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Betta splendens is a very important ornamental species. The current paper describes the embryonic and larval development of B. splendens under stereomicroscopy and scanning electron microscopy. Eggs and larvae from natural spawning were collected at different developmental stages at previously established intervals and analysed. The eggs of B. splendens are yellowish, clear, spherical, demersal, translucent and telolecithal with a large amount of yolk. Between 0–2 h post-initial collection (hpIC), the eggs were at the egg cell, first cleavage and morula stages. The blastula stage was identified at 2–3 hpIC and the early gastrula phase was observed at 3–4 hpIC with 20% epiboly, which was finalized after 13–18 hpIC. When the pre-larvae were ready to hatch, the appearance of somites and the free tail were observed, at 23–25 hpIC. At 29 hpIC, the majority of larvae had already hatched at an average temperature of 28.4 ± 0.2°C. The newly hatched larvae measured 2.47 ± 0.044 mm total length. The mouth opened at 23 h post-hatching (hPH) and the yolk sac was totally absorbed at 73 hPH. After 156 hPH, the heart was pumping blood throughout the entire larval body. The caudal fin, operculum and eyes were well developed at 264 hPH. When metamorphosis was complete at 768 hPH, the larvae became juveniles. The current study presents the first results about early development of B. splendens and provides relevant information for its reproduction, rearing and biology.
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Zygote: page 1 of 10 c
Cambridge University Press 2013
Early development of Betta splendens under stereomicroscopy and
scanning electron microscopy
Fernanda Nogueira Valentin2,Nivaldo Ferreira do Nascimento2,Regiane Cristina da Silva2,
João Batista Kochenborger Fernandes2,Luiz Gustavo Giannecchini and Laura Satiko Okada Nakaghi1
Laboratório de Histologia e Embriologia do Departamento de Morfologia e Fisiologia Animal, Faculdade de Ciências
Agrárias e Veterinárias, Universidade Estadual Paulista, Jaboticabal, São Paulo, Brazil; and Centro de Aquicultura da
Universidade Estadual Paulista (CAUNESP), Jaboticabal, São Paulo, Brazil
Date submitted: 8.5.2013. Date revised: 20.6.2013. Date accepted: 9.9.2013
Betta splendens is a very important ornamental species. The current paper describes the embryonic and
larval development of B. splendens under stereomicroscopy and scanning electron microscopy. Eggs
and larvae from natural spawning were collected at different developmental stages at previously
established intervals and analysed. The eggs of B. splendens are yellowish, clear, spherical, demersal,
translucent and telolecithal with a large amount of yolk. Between 0–2 h post-initial collection (hpIC),
the eggs were at the egg cell, first cleavage and morula stages. The blastula stage was identified at 2–
3 hpIC and the early gastrula phase was observed at 3–4 hpIC with 20% epiboly, which was finalized
after 13–18 hpIC. When the pre-larvae were ready to hatch, the appearance of somites and the free tail
were observed, at 23–25 hpIC. At 29 hpIC, the majority of larvae had already hatched at an average
temperature of 28.4 ±0.2°C. The newly hatched larvae measured 2.47 ±0.044 mm total length. The
mouth opened at 23 h post-hatching (hPH) and the yolk sac was totally absorbed at 73 hPH. After
156 hPH, the heart was pumping blood throughout the entire larval body. The caudal fin, operculum
and eyes were well developed at 264 hPH. When metamorphosis was complete at 768 hPH, the larvae
became juveniles. The current study presents the first results about early development of B. splendens
and provides relevant information for its reproduction, rearing and biology.
Keywords: Eggs, Embryos, Fish, Larvae, Ontogeny, Reproduction
Betta splendens, widely cultivated as an ornamental
fish, is a very important species for fish farming.
The betta fish has an auxiliary breathing organ
known as the labyrinth, which allows them to
breathe atmospheric oxygen and tolerate low levels
of dissolved oxygen in the water (Damazio, 1992;
Faria et al., 2006). Certain features of B. splendens,
such as colour, fin length and temperament, have
1All correspondence to: Laura S.O. Nakaghi. 2Laboratório
de Histologia e Embriologia, Departamento de Morfologia
e Fisiologia Animal, Faculdade de Ciências Agrárias e
Veterinárias, Universidade Estadual Paulista, Jaboticabal,
Via de Acesso Prof. Paulo Donato Castellane s/n, ZIP code-
14884–900, Jaboticabal–São Paulo, Brazil. Tel:/Fax: +55 16
3209 2654 (ext. 232). e-mail:
2Centro de Aquicultura da Universidade Estadual Paulista
(CAUNESP), Jaboticabal, São Paulo, Brazil.
been selected by breeders for centuries for ornamental
purposes and fighting. Therefore, in the wild, bettas
are less aggressive with less colouring and shorter fins
(Monvises et al., 2009).
Among the several Betta species, the best known
is Betta splendens (Faria et al., 2006; Monvises et al.,
2009). However, despite its commercial importance,
little information is known about its ontogeny and
it is during the early developmental stages, such as
the onset of exogenous feeding, that high mortality
rates are observed (Yúfera & Darias, 2007). Therefore,
study of the early developmental stages is necessary
to establish good production methods (Maciel et al.,
The embryonic developmental stage lasts from
fertilization of the oocyte by the sperm until hatching
of the larvae (Matkovic et al., 1985; Solnica-Krezel,
2005; da Rocha Perini et al., 2010). The larval
development stage begins with hatching and lasts
until metamorphosis is complete, when the larvae
2Valentin et al.
acquire morphological characteristics similar to those
of the adults and are known as juveniles (Kendall
et al., 1984). Meanwhile, a series of morphological
changes essential to survival are observed, such as
the development of fins, breathing and feeding (Osse,
1989; da Rocha Perini et al., 2010).
