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Cytogenetic Characterization of Thalassophryne maculosa Gunther, 1861 (Pisces: Batrachoididae) from Margarita Island, Venezuela.

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Mauro Nirchio at Universidad Técnica de Machala (Ecuador)
Mauro Nirchio
  • Universidad Técnica de Machala (Ecuador)
Alberto Fenocchio at National University of Misiones
Alberto Fenocchio
Ana Claudia Swarca at Universidade Estadual de Londrina
Ana Claudia Swarca
Ana Lúcia Dias at Universidade Estadual de Londrina
Ana Lúcia Dias
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A cytogenetic analysis by conventional Giemsa staining, C-banding, G-banding, Silver staining, and Fluorescence in situ Hybridization (FISH) was carried out on Thalassophryne maculosa from Margarita Island, Venezuela. T. maculosa exhibits a diploid complement 2n=46, consisting of 12 metacentrics, 6 submetacentrics, 20 subtelocentrics and 8 acrocentrics with an NF of 84. Constitutive heterochromatin was not abundant and was located only in the centromeric region in all chromosomes. A clear G-band pattern that aids in a better chromosome pairing was obtained. Localization of Nucleolus Organiser Regions (NORs) on the short arms of a medium-sized subtelocentric chromosome pair was detected with Ag-staining and was also confirmed by FISH, using probes of 18S rDNA (from fish) and 45S rDNA (plant origin).
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Cytogenetic Characterization of Thalassophryne maculosa Gu¨ nther,
1861 (Pisces: Batrachoididae) from Margarita Island, Venezuela
M. NIRCHIO
1
,A.S.FENOCCHIO
2
,A.C.SWARC¸A
3
,A.L.DIAS
3
,L.GIULIANO−CAETANO
3
,
E. R
ON
1
,J.I.GAVIRIA
1
, AND J. E. PE
´
REZ
4
1
Escuela de Ciencias Aplicadas del Mar. Universidad de Oriente, Isla de Margarita, Venezuela.
Phone: ++58-295-2626003. Corresponding author: nirchio@cantv.net
2
Universidade Federal do Parana´, Departamento de Gene´tica, Centro Polite´cnico, Curitiba, Brazil.
afenocch@fceqyn.unam.edu.ar
3
Universidade Estadual de Londrina, Departamento de Biologia Geral, Caixa Postal 6001, CEP 86051-990,
Londrina, PR, Brasil. szwarcy@sercomtel.com.br
4
Instituto Oceanogra´fico de Venezuela, Universidad de Oriente, Apartado 243, Cumana´, Venezuela. jperez@telcel.net.ve
A
BSTRACT.—A cytogenetic analysis by conventional Giemsa staining, C-banding, G-banding, Silver stain-
ing, and Fluorescence in situ Hybridization (FISH) was carried out on Thalassophryne maculosa from Mar-
garita Island, Venezuela. T. maculosa exhibits a diploid complement 2n = 46, consisting of 12 metacentrics,
6 submetacentrics, 20 subtelocentrics and 8 acrocentrics with an NF of 84. Constitutive heterochromatin was
not abundant and was located only in the centromeric region in all chromosomes. A clear G-band pattern that
aids in a better chromosome pairing was obtained. Localization of Nucleolus Organiser Regions (NORs) on
the short arms of a medium-sized subtelocentric chromosome pair was detected with Ag-staining and was
also confirmed by FISH, using probes of 18S rDNA (from fish) and 45S rDNA (plant origin).
K
EYWORDS.—cytogenetics, marine fishes, chromosome banding, FISH
INTRODUCTION
Thalassophryne maculosa belongs to the
family Batrachoididae (Order Batrachoidi-
formes), a small group known as toadfishes
that contains over 70 species, with 21 gen-
era and three subfamilies (Greenfield et al.
1994; Greenfield 1997; Collette 2001; Froese
and Pauly 2003). T. maculosa is distributed
on the northern coast of South America off
Colombia and Venezuela, Aruba, Curac¸ao,
Margarita, Cubagua, Trinidad, and To-
bago, but not extending into the Antilles
(Collette 1966). It is one of the five recog-
nized species of toadfish reported for Ven-
ezuela and is common along sandy beaches
on the leeward side of Cubagua Island, and
on the southern side of Margarita Island
(Cervigo´n 1993).
Details about the karyotype of toadfishes
have been reported for Porichthys notatus
(Chen 1967; in Gold et al. 1980), Halobatra-
chus didactylus (Palazo´n et al. 2003), Ba-
trachoides pacifici (Nirchio et al. 2001), Am-
phichthys cryptocentrus, Batrachoides manglae,
Thalassophryne maculosa (Nirchio et al. 2002)
and Porichthys porosissimus (Brum et al.
2001). Most of these chromosomal studies
dealt with karyotype determined by Gi-
emsa staining, a conventional method that
allows describing chromosome number
and types. The use of special techniques
has not been extensively applied in this
group. Our objective was to determine con-
stitutive heterochromatin, G-Band pattern
and NOR localization and number of T.
maculosa using silver impregnation and
Fluorescence in situ Hybridization.
