Download Chromosome banding studies in an Indian mullet: Evidence

Indian Journal of Experimental Biology
Vol. 38, May 2000, pp. 467-470
Chromosome banding studies in an Indian mullet: Evidence of structural
rearrangements from NOR locations
J Chakrabarti & A R Khuda-Bukhsh*
Department of Zoology, University of Kalyani, Kalyani, Nadia-741 235, W.B.
Received 22 November 1999; revised 24 January 2000
C- , G- and NOR bands have been studied in the female sex of Rhinomugil corsula. (Mugilidae, Pisces) by deploying
the conventional methodologies with suitable modifications of minor nature. The diploid metaphase complements contained
48 acrocentric chromosomes. The localization of C-band heterochromatin was found to be mostly at or near the centromeric
regions of the acrocentric chromosomes. The G-type bands were not so well defined, but some of the G-banded
chromosomes also contained C-bands. Interestingly, silver-positive NORs were found at the telomeric ends of five
acrocentric chromosomes, including one homologous pair having NORs in both chromatids, while one chromosome showed
NORs in both of its chromatids and the other two had only one NOR localized at one of its chromatids. This would suggest
th at one homologue of the second pair of NOR-bearing chromosomes possibly underwent a chromatid exchange with a nonNOR bearing chromosome. This is quite a unique situation not reported earlier in any species of fish. , though some other
form of NOR-polymorphism /heteromorphism has rarely been reported. Therefore, further exploration in natural populations
of this species to examine the other sex and to verify if there also exists other chromosomally polymorphi c races (in respect
of NOR-polymorphism) of this species, would be rewarding.
Chromosome banding studies in fish, particularly the
C- and G- bands, have proved to be relatively difficult
in fish because of the small size and large number of
chromosomes in most of them. In course of our
chromosome banding studies on in vivo metaphase
chromosomes of fish I-S an interesting situation was
encountered in an Indian mullet, Rhinomugil corsula
(Fam: Mugilidae) where structural rearrangements
between chromosome pairs could be inferred from
their NOR locations .
Materials and Methods
In an effort to obtain adequate number of good
metaphase spreads and to deterrrline if a particular
concentration and dose of colchicine treatment could
be ascribed as optimal, different specimens were
injected intramuscularly with 0.03, 0.05, 0. 1, 0.2, 0.3
and 0.4% colc hi c ine solution @ I mill 00 g body
weight separately and specimens injected with each of
these doses were sacrificed at 3, 4, 5, 6, 8 and 12 hr,
respectively, after injection. Although over 50 living
specimens of both sexes of R.. corsula of varying
weights and sizes collected from brackish water
cisterns of Central Institute of Brackishwater
Aquaculture, Kakdwip, were used for the present
study, suitable metaphase spreads could be obtained
*Correspondent auth or
only in one female specimen injected with 0.3 %
colchicine and sacrificed at 3 hr, permitting us to
induce C-, G- and NOR bands. However, some other
specimens treated with the same or other dose and
concentration did occasionally yield some fairly good
plates but those were not found suitable for banding
studies, because only very good plates give
reproducible results. But since the unique location of
NORs in this specimen apparently showing no
morphological distinction from the other specimens
used, clearly demonstrated structural rearrangement
between two chromosomes of sirrlilar size, one NOR
bearing and one non NOR bearing, it was felt that thi s
information might be cytologically interesting and
useful to other researchers.
For C- and G- band studies the method of Sumner
·
·
. 5, 6, 10, II was use d .
mo d 1'f'tcattons
wtt h mmor
9
For NOR location the one-step silver nitrate
method of Howell and Black
12
was followed.
Results
The typical diploid Giemsa stained metaphase
spreads (Fig 1) contained 48 rod like chromosomes
alignable into 24 homomorphic pairs (Fig 2). The Cbanded metaphase spreads (Fig 3) showed the
distribution of constitutive heterochromatin mostl y at
or near the centromeres excepting a few pairs which
INDIAN 1 EXP BIOL, MAY 2000
468
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Figs 1-6 - Showing Giemsa stai ned so matic metaphase co mpl ement (Fig. I) of female Rhinomugil corsula; karyotype (Fig. 2) prepared
from Fig. I ; C-banded metaphase complement (Fig. 3); karyotype (Fig. 4) prepared from Fig. 3; G-banded metaphase co mplement (Fig.