Stereomicroscopy and scanning electron microscopy
show structural differences during the development
of eggs and larvae (Paes et al., 2011), which provide
important information about betta biology. Therefore,
due to the importance and the lack of knowledge
about the ontogeny of this species, the current study
analyses the embryonic and larval development of
B. splendens using stereomicroscopy and scanning
electron microscopy.
Materials and methods
Site, animals and sampling
The experiment was carried out at the Ornamental
Fish Laboratory of the Aquaculture Center of UNESP
(CAUNESP), in Jaboticabal, São Paulo, Brazil. The av-
erage physico-chemical water parameters monitored
in the tanks were as follows: average temperature
28 ±0.2°C; dissolved oxygen 5.3 mg/L and conduct-
ivity 44 S/cm2. Four animals (two males and two
females) were kept in individual 2-litre tanks equipped
with a water recirculation system for 10 days. After
this period, the females with visible oviducts were
transferred to the tanks with the males, where they
remained inside plastic cups with holes and were
released after a day to initiate reproductive behaviour.
Gamete release started after 3 days. This process was
slow and lasted approximately 3 h (the two couples
spawned and the eggs were pooled). The sampling
started at pre-established times: initial collection (IC;
as soon as eggs were observed in the nests), hourly
until 6 h post-initial collection (hpIC), every 3 h until
hatching, 1 h post-hatching (hPH), every 2 h until 19
hPH, every 4 h until 43 hPH, every 6 h until 91 hPH
and every 7 days until 936 hPH. Sampling consisted of
collecting 10 eggs and larvae at a time. From 72 hPH,
the larvae were fed with Artemia sp. twice daily. The
samples were fixed in 4% formaldehyde and 0.1 M
phosphate buffer, pH 7.4 and modified Karnovsky
(2.5% glutaraldehyde and 1.0% paraformaldehyde,
with 1.0 M cacodylate buffer and pH 7.2).
Eggs and larvae were examined under a LEICA
MZ8 stereomicroscope equipped with a LEICA DFC
280 camera, using the IM 50-LEICA software. Egg
diameter and total length of the larvae (n=10) were
measured in each sample. For scanning electronic
microscopy analysis, the samples were post-fixed in
1% osmium tetroxide for 2 h and washed in sodium
phosphate buffer. Subsequently, they were dehydrated
in graded series of ethanol at 30, 50, 70, 80, 90 and
95% concentrations plus three washes at 100% (10 min
each). Soon after, the samples were dried to the critical
point in a liquid CO2drier, mounted on a copper
grid, coated with gold–palladium ions, observed and
photographed under a scanning electron microscope
(JEOL-JSM 5410).
Embryonic development
Betta splendens exhibited external fertilization and
partitioned spawning, with gamete release that
extended up to 3 h. Once spawning had finished, the
females were removed due to the aggressive behaviour
of the males.
Eggs from a single collection exhibit different
development stages due to partitioned spawning and
the pooling of eggs from two different spawning.
Therefore, eggs from the initial collection were
light yellow, spherical, translucent, telolecithal and
demersal. At this time, the eggs were at the egg cell,
first cleavage and morula stages, with an average
diameter of 1.08 ±0.038 mm (Fig. 1A–C). The early and
late blastula stages were identified between 2–3 hpIC
while the cells continued to divide mitotically (Fig. 1D,
E). The early gastrula was observed between 3–4
hpIC with 20% epiboly (Fig. 1F). Gastrula presented
between 30 and 50% epiboly at 5–6 hpIC (Fig. 1G).
In B. splendens, the blastoderm does not completely
surround the yolk; therefore, no yolk plug is formed
and approximately 50% of the yolk is covered. At 11–17
hpIC, thickening of the dorsal epiblast was observed,
which would give rise to the head of the embryo
(Fig. 1H).
At 21 hpIC, the head and tail were differentiated
and the first melanophores were present in the yolk,
found mainly near the ventral region of the embryo
(Fig. 1I). Over time, between 23–25 hpIC, the embryos
became pre-larvae. During this period, pairs of somites
occupied the entire notochord from the occipital to
the caudal region. The pre-larva was ready to hatch
and presented a free tail with strong and continuous
movements. This period, propagation and increase of
melanophores in the yolk toward the dorsal region of
the pre-larva was also observed (Fig. 1J). The hatching
began at 28 hpIC and the larvae presented poorly
pigmented eyes (Fig. 1K). After 32 hpIC, 90% of the
larvae had already hatched and after 38 hpIC, 100%
of the larvae had hatched. The total average length of
newly hatched larvae was 2.47 ±0.04 mm. Table 1 and
Early development of B. splendens 3
Figure 1 Main stages of the embryonic development of Betta splendens.(A) Egg cell; (B) 0–2 h post-initial collection (hpIC):
first cleavage; (C) 0–2 hpCI: morula; (D) 2–3 hpIC: blastula begins; (E) 2–3 hpIC: blastula ends; (F) 4–5 hpIC: beginning of the
gastrula with 20% of epiboly; (G) 6–9 hpIC: gastrula with 50% of epiboly; (H) 12–18 hpIC: gastrula at the end of epiboly; (I)21
hpIC: beginning of the formation of embryo and yolk pigmentation (arrow); (J) 24–26 hpIC: pre-larva, ready to hatch, showing
the cephalic and tail regions (arrows) and the presence of somites (arrowheads); (K) 29 hpIC: hatching of the larva and eye
Fig. 1 show the main embryonic development stages of
B. splendens.
Larval development
At the beginning of hatching, B. splendens measured
2.47 ±0.044 mm total length (TL) and presented
melanophores in the yolk at the antero-ventral axis.
