Cytotogia
Irradiation
Studies
on
the
II.
Karyology
Ultraviolet
Y. S. R. K. Sarma Laboratory
Received
October
of
42: 291-295,
1977
Charophyta
light
and
S. B. Singh
of Algal Cytology and Cytogenetics, Department of Botany, Banaras Hindu University, Varanasi-221005, India
18, 1975
The effects of ultraviolet light on green algae have been studied by many work ers interested in different fields but the studies of UV particularly in relation to algal karyology are very few. Cytological studies of UV irradiated chromosomes have been carried out in Prorocentrum micans (Dinophyceae) by Dodge (1964). Chromo some and chromatid exchanges giving rise to anaphase bridges were reported by him. Other UV studies made on Oedogonium cardiacum by Parker and Harsley (1972) and in Chlamydomonas by Davies (1965). Kimball (1966) pertained mainly to radiosensitivity studies in relation to different stages of mitotic cycle. But so far as the authors are aware, the present work is the first study to demonstrate that UV is capable of inducing chromosomal aberrations in charophytes. Material Two and
species
one
mental
in
the
were
same
carefully
water
which
were
charophyte
taxa in
were
were a mixture
terials
were
nique
(Godward
the
ergs/mm2),
24hr
at of
cytologically 1948)
to
different
3:1
absolute studied
throughout
sizes
by
effects
24, and
48, glacial
Godward's course
of
of
of 72
and
taxon pond
23cm
(51 time
3mt
96
materials
irradiation
and
of were The
following
acid. alum
ex
(9.18•~
the
hr
ergs/ of
photoreactivation.
acetic iron
study.
filtered
ergs/mm2),
after
labo
each
the
All
the
further
of
increasing
W.)
experi
environment
of
a distance
medium
the of
alcohol
for
suitable
D.
as to
natural
respectively.
eliminate
employing
brought
5ml
R.
used
medium
(6.12•~103
culture
intervals
at
achieved 2 mt
fresh
A)
Br.)
were
in
containing
ergs/mm2)
the
of
(2537
was
W.)
were
culture
light
(A.
D.
growing
antheridia
doses
to
R. taxa
were
petridishes
UV
(12.24•~103
for fixed
in
flagelliformis em
different they
light,
to
transferred
darkness
antheridia in
4mt
N.
Bruz.
(soil-water)
exposed
(3.06•~103 and
and
of
which
placed
in
methods
ex.
plants in
biphasic
variation
Ag. Ag.
ultraviolet and
then
from 1mt ergs/mm2)
placed
water
with
The
posure
selected
to
opaca
.fibrosa
The
selected
mm2/sec).
(N. (C.
pond
irradiation
were
ation
Nitella Chara
transferred
For
103
of
materials.
ratory and
of
species
and
The
acetocarmine
irradi fixed
ma tech
investigation.
Observations
The karyology of the spermatogenous filaments of N. opaca (n=6), N. flagelli formis (n=9) and C. fibrosa (n=14) was critically examined and the cytological
292
Y. S. R. K. Sarma and S. B. Singh
effects produced ities
in dividing
frequently were
as a result cells
observed
stickiness
and
of UV irradiation
were effects
scored commonly
clumping
were recorded.
at metaphase met
Cytologia
with
of chromosomes
and
The mitotic
anaphase
in all the
at metaphase
at
anaphase,
chro
chromosome
metaphase
formation
chromosomes,
most
degrees,
anaphase
erosion,
breakage
The
to varying and
mosome
abnormal
stages.
taxa,
42
and of
anaphase
ring
bridges,
laggards, unequal separation chromosomes, granulation
of and
vacuolization of interphase nu clei and degenerating nuclei. These
effects
were
those
obtained
with
gamma-rays, illustrated
in
similar X-rays
to and
as described and the preceeding
paper.
The most frequently ob served effects in all the three taxa used for experimental work were stickiness and clumping of chromosomes even in materials exposed to 1mt. Besides, the chromosomes appeared faintly stained with fuzzy outlines. With a dose of 1mt treatment, breakage of chromosomes was observed though infrequently at metaphase and anaphase stages in N. opaca and N. flagelliformis but when the dose was increas ed to 2mt, generally one to two fragments in C. fibrosa and upto three fragments in N. opaca and N. flagelliformis were recorded. It was observed that with the Figs. H1-H3. H1,histogram show ing percentage of affected cells in Nitella opaca subjectedto UV light. H2a histogram showing percentage of affectedcells in Nitellaflagellifor missubjectedto UV light. H3ihisto gram showingpercentageof affected cells in Chara fibrosa subjected to UV light.