M
ATERIALS AND METHODS
We analyzed nine adult specimens (2 fe-
males and 4 males) of T. maculosa caught
near Margarita Island, Venezuela. Voucher
specimens were deposited at the Ichthyol-
ogy Collection of the Escuela de Ciencias
Aplicadas del Mar, Universidad de Ori-
ente. Chromosome preparation was per-
formed according to Nirchio et al. (2002)
Caribbean Journal of Science, Vol. 40, No. 2, 218-222, 2004
Copyright 2004 College of Arts and Sciences
University of Puerto Rico, Mayagu¨ez
218
but exposure time to colchicine was re-
duced from 5 to 3 hours. Conventional
karyotype was visualized by staining the
preparation for 20 minutes with a 10% Gi-
emsa solution in a phosphate buffer, pH
6.88. NOR silver staining was performed
using the method of Howell and Black
(1980). C- and G-banding were performed
using the methods of Sumner (1972) and
Cano et al. (1996), respectively. FISH with
18S rDNA probes from Oreochromis niloti-
cus and 45S from Triticum was performed
following Swarc¸a et al. (2001). At least 10
well-spread mitotic metaphases from each
individual were examined for each cytoge-
netical method applied.
Chromosomes were photographed
using a Nikon COOLPIX® 995 Digital
Camera following the microscopy set-
tings recommended by the manufac-
turer. (http://www.nikonusa.com/pdf/
CoolpixSeriesMtngInstructions.pdf). Im-
ages were stored as *.tif files and digitally
processed with ADOBE PHOTOSHOP
v.7.0. This software was also used for mea-
suring long arm (L), short arm (S) and con-
structing karyograms. Chromosomes were
classified according to the arm ratio criteria
(Levan et al. 1964).
R
ESULTS AND DISCUSSION
The chromosomes (2n = 46) were ar-
ranged in decreasing size and centromere
position and distributed in four types: 12M,
6SM, 20ST, 8A (NF = 84) (Fig. 1). These data
do not agree with Nirchio et al. (2002) who
reported 2n = 46 but 8M, 6SM, 32A (NF =
60). For classifying the karyotype of T.
maculosa, Nirchio et al. (2002) used a con-
ventional caliper (precision = 0.05 mm), to
measure chromosome arm length, whereas
in the present report, these lengths were
obtained with the measure tool (precision =
0.01 mm) of the software used for process-
ing the digitalized images of metaphase
spreads. Additionally, the reduction in ex-
position time to colchicine (see materials
and methods), produced better chromo-
some preparations, morphology identifica-
tion and arms length determination. These
features explain the differences in the com-
position of chromosome types, mainly in
FIG.1. Thalassophryne maculosa: Conventional Giemsa stained karyotype A: male, B: female. In box: Ag-stained
NOR-bearing chromosomes identified by sequential staining.
KARYOLOGY OF THALASSOPHRYNE MACULOSA 219
the number of bi-armed (metacentrics, sub-
telocentrics) elements, and consequently,
the NF values.
The C-bands were present in centromeric
regions in almost all chromosomes; the
band pattern indicated the presence of het-
erochromatic regions restricted to centro
meres (Fig. 2). As far as we know, Porichthys
porosissimus (2n = 44), a toadfish recently
karyotyped by Brum et al. (2001), is the
only species whose chromosomes were also
C-banded and the bands appeared as pale
pericentromeric blocks, except for the two
larger chromosome pairs in which a large
heterochromatic segment along both arms
was present, beside the centromeric region.
This different feature between Talasophryne
and Porichthys suggests that C-bands could
be a good cytotaxonomic marker in the
family Batrachoididae.
An accurate classification of homologous
chromosomes, using only Giemsa staining,
is not always possible because differences
in chromosome size and arm ratios are of-
ten small between adjacent pairs within a
size-graded series. In these cases G-
banding is helpful to improve pairing. A
consistent G-band pattern that allowed
pairing practically all chromosomes (Fig. 3)
was obtained in T. maculosa. This result
suggests the existence of genome compart-
mentalization in this species, but chromo-
some types, arranged in decreasing size or-
der, were difficult to identify. G-banding
has been reported for several fish species
(Gold and Li 1990; Abuı´n et al. 1996; Ber-
tollo et al. 1997), which indicates that fail-
ure to obtain it could be due to technical
problems in the use of banding protocols
rather than to total absence of structural
compartmentalization in the fish genome
as proposed by Saitoh and Laemmli (1994)
and Maistro et al. (1999). As in most tel-
eostean fish (Gold and Amemiya 1987; Vit-
turi et al. 1995), Ag-stained metaphases
from T. maculosa exhibited positive signals
in only one chromosome pair (N° 16) of
medium-sized subtelocentric chromo-
somes; the black dots were located termi-
nally on the short arms of these chromo-
somes (Fig. 1). Palazo´n et al. (2003) also
reported in Halobatrachus didactylus a single
pair of NOR-bearing chromosomes but
submetacentric instead of subtelocentric.
Brum (2001) attempted to describe the
NOR patterns of P. porosissimus but, al-
though no satisfactory results were ob-
tained, they inferred the occurrence of
more than a single NOR-bearing pair on
the base of the presence of one to three
nucleoli per nucleus when the silver nitrate
technique was applied.
FIG.2. Thalassophryne maculosa: C-banded karyotype
M. NIRCHIO ET AL.220
Silver staining is a method, which only
demonstrates the residues of the Ag stain-
able rRNAprotein complex synthesized by
the active NORs in the preceding inter-
phase (Howell and Black 1980), thus, the
Fluorescence in situ Hybridization (FISH)
approach was applied in the chromosomes
of this group of fish, and bright signals
were obtained. FISH applied to study the
NORs using probes of 18S rDNA from fish
(Oreochromis niloticus) and of 45S rDNA
from plants (Triticum) revealed fluorescent
signals in the same regions on the short
arms of the two subtelocentric chromo-
somes identified using silver salts (Fig. 4),
and demonstrates that T. maculosa does not
possess additional NORs. The strongest
signal was obtained when the 45S rDNA
probe was applied probably because the
plant probe is constituted by the entire
gene sequence, a longer probe, whereas the
fish obtained probe (18S rDNA) is only the
sequence of the ribosomal gene.