5); karyotype (Fig. 6) prepared from Fig. 5. Arrows indicate lateral asymmetry in stri ations. Bar= I 0 J.lm.
CHAKRABARTI & KHUDA-BUKHSH : CHROMOSOME BANDING STUDIES IN AN INDI AN MULLET
8 & I 0), however, it was revealed th at one pair was
perfectly homologous hav ing the ir NOR located at
both chromatids in the ir telomeric ends (chromosome
no. 2). Interestingly enough while both chromatid
ends of chromosome pair no. 3 contained NORs in
one homologue, the other homologue (chosen by its
similarity in size) did not conta in any NOR at either
chromatid. On the other hand , the chromosome pair
arranged at no. 4 in the karyotype comprised
homologues, both of which had only one NOR
located at one chromatid only at its terminal end .
showed blocks of heterochromatin either at one end
(chromosome no . 3, chromosome no . 5, chromosome
no. 6, chromosome no. 9, chromosome no . 17,
chromosome no. 23 and chromosome no. 24) or both
ends of chromatids (chromosome no . 4) or whole
length of chromosome (chromosome no. 2) (Fig 4).
Interestingly, a rather unequal di sposition of
heterochromatin was noted between homologues of
pair no . 3 and 4 (Fig 4).
The G-banded metaph ase spreads (Fig 5) showed
lateral stri ation s of dark and light bands which when
karyotyped (Fig 6) also showed a little asymmetry in
banding pattern in respect of a few individual pairs
(chromosome
no. 3, chromosome
no.
15,
chromosome no. 19, chromosome no. 20), some of
which might be due to unfavourable deposition , if not
inherent in the material. A few chromosome pairs also
showed C-bands (chromosome no. I , chromosome no.
6, chromosome no. 13 to 16, chromosome no. 22).
The sil ver stained metaphase preparations (Figs 7
& 9) showed five chromosomes showing NOR
positi ve regions. From the respective karyotypes (Figs
Discussion
The diploid karyotype of 2n=48 rods and n=24 had
13
earlier been reported by Khuda-Bukhsh and Manna
in Rhinomugil corsula . The present study would
confirm the diploid number of 48 chromosomes in
thi s species. A careful analysis of the C-banded
chromosomes revealed at least one pair of
chromosomes which were C-heteroc hromatin positive
for the whole length (c hromoso me no. 2, Fig-4) and
another pair which had the greater part (chromosome
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Figs 7-10- Showing silver stai ned somatic metaph ase complements (Figs. 7 and 9) of fema le Rhinomugil corsula and karyotypes (Figs.
8 and I 0), prepared fro m Fig. 7 and Fig. 9, respectively. NOR-bearing chromosomes have been indicated by "arrows" and und erlined in
karyotypes. (==?=NOR in both chro m atid s,~ = NOR in single chromatid), Bar= I 0 )lm.
INDIAN J EXP BIOL, MAY 2000
470
no . 4, Fig-4) of the chromosome localizing Cheterochromatin . Interestingly, two other pairs tended
to show unequal distribution of heterochromatin
(chromosome nos . 3 and 5, Fig-4) . Therefore, it
appeared that the C-heterochromatin possibly
incftided or was confluent with the NOR bearing
zones. However, in our earlier study on the
localization of C-band heterochromatin in the male
sex of this species collected from Haldia, West
Bengal' the C-bands were apparently all localized at
or near the centromeres. But since in our present
study male specimens did not yield suitable plates for
banding, the C-band distribution could not be
compared between the sexes of this population of R.
corsula.
Lateral striations of the G-banded chromosomes
could not throw any additional light possibly because
of the smaller size and rather unsuitable resolution of
the banded structure, characteristic of the fish in
genera1
5 14
'
or in the absence of well defined "isocore"
. f ts
' h 15 .
structure m
However, what makes the present study interesting is
the location of NORs which clearly suggests that there
had been a chromatid exchange occurring between one
homologue of an NOR bearing chromosome and a nonNOR bearing chromosome. Incidentally, Khuda-Bukhsh
and Chakrabarti 16 recently reported the association of
Ag-NORs with sex chromosomes in another brackish
water fish, Scatophagus argus showing morphologically
distinguishable
(heteromorphic)
sex
elements.
Therefore, it was not conclusive if this unique NOR
location in R. corsula has anything to do with the sex
chromosome
mechani sm,
particularly
because
difference in NOR numbers and locations between sexes
has also been reported in case of a mosquito fish,
Aplocheilus panchax7. Further, because R. corsula can
thrive well in both brackish and fresh waters, the
unorthodox NOR location in thi s species may have some
added significance.
