Brief communication
Male function for ensuring pollination and reproductive success
in Berberis lycium Royle: A novel mechanism
SUPRIYA SHARMA* and SUSHEEL VERMA
Conservation and Molecular Biology Lab, Centre for Biodiversity Studies,
Baba Ghulam Shah Badshah University, Rajouri, Jammu and Kashmir, India
*Corresponding author (Email, [email protected]; [email protected])
In Berberis lycium anthers on alternate stamens dehisce, thus prolonging the male function so that pollination is
affected and reproduction is ensured. The large pollen sac of each bithecous anther after the appearance of longitudinal
dehiscence slit moves away from the filament while remaining attached at the tip of the connective and then orients in
such a way that pollen-laden surface faces the stigma. No pollen is available to receptive stigma as pollen grains
remain stuck to the anther sac. They do not get dispersed even by wind. Pollination and consequently reproduction is
ensured through the intervention of insect, which does not affect pollen transfer to the stigma directly but by touching
the base of the staminal filament while foraging nectar secreted by nectaries at the base of corolla, thus leading to
staminal movement. This makes the dehisced anthers stick to the stigma and deposit pollen there.
[Sharma S and Verma S 2016 Male function for ensuring pollination and reproductive success in Berberis lyceum Royle: A novel mechanism.
J. Biosci. 41 21–25] DOI 10.1007/s12038-015-9581-x
1.
Introduction
Production of viable pollen and its efficient transfer to the
stigmatic surface determines the success of male function in
plants (Carr and Dudash 1995; Ashman 1998; Verma et al.
2008). Plants have also evolved certain mechanisms to extend the period of male fitness. To maximize this, many
plants show temporal separation in anther dehiscence (Lloyd
and Yates 1982; Thompson et al. 2000; Castellanos et al.
2006). Mechanism of anther dehiscence is conserved among
angiosperms and may also influence pollination mechanism
(Buchmann 1983; Bernhardt 1996; Endress 1996a).
Intervention of insects is required in certain cases for
pollen release from anthers (Cutting 1921; Verma et al.
2008; Yi Han et al. 2008). While in most of the plants they
transfer pollen to the stigmatic surface from the anthers and
affect pollination (Meeuse 1961; Polhill 1976; Faegri and
Van der Pijl 1979; Broyles and Wyatt 1991), in Berberis
lycium, which produces deep yellow flowers in dense racemes, insects affect pollination by triggering the movement
of staminal filaments so that dehisced anthers strike to the
Keywords.
stigma and shed pollen there. The present study was conducted to understand the mechanism of male function and its
efficiency in bringing about pollination and ensuring reproductive success in Berberis lycium, an endangered plant of
high medicinal value.
2.
Materials and methods
Flowers of B. lycium were studied to determine the mechanism of anther dehiscence and pollination. Flower dependency
on insect visit to affect pollination was determined by pollination experiments including unassisted selfing, manual crosspollination and manual self-pollination, and percent fruit set
was determined when flowering was over using the formula:
Percent fruit set ¼
No: of fruits formed
100
No: of flowers treated
Stigmata in case of bagged and open pollinated flowers were
also scanned for the presence of pollen. Pollinators were collected using nets and were then etherized. Pollen load on their body
Berberis lycium; large pollen sac; male function; pollination; staminal filament
http://www.ias.ac.in/jbiosci
Published online: 22 January 2016
J. Biosci. 41(1), March 2016, 21–25, * Indian Academy of Sciences
21
Supriya Sharma and Susheel Verma
22
parts was then studied by scratching each body part in Lewis
stain and then observing under the microscope (Nikon 80 i
eclipse). The pollen–ovule ratio was calculated by calculating
Pollen−Ovule Ratio ¼
No: of pollen produced by one anther total no: of anthers in a flower
Total number of ovules produced by a single flower
To check anemophily, slides smeared with Mayer’s albumin were suspended at varying distances from plants. These
slides were examined after 24 h by staining in Lewis stain for
presence or absence of the pollen of the plant under study.