The present report represents the first cy-
FIG.3. Thalassophryne maculosa: G-banded karyotype
FIG.4. Thalassophryne maculosa: FISH with 18S rDNA from fish (A) and 45S rDNA from a plant (B).
KARYOLOGY OF THALASSOPHRYNE MACULOSA 221
togenetic study that used banding, fluores-
cent and molecular techniques of a marine
Neotropical fish species. It could contribute
to cytotaxonomic and evolutionary studies.
Acknowledgements.—The authors are
grateful for the valuable comments and
suggestions on the manuscript given by Dr.
Angelo Libertini (CNR-ISMAR − Sezione di
Venezia) and Dr. Claudio Oliveira (Univer-
sidade Estadual Paulista, Botucatu, SP, Bra-
zil). This work was supported by Consejo
de Investigacio´n of Universidad de Oriente
and Conselho Nacional de Desenvolvim-
ento Cientı´fico e Tecnolo´gico (CNPq) and
to CAPES for fellow to ACS.
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M. NIRCHIO ET AL.222
... The FN, together with the large number of subtelocentric chromosomes and the conservation of 2n = 46, reflect the role of pericentric inversions in the modeling of the karyotype of the extant Batrachoididae species, which is consistent with the findings of previous studies (Brum et al., 2001;Palazón et al., 2003;Nirchio et al., 2004a, 2004bCosta and Molina, 2009). This considerable chromosomal diversity in comparison with more marine conservative groups of fish (Molina, 2007) makes this genus an excellent model for the understanding of the evolutionary mechanisms underlying the considerable genome diversity found in these vertebrates. ...
... Sousa et al. Estuarine, Coastal and Shelf Science 213 (2018) 253-259 heterochromatin distribution is highly varied, with complex and inconclusive patterns, as in Batrachoides pacifici and Thalassophryne maculosa (Nirchio et al., 2004a(Nirchio et al., , 2004b2004c). In addition, the numerical and morphological diversity of karyotypes are not always accompanied by equivalent differentiation in the heterochromatic regions (Galvao et al., 2011). ...
... In fact, besides their occurrence in all pericentromeric regions, they are also highly accumulated in some pairs, as in the long arms of the 3rd pair (Fig. 2B). Although high heterochromatin content is a shared trait with other cofamiliar species (e.g., Nirchio et al., 2004aNirchio et al., , 2004b2004c), the heterochromatic distribution in B. surinamensis is shown to be quite distinct from B. pacifici, in which the heterochromatin is mainly restricted to pericentromeric regions, as well as from other Batrachoididae species (Fig. 3). Therefore, according to the data available so far, the distribution of the constitutive heterochromatin provides species-specific patterns for B. surinamensis. ...
Cytogenetics description in Batrachoides surinamensis, (Batrachoididae: Batrachoidiformes): What does the estuary have to say?
Article
  • Aug 2018
The Batrachoididae (Batrachoidiformes) is a diverse fish family (84 species) of considerable medical interest, responsible for large numbers of injuries in fishermen and bathers in northern Brazil. Batrachoides surinamensis (the Pacuma toadfish) is the most common Batrachoididae species in the Amazon coastal zone. The capacity of this species to adapt to the dynamic environment of estuaries, its sedentary behavior, and benthic spawning all contribute to the interest in this fish as a model for the study of patterns of chromosomal diversification. The present study investigated the chromosomal features of this species through conventional (Giemsa, C-banding, and Ag-NOR) and molecular (FISH mapping of the 18S and 5S rRNA, and the telomeric sequences) cytogenetic approaches. This is the first description of the karyotype of this species, which has a diploid number (2n) of 46 chromosomes (6m+8sm + 20st+12a) and a fundamental number (FN) of 80. All the chromosomes presented C-banding in the pericentromeric region. The AgNOR/18S rRNA sites were observed exclusively on the short arms of pair 13, whereas multiple 5S rRNA sites were found, on pairs 7 (submetacentric) and 20 (acrocentric). The telomeric probes revealed interstitial telomeric sequences (ITSs) in metacentric pair 3, indicating the occurrence of chromosome fusion. The karyotype of B. surinamensis presents a number of derived karyotypic features, and is differentiated primarily by its pericentric inversions and the apparent occurrence of at least one chromosome fusion event. The collection of additional cytogenetic data on other Batrachoididae species, and other populations of B. surinamensis, will provide further insights into the role of estuarine environments in the chromosomal diversification of this fish group.
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... Cytogenetic studies in the Batrachoididae family are few, and classic cytogenetic techniques have been applied, mainly to determine chromosome number and karyotype in Porichtys notatus (Gold et al. 1980), Batrachoides pacifici (Nirchio et al. 2001), Porichtys porosissimus (Brum et al. 2001), Amphichtys cryptocentrus (Nirchio et al. 2002), Batrachoides manglae (Nirchio et al. 2002), Thalassophryne maculosa (Nirchio et al. 2002(Nirchio et al. , 2004a, Halobatrachus didactylus (Palazón et al. 2003;Merlo et al. 2007), Porichthys plectrodon (Nirchio et al. 2004b) and Halophryne trispinosus (Magtoon and Donsakul 2008). However, studies using more advanced cytogenetic techniques are very scarce. ...
