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Cytological Changes in Trigonella
Foenum-graecum (l.) under the Cadmium Stress.
Kailash P. Patel and Kalpesh M. Patel Veer Narmad South Gujarat University, Surat, Gujarat, India- 395007.
Email: {kailash.patel, incredibleever}@yahoo.com
Abstract—The experiment was carried out to the study of
cytological change in Trigonella foenum-graecum under the
cadmium stress. Different kind of chromosomal
abnormalities like chromosome fragment, ring chromosome,
micronuclei, vagrant chromosome, C-metaphase etc. were
observed during cytological studies. The mitotic index was
frequently reduced and percentage of aberrant cells increase
due to the concentrations of cadmium was increase and
finally plant growth reduced.
Index Terms—Abnormalities, cadmium, chromosome,
mitotic index, micronuclei.
I. INTRODUCTION
Cadmium (Cd) is released into the environment by
human activities such as phosphate fertilizers, disposal of
household, municipal and industrial wastes. These sources
may cause enhanced soil and hence crop cadmium levels,
which may lead to increases in dietary cadmium exposure.
Cadmium is a particularly dangerous pollutant due to its
high toxicity and great solubility in water [1] and [2].
Exposures to high cadmium concentration have been
found to be carcinogenic, mutagenic and teratogenic for a
large number of animal species [3]. Characteristically
inhibits root growth and cell division in plants such as
onion, tradescantia, Vicia faba [4], Nicotiana tabacum [5]
and Allium sativum [6]. Numerous experimental studies
have shown the genotoxicity of cadmium salts [5] and [7].
The purpose of the present investigation was to evaluate
the influence of cadmium concentrations on somatic cells
of Trigonella foenum-graecum, since the most pronounced
effect of heavy metals on plant development is growth
inhibition, which is inseparably connected with cell
division.
Fenugreek (Trigonella foenum-graecum L.) is
undoubtedly one of the oldest cultivated medicinal plants.
It is an erect annual herb native to Southern Europe and
Asia, belonging to the family Leguminosae. Over 80% of
the total world's production of this seed is contributed by
India, one of the major producers and exporters of
fenugreek legume in the world. Fenugreek seeds can be a
good supplement to cereals because of its high protein
(25%), lysine (5.7 g / 16 g N), soluble (20%) and insoluble
Manuscript received October 16, 2012; revised Jan 28, 2013.
(28%) dietary fiber besides being rich in calcium, iron and
beta-carotene [8].
The climate of India being arid to semiarid necessitates
artificial irrigation for successful agriculture. There is very
less canal water available, which does not fulfill the need
for rising crops. Therefore, farmers near the cities use
industrial effluents for raising crops. Most of the sewage
wastewater contains heavy metal ions like Chromium,
Cadmium, Nickel, Mercury and Iron. Their concentrations
are variable in time and space. Therefore, soil near the
factories and cities are being contaminated the regular use
of effluent water for raising crops.
Keeping in view the above mentioned aspects of
industrial set up in India and its impact in creating stressful
environments for agriculture through industrial effluents
and the importance of Trigonella foenum-graecum as
nutritionally important crop, experiments were conducted
to evaluate the effects of cadmium toxicity on this crop.
II. MATERIALS AND METHODS
The germination was carried out in petri dishes. Seeds
were surface sterilized with H2O2 for the prevention of
surface fungal/bacterial contamination. Different ppm
solutions were prepared in pure distilled water in
laboratory by using Cd (NO3)2 (cadmium nitrate) pure
distilled water was used as control for the study. Ten seeds
were placed on cotton in each petri dish and 40 ml solution
of each concentration was supplied once for seed
germination. Distilled water was applied every alternate
day after this treatment. The petri dishes were monitored
daily for fungal and other inspections. The cytological
study was carried out control, 1ppm, 3ppm and 5ppm of
cadmium concentrations.
The staining procedure followed was that of [9]. The
staining with aceto orcine as well as 1:1 aceto orcine and
aceto carmine was found suitable for present study.
Mitotic index was calculated by observing the slides of
root tip of control and treated plants at 3rd, 5th and 7th days
of growth. Total 100 cells were observed on slide. The cell
in the stage of prophase, metaphase, anaphase and
telophase were counted. The mitotic index and percentage
of aberrant cells were calculated using following
formulae;
.