1977
Irradiation
Studies
on the Karyology
of Charophyta
II
293
administration of higher dose of 3mt and 4mt more than two fragments were noticed in C. fibrosa and more than three in both the species of Nitella. One the whole the percentage of affected cells scored at metaphase and anaphase at lowest dose of 1mt in N. opaca and N. flagelliformis to be 2.00 to 3.11 and 3.85 to 4,75 respectively, whereas no visible abnormalities were scored in C. fibrosa at the same dose level. However, at higher dose of 4mt the percentage of affected cells in creased to 14.44 to 17.78 in N. opaca 16.47 to 18.38, in Ns flagelliformis and 10.00 to 13.79 in C. fibrosa (cf. H1-H3). Apart from the chromosome fragments, anaphase bridges were also noticed. But the nature and shape of anaphase bridges varied from cell to cell at different doses. Among the anaphase bridges, sticky bridges were generally found at low doses of 1mt and 2mt in all the test materials. But at higher doses of 3mt and 4mt most of the bridges were formed of long continuous strands associated with some fragments. In some cases anaphase bridges were constituted by chains of fragments. Formation of ring chromosomes, though infrequent were also recorded in N. opaca and N. flagelliformis both at metaphase and anaphase stages. The other chromosomal abnormalities which were common in all the taxa were laggards, unequal separation of daughter chromosomes to the two poles, and chromosome erosion. Irradiated samples fixed after 96 hr following irradiation with high dose of 4mt showed very few mitotic figures and the frequency of vacuolated and granu lated interphase nuclei was very high. Most of the nuclei in a degenerating con dition. Discussion
The UV induced qualitative effects on the chromosomes of three charophyte taxa studied here are very much similar to the results obtained with X-rays and gamma-rays (see first paper in the series). Stickiness and clumping of chromosomes were found to be more frequent upto 24hr after irradiation at all doses ranging from 1mt to 4mt. Such types of ab normalities were frequently observed in Wada and Izutsu (1961) in Tradescantia cells, Bajer and Bajer (1961) in Haemanthus endosperm which were again con firmed by the observations of Zirkie, Bloom and Urtz (1956). Similar observations also made by Stadler and Uber (1938), Bloom et al. (1955), Gaulden and Perry (1958), Izutsu (1959) and Takeda and Izutsu (1961) pointed out that stickiness was usually "confined to the irradiated pole side of the chromosomes or those lying near the irradiated areas" in Tradescantia. However, in the present study, stickiness was found to be general along the length of the chromosomes. Chromosome breakages were also observed though infrequently in species of N. opaca and N. flagelliformis at a low dose of 1mt whereas generally one to two rarely three fragments were recorded at a dose of 2mt and more than three at higher dose levels. Sticky bridges were more common in comparison to other types of bridges. At high doses of 3 and 4mt anaphase bridges were made up of long con tinuous strands with some fragments. The three taxa viz. N. opaca, N. flagelli
294
Y. S. R. K. Sarma and S. B. Singh
formis
and
ference
C.
in
scored
it
to
light
UV
may
to to
ergs/cm2,
and
dose
of
were
made
of
2•~103
108
ergs/cm2.
appear
at
of
the
more
or
of
of
effects
with
percentage
of
flagelliformis
were
slight
dif
affected more
of
cells sensitive
dose
of
5•~108
the
observations
of
it
seems
the
earlier the
at to
at
on of
UV
dose
uniform
30 of
Tradescantia on
of
Stad
a
reported
a higher
workers effects
in a dose
observations
who
48%
too
25%. to
Similar
Tradescantia
and
that
were
chromosomes
Tradescantia
deficiency.
in
ergs/cm2
of
it increased of
of
Tradescantia
abnormalities
ergs/cm2,
(1957)
0.52•~108
shattering
endosperms
Criag
of fragments
chromosomal
11.6•}1.6%
study,
chromosomes chromosome
reported
cent
and of
tube
per
scored
a dose
less
N.
ergs/mm2
subjecting
the
type
basis
pollen
(1953)
a higher
present
similar the and
2.5•~103
one
Smith
From
basis
of
after
Kirby,
cells
when
of
ergs/mm2,
by
abnormal
the
at
(1942)
opaca
Lovelace
extent
but
Uber
N.
that
a dose
scored. the
less On
42
C. fibrosa.
noticed
be
or
cells.
that to
with
Tradescantia
ler
concluded
(1964)
numerous
more
affected
compard
irradiated
5•~106
showed of
be as
Bailey were
fibrosa
percentage
Cytologia
1.18•~ and
charophyte
on taxa
similar.