At this moment, the newly hatched larvae showed a
closed mouth, large yolk sac, non-differentiated and
slightly pigmented eyes (Fig. 2A), a pectoral fin bud
and a well developed caudal fin (Fig. 3B).
At this stage, the larvae had low swimming capacity
and remained attached to the bubble nest under the
parental care of the male. For this reason, they had
4Valentin et al.
Table 1 Embryonic development of the Betta splendens at 28.4 ±0.2 °C
Time post-fertilization
Developmental stage (h) Note
Egg cell 0 Fig. 1A
First cleavage 0–2 Fig. 1BBlastodisc divided to form two equal cells
Morula 0–2 Fig. 1C
Blastula begins 2–3 Fig. 1D
Blastula ends 2–3 Fig. 1E
Gastrula with 20% of epiboly 4–5 Fig. 1FBlastoderm cells begin to spread over the yolk.
30–50% of epiboly 6–9 Fig. 1GGerm ring epiboled ½of yolk sac
End of epiboly 12–18 Fig. 1F
Organogenesis begins 21 Fig. 1IBeginning of embryo formation
Preparation for hatching 24–26 Fig. 1J, Fig. 3APre-larva with free tail and the presence of somites
70% Hatching 29 Fig. 1K
adhesive glands that were identified in the dorsal
region of the head, just above the eye (Fig. 4A,B).
1–11 hPH
The olfactory cavity was formed after 1 hPH,
surrounded by mucus-producing cells (Fig. 4C), and
after 7 hPH it was deeper (Fig. 4D). It was observed
that the eyes were more pigmented; the notochord
more visible and the yolk volume reduced after 11
hPH (Fig. 2B). The melanophores were more evident
in the entire yolk and concentrated mainly on the
larval antero-ventral axis (Fig. 2B). At this stage,
the disappearance of the adhesive gland was also
11–43 hPH
Cilia were identified on the upper lip and the mouth
opening at 17 hPH (Fig. 5A), when the larva TL was
2.66 ±0.068 mm. At 43 hPH, the opercle covered the
gills (Fig. 3E). Despite the mouth opening was present,
we did not observe larvae feeding at this stage.
43–65 hPH
At 49 hPH the eyes and body were more pigmented
(Fig. 2D). At 65 hPH, a large amount of neuromasts
was observed in the lateral line and around the
eye (Fig. 5D–F). At this stage, a greater swimming
ability was observed, with the larvae displaying
morphological and sensory structures that enabled
greater mobility and perception of the surroundings.
65–264 hPH
At this stage, although the larvae still had yolk,
exogenous feeding had begun, as Artemia sp. nauplii
could be seen inside their bodies at 72 hPH (Fig. 6A).
The yolk was completely absorbed at 73 hPH, when
the larvae measured 3.20 ±0.176 mm in length. From
this phase onwards, the larvae exclusively obtained
food via exogenous feeding, by actively chasing
the nauplii (Artemia sp.) (Fig. 6B). After 86 hPH,
neuromasts were observed in the lower jaw (Fig. 5G).
At 156 hPH, the heart was pumping blood throughout
the entire larval extension (Fig. 6D). At 264 hPH, the
caudal fin was fully formed, with opercle and eyes well
developed (Fig. 3H).
264–768 hPH
At 432 hPH, the dorsal and anal fins were being
formed and the caudal fin rays were clearly evident
(Fig. 2F). Characteristics similar to the adults were
observed when larval TL was 17.24 ±2.064 mm, which
characterized the end of the larval stage (Fig. 2G),
after 768 hPH. From this point on, the animals are
considered juveniles. Table 2 describes the main stages
of larval development.
Betta splendens are sedentary fish with parental
care, whose eggs adhere to the bubble nest. Betta
eggs are non-adhesive as, morphologically, they do
not present any of the structures that define this
characteristic such as zona radiata with hexagonal
pore canals, filaments, villous blood cells or gelatin
covers (Godinho & Godinho, 2003). However, the eggs
remain and develop in bubble nests constructed by
the male, who produces in his mouth a thick mucus
that consists of glycoproteins that help to maintain
the permanence of the bubbles in the nest (Kang &
Lee, 2010); thus, it is believed that this mucus may
also be linked to egg permanence in the bubble nest.
However, adhesive eggs are described for sedentary
fish such as Acestrorhynchus britskii,Acestrorhynchus
lacustris,Serrasalmus spilopleura (Rizzo et al., 2002),
Astronotus ocellatus (Paes et al., 2011) and Franciscodoras
marmoratus (Alberto Weber et al., 2012).
Early development of B. splendens 5
Figure 2 Larval development of Betta splendens.(A) newly hatched larva (29 h post-fertilization); (B) 11 h post-hatching (hPH);
(C)23hPH;(D)49hPH;(E)73hPH;(F) 432 hPH; (G) 768 hPH.
Betta splendens eggs do not have oil droplets and
are telolecithal. The yolk is most concentrated at the
vegetal pole while the organelles and cytoplasm are
concentrated in the animal pole (Kunz, 2004; Ninhaus-
Silveira et al., 2006). The eggs are demersal because
their specific gravity is greater than water (Godinho
& Godinho, 2003) and the animal pole is oriented
upward. This event occurs because the yolk sac has
a greater relative gravity than the blastodisc (Kunz,
Egg average diameter in the current study was
approximately 1.08 ±0.038 mm. This value is close to
the 0.8 mm reported by Watson & Chapman (2002) as
the average for ornamental species. The egg diameter
6Valentin et al.
Figure 3 Electron micrographs of Betta splendens.(A) 25 h post-fertilization (hPF): Pre-larva ready to hatch; (B) 29 hPF: newly
hatched larva; (C) 15 h post-hatching (hPH); (D)17hPH;(E)43hPH;(F)61hPH;(G)85hPH;(H) 264 hPH. FF: finfold. Y: yolk.
