads in a diameter of 25 meter or more. The plant body is
distinctly differentiated into rhizome, leaves and roots (Fig. 1 A).
1. Rhizome:
All the species possess a rhizome which creeps on or just beneath the
soil surface. It is slender, dichotomously branched with distinct nodes
and internodes and is capable of indefinite growth in all directions as a
result of which it occupies an area of 25 metre or more in diameter.

In aquatic species the internodes are long while in sub-terrestrial

species they are short. Usually from the upper side at nodes, the leaves
are given out while from their lower side, the roots.

2. Leaves:
They are borne alternately on upper side of rhizome at nodes, in two
rows. Young leaves show circinate vernation (like ferns) (Fig. 1 A). In
some species young leaves are covered with multicellular hairs. The
leaves are compound, with basal petiole and terminal lamina.

In submerged plants the petiole is a long and flexible structure and the
lamina floats over the surface of water but in muddy or marshy plants
the petiole of the leaf is short and rigid with short lamina spreading in
the air.

The lamina consists of 4 leaflets (pinnae) which are present at the apex
of petiole. The 4 leaflets arise as a result of 3 dichotomies of the lamina
in close succession to each other i.e., 2 leaflets arise slightly higher
than other two (Fig. 1B).

Puri and Garg (1953) suggested that the leaf consists of single pinna
consisting of 4 pinnules. Pinnae have got a dichotomously branched
vein system with cross connections (Fig. 1C). The veinlets at the
margin are connected with loops thus forming a reticulum. The shape
of pinna varies from obovate to obcuneate and margin also varies from
entire to crenate or crenate to lobed.

Sometimes the pinnas are once or twice deeply dichotomously lobed

(M. biloba) or toothed (M. minuta). At night the pinna are folded
upwardly. This is known as sleeping movement of pinna. Near the
base of petiole the stalked bean-shaped sporocarps are borne.
3. Roots:
The roots are adventitious, arising from the underside of the node of
rhizome, either singly or in groups. In certain cases the roots are given
out even from the internodes (M. aegyptiaca).

Internal Structure of Marsilea:

1. T. S. Rhizome (stem):
A T. S. of the young rhizome shows a protostelic structure i.e., pith is
absent and xylem is completely surrounded by phloem but in the old
stem pith is developed in the centre and the stele is amphiphloic
siphonostelic type.

A. T. S. of the old stem is somewhat circular in outline and

shows the following structures:
(i) Epidermis:
It is the outermost limiting layer of single celled thick parenchymatous
cells. The stomata are absent.

(ii) Cortex:
It is differentiated into three regions – the outer cortex, the middle
cortex and the inner cortex.
(a) Outer cortex:
It is present just below the epidermis (also called hypodermis). It is
parenchymatous and may be one to several cells thick. Some of its
cells contain tannin.

(b) Middle cortex:

It is also called aerenchyma. It lies below the hypodermis. It consists
of large air spaces (chambers) separated by one cell thick
parenchymatous septa. In the xerophytic species e.g., aegyptiaca the
air chambers are obliterated.

(c) Inner cortex:

It is a solid tissue of several cells thickness. The outer layers are thick
walled (sclerenchymatous) while the inner layer of cells is thin walled
(parenchymatous) and compactly arranged. Some of these cells are
filled with starch or tannin.

(iii) Stele:
Stele is amphiphloicsiphonostele i.e., in the centre there is a pith
which may be either parenchymatous (aquatic species) or
sclerenchymatous (terrestrial muddy species). Xylem is present in the
form of a complete ring which is surrounded on both sides by a
complete ring of inner and outer phloem, pericycle and endodermis.

In this way the continuation of different tissues in the form of

complete ring in stele is as follows—outer endodermis, outer pericycle,
outer phloem, xylem, inner phloem, inner pericycle and inner
endodermis. The protoxylem may be well defined exarch (M.vestita)
or mesarch (M.aegyptiaca) or ill defined (M.quadrifolia).