... For example, the technique of fluorescence in situ hybridization (FISH) has been used to localize the major ribosomal genes (18S-5.8S-28S rDNA) in studies of T. maculosa (Nirchio et al. 2004a) and to localize four sequences of repetitive DNA: telomeric, GATA, 5S rDNA and 45S rDNA sequences in H. didactylus (Merlo et al. 2007). ...
... Double-color FISH with digoxigenin-labelled 18S rDNA probe (green, arrow heads) and rhodamine-labelled 5S rDNA probe (red, arrows): a Amphichthys cryptocentreus, b Batrachoides manglae, c Porichthys plectrodon, and d Thalassophryne maculosa as follows: A. cryptocentrus (2n = 46) 4M, 2SM, 40A(Nirchio et al. 2002), B. manglae, (2n = 46) 6M, 6SM, 34A(Nirchio et al. 2002), P. plectrodon (2n = 44) 8M, 10SM, 6ST, 20A(Nirchio et al. 2004b), and. T. maculosa (2n = 46) 12M, 6SM, 20ST, 8A(Nirchio et al. 2004a). ...
Chromosomal mapping of the major and minor ribosomal genes, (GATA)n and U2 snRNA gene by double-colour FISH in species of the Batrachoididae family
Article
Full-text available
  • Jul 2010
  • GENETICA
In the present study dual-colour fluorescence in situ hybridization (FISH) was performed to study the chromosomal distribution of 18S and 5S rDNAs, (GATA)n and 5S rDNA, and U2 snRNA and 18S rDNA in four species of Batrachoididae family: Amphichthys cryptocentrus, Batrachoides manglae, Porichthys plectrodon and Thalassophryne maculosa. The 18S rDNA signals were present in only one pair of chromosomes in all the four Batrachoididae species. The 5S rDNA was mapped on one pair of chromosomes, except in B. manglae, which showed a hybridization signal in two pairs. The two ribosomal genes are located on different chromosome pairs, except in A. cryptocentrus, in which they appear co-located. In all the cases, the (GATA)n probe produced disperse hybridization signals in all four species. The U2 snRNA signals appear very widely scattered in A. cryptocentrus, P. plectrodon, but show a degree of clustering in a specific chromosome pair in B. manglae. In T. maculosa, they are thinly dispersed and strong hybridization signals are observed co-located to the 18S rDNA-bearing chromosomes. Finally, a double-colour FISH with U2 snRNA and 5S rDNA probes was performed in B. manglae, and this showed that these genes were not co-located. These results have been compared with those from another Batrachoididae species, and evolutive processes of these species are discussed.
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... The main difference in the karyotypic macrostructure between T. nattereri and T. maculosa from the Venezuela coast (Nirchio et al., 2004a ) seems to involve the occurrence of pericentric inversions leading to distinct karyotypic formulae. Besides the oceanic distance between the collection sites, both species are isolated by the Amazon River outflow, originating 6 to 10 million years ago (Floeter et al., 2007), and recognized as an important biogeographic barrier for several taxa, causing the disruption between Caribbean and Brazilian fauna (Rocha et al., 2002). ...
... Two centric fusions among four chromosome pairs would be required for the establishment of 2n = 44 karyotypes, thereby representing an apomorphic trait, exclusively found in the genus Porichthys. Contradictory karyotypic formulae have been reported in a single species of Batrachoididae (Palazón et al., 2003; Merlo et al., 2007), and some of them are quite discrepant (Nirchio et al., 2002Nirchio et al., , 2004a). However, the relatively large number of subtelocentric chromosomes in the karyotypes coupled with an imprecise differentiation between subtelocentric and acrocentric chromosomes should account for most of the differences in the number of chromosome arms observed within this family. ...
... The heterochromatic blocks are more conspicuous in the centromeric region of the first and second chromosome pairs (metacentric). Such distribution pattern and amount of heterochromatin is similar to that reported in T. maculosa (Nirchio et al., 2004a). On the other hand, the cytogenetic data described for Thalassophryne differ from the pattern observed in the genus Porichthys (Brum et al., 2001; Nirchio et al., 2004b), i.e., large heterochromatic blocks restricted to some chromosome pairs, besides conspicuous pericentromeric heterochromatic segments in the first and second chromosome pairs. ...
Karyoevolution of the toadfish Thalassophryne nattereri (Batrachoidiformes: Batrachoididae)
Article
  • Sep 2009
  • GENET MOL RES
The Batrachoididae includes some venomous brackish and marine fish found in the Atlantic, Indian and Pacific oceans. This family is composed of 69 species, distributed among 19 genera. Species of the genus Thalassophryne have been reported along the coast of Rio Grande do Norte (Brazil); T. nattereri has been responsible for a large number of human injuries. Little is known about the cytogenetic features of this family. We made a karyotypic characterization of T. nattereri collected from the estuary of the Apodi/Mossoró River, using conventional Giemsa staining, C-banding and silver nitrate-nucleolar organizer region technique. There was a modal diploid value of 2n = 46 chromosomes (8m + 8sm + 24st + 6a; fundamental number = 86). Single ribosomal sites were detected in the terminal region on short arms of a subtelocentric pair (19th). Heterochromatin segments were preferentially located over centromeric regions in some chromosome pairs. Pericentric inversions and Robertsonian rearrangements seem to have played a major role in karyotype evolution within this genus of toadfish.