Arrowheads: mouth. Black arrows: pectoral fin. White arrows: opercle.
and quality are related to factors such as parental care,
breeder nutrition, ecological strategy, water quality,
photoperiod, animal welfare and genetic influence
(Brooks et al., 1997; Kolm & Ahnesjo, 2005). Parental
care is directly related to egg size: the larger the egg,
the greater the parental care (Kolm & Ahnesjo, 2005).
Usually, migratory fish exhibit another strategy, which
does not present parental care and produce a large
number of small eggs (Godinho et al., 2010).
The size and shape of the eggs may be important
for systematic and phylogenetic studies, as observed
in the identification of species of the genus Gobius
(Borges et al., 2003). Furthermore, it is also relevant
to identify spawning areas and to implement pro-
grammes to protect and preserve the species (Nakatani
et al., 2001).
The cleavage process, which consists of the division
of the egg into smaller cells named blastomeres, starts
Early development of B. splendens 7
Figure 4 Electron micrographs of Betta splendens showing some structures during development. (A,B) Newly hatch larvae,
adhesive glands. (C) 1 h post-hatching (hPH): olfactory cavity surrounded by mucus-producing cells. (D) 7 hPH: olfactory
cavity. Arrow: pectoral fin. Arrowhead: adhesive glands.
Figure 5 Electron micrographs of Betta splendens showing some structures during development. (A) 17 h post-hatching (hPH);
(B)23hPH;(C)85hPH;(D–F) 65 hPH: neuromasts on the lateral line region and around the eye; (G) 86 hPH: neuromasts on
the lower jaw region; (H) 936 hPH: detail showing scales in juveniles. Thick arrows: ciliated cells. Thin arrows: neuromasts.
after fertilization and zygote formation (egg cell).
This process varies greatly among vertebrates and
depends on the amount of egg yolk (Gilbert, 2003).
The eggs of B. splendens undergo meroblastic or partial
cleavage because the mitotic divisions occur only in
the animal pole of the egg. This type of cleavage
is typical of fish that accumulate a large amount
of yolk (Leme dos Santos & Azoubel, 1996; Gilbert,
2003; Takeuchi et al., 2008), such as Gymnocorymbus
ternetzi (Celik et al., 2012), F. marmoratus (Alberto
8Valentin et al.
Figure 6 Photomicrographs of Betta splendens.(A) 72 h post-hatching (hPH). (B)96hPH.(C) 156 hPH. (D) 156 hPH. Circle:
Heart. Black thick arrow: Artemia sp. ingested by the larva. Black thin arrow: mouth opened for respiration.
Table 2 Main events during larval development of B. splendens
stage Main events
29 hpf Hatching begins. Displays total length of 2.47 ±0.044 mm, little pigmented eyes, closed mouth, adhesive
glands and the presence of melanophores in the anterior region (Fig. 2A, Fig. 3Band Fig. 4A,B)
11 hPH Eyes more pigmented, reduced yolk volume, evident notochord and the presence of melanophores in a
region of the yolk (Fig. 2B)
17hPH Total length of 2.66 ±0.068 mm and mouth opening (Fig. 5A)
49 hPH Eyes and body were more pigmented (Fig. 2D)
73hPH Yolk totally absorbed and total length of 3.20 ±0.176 mm (Fig. 2E)
432 hPH Dorsal and anal fins appear (Fig. 2F)
768 hPH Characteristics similar to adult fish and total length of 17.24 ±2.064 mm (Fig. 2G)
hpf: hours post-fertilization; hPH: hours post-hatching.
Weber et al., 2012) and Brycon gouldingi (Faustino et al.,
The morula stage is reached once the zygote has
divided into 64 blastomeres. Through these divisions,
the number of cells increases while the volume of each
individual cell decreases (Wolpert et al., 2000; Gilbert,
2003), as observed for B. splendens. Immediately after
the morula stage, the blastula, which is characterized
by the blastoderm, is formed (Marques et al.,
Early development of B. splendens 9
The gastrulation process is characterized by epiboly
and involution movements (Wolpert et al., 2000;
Gilbert, 2003; Kunz, 2004). The epiboly movement
consists of the spreading of the blastoderm toward
the vegetal pole (Gilbert, 2003; Faustino et al., 2010a).
After the blastoderm has engulfed at least the half of
the yolk, the involution movement initiates (Gilbert,
2003; Kunz, 2004). Wolpert et al. (2000) and Kunz (2004)
show that this process leads to the formation of two
layers: the epiblast, which gives rise to the ectoderm,
and the hypoblast, which gives rise to mesoderm and
Temperature strongly influences the duration of
embryonic and larval development (Morrison et al.,
2001; Martell et al., 2005): the higher the temperature,
the shorter developmental time and vice versa (Leme
dos Santos & Azoubel, 1996; Martell et al., 2005).
The embryonic development of B. splendens was
slower compared with other ornamental species. The
cardinal tetra, Paracheirodon axelrodi, hatches 19–20 h
post- fertilization (hpf) at an average temperature of
26 ±1ºC (Anjos & Anjos, 2006); Gymnocorymbus ternetzi
hatches 20–21 hpf at 24 ±0.5ºC (Celik et al., 2012);
however, A. ocellatus development was even slower, as
it hatched after 46–58 hpf at a temperature of 27.5ºC
(Paes et al., 2011). This variability can be explained
by factors such as temperature and interspecific
Fish species can be classified either as precocial or
altricial depending upon the strategy adopted. The
larvae of precocial species hatch from the eggs in the
juvenile stage, while the altricial species hatch before
this stage and the larvae exhibits an undifferentiated
developmental stage (Bejarano-Escobar et al., 2010).