A T. S. of the nodal region shows an amphiphloicsolenostelic condition

and is provided with one leaf gap.
2. T. S. of Petiole:
A T. S. of the petiole is somewhat circular in outline and is
differentiated into epidermis, cortex and stele.

(i) Epidermis:
It is the outermost layer of single cell thickness. The cells are
parenchymatous and slightly elongated.

(ii) Cortex:
It is differentiated into three regions: The outer cortex, the middle
cortex and the inner cortex.

(a) Outer cortex:

It is present just below the epidermis, (also called hypodermis). It is
made of thin walled cells (parenchymatous).

(b) Middle cortex:

It lies below the hypodermis and called aerenchyma. It consists a ring
of air chambers. The air chambers are separated by single layered
partitions of thin-walled parenchymatous cells.

(c) Inner cortex:

It is a solid tissue of several cells thickness. The cell layers are
parenchymatous and contain starch and tannin filled cells. In
M.minuta few sclerenchymatous layers are also present just inner to
middle cotex.

(iii) Stele:
It is somewhat triangular in outline and is of protostelic type i.e. pith
is absent. Xylem is “V” shaped with 2 distinct arms. Each arm is
provided with metaxylem elements in the centre and protoxylem is
situated at both the margins i. e., protoxylem is exarch.

The xylem is surrounded on all sides by phloem. Phloem is externally

surrounded by a single layer of parenchymatouspericycle which, in
turn, is bounded by a single layered endodermis.

3. Transverse Section of Leaflet:

A. T. S. of the leaflet shows epidermis, mesophyll and vascular
bundles.

(i) Epidermis:
It is the outermost surrounding layer and is only one cell in thickness.
It is differentiated into upper and lower epidermis. In floating leaflets
the stomata are present on the upper epidermis but in case of plants
growing in mud or moist soil where the leaves are aerial, the stomata
are present both on upper as well as lower epidermis.

(ii) Mesophyll:
It occupies a wide space between upper and lower epidermis. It is
usually differentiated into an upper palisade tissue and lower spongy
parenchyma. The palisade tissue is made up of elongated cells
provided with chloroplast. The spongy tissue is made up of loosely
arranged parenchymatous cells with large air spaces separated by
single layered septa. In submerged species, however, the mesophyll is
not differentiated into palisade and spongy parenchyma.
(iii) Vascular bundles:
In between the mesophyll tissue are present several vascular bundles.
Each vascular bundle is concentric and amphicribal type i. e., made up
of a centrally situated xylem, surrounded on all sides by phloem. The
phloem is enclosed by a single layered thick endodermis.

4. T. S. Root:
A T. S. of root is somewhat circular in outline and can be differentiated
into epidermis or piliferous layer, cortex and stele (Fig. 5A, B).

(i) Epidermis:
It is the outermost, parenchymatous, single layered covering.

(ii) Cortex:
It can be differentiated into two parts: outer cortex and inner cortex.
The outer cortex consists of large air chambers arranged in the form of
a ring (parenchymatous). These chambers are separated from each
other by longitudinal septa. The inner cortex is differentiated into
outer parenchymatous and inner sclerenchymatous regions. The inner
cortex is delimited by single layered thick endodermis.

(iii) Stele:
It is of protostelic type and occupies the central position. It is devoid of
pith. Xylem is situated in the centre which is diarch and exarch. It is
surrounded by phloem. The phloem is bounded externally by a single
layer of pericycle.

Reproduction in Marsilea:
Marsilea reproduces by two methods:
(i) Vegetative reproduction

(ii) Sexual reproduction.

Vegetative reproduction:
It takes place by means of tubers which are produced in dry conditions
from the rhizome. First a branch is given out from the rhizome, which
later on swells up due to the accumulation of food material. The
structure is termed as tuber and is capable of tiding over the
unfavourable conditions. On the return of favourable conditions it
germinates to produce a new sporophytic plant, e.g. ,M. hirsuta, M.
quadrifolia.