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... It was one of the first animals to be used as a model for research in outer space (Boyle et al. 2001 ). In spite of this, genetic knowledge of Batrachoididae family is scarce and cytogenetical studies have been limited to the analysis of the karyotypes of some species, such as Porichthys notatus (Gold et al. 1980), Batrachoides pacifici (Nirchio et al. 2001), Amphichthys cryptocentrus, Batrachoides manglae, Thalassophryne maculosa, Porichthys plectrodon (Nirchio et al. 2002Nirchio et al. , 2004a, b), and Porichthys porosissimus (Brum et al. 2001). In H. didactylus, only the karyotype has been described (Gutié rrez et al. 1984) and localization of the nucleolar organizer regions has been obtained by means of silver staining (Ag-NORs) (Palazó n et al. 2003). ...
... The FISH analysis has only been applied to the toadfish Thalassophryne maculosa for location of the ribosomic genes 18S-5.8S-28S (Nirchio et al. 2004a). Sequences (TTAGGG) n , are present in the telomeres of the chromosomes of all vertebrates and their study allows one to establish the presence of chromosomal rearrangements , such as Robertsonian fusions or inversions , which are involved in the evolution of the chromosomes. ...
... FISH localization of the major ribosomal genes in a submetacentric pair of large size is coincident with the Ag-NOR location reported by Palazó n et al. (2003). In other toadfish species, Ag-NORs have been located in centromeric position in an acrocentric chromosome in Porichthys plectrodon (Nirchio et al. 2004b) and in terminal position of a subtelocentric pair, in Thalassophryne maculosa (Nirchio et al. 2004a). The presence of NORs, either in the terminal position or close to the centromeres (but not in the interstitial positions) is considered to be the plesiomorphic character (Vitturi et al. 1995). ...
Chromosomal mapping of the major and minor ribosomal genes, (GATA) n and (TTAGGG)n by one-color and double-color FISH in the toadfish Halobatrachus didactylus (Teleostei: Batrachoididae)
Article
Full-text available
  • Nov 2007
The karyotype of Halobatrachus didactylus presents 46 chromosomes, composed of eight metacentric, 18 submetacentric, four subtelocentric, and 16 acrocentric chromosomes. The results of FISH showed that the major ribosomal genes were located in the terminal position of the short arm of a large submetacentric chromosome. They also showed a high variation in the hybridization signals. The products of amplification of 5S rDNA produced bands of about 420 pb. The PCR labeled products showed hybridization signals in the subcentromeric position of the long arm of a submetacentric chromosome of medium size. Double-color FISH indicated that the two ribosomal families are not co-located since they hybridizated in different chromosomal pairs. Telomeres of all the chromosomes hybridized with the (TTAGGG)n probe. The GATA probe displayed a strong signal in the long arm of a submetacentric chromosome of medium size, in the subcentromeric position. The double-color FISH showed that the microsatellite GATA and the 5S rDNA gene are located in different chromosomal pairs. The majority presence of GATA probes in one pair of chromosomes is unusual and considering its distribution through different taxa it could be due to evolutionary mechanisms of heterochromatine accumulation, leading to the formation of differentiated sex chromosomes.
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... Adicionalmente, los procesos espinosos de los opérculos también son utilizados como órganos defensivos venenosos, pudiendo infligir, con tales estructuras, punciones dolorosas en sus víctimas (Haddad Junior et al. 2003). Thalassophryne maculosa conocido como "pez sapo cano" es una de las cinco especies reconocidas de pez sapo reportadas para Venezuela y es muy común en las playas de las islas de Cubagua y Margarita (Cervigón 1993, Nirchio et al. 2004) y en las costas del norte de Venezuela (Machado-Allison y Rodríguez-Acosta 1997, Rodríguez-Acosta y Reyes-Lugo 1999). ...