Betta splendens is an altricial species, as it hatches with
several organs and systems in differentiation.
The newly hatched B. splendens larvae had an
average TL of 2.47 ±0.044 mm, higher than other
sedentary species such as F. marmoratus (Alberto Weber
et al., 2012) that hatches at 1.27 ±0.4 mm and G. ternetzi
(Celik et al., 2012) at 1.44 mm, but smaller than P.
axelrodi (Anjos & Anjos, 2006) which hatches at 2.9 ±
0.2 mm. Coleman & Galvani (1998) stated that there
is a relationship between egg size and the length of
newly hatched larvae and analysed it in a wide range
of tropical species, concluding that the larger the egg
size, the longer would be the newly hatched larvae.
However, there are few studies available on this topic.
After hatching, B. splendens larvae have adhesive
glands that consist of mucous cells present in the head.
These glands enable the larvae to remain attached
to the nest, to increase parental efficiency (Araújo-
Lima & Bittencourt, 2001). These glands are similar
to those described for A. ocellatus (Paes et al., 2011),
Cichlasoma dimerus (Meijide & Guerrero, 2000) and
Hoplias malabaricus (Araújo-Lima & Bittencourt, 2002).
In the current study, the mouth opened very
quickly, while depletion of the yolk sac happened
later. According to Yúfera & Darias (2007), when
the larvae initiates exogenous feeding it is important
that all structures related to ingestion, digestion and
assimilation are ready. Furthermore, it is necessary
to highlight the importance of sensory structures
such as neuromasts and eyes (Bilotta & Saszik,
2001). Along with the development of fins, these
structures are essential for sensing and chasing food.
Therefore, describing these ontogenetic events will
help improve husbandry practices by making it
possible to determine the real needs of animals at
different developmental stages.
The larval stage was completed when the larvae
reached an average TL of 17.20 mm, when their
body features are similar to adults and become
juveniles (Kendall et al., 1984). The results of the
initial developmental stages of B. splendens provide
important information for the biology, breeding and
rearing of the species as well as a basis for further
We would like to thank Drs Maria do Carmo Faria Paes
and Sheryll Corchuelo for revising the manuscript
and the ornamental fish laboratory of CAUNESP for
supplying the animals.
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... Berdasarkan data pada Tabel 1 dapat dilihat bahwa lama waktu pengeraman telur di dalam mulut induk jantan dari mulai memijah sampai menetas adalah enam hari. Waktu penetasannya cukup lama, berbeda dengan B. imbellis yang memerlukan waktu sekitar 29 jam 04 menit (Cindelaras et al., 2015), 35 jam (Narwati, 2012), dan pada B. splendens sekitar 38 jam (Duarte et al., 2012;Valentin et al., 2013;Annur et al., 2016). ...
... Selain pergerakan aktivitas enzim juga berperan dalam pelunakan chorion sehingga mudah untuk dipecahkan oleh embrio (Blaxter, 1969). Pada ikan B. imbellis, larva cupang menetas setelah mengalami perkembangan selama 29 jam 04 menit (Cindelaras et al., 2015), hal yang sama seperti pada B. splendens menetas selama 29 jam (Valentin et al., 2013). ...
... Periode perkembangan larva berlangsung sampai umur 39 hari. Periode ini berlangsung lebih lama dibandingkan dengan periode perkembangan larva B. imbellis yang berlangsung sampai umur 23 hari pada suhu ruang (Rahmawati & Kusrini, 2013) dan 32 hari pada B. splendens (Valentin et al., 2013 Sehari setelah menetas, larva mempunyai kuning telur yang cukup besar dengan diameter 1,07 mm; sudah ada pigmentasi, bintik mata sudah terlihat namun belum berfungsi, mulut belum membuka, organ pencernaan belum terbentuk, dan sirip belum terbentuk (Gambar 1a); dan pada hari kedua setelah menetas, sirip ekor larva mulai berkembang dan pigmentasi sudah lebih jelas (Gambar 1b). Larva berwarna transparan agak kekuningan terutama pada kuning telur, pembuluh darah terlihat jelas terutama di bagian depan kuning telur. ...