(ii) Sexual Reproduction:

1. Sporophytic Phase:
Spore producing organs:
Marsilea is heterosporous i. e., it produce two types of spores—
microspores and megaspores. These spores are produced in
microsporangia and megasporangia, respectively. These sporangia are
borne in special type of spore producing organ called sporocarp. The
sporocarp are born laterally on the short and lateral branches of the
(called the peduncles or pedials) petiole of leaf either near the base or
a little higher up.
They arise solitary or in clusters. The peduncle is usually unbranched
but it may be branched also. Number of sporocarp differs in different
species and varies from 1 to 20 or more. In M. vestitasporocarparises
single, in M. quadrifolia the peduncle is dichotomously branched
bearing 2-4 sporocarps, in M. polycarpa several sporocarps arise in a
linear row. The attachment of the pedicel sporocarp varies in different
species.

Mainly it is of three types (Gupta 1962):

(i) Pedicel of the solitary sporocarp is directly attached to the base of
the petiole (e.g., M. coromendelica Fig. 6A) or pedicels of many
sporocarps are attached on the petiole in a linear sequence on the
same side (e.g.,M. polycarpa; Fig. 6B).

(ii) Pedicels first become united themselves with one another and then
are attached to the petiole by a common stalk e.g., M. quadrifolia; (Fig.
6C).

(iii) Pedicels are free or slightly united and attached to the petiole by a
single point (e.g.,M. minuta; Fig. 6D).

External Morphology of Sporocarp:

Each sporocarp is an oval or bean shaped biconvex, flattened

structure. It is green and soft when it is young but at maturity it
becomes very hard and brown in colour. It is made up of a short stalk
like structure known as peduncle and the body.

The point of attachment of peduncle with the body is called raphe (Fig.
7A). Slightly above the raphe in a median plane are present 1 or 2
protuberances called tubercles. They are unequal in size and lower one
is stouter than the upper one. In some cases the tubercles are absent
e.g.,M. polycarpa.

Internal Structure of Mature Sporocarp:

The sporocarp is a bivalved structure. It can be split open in the
dorsiventral plane into two halves (valves).

Wall of sporocarp:
It is very hard, thick and highly resistant to mechanical injury. It can
be differentiated into three zones—outer epidermis, middle
hypodermis and inner parenchymatous zone. Epidermis is single
layered made up of broad and columnar cells. Its continuity is broken
by the presence of sunken stomata (Fig. 7C).

Some of the epidermal cells develop into multicellular hairs (Fig. 7D).
Hypodermis consists of two layers of radially elongated palisade like
cells. Both the layers are without intercellular spaces and have
chloroplast in their cells. Next to hypodermal layers is the
parenchymatous zone (Fig. 7B). In mature sporocarp the cells of this
zone gelatinise and form a gelatinous ring which helps in the
dehiscence of the sporocarp.
Cavity of sporocarp:
The alternating rows of sori (sing, sorus, a group of sporangia is called
sorus), one along each side lies transversely-dorsiventrally to the long
axis of the sporocarp. The sori on either side alternate with each other.
The number of sori inside the sporocarp varies from species to species.
It may be from two (e.g., M. aegyptiaca) to twenty (e.g.,M. vestita).
Each sorus bears both microsporangia and megasporangia.

Their number also varies from species to species. In M. minuta a sorus

has 4-8 megasporangia and 8-13 microsporangia. In M. aegyptiaca
each sorus has 5-16 megasporangia and 9-19 microsporangia.

In M. minuta, M. vestita, M. rajasthanensis, sometimes

megasporangia are absent in sorus. Each sorus arises on a ridge like
placenta or receptacle formed on the sporocarp wall. Each sorus is
surrounded by a thin, membranous two layered true indusium. The
indusia of adjacent sori are partially fused.

Vascular supply of the sporocarp:

It is supplied by a main dorsal vein which runs along the narrow side
facing the peduncle. From the dorsal vein, lateral branches are given
alternatively right and left, at right angle to the dorsal vein which
supplies laterally (Fig. 8). These lateral veins at their middle divide
dichotomously. In the region here lateral vein forks, arises a placental
bundle which too branches dichotomously. The first and the last
lateral veins do not possess placental bundles.