% 100.
No of dividing cellsMitotic index
Total no of cells studies (1)
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Journal of Life Sciences and Technologies Vol. 1, No. 1, March 2013
©2013 Engineering and Technology Publishing doi: 10.12720/jolst.1.1.10-13
.100
.
No of aberrant cellsPercentage of aberrant cells
No of dividing cells (2)
Photomicrographs were taken on Carl-Zesis
photomicroscope with planophotochromatic objectives
using Kodak 100 ASA-35mm color film. Daylight, yellow
or green filter were used.
III. RESULTS
Trigonella foenum-graecum has 2n=16 number of
chromosomes. When the roots of Trigonella
foenum-graecum were treated with different concentration
of cadmium, the cells exhibited various abnormalities
showing its toxic effect on mitotic divisions. A number of
cytological abnormalities viz., chromosome fragments
(Fig.1), micronuclei (Fig. 1), vagrant chromosome (Fig. 1)
and ring chromosome etc. observed in the treated plants.
There was ‗stickiness of chromosomes‘ in the metaphase
some cells also seen with temporary chromosome bridge.
Difference and decline of mitotic index, compared to
controls were noticed in the cells of roots of treated plants.
The mitotic index and percentage of aberrant cells in
control and cadmium treated plants on 3rd, 5th and 7th day
of growth in main roots as well as in side roots (Table I,
Table II and Table III). At 3rd day mitotic index was
reduced in all treatment, compared to control. The
maximum reduction was observed in 5 ppm. At 7th day it
was almost nil in 1 ppm and 3 ppm treatment. The
percentage of aberrant cells was increased with increase in
the cadmium concentration.
Figure 1. Different kind of chromosomal abnormalities.
TABLE I. EFFECTS OF CADMIUM ON ROOT TIP CELLS OF TRIGONELLA FOENUM-GRAECUM ON 3RD DAY OF GROWTH.
Concentration of cadmium
Control 1 ppm 3 ppm 5 ppm
Main root Side root Main root Side root Main root Side root Main root Side root
Mitotic index
(%)
28.66
2.16 ND
15.33
1.25 ND
10.66
0.86 ND
8.33
0.56 ND
Aberrant
cell (%)
4.64
0.46 ND
26.09
2.18 ND
40.61
3.26 ND
48.02
4.14 ND
TABLE II. EFFECTS OF CADMIUM ON ROOT TIP CELLS OF TRIGONELLA FOENUM-GRAECUM ON 5TH DAY OF GROWTH.
Concentration of cadmium
Control 1 ppm 3 ppm 5 ppm
Main root Side root Main root Side root Main root Side root Main root Side root
Mitotic index
(%) 27.33 1.48 ND 8.66 0.32 ND 4.33 0.28 ND NS ND
Aberrant cell
(%) 4.86 0.60 ND 46.18 3.29 ND 61.43 4.43 ND NS ND
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Journal of Life Sciences and Technologies Vol. 1, No. 1, March 2013
TABLE III. EFFECTS OF CADMIUM ON ROOT TIP CELLS OF TRIGONELLA FOENUM-GARECUM ON 7TH DAY OF GROWTH.
Concentration of cadmium
Control 1 ppm 3 ppm 5 ppm
Main root Side root Main root Side root Main root Side root Main root Side root
Mitotic index (%) 25.66 1.33 21.66 1.12 7.66 0.36 15.66 1.06 2.33 0.18 10.33 0.83 NS ND
Aberrant
cell (%) 2.57 0.42 1.52 0.28 52.21 4.56 27.65 2.68 57.08 4.88 35.43 3.59 NS ND
All the values are means S.D.
ND- Not developed during study period.
NS- Not studied due to bursting and decay.
IV. DISCUSSION
From these investigations it was revealed that cadmium
was mitodepressive and induced various types of
chromosomal aberrations in root tip cells of Trigonella
foenum-graecum. Different concentration of cadmium
solution induced different types of chromosomal
aberrations at various stages of cell division. Ring
formation, C-metaphase, vagrant chromosome,
micronuclei and chromosomal fragment were common
aberrations during cells division. Mitotic index was
decreased with the increase in concentration of cadmium.