Salah satu jenis ikan cupang alam yang menarik perhatian adalah cupang Betta rubra Perugia, 1893; yang merupakan jenis endemik dari perairan Banda Aceh. Status B. rubra di habitat aslinya sudah mulai sulit diperoleh sedangkan budidayanya belum berkembang. Oleh karena itu, informasi tentang embriogenesis dan perkembangan stadia awal ikan B. rubra sangat diperlukan untuk mendukung keberhasilan pengembangbiakannya. Penelitian ini bertujuan untuk mengetahui embriogenesis dan perkembangan larva ikan B. rubra. Telur dan larva yang digunakan berupa hasil pemijahan alami B. rubra di Balai Riset Budidaya Ikan Hias, Depok. Parameter yang diamati yaitu fase embriogenesis, perkembangan larva, dan benih ikan B. rubra. Pengamatan embriologi di mulai setelah ikan memijah sampai telur menetas, sedangkan perkembangan larva di mulai dari larva menetas sampai menjadi benih atau perkembangan telah sempurna. Pengamatan dilakukan setiap hari di bawah mikroskop binokuler Olympus SZX9 perbesaran 8-25 kali. Hasil penelitian menunjukkan bahwa perkembangan embrio telur B. rubra terjadi selama enam hari atau 144 jam hingga menetas menjadi larva pada suhu 27°C-28°C. Perkembangan embriogenesisnya yaitu hari pertama setelah memijah di mulai dengan pembelahan awal; hari kedua: blastula; hari ketiga: gastrula; hari keempat: pembentukan bakal kepala dan ekor; hari kelima: pembentukan ovtic vesicle dan notochorda; hari keenam: menetas. Perkembangan larva H-3—H-4 setelah menetas mata dan mulut mulai membuka; H-5—H-6: terbentuk anus dan kuning telur habis; H-7: peralihan pakan (indogeneous ke exsogeneous) dan metamorfosis terjadi 39 hari atau 936 jam setelah menetas.Betta rubra Perugia, 1983 is an endemic ornamental fish found in the swamp areas of Banda Aceh. Due to its appealing physical appearance, B. rubra wild population has been heavily exploited. Current aquaculture technology of the species is not yet available which implies an imminent threat to the conservation of this species. Therefore, the domestication the fish species is the first important step toward developing the aquaculture technology of the species which requires specific information on embryogenesis and the development of the early stadia B. rubra. This study aimed to determine embryogenesis and larval development of B. rubra. The eggs and larvae used from the natural spawning of B. rubra wild parents reared in the facility of the Ornamental Fish Cultivation Research Center, Depok, Indonesia. The parameters observed were the embryogenesis and early stages development of the fish from larvae to juvenile. Observation of embryogenesis started from eggs fertilization until hatching. The development of larvae was observed from post hatching until fully developed as fish juvenile. Embryonic and larval development were monitored daily using an Olympus SZX9 binocular microscope with 8x-25x magnification. The results showed that the embryogenesis of B. rubra lasted for six days or 144 hours until it hatched. The development stages of the embryogenesis after fertilization are as follow: division phase on the first day; blastula on the second day; gastrula on the third day; formation of heads and tails on the fourth day; formation of ovtic vesicles and notochordas on the fifth day; hatch on the sixth day. Larval development consists of: eyes and mouth begin to open at three to four days after hatching; anus is formed and the yolk is gone between the fifth and sixth days; intermediate feed (indogeneous to exsogeneous) at seventh day and metamorphosis at 39 days or 936 hours after hatching.
... Establishing genetic tools in betta will advance their use as a powerful experimental system to study developmental processes and behavioral traits. Genetic manipulation of betta is facilitated by their reproductive biology: betta fertilize externally and produce clutches of 250 eggs, each with a relatively large diameter of 1 mm (Valentin et al., 2015;Lichak et al., 2022). This enables the microinjection of zygotes in a similar manner to methods for genetic manipulation of zebrafish (Danio rerio) and medaka (Oryzias latipes). ...
... This enables the microinjection of zygotes in a similar manner to methods for genetic manipulation of zebrafish (Danio rerio) and medaka (Oryzias latipes). Nevertheless, the asynchronous egg fertilization resulting from a protracted mating process that can last many hours, coupled with a short interval between fertilization and cell division, as well as a thick chorion, constitute significant challenges for genetic manipulation (Valentin et al., 2015;Lichak FIGURE 1 CRISPR/Cas9-mediated knockout in betta (A) Experimental scheme for generating knockout betta. (B) Efficiency of knockout generation in P0, as determined by a T7EI assay. ...
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Betta splendens, also known as Siamese fighting fish or “betta,” is a freshwater fish species renowned for its astonishing morphological diversity and extreme aggressive behavior. Despite recent advances in our understanding of the genetics and neurobiology of betta, the lack of tools to manipulate their genome has hindered progress at functional and mechanistic levels. In this study, we outline the use of three genetic manipulation technologies, which we have optimized for use in betta: CRISPR/Cas9-mediated knockout, CRISPR/Cas9-mediated knockin, and Tol2-mediated transgenesis. We knocked out three genes: alkal2l, bco1l, and mitfa, and analyzed their effects on viability and pigmentation. Furthermore, we knocked in a fluorescent protein into the mitfa locus, a proof-of-principle experiment of this powerful technology in betta. Finally, we used Tol2-mediated transgenesis to create fish with ubiquitous expression of GFP, and then developed a bicistronic plasmid with heart-specific expression of a red fluorescent protein to serve as a visible marker of successful transgenesis. Our work highlights the potential for the genetic manipulation of betta, providing valuable resources for the effective use of genetic tools in this animal model.
... With regard to reproduction, the male constructs a bubble nest in which he attaches the eggs and the embryos develop under his care [4,38,44]. The hatching takes place between 29-and 38-h-post-fertilization (hpf) at 28 • C, according to different authors [12,47]. ...
... B. splendens is native for South East Asia and was domesticated for centuries [38]. Its selective breeding aims fighting and ornamental purposes like looking constantly for new color patterns and fin shapes [32,47]. This last aspect is very important due to the keeping of ornamental fishes is one of the most popular hobbies around the world [50]. ...
The Siamese fighting fish (Betta splendens) has great importance as an ornamental aquarium fish as well as laboratory model species. Due to its rapid development, a cooling-embryo protocol could provide some advantages in their transportation, embryonic synchronization, and optimization of hatcheries. In this context, this work aimed to develop a protocol to storage B. splendens embryos at two temperatures (5 and 14 °C), testing three cryoprotective solutions (S1: 0.5 M sucrose, 1.5 M methanol; S2: 0.25 M sucrose, 0.75 M methanol; and S3: 0.125 M sucrose, 0.375 M methanol) and evaluating the quality of the larvae obtained. Moreover, a method to isolate the embryos from the bubble nest constructed by the male and to incubate them without parental care was applied in this study. The cooling assays were done using embryos of 24-hours-post-fertilization at 26 °C and the results demonstrated that it is possible to store these embryos deprived of cryoprotectants at 5 °C for at least 6-hours without negative effects. Meanwhile, S2 and S3 were the most suitable solutions for its storage for 9-hours at 5 °C or 24-hours at 14 °C, obtaining 77% hatching and 52% normal larvae in the first case or 88% hatching and 81% larvae with mild abnormalities in the second one. Indeed, type and frequency of larval abnormalities were evaluated and, remarkably, a partial recovery was described on malformed larvae from embryo cooled at 14 °C. Finally, this work is the first report about the cooling of B. splendens embryos and establishes the conditions for further studies on this field with this species.