The entire internal structure of the sporocarp can be best

seen in section cut in three plains:
(i) Horizontal Longitudinal Section (H.L.S.): Section is cut
horizontally but the sporocarp is cut longitudinally.

(ii) Vertical Longitudinal Section (V.L.S.): Section is cut vertically but

the sporocarp is cut longitudinally.

(iii) Vertical Transverse Section (V.T.S.): Section is cut vertically but

the sporocarp is cut transversely.

(i) Horizontal Longitudinal Section (H.L.S.):

The section passes through the peduncle and cuts it transversely.
Peduncle shows characteristic ‘V’ shaped xylem (Fig. 9 A, B). A H.L.S.
of sporocarp shows the usual wall layers. The gelatinous ring is cut
transversely and it appears in the form of dorsal and ventral mass at
its proximal and distal ends.

The dorsal mass is more prominent than transversely along with their
two layered inducia. Sori show their alternate arrangement in the two
rows. Each sorus has a receptacle which has a central terminal
megasporangium and two lateral microsporangia, one on either side.
The lateral bundle is also cut transversely below each sorus.

(ii) Vertical Longitudinal Section (V.L.S.):

A V.L.S. of the sporocarp shows the usual wall layers. The peduncle
along its vascular bundle is cut longitudinally. The entire gelatinous
ring is cut vertically and it appears as a complete ring around the sori.
The section cut the sori longitudinally, which are arranged in many
vertical rows.

If the section passes strictly through the median plane of sporocarp,

only megasporangia are visible (Fig. 10A, B) but if it passes slightly
away from the median line, only microsporangia are visible (Fig. 10C).
(iii) Vertical Transverse Section (V.T.S.):
A V.T.S. of the sporocarp shows the usual wall layers. Peduncle is not
cut in the section. The gelatinous ring appears in the form of dorsal
and ventral mass (as in H.L.S.). The gelatinous mass on the dorsal side
is much more prominent.

In V.T.S. only two sori covered with indusia are visible on the inner
side and attached to the placental ridge on the outer side (Fig. 11 A, B).
The sori reveal many megasporangia and only two or three
microsporangia at the sides. The dorsal bundles, the lateral bundles
and the placental bundles are clearly visible.
Morphological Nature of the Sporocarp:
Two main views have been put forward by different
morphologists to explain the morphological nature of the
sporocarp of Marsilea which are discussed below:
(1) The laminar or leaf segment hypothesis.

(2) Petiolar or whole leaf hypothesis.

The laminar or leaf segment hypothesis. According to the

supporters of this hypothesis the sporocarp is a lateral
modified fertile segment of the leaf but their interpretations
are different which are given below:
(i) Russow (1872) and Busgen (1890) regarded the sporocarp to be
made up of 2 leaflets with ventral surface facing each other (Fig. 121).

(ii) Goebel (1882, 1905, 1930) considered the sporocarp as a fertile

leaflet (pinna), comparable with one or more leaflets.

(iii) Campbell (1893, 1928, 1940) regarded the sporocarp as a folded

pinnate leaf formed by the fusion of several pairs of pinnae.

(iv) Bower (1926) stated “the hypothesis seems to be tenable that the
sporocarp consists of rachis, bearing 2 rows of pinnules, this is
indicated by the veination (Fig. 12 G, H).”

(v) Eames (1936) compared it to the tip of the leaf with 4 leaflets. He
was of the opinion that the body of the sporocarp represents the two
distal leaf-lefts and the region of 1st and 2nd protuberance represent
the remaining 2 proximal leaflets (Fig. 12 D, E).

(vi) Smith (1938, 1955) considered the sporocarp as a modified pinna

with one midrib and several lateral branches (Fig. 12 A-C).

(vii) Takhtajan (1953) considered the sporocarp as a more specialised

fertile segment of a leaf as in Schizaeaceae.

(viii) Puri and Garg (1953) regarded the sporocarp equivalent to a

single leaflet which consists of as many pinnules as the number of
lateral bundles (Fig. 12F).