Effect of heavy metal on chromosomal aberration and
mitotic index was earlier studies by [10]-[16]. Reduction
in mitotic index by the treatment of copper chloride on the
root tips cells of Helianthus annus was reported by [17].
Presence of more than one nucleus in proliferating
following heavy metal treatment is a structural
abnormality [18]. In the present work the micronuclei
were observed in Trigonella foenum-graecum due to the
treatment of cadmium. These micronuclei arose from
acentric chromatid or chromosome fragments induced by
heavy metal cadmium. Evans et al. [19] studied the
production of micronuclei after irradiating the Vicia faba
roots. According to them all chromatid breaks,
chromosome breaks, isochromatid breaks, asymmetrical
exchanges and incomplete symmetrical exchanges would
give rise to acentric fragments at mitosis. At the later stage
of mitosis these fragments are frequently excluded from
the two daughter nuclei and in the following interphase
they appear as micronuclei in one or both the daughter
cells. The micronucleus test has been suggested by [20] as
an in vitro cytogenetic screening procedure for the
detection of freshly induced structural chromosomal
aberrations and for revealing chromosome loss due to
partial impairment of spindle apparatus. He had suggested
that micronuclei originate from chromatin lagging at
anaphase. In the course of telophase this material included
into one or the other daughter cell where it can either fuse
with the other daughter cell where it can either fuse with
the main nucleus or from one or several secondary nuclei.
Another interesting feature noticed after treatment with
cadmium was strong C-mitotic effect in the investigated
plants. The normal mitotic stages were modified in to
C-metaphase due to inactivation of spindle followed by
random scattering of chromosomes. Ring formation was
very rarely reported abnormalities in these studies. Younis
et al. [21] reported it in Vicia faba by the treatment of
nuvacron.
In Allium cepa vagrant was also noticed due to the
treatment of an herbicide avenoxan by [22]. The
abnormalities vagrant was results of impaired spindle
function [18]. Laggards may produce micronuclei if they
fail to reach the poles in time to be included in the main
nucleus [23].
In conclusion, as has been stated above, cadmium has
harmful effects on the root tip cells of Trigonella
foenum-graecum. In addition to these findings, the
increase in soil and water pollution can lead to certain
irreversible cytogenetic effects in plants and even in
higher organisms and reduced growth.
ACKNOWLEDGEMENT
We are acknowledges the University Grants
Commission, New Delhi, for granting minor research
project [F.No. 32-425-2006(SR)].
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13
Dr. Kailash P. Patel was born in 1974 has done
his M.Sc. (Botany) at VNSG University, Gujarat,
India in 1997. She completed his B.Ed (Science
–Math) from same university in 1998. She also got
Ph.D. in 2004 from VNSG University, Gujarat,
India.
Presently she is working as a Assistant
Professor in Botany department of VNSG
University, Gujarat, India. She has published
about 12 research papers in various field like plant science and
environmental science, also she has published on book titled ―Effect of
Cd & Zn on seed germination and growth of Cajanus cajan & Trigonella
foenum and their removal by Various method‖ (LAP Lambert Academic
Publishing GmbH & Co. KG, Germany, 2011). Also 3 research students
have completed their M.Phil and 1 research student has completed his
Ph. D under her guidance.
Dr. Kalpesh Patel was born in 1983 has done his
M.Sc. (Botany) at VNSG University, Gujarat,
India in 2005. He completed his M. Phil from same
university in 2009. He also got Ph.D. in 2012 from
VNSG University, Gujarat, India.
Presently he is working as a Lecturer
In Botany department of VNSG University,
Gujarat, India. He has published about 6 research
papers in various field like plant science and
environmental science, he is also a co-author of a book titled ―Effect of
Cd & Zn on seed germination and growth of Cajanus cajan & Trigonella
foenum and their removal by Various method‖ (LAP Lambert Academic
Publishing GmbH & Co. KG, Germany, 2011)
Author‘s formal
photo
Author‘s formal
photo
Journal of Life Sciences and Technologies Vol. 1, No. 1, March 2013