ACTIVIDAD BIOLÓGICA IN VITRO Y DOSIS LETAL CINCUENTA DEL VENENO DE Thalassophryne maculosa (PISCES, BATRACHOIDIDAE) IN VITRO BIOLOGICAL ACTIVITY AND FIFTY LETHAL DOSES OF THE VENOM OF Thalassophryne maculosa (PISCES, BATRACHOIDIDAE)
Article
  • Jan 2022
RESUMEN El envenenamiento causado por Thalassopryne maculosa (Pisces, Batrachoididae) en humanos, origina manifestaciones clínicas fundamentalmente inflamatorias, conllevando en ciertas ocasiones a lesiones muy severas, temporalmente incapacitantes, aunque aparentemente no letales. A pesar de ello, muchos de los componentes responsables de tales actividades no han sido estudiados ni identificados con precisión; por lo cual, estudiar algunos de los efectos biológicos in vitro e in vivo del veneno de esta especie marina es considerado necesario desde un punto de vista clínico. El espectro de absorción dejó en evidencia el predominio de componentes proteicos del veneno, mientras que la concentración de proteínas totales en la fracción soluble fue determinada a 280 nm. Veinte ejemplares produjeron 79,41 mg de proteínas usados en los ensayos para la evaluación de los efectos biológicos in vivo y la caracterización in vitro. La actividad biológica in vivo fue ensayada luego de la inyección intraperitoneal del veneno en ratones hembra C57BL/6, calculándose la dosis letal cincuenta (DL50), según el método de Dixon y Mood, en 70,80 (66,95-75,12) mg.kg-1 , para 60 minutos de experimentación, y simultáneamente valorando el curso temporal de toxicidad aguda experimental. La aparición de manifestaciones clínicas de toxicidad fue gradual; iniciando con hiperactividad y taquipnea, seguida de hipoactividad y distensión abdominal para suceder, alrededor de los 30 minutos, periodos de bradipnea/apnea a la cual sobreviene la muerte o supervivencia de los ratones. La actividad proteolítica, fosfolipasa, hemolítica y antibacteriana (sobre Aeromonas hydrophila, A. sobria y Staphylococcus aureus) se evaluó in vitro. El ensayo de actividad gelatinasa, dejó en evidencia la capacidad proteolítica del veneno hasta una dilución 1:64; el efecto hemolítico se observó en ensayos con agar sangre desde cantidades iguales o superiores a los 50 µg.µL-1 , mientras que una ligera actividad fosfolipasa ha sido sugerida con la mayor concentración de veneno utilizada (8,0 mg.mL-1). Se demostró ausencia de actividad antibacteriana con las distintas concentraciones ensayadas del veneno sobre las cepas bacterianas utilizadas. ABSTRACT Envenoming caused by Thalassopryne maculosa (Pisces, Batrachoididae) in humans causes mainly inflammatory clinical manifestations, sometimes leading to very severe, temporarily disabling, although apparently non-lethal lesions. Despite this, many of the components responsible for such effects have not been precisely studied or identified; therefore, studying some of the in vitro and in vivo biological effects of the venom of this marine species is considered necessary from a clinical point of view. The absorption spectrum revealed the predominance of protein components of the venom, while the concentration of total proteins in the soluble fraction was determined at 280 nm. Twenty specimens produced 79.41 mg of protein used in the tests for evaluation of biological effects in vivo and characterization in vitro. Biological activity in vivo was tested after intraperitoneal injection of the venom in C57BL/6 female mice, calculating the fifty lethal doses (LD50), according to the Dixon and Mood method, at 70.80 (66.95-75.12) mg.kg-1 , for 60 minutes of experimentation, and simultaneously assessing the temporal course of experimental acute toxicity. The appearance of clinical manifestations of toxicity, was gradual; beginning with hyperactivity and tachypnea, followed by hypoactivity and abdominal distension to succeed, around 30 minutes, periods of bradypnea/apnea which resuled in death or survival of the mice. The proteolytic, phospholipase, hemolytic and antibacterial activity (against Aeromonas hydrophila, A. sobria and Staphylococcus aureus) was evaluated in vitro. The gelatinase activity assay revealed the proteolytic capacity of the venom up to a 1:64 dilution; the hemolytic effect was observed in assays with blood agar from amounts equal to or greater than 50 µg.µL-1 , while a slight phospholipase activity has been suggested with the highest concentration of venom used (8.0 mg.mL-1). Absence of antibacterial activity was demonstrated with the different concentrations of the venom tested on the bacterial strains used.
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... Accordingly, P. plectrodon exhibited positive signals on the telomeric regions of the short arm of pair 21 (Fig. 1A); Halobatrachus didactylus possesses a single pair of NOR-bearing chromosomes but submetacentric instead of subtelocentric (Palazo´n et al. 2003). Unsatisfactory results were obtained by Brum et al. (2001) when attempting to detect NORs by silver impregnation in P. porosissimus, but they inferred the occurrence of more than a single NOR-bearing pair on the basis of the presence of one to three nucleoli per nucleus, whereas Nirchio et al. (2004) established that T. maculosa does not possess additional NORs since fluorescence in situ hybridization revealed fluorescent signals on the telomeric region of the short arm of only one medium-sized subtelocentric chromosome pair (pair 16), the same that was identified using silver salts. Although more information is required for inferring phylogenetic relationships between batrachoidid species, available data suggest that chromosome translocation events involving the active NOR sites detectable by silver impregnation seem to be associated with the diversification in the group; hence NOR sites have a potentially high cytotaxonomical value within this fish group. ...
Karyology of the toadfish Porichthys plectrodon (Jordan and Gilbert, 1882) (Batrachoididae) from Margarita Island, Venezuela
Article
Full-text available
  • Dec 2004
This paper reports the results of cytogenetic analyses carried out on Porichthys plectrodon using conventional Giemsa staining, C-banding and silver staining techniques. A diploid chromosome count of 2n=44 was observed, consisting of 8 metacentric, 10 submetacentric, 6 subtelocentric and 20 acrocentric chromosomes. Differences in length made it possible to identify homologous chromosomes within the metacentric group. Constitutive heterochromatin was distributed as large pericentromeric blocks in pairs 1 and 2, while the rest of the chromosomes were marked in centromeric regions, some more conspicuously than others. One pair of small-sized acrocentric NOR-bearing chromosomes (21) was identified by the nucleolar regions located terminally on their short arms.
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Chromosome characterization of the toadfish Halobatrachus didactylus (Schneider, 1801) (Teleostei: Batrachoididae) by fluorescent in situ hybridization
Article
  • Jan 2005
Halobatrachus didactylus (Schneider, 1801) has a chromosome number of 2n = 46 (8 metacentrics, 12 submetacentrics and 26 acrocentrics). Specimens studied were collected from waters of the Bay of Cadiz. Chromosome preparations were made from the cephalic portion of the kidney. The telomeric probe hybridized on the terminal position of all toadfish chromosomes. (GATA), hybridized preferentially on the subcentromeric region of only one submetacentric chromosome pair, although additional weak signals dispersed throughout the chromosomes were found. Major ribosomal genes were also located on a submetacentric pair, but at a telomeric position. Finally, the 5S rDNA probe produced a hybridization pattern similar to the GATA probe. These results provide interesting information which describes chromosomal markers that may be of utility for the management of natural populations of this species.