Acknowledgement
Grateful
acknowledgements
are
made
to
Department of Environment and Forests, Government
of India, New Delhi, for their financial support.
Authors are also thankful to the staff of Central
Institute of Brackish-water Aquaculture, Kakdwip
Research Centre, Kakdwip, West Bengal, India, for
their help in collection of the specimens.
References
I Nayak K & Khuda-Bukhsh A R, Chromosome co mpl ements
in Mugil corsula (Mugilidae, Pisces) with reference to
somatic C-band localization. Proc Na~ Acad Sci Letters, 14
(1991) 195.
2 Khuda- Bukhsh A R & Tiwary S, Locali zation of Nucl eolar
Organizer Regions (N ORs) in the met a_ hase chromosomes of
the Indian climbing perch, Anabas testudineus. Proc Zoo/ Soc,
44 (1991 ) I 0 I.
3 Khuda-B ukhsh A R & Tiwary S, Localization of Nucleolar
Organizer Regions (NORs) in the chromosomes of nine
species of teleosts (Pisces) from India. Systematics and
Evolution of lndoPacijic Fishes: Proc 4th lndoPacijic Fish
Conf. Bangkok, 1994,22.
4 Khuda- Bukhsh A R & Tiwary S , NOR locations in metaphase
chromosomes of six species of teleosts. Nucleus, 39 ( 1996)
128.
5 Khud a-Bukhsh A R & Chakrabarti ", Indu ction of serial
bands akin to G-type on metaphase chromosomes of two
species of fish , Rita rita (Bagridae) a.nd Hypophthalmicthys
molitrix (Cyprin id ae). Indian J Exp Bioi, 34 ( 1996) 1271.
6 Khuda-Bukhsh A R & Chakrabarti C, D istribution of C-band
heterochromatin in metaphase c hromoso mes of the sil ver
carp , Hypophthalmichthys molitrix (C yprinidae, Pisces). Proc
Zoo/ Soc, 52 ( 1999) I .
7 Khuda-Bukhsh A R & Datt a S , Sex-specific differences in
NOR-location on metaphase chromoso mes of the mosquito
tish, Aplocheilus panchax (Cyprinod•)n tidae). Indian J Exp
Bioi, 35 (1997) II II.
8 Mallick P, Chakrabarti J & Khuda-Bukhsh A R , Ag- NOR
locations on metaphase chromosomes of two species ol
estuarine fi shes , Lates calcarifer (Fam: Centropomidae) and
Etroplus suratensis (Fam: Cichlidae). Nucleus, 41 , ( 1998) 43 .
9 Sumner A T, A simpl e tec hn iqu e for d e mon s tra tin ~
centromeric heterochromatin . Exp Cel! Res, 75 ( 1972) 304.
I 0 Datta S & Khud a-Buk hsh A R , Unorthodox distribution of C.
band heterochromatin in mosquito fi sh, Aplocheilus pancha;
with female heterogamety (Fam: Cyp rinodontidae). Uttw
Pradesh J Zoo/, 18 ( 1998) 165.
II Khuda-Bukhsh A R & Datta S, Induc tion of C- and G-typ(
bands on in vivo metaphase chromosomes of three species o
larv i vorou s fi shes. Uttarpradesh J Zoo/, 19 ( 1999) 143.
12 Howell W M & Blac k D A, Controlled silver staining o
nucleolus organ izer regions with a protective colloida
developer: a one-step method. Experientia, 36 ( 1980) I 014.
13 Khuda- Bukhsh A R & Manna G K, Chromosome analysis ir
two species of Indi an mullets (Pisces). Indian Biologist, l
( 1976) 23 .
14 Gold J R , Li Y C , Shipley N S ·& Powers P K, lmprovec
methods for working with fish chromoso mes with a review o
metaphase chromosome banding. J Fish Bioi, 37 ( 1990) 563 .
IS Sumner A T , in Chromosome Banding, (Unwin Hyman Ltd
London , UK) 1990 I .
16 Khud a-Bukhsh A R & Chakraban i 1, Indi cation of se:
chromosome pair bearing Ag-NORs in a brackish water fi sh
Scatophagus argus (Fam: Scatop hagi dae) showing mall
heterogamety. Indian J Exp Bioi, 37 ( 1999) 793 .