3.
3.1
Results
Another dehiscence and pollination mechanism
In B. lycium, the androecium is represented by 6 adnate and
anti-petalous stamens (figure 1A and B). Anther dehiscence
in this species is temporally separated. Anthers on alternate
stamens dehisce first. Also the lobes on alternate stamen
dehisce sequentially one by one (figure 1C and D). Anther
dehiscence starts with the opening of flower and being
temporally separated, dehiscence covers the day of the anthesis when the stigma is also receptive. In some flowers,
successive and sequential anther dehiscence also occurs. Out
of the 88 flowers studied, alternate dehiscence was observed
in 74 flowers and in 8 flowers anthers dehisce successively.
Each anther has two lobes. Each lobe is further divided
into a large pollen sac (LPS) and small pollen sac (SPS)
(figure 1E– G). The stomium extends longitudinally between
the two pollen sacs and along the basal and dorsal margins of
large pollen sac.
In the bud condition, stamens are closely adpressed to the
pistil (figure 1B). As the flower opens, stamens move away
from the stigma. In a fully opened flower, stamens are placed
against the corolla lobes, one each. One lobe of the dehiscing
anther starts moving away from the filament and becomes
almost perpendicular to the long axis of the stamen
(figure 1H–J). This is achieved in 10–15 min after the formation of dehiscence slit. It then turns inwards and upwards
exposing the inner pollen-laden surface to the stigma assuming
an angle of 180° vis-à-vis the axis. This is accomplished in 3–5
min. Finally, it curves towards the tip of connective
(figure 1K– M). At this stage anther lobe is at level with the
stigma but apart from the stigmatic surface (3 mm).
Pollen deposition on the stigmatic surface requires the
intervention of insects (figure 1N). When the proboscis of
the nectar foraging insect touches the base of the stamen with
the dehiscing anther, the staminal portion with the dehiscing
anther gets a sudden push towards the stigma, thus depositing huge amount of pollen at the stigmatic surface (figure 1O
J. Biosci. 41(1), March 2016
number of pollen produced by a single anther and multiplying it
by total number of anthers per flower and dividing the product
by total number of ovules produced per flower as
and P). The anther remains stuck to the stigma for about half
a minute and slowly and gradually stamen brings it away
from the stigma, taking between 10–15 min.
This was experimentally tested by touching the base of
the staminal filament with the needle tip and it was observed
that the stamen with the dehiscing anther lobes gets pushed
towards the stigma instantaneously, thus affecting the pollen
deposition on the stigma. The stigma is wet and pollen grains
are sticky and occur in clumps. In a single contact, stigma
receives about 280–412 pollen (X̄=360). Wind pollination
experiments did not give any indication of pollen dispersal
through wind, whereas the pollen load carried by pollinators
varied between 25 and 74 pollen.
Many times, the insect inserts its proboscis through the
gap between the petals laterally at the base to forage the
nectar, as the petals are obovate, broader distally and
narrower proximally with respect to their attachment point.
A mere contact is enough to tilt the staminal filaments, thus
resulting into the movement of dehisced anthers towards the
receptive stigmatic surface. This way pollination is
accomplished.
3.2
Pollen–ovule ratio and reproductive success
The pollen–ovule ratio for the species stands at 1993:1. The
results of different pollination experiments are shown in the
table 1. The stigma becomes receptive in the bud condition
but pollen deposition onto the stigma is brought about only
after the visit of insects, as revealed by unassisted selfing
experiments. There was no pollen load on the stigma of
bagged flowers. In the case of open pollinated flowers,
pollen load on the stigma varied from 68 to 884 pollen and
the percent fruit set was 71.92.
4.