... For male Siamese fighting fish, Betta splendens, territories are a key resource because they provide space to build bubble-nests for their offspring, which plays a significant role in reproductive success [27][28][29][30]. The high fitness value of territory for males looking to build nests, and the even higher value for males protecting existing nests, underlies much of the B. splendens well-defined agonistic repertoire, including their display, attack and use of visual cues from opponents to assess their ability (RHP) and compare it to their own [31][32][33][34][35][36]. ...
... Bars indicate the standard error and effect sizes are indicated by Cohen's d where appropriate [**P ≤ 0.01] exceeds resource and contest costs. This relies on the contribution of bubble nests to reproductive success by preparing fish for mating and constituting investment in parental care [27][28][29][30]. The effects of nest presence on contest motivation are in line with previous evidence showing that the defence of territory with nests is a paternal priority for male Siamese fighting fish [28,31] and that bubble-nest holders exhibit elevated defensive aggressiveness against other males [32,33]. ...
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Background Competition is considered to rely on the value attributed to resources by animals, but the influence of extrinsic stressors on this value remains unexplored. Although natural or anthropogenic environmental stress often drives decreased competition, assumptions that this relies on resource devaluation are without formal evidence. According to theory, physiological or perceptual effects may influence contest behaviour directly, but motivational changes due to resource value are expected to manifest as behavioural adjustments only in interaction with attainment costs and resource benefits. Thus, we hypothesise that stressor-induced resource devaluations will impose greater effects when attainment costs are high, but not when resource benefits are higher. Noise may elicit such effects because it impacts the acoustic environment and imposes physiological and behavioural costs to animals. Therefore, we manipulated the acoustic environment using playbacks of artificial noise to test our hypotheses in the territorial male Siamese fighting fish, Betta splendens. Results Compared to a no-playback control, noise reduced defense motivation only when territory owners faced comparatively bigger opponents that impose greater injury costs, but not when territories also contained bubble nests that offer reproductive benefits. In turn, nest-size decreases were noted only after contests under noise treatment, but temporal nest-size changes relied on cross-contest variation in noise and comparative opponent size. Thus, the combined effects of noise are conditional on added attainment costs and offset by exceeding resource benefits. Conclusion Our findings provide support for the hypothesised modulation of resource value under extrinsic stress and suggest implications for competition under increasing anthropogenic activity.
... Traditional morphometric and meristic methods and visual observation are not discriminatory for many larval species (Matarese & Sandknop, 1984;Watson et al., 1998;Victor et al., 2009;Ko et al., 2013;Kwun, 2018). Furthermore, the pigmentation pattern of eggs and larvae changes in the later stages of development, making the species identification very challenging (Taylor et al., 2002;Valentin et al., 2015). Additionally, the succession of changes in eggs and larvae is very quick, and it leads to incorrect identification of early stages of commercially important fish species (Fox et al., 2005;Fox et al., 2008). ...
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Identifying the breeding grounds of fishes is crucial for the sustainable management of fisheries resources. The present study is aimed at identifying the potential breeding ground of Mugil cephalus along the estuary of the North Mumbai coast. A total of 1197 specimens of M. cephalus, including 546 eggs, 271 larvae, 235 juveniles, and 235 adults, were collected from four sampling stations in the Karanja estuary between January to October 2022. Water quality parameters, plankton dynamics in the estuary, and the reproductive and feeding biology of M. cephalus were also examined. The eggs, larvae, juveniles, and adults were identified using traditional morpho-meristic and DNA barcoding techniques. The results revealed a potential spawning ground of M. cephalus in the Karanja estuary. The results of reproductive biology also confirmed the occurrence of matured fishes during May–July. The abundance of eggs and larvae at the estuary’s mouth and the presence of juveniles and mature individuals of M. cephalus dominantly in the Karanja estuary from May to July infer the presence of a spawning site. It is also recorded that M. cephalus spawn in higher salinity (35 ppt) and seawater temperature (33 °C) where the hatching of offspring takes place successfully. This study emphasizes the significance of DNA barcoding in guiding routine monitoring surveys and demonstrates its usefulness when combined with these techniques in identifying fish spawning grounds. The study findings will serve as baseline information to develop effective conservation and management strategies and protect the ideal spawning stock.
... Moreover, morphological taxonomy of fish species is challenging and can provide erroneous identification status that would be deterrent towards the management of commercially important fish species (Fox et al., 2008). For instance, the morphological taxonomy of fish eggs and larvae have been in practice for the identification of spawning fish species (Harada et al., 2015), however, due to limited morphological development at this stage, the morphological taxonomic keys have been found mostly irrelevant in correct identification of fish specimens (Valentin et al., 2015). Thus, to complement the traditional morphology-based taxonomic identification, alternative techniques as molecular barcoding has been advocated (Ahern et al., 2018). ...