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Chromosome evolution in fishes: A new challenging proposal from Neotropical species
Article
Full-text available
  • Oct 2014
  • NEOTROP ICHTHYOL
Se presenta una base de datos que contiene los datos citogenéticos de peces Actinopterigios Neotropicales de Venezuela obtenidos por primera vez en un solo laboratorio. Los resultados de este estudio incluyen 103 especies pertenecientes a 74 géneros de 45 familias contenidas en 17 de los 40 órdenes de teleósteos. En el grupo de peces marinos, el número diploide modal fue 2n=48 representado en 60% de las especies estudiadas, mientras que en el grupo de peces de agua dulce el complemento diploide modal fue 2n=54, representado en el 21,21% de las especies estudiadas. El número de cromosomas y FN promedio fueron estadísticamente superiores en peces dulceacuícolas. El grado de diversificación y variación en el cariotipo también fue mayor en peces de agua dulce en contraste con un patrón citogenético más conservado en peces marinos. En contraposición a la suposición según la cual 48 cromosomas acrocentricos era el número cromosómico basal en los peces, los datos aquí presentados muestran que existe una evidente tendencia hacia la reducción del número de cromosomas desde valores cercanos a 2n=60 con alto número de cromosomas birrámeos en las especies más basales a 2n=48 elementos acrocéntricos en los actinopterigios más derivados.
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A Review of the Venomous Toadfishes, Subfamily Thalassophryninae
Article
Full-text available
  • Dec 1966
  • COPEIA
The subfamily Thalassophryninae differs from the other two subfamilies of Batrachoididae (Batrachoidinae and Porichthyinae) in having two hollow dorsal spines and a hollow opercular spine that connect with venom glands. Thalassothia Berg is shown to be a synonym of Thalassophryne Günther. The eastern Pacific Daector Jordan and Evermann differs from the western Atlantic Thalassophryne in having discrete glands between the bases of the upper pectoral fin rays and in having more precaudal and caudal vertebrae and more second dorsal and anal fin rays. Three species of Daector are recognized: reticulata Günther; gerringi Rendahl; and dowi Jordan and Gilbert (including depressa Hildebrand). Six species of Thalassophryne are recognized: amazonica Steindachner; megalops Bean and Weed; maculosa Günther (including quadrizonatus Eigenmann and wehekindi Fowler); nattereri Steindachner (including branneri Starks); punctata Steindachner; and montevidensis Berg (including platensis Devincenzi). The presence or absence of the axillary and pectoral fin glands are considered to be of generic value. Thalassophryne, Amphichthys, and Halophryne lack both glands. Daector, Aphos, Porichthys, and Batrachoides lack the axillary gland but have discrete pectoral fin glands. Batrachomoeus and Sanopus have the axillary gland and lack pectoral fin glands. Opsanus has both types of glands. Opsanus astrifer is placed in Sanopus along with O. barbatus.
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Chromosome Formulae of North American Fishes
Article
Full-text available
  • Jan 1980
Chromosome formulae (diploid chromosome and chromosome arm number) are listed for 309 North American fish species, including representatives from 145 genera in 54 families. This updates earlier lists by more than 130 entries. In addition the list displays the presence of microelements or sex chromosomes in the complements of several species. The source of taxa was the Third Edition (1970) of the list of common and scientific(names of fishes prepared by the American Fisheries Society's Committee on Names of Fishes.
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Trypsin G-banding of North American Cyprinid Chromosomes: Phylogenetic Considerations, Implications for Fish Chromosome Structure, and Chromosomal Polymorphism
Article
Full-text available
  • Jan 1991
Trypsin G-banding of chromosomes from North American cyprinid fishes, was used to address phylogenetic problems within the group and to demonstrate the occurence of G-bands which are not differentially rich in AT DNA base pairs. G-band homology of the long arm of the F′, C′, and E′ NOR chromosomes found among six North American cyprinid species, in concert with a hypothesis of species relationships based on morphology, suggests that a NOR situated terminally on the largest chromosome in the complement may represent the plesiomorphic NOR character state within the large “Notropis”-like shiner assemblage. Outgroup comparison suggests that this chromosome may also represent a synapomorphy for the same lineage. Evidence exists which suggests that the F′ NOR is ancestral and that the C′ and E′ NORs are derived. The occurrence in cyprinids of trypsin-induced G-bands which are not differentially rich in AT-/GC-DNA base pairs may indicate that the evolution of trypsin induced G-bands preceded the evolution of differential AT-/GC-richness. The use of trypsin G-banding in cyprinids is expected to permit the identification of previously cryptic chromosomal rearrangements both within and among species. © 1991, Japan Mendel Society, International Society of Cytology. All rights reserved.