Summary The N.
experimental
opaca,
district. to
tative A and less
N.
flagelliformis
The
exposure
12.24•~103
titatively
changes
were
similar
on
were
very
charophyte
radioresponse
similar
to
those
UV
recorded
in
local
to
4mt
each
and
anaphase with
on the
light
under
found
effects
UV
from 1mt
metaphase
chromosomal taxa
to
from effects
at
to
collected
ranged
cytological
much of
subjected
fibrosa,
UV
estimated
assessment
those
C.
of The
qualitatively
comparative
that
and time
ergs/mm2).
and
of
materials
antheridium
ponds
of
(3.06•~103
stages.
X-rays
study
and by
of
Varanasi ergs/mm2
experiment
Tradescentia present
are
were
quan
The
quali
gamma-rays. earlier
revealed
workers more
or
light.
Acknowledgements
of
The
authors
wish
Botany
for
providing
to the second
author
to express
their
laboratory
by the
gratitude
facilities.
authorities
of Banaras
to the The
Head
financial Hindu
of the
Department
assistance
University
provided
is also
grate
fully acknowledged.
Literature
cited
Bajer, A. and Mole-Bajer, J. 1961. UV microbeam irradiation of chromosomes during mitosis in endosperm. Exp. Cell Res. 25: 251-267. Bailey, P. C. 1964. A comparison of X-rays and UV induced aberrations in pollen tube chromo somes of Tradescantia I. Dose curve response to varying moisture conditions. Rad. Bot. 4: 115-119. Bloom, W. and Leider, R. J. 1955. Changes in refractive index of chromosomes by UV or proton microbeam irradiation. Rad. Res. 3: 214-215. Davies, D. R. 1964. Repair mechanism and variations in UV sensitivity within the cell cycle. Mut. Res. 2: 477-486.
1977
Irradiation
Studies on the Karyology
of Charophyta
II
295
Dodge, J. D, 1964. Effects of UV light on the survival and nuclear division of a dinoflagellate . Protoplasma 59: 485-493. Gaulden, M. E. and Perry, R. P. 1958. Influence of the nucleolus on mitosis as revealed by UV microbeam irradiation. Proc. Nat. Acad. Sc. 44: 553-559. Godward, M. B. E. 1948. The iron alum acetocarmine method for algae. Nature (London). 161: 203. Izutsu, K. 1959. Irradiation of parts of single mitotic apparatus in grasshopper spermatocytes with an ultraviolet microbeams. Mie Medical Jour. 9: 15-29. Kimball, R. F. 1966. A repair premutational damage in L. G. Augen Stein, Ronald Mason and Max Zella (Eds). Advances in Radiation Biology, Acad. Press, New York. 2:135-166. Kirby, Smith, J. S. and Criag, D. L. 1957. The induction of chromosome aberrations in Tradescan tia by UV radiation. Genetics 42: 176-187. Lovelace, R. 1954. Chromosome shattering by UV radiation (2650 A). Proc. Nat. Acad. Sc. 40: 1129-1135. Parker, K. E. and Horsley, R. J. 1972. The UV radiosensitivity of Oedogonium cardiacum cells at various stages of the cell cycle. Rad. Bot. 12: 239-248. Sarma, Y. S. R. K. and Singh, S. B. 1977, Irradiation studies on the karyology of Charophyta I. X-Rays and gamma-rays. Cytologia 42: 279-290. Stadler, L. J. and Uber, F. 1938. Genetic effects of UV light on maize. Genetics 23: 171. - and 1942. Comparison of genetic effects of different wavelength of UV light on maize. Genetics 27: 84. Takeda, S. and Izutsu, K. 1961. Effect of UV microbeam irradiation upon cell division. Jap. J. Genetics 36: Supl. 1: 105-114. Wada, Bungo and Izutsu, K. 1961. Effects of UV microbeam irradiations on mitosis studied in Tradescantia cells in vivo. Cytologia 26: 480-491. Zirkle, R. E., Bloom, W. and Uretz, R. B. 1956. Proc. Int. Conf. Peaceful Uses of Atomic Energy. Geneva 1955. 11: 273.