(ix) Gupta K. M. (1962) regarded the sporocarp to be leaflet with as

many lobes as the number of lateral bundles. The sori are situated
marignally in the depressions of the lobes.
The following evidences support the above hypothesis:
(1) The sporocarp initial and the leaf initial both are two sided apical
cells and they behave in the same way during early segmentation.

(2) In M. polycarpa, 10-15 sporocarps arise acropetally on one side of

the petiole.

(3) In M. quadrifolia, branching of peduncle is similar to the formation

of pinnules.

(4) The vascular supply of the sporocarp is similar to that of the sterile
leaflets.

(5) Presence of epidermis with stomata, hypodermis and parenchyma

with air spaces also supports the laminar hypothesis of sporocarp.

Petiolar or whole leaf hypothesis:

Johnson (1898, 1933) proposed this hypothesis and regarded the
sporocarp as homologous to the swollen end of the petiole. The
marginal cells here develop sporangia instead of lamina. The evidence
in support of this theory is that sometimes the primary sporocarp
gives rise to secondary sporocarp which develops from the initial
arising from the marginal cells of the stalk of primary sporocarp.
Structure of Microsporangium:
It is somewhat oval structure with a long stalk and is present laterally
on the receptacle. It is smaller in size. It has a single layered jacket
followed by two layers of tapetal cells. In the centre is present a cavity
filled with microspore mother cells (Fig. 14H).

At maturity the tapetal cells disintegrate and each microspore mother

cell divides reductionally forming 4 haploid microspores (Fig. 141).
The microspores are usually 32-64 in number and are liberated by the
disintegration of the microsporangial wall (Fig. 14J).
Structure of Megasporangium:
It is a spherical structure with a short stalk and is present on the top of
the receptacle (Fig. 14A). It is bigger in size than the microsporangium
(Fig. 14A). Its structure is similar to microsporangium except that only
one megaspore is present per megasporangium at maturity. The
megaspore is liberated by the disintegration of the megasporangial
wall.
Dehiscence of Sporocarp and Liberation of Spores:
The decaying of the wall of the sporocarp takes place due to bacterial
action and thus the sporangia and spores are liberated. The sporocarp
bursts open only in water in valvecular manner along the ventral side
and apex. The gelatinuous ring absorbs water and extends greatly
through the open margins of the sporocarp thus dragging out sori
along with it.

It straightens and behaves as sporophore. The gelatinous ring bears

two alternating rows of sori. The delicate mucilage wall of the
sporangia (micro-or mega) opens in water and the spores (micro-or
mega are liberated which germinate soon (Fig. 15 A, E).

2. Gametophytic Phase:
The microspores and the megaspores are the unit of male and female
gametophytes respectively.

They germinate to produce the respective gametophyte in

the following ways:
Development of male gametophyte:
The microspore is the initial stage in the development of male
gametophyte. Each microspore is a unicellular, uninucleate, thick
walled globose and haploid structure, ranging from 0- 060 to 0- 075
mm in diameter.

The cytoplasm is surrounded by inner wall called endosporium and

outer wall called exine or exosporium. The microspore germinates just
after its liberation. The first division is in a lenticular plane to form a
small lens shaped prothallial cell and a apical cell (Fig. 16A, B).

The apical cell divides transversely to form 2 equal antheridial initials

(Fig. 16C). Each antheridial initial divides periclinally to form an outer
first jacket cells (initial) and inner wedge-shaped sister cells (3-3) (Fig.
16D). The inner cell further divides by periclinal division to form a
second smaller jacket cell and a large outer cell (Fig. 16E).

The large cell again divides by a periclinal wall to form an outer or

peripheral third jacket layer and a central cell or the primary
androgonial cell (Fig. 16F). Each primary androgonial cell divides to
form 16 androcytes (Sperm mother cells) and finally metamorphosises
into antherozoids. In all 32 antherozoids are produced (Fig. 16 G-I).