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Allenbatrachus, A New Genus of Indo-Pacific Toadfish (Batrachoididae)l
Article
Full-text available
  • Jul 1997
Allenbatrachusis described as a new genus for two Indo-West Pacific species previously known as Batrichthys grunniens (Linnaeus, 1758) and Batrachus reticulatus (Steindachner, 1870). It is included in the subfamily Batrachoidinae and is separated from other genera on the basis of the following combination ofcharacters: a dorsocranium foramen behind each eye; two subopercular spines; no pectoral-fin axil foramina; raised flange on dorsal surface of maxilla; and protruding lower jaw. The two species of Allenbatrachus are redescribed and a neotype designated for Batrachus reticulatus. NEARSHORE COLLECTIONS FROM the Indo-Aus­ tralian Archipelago to the Ganges River area of India may yield two species of toadfish that lack a foramen in the pectoral-fin axil. One of these species has been identified as Batrichthys grun­ niens by Hutchins (1981). The second species has masqueraded under the name of B. grun­ niens, but in fact is the species described as Batrachus reticulatus by Steindachner (1870); it has blunt, rounded teeth rather than the pointed teeth of Batrichthys grunniens. Hutchins (1981) unraveled the complex nomenclature of the toadfish species of India and correctly determined that the species described by Linnaeus (1758) as Cottus grun­ niens had priority over Batrachoides gangene described by Hamilton (1822). Hutchins fol­ lowed Menon (1963) in utilizing South African genera (Smith 1952) for the Indian species he treated and placed grunniens in the genus Batrichthys; however, he commented that of the recognized batrachoidid genera, "... none is apparently suitable for either of the above two Indian species." As part of a revision of the batrachoidid gen­ era, I have cleared and stained representatives of all described genera in the family. Examina­ tion of this material has demonstrated that Hutchins' misgivings about the use of the genus Batrichthys for grunniens is correct and neither grunniens nor reticulatus can be included in that
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Karyotypes of three species of toadfish (Batrachoididae: Teleostei) from Venezuela
Article
Full-text available
  • Mar 2002
  • SCI MAR
Karyotypes of three species of toadfish (Amphichthys cryptocentrus, Batrachoides manglae and Thalassophryne maculosa), reported here for the first time revealed diploid complements of 46 chromosomes. Karyo-evolutionary trend of these species is discussed. Based on differences in arm number, we suggest that T. maculosa possesses the most evolved karyotype among the three species, hence it would be the more recent one, followed by B. manglae and A. cryptocentrus.
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Conventional karyotype and nucleolar organizer regions of the toadfish Halobatrachus didactylus (Schneider, 1801) (Pisces: Batrachoididae)
Article
Full-text available
  • Dec 2003
For this report, we studied the conventional karyotype and the NOR-bearing chromosomes in the toadfish Halobatrachus didactylus. We found a karyotype of 46 chromosomes made up of 8 metacentric, 12 submetacentric, and 26 acrocentric elements (FN = 66). No heteromorphic sex chromosomes were observed in the species. Metacentric chromosomes were easily classified as homologous pairs according to their morphology and L/S ratio. The rest of the chromosomes could not be accurately classified as homologous pairs because differences in chromosome size and L/S ratio were too slight between adjacent pairs within a size-ranged series. A single pair of NOR-bearing chromosomes was found. Active NORs were terminally located in a submetacentric pair of chromosomes. Se estudió el cariotipo del pez sapo marino. El número de cromosomas. La ausencia de cromosomal sexuales heteromórficos. Lso cromosomas metacéntricos fueron facilmente clasificados y el resto de cromosomas no pudo ser clasificado con precisión. Peer reviewed
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R- and G-band patterns in Astyanax scabripinnis paranae (Pisces, Characiformes, Characidae)
Article
Full-text available
  • Jun 1999
The absence of longitudinal bands in fish chromosomes has been associated with technical problems in chromosome preparations or the absence of a structural compartmentalization in the fish genome. In the present study, a R-banding pattern was obtained using a replication banding technique by in vivo treatment with 5-bromodeoxyuridine (5-BrdU). G-banding patterns were obtained after trypsin treatment and also after chromosome cleavage by in situ treatment with the restriction endonuclease BamHI. A similar G-banding pattern was also obtained after cleavage with the endonuclease HinfI. Presence of a resolute R- and G-banding patterns shows that Astyanax scabripinnis paranae chromosomes could present an isochore-like structure similar to that found in other vertebrates.
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Determination of early stages of sex chromosome differentiation in the sea bass Dicentrarchus labrax L. (Pisces: Perciformes)
Article
  • Jan 1996
  • Cytobios
Dicentrarchus labrax from two populations, sea farm, and a wild population were cytogenetically characterized. The diploid chromosome number was 2n = 48 (NF = 48). One pair of Ag-NORs were detected at the terminal or near-terminal site on the short arms of the acrocentric chromosome pair 22. A polymorphism, affecting this NOR-bearing chromosome pair, was found with many individuals from the wild population being involved. A chromosome variation was discovered in the specimens from the wild population, involving the amount and the patterns of heterochromatin distribution in one of the two smallest chromosome pairs (24) of the mitotic complement. In females this pair comprised two chromosomes which displayed roughly the same C-band patterns. In males, both homologues differed from one another as to the amount of heterochromatin and the pattern of distribution (C-bands). This differentiation was interpreted as an indication of the likely type of sex chromosome determining mechanism, namely XX/XY. Similarly, the smallest chromosome pair in Serranus cabrilla displayed a heteromorphism in respect of the size and the pattern of heterochromatin distribution. The significance of some cytogenetic peculiarities and the results of their comparative analysis in both species are discussed.
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Comparative karyology of selected deep-sea and shallow-water teleost fishes.
Article
Microfilm of typescript. Ann Arbor, Mich., University Microfilms, 1968. 1 reel. 35 mm. Thesis--Yale University, 1967. Bibliography: leaves 135-153.
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