Discussion
This mechanism of anther dehiscence is attributed to the size
difference between the two pollen sacs of each pollen lobe
(Endress and Hufford 1989). The insect-induced movements
of the male reproductive part (stamen) are also reported from
Berberis vulgaris (Lechowski and Białczyk 1992) Opuntia
brunneogemmia, O. viridibura and O. spinosissima
(Schlindwein and Wittmann 1997a; Negrón-Ortiz 1998)
Male function for ensuring pollination and reproductive success
23
LPS
A
B
C
SPS
SPS
F
K
H
L
I
J
M
N
Pl
Pln
St
E
Slit
LPS
G
D
Al
O
P
Pl
Q
R
Figure 1. (A) Position of stamens in the bud condition; (B) a fully opened flower; (C–D) anther dehiscence on alternate stamens; (E–F) back and
frontal view of anther respectively; (G) T.S. of anther showing unequal pollen sacs of an anther lobe; (H–I) dehiscence slit as it appears on the back
and frontal surface of anther; (J) large pollen sac (LPS) as separated from the filament; (K–M) LPS as moved upwards exposing the pollen to the
stigmatic surface; (N) pollinators visiting the flowers; (O–P) dehisced anther in contact with the stigma and Pollen deposition on the stigma; (Q)
papillae at the base of staminal filament (400×); (R) T.S. of the filament showing papillae (1000×) (SPS, small pollen sac; LPS, large pollen sac; St,
stigma; Al, anther lobe; Pln, pollen; Pl, papillae).
Table 1. Reproductive output under natural and different experimental conditions
S. No.
Treatment
No. of flowers under observation
No. of fruits formed
Percent fruit set
1
2
3
4
Open pollination
Manual self-pollination
Manual cross pollination
Unassisted selfing
228
144
158
796
164
108
114
0
71.92
75
72.15
0
J. Biosci.41(1), March 2016
Supriya Sharma and Susheel Verma
24
and Cajophora arechavaletae (Schlindwein and Wittmann
1997b), but the adaptive significance varies depending upon
the floral architecture. In B. vulgaris, stamens are longer than
the pistil and when touched upon by the visiting insects,
anthers make contact with the body of pollinators. This leads
to pollen deposition on body parts of pollinators. It thus
promotes outcrossing. In Berberis lycium, the dehisced anther lies at level with the stigma and makes contact with the
stigma when touched at the base by the pollinator. Thus,
insect-induced staminal movement in this species ensures
self-pollination. It is also supported by the pollination experiments and pollinator behavior.
As reported by Cota-sanchez et al. (2013), the sensitive
stamens do possess some special morphological and anatomical traits. One such character is the presence of papillae on
the sensitive stamens or on the sensitive part of the stamen.
B. lycium also possess papillae at the base of the staminal
filament (figure 1Q and R) as they might be playing some
role in its sensitivity to touch.
High pollen–ovule ratio indicates xenogamy as the preferred breeding system (Cruden 1977). Herein, P/O ratio
does not bear any correlation with the type of breeding
system as revealed by the results of manual self- and manual
cross-pollination experiments. Fruit set in manual crosspollination is slightly lower than that in manual selfpollination (table 1). Also, the polleniferous plants have
higher P/O ratios than nectariferous plants (Lopez et al.
1998), which is also not the case with this species because
of very little presence of pollen on the pollinators’ bodies. As
such, this species exhibits autogamy in spite of having bright
coloured fragrant flowers with nectar, contrivances promoting cross-pollination. Moreover, the mechanism of anther
dehiscence, presentability of pollen to the receptive stigma,
alternate dehiscence of anthers and role of insects point
towards extension of male function to ensure pollination
and reproductive success in B. lycium. Also, the stigma is
wet and receptive at its periphery, where the dehisced anthers make contact, which shows synchrony between male
and female functions in this species.
Acknowledgements
Thanks are due to the Director, Centre for Biodiversity
Studies, BGSBU, Rajouri, for providing necessary facilities
to carry out this work. DBT is also acknowledged for providing funds to undertake research on reproductive biology.
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MS received 01 September 2015; accepted 23 November 2015
Corresponding editor: MAN MOHAN JOHRI
J. Biosci.41(1), March 2016