The male gametophyte is developed inside the microspore and

produces 32 antherozoids with usually one prothallial cell. Sometimes
2 prothallial cells are also produced. By breaking of the jacket cells and
disintegration of male gametophytic tissue the antherozoids are
liberated.
Each antherozoid is a cork screw shaped, spirally coiled
multiflagellate, structure (Fig. 16 J, K). It is characterised by the
presence of a prominent terminal vesicle. The cilia are attached only to
the posterior coils. The coiling looses at the time of fertilization.

Development of female gametophyte:

The megaspore is the initial stage in the development of female
gametophyte. Each megaspore is a unicellular, uninucleate, ellipsoidal
structure with an apical papilla (Fig. 17A). The mucilaginous wall is a
thick structure except in the papillate region.

The wall has 2 covering layers, outer one is known as exospore and
inner one as endospore. The nucleus lies in the apical papilla (Fig.
17A). The rest of the basal portion of the megaspore contains granular
starch, oil globules and albuminous substances. The first division is in
a transverse plane at the base of papilla, thus forming an upper small
cell and a basal bigger prothallial cell (Fig. 17B).

The prothallial cell serves as a nutritive cell and provides nutrition to

the developing gametophyte while the apical cell forms the female
gametophyte. The upper smaller cell again divides transversely to
form an upper apical cell and lower basal cell (Fig. 17C). The apical cell
divides by three vertical divisions so as to form 3 lateral cells
surrounding a central cell or archegonial initial (Fig. 17D).

The archegonial initial divides by a periclinal wall forming an outer

primary cover cell and an inner central cell (Fig. 17E). The 3 lateral
cells and a basal cell by further horizontal and vertical divisions form a
jacket round the archegonium. The cover cell divides vertically by 2
divisions at right angle to each other to form 4 neck initials (Fig. 17F)
which by transverse division forms a neck of 2 tiers of 4 cells each.

The inner cells first divides transversely to form an upper primary-

neck canal cell and lower primary venter cell. The former may or may
not divide to form 1 or 2 neck canal cells. The latter divides to form an
upper ventral canal cell and a basal egg. Thus, at maturity the mature
archegonium has 8 celled neck, 1 neck canal cell, a small venter canal
cell and a large egg. Each megaspore produces a single archegonium
(Fig. 17G).

Fertilization:
The free swimming antherozoids are attracted chemotactically
towards the neck of a mature archegonium but only one enters the
neck and reaches the egg. The male and female nuclei fuse to form a
diploid structure called oospore or zygote. Thus the gametophytic
generation ends and the unit of sporophytic generation is formed. In
some species e.g.,M. drummondii (Strasburger 1907), parthenogenesis
has been observed.

Development of embryo:
Oospore is the initial stage of sporophytic generation. The first
division of the oospore is in a vertical plane (parallel to the long axis of
archegonium) to form 2 unequal cells. The bigger one is known as
epibasal cell and the smaller one as hypobasal cell (Fig. 18 A, B). This
is followed by a second transverse division to form 4 cells (quadrant
stage) (Fig. 18C).

The epibasal half gives rise to shoot and leaf whereas the hypobasal
half gives rise to root and foot. The cell of epibasal half near the neck
gives rise to cotyledon and other away from the neck, to the stem.

In the same way the cell of the hypobasal half near the neck gives rise
to root and other away from the neck, to the foot. Simultaneously, the
tissue surrounding the archegonium divides to form a 2 or 3 celled
thick calyptra which protects the embryo in young stage. The embryo
later on gives rise to an adult plant.

Life Cycle Patterns of Marsilea:

Mature plant of Marsilea is diploid. Marsilea is a heterosporous fern
because it produces 2 different types of spores i. e., microspores and
megaspores. Micro- and megaspore mother cells are produced inside
micro- and megasporangium respectively which represents the late
stage of sporophytic generation. After reduction division microspores
and megaspores are produced which represent the initial stage of
gamestophytic generation.

Microspore gives rise to make gametophyte which, in turn, produces

archegonium and egg. Both antherozoid and egg fuse to from a diploid
oospore (2x).The oospore is the initial states of sporophytic
generation. Hence, in the way the sporpytic andgametophytic
generation alternate with each other although the sporophytic phase is
dominant over gametophytic phase (fig 19,20).