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Aim: In this study we characterized the fish Cynoglossus lingua collected from Chandipur Coast of Odisha using the mtDNA markers cytochrome c oxidase I (COI) and cytochrome b (Cyt-b) genes. Methodology: Samples of flat fish were collected from Balaramgadi fish landing center, Chandipur sea beach, Odisha coast. The collected specimens were initially identified based on the morphology using standard taxonomic keys. Molecular characterization was accomplished through PCR amplification of COI and Cyt-b genes. The sequences were subjected to BLAST analysis and aligned using Clustal W for homology mapping. Phylogenetic status was inferred by the neighbor-joining method using MEGA v7.0. Results: The taxonomic keys and the COI and Cyt-b gene sequences identified the specimens as C. lingua. The COI and Cyt-b gene sequences showed 98% and 99% sequence similarity with C. lingua isolate FMU.CIFRI.FF-7 COI gene (GenBank accession: MN889532.1) and the C. lingua isolate FM.CIFRI.CYTB.FF-7 Cyt-b gene (GenBank accession MT362541.1), respectively. Interpretation: The study supports the reliability of combining classical taxonomic approaches with molecular analysis using COI and Cyt-b genes for the identification of C. lingua. Key words: Cynoglossus lingua, COI gene, Cytochrome b gene, DNA barcoding, Molecular taxonomy
... Kematian ikan tidak ditemukan setelah umur 60 hari setelah menetas (hsm) (Gambar 5). Hal ini diduga berkaitan dengan perkembangan ikan cupang yang mencapai stadia juvenil pada umur 32 hsm (Valentin et al. 2013) dan 40 hsm (Poungcharean & Limpivadhana 2022). Karena itu setelah umur 60 hsm ikan lebih kuat dan diduga organ labirin juga telah berfungsi dengan baik. ...
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Betta splendens males have colours and shapes that are popular in the ornamental fish market compared to females. Because of that, betta cultivation can be done through male production using sex reversal technology in directing the sex development of fish to become male (masculinization). The natural ingredient that has been used to masculinize fish is honey. So the research objective was to examine the use of honey by immersing the embryo to masculinize betta. Analyzed masculinization success through characteristics of honey, percentage of male fish, egg hatching rate, mortality every 15 days, and survival at the end of rearing. The embryos used were 20 hours post-fertilization. The research treatment was immersion of betta embryos in honey solution (mL L-1) 5, 10, 15, 20, and 25. Soaking was carried out for 7 hours. The results showed that the honey used had 0.31% potassium and a pH of 4. In this study, the administration of honey did not affect the number of male betta. Giving honey 25 mL L-1 water produced 56.98 ± 4.58% of males, egg hatching rate 99.17 ± 1.67%, and survival at 90 days after hatching 79.89 ± 4.50%. Mortality occurs at the start of larval rearing. After the age of 60 days after hatching, there is no death in betta. The high values of egg hatching and survival rates indicate that honey is a natural material safe for masculinizing fish in mono-sex aquaculture.
... This lab space is lighted for roughly 12 h per day (from around 7:00-19:00) with white and yellow LEDs, and has windows to the outside of the building. Larval fish begin hatching between 29 and 44 h after fertilization 19,30,75 and begin swimming by 72 h after fertilization. 30 We use a glass transfer pipet daily to remove unfertilized eggs or embryos that are not developing or that are clearly malformed. ...
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Betta splendens, also called Siamese fighting fish or 'betta,' are a popular species in the fishkeeping hobby. Native to Southeast Asia, betta have been selectively bred for their fighting ability for hundreds of years, which has resulted in the species' characteristic male aggression. More recently, betta have been bred for a number of ornamental traits such as coloration, fin morphology, and body size. Betta have unique characteristics and an evolutionary history that make them a useful model for studies in the fields of behavior, endocrinology, neurobiology, genetics, development, and evolution. However, standard laboratory procedures for raising and keeping these fish are not well established, which has limited their use. Here, we briefly review the past and present use of betta in research, with a focus on their utility in behavioral, neurobiological, and evolutionary studies. We then describe effective husbandry practices for maintaining betta as a research colony.
... However, morphological taxonomy on fish eggs and larvae still remains challenging due to their small size, limited morphological development, scarcity of comparative material of known origin, and reliance on few characteristics such as egg diameter and presence of oil globules for species determination (Ahlstrom and Moser 1980;Moser 1996;Richards 2006;Richardson et al. 2007). Furthermore, some of the embryonic characters, such as pigmentations on the embryo, are only applicable in late stages of the egg development making earlier species determination difficult or impossible (Taylor et al. 2002;Valentin et al. 2013). In some cases, this have led to errors in the identification of commercially important fish species with consequences for management (Fox et al. 2005;Fox et al. 2008). ...
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The North Sea is an important spawning and nursery ground for many demersal and pelagic fishes whose spawning areas are largely overlapping in time and space. This makes ichthyoplankton visual identification from the various species particularly challenging. Despite historically intensive research in the area, detailed information on spawning sites and times for many taxa, are incomplete. To update and detail the mapping of fish spawning performance and distribution in the central and northern regions of the North Sea, the performance of a visual method and a molecular taxonomic approach used for taxonomic classification of ichthyoplankton was evaluated. Samples of fish eggs and larvae were collected regularly and in parallel at different latitudinal locations from the central to the northern North Sea, including a sample with 78 larvae used for direct comparison between both methods. A total of 5332 individuals were inspected and 36 different species were identified. The visual processing identified 89% of the collected larvae to species level, however, for the eggs the taxonomic resolution was lower with only 5% identified to species level. In comparison to visual identification , molecular barcoding gave higher precision of identification for larvae and especially for the eggs. For the larvae, 98% were assigned to species level, and for the eggs 94% were assigned to species level. We find that molecular barcoding is more effective and precise in taxonomic identification of both eggs and larvae to species level. However, visual identification is still needed to provide information on the developmental stages.
The fighting fish Betta splendens is a freshwater species from Thailand and other Southeast Asian countries. This fish has been domesticated for 1000 years and bred for fighting, various colours, body size and fin types for 600 years. It is one of the most important fish species cultured for the world ornamental fish market. This fish is easy to culture, highly fecund and displays great morphological diversity. Its biology has been studied for over 100 years. Recently, its compact genome and transcriptomes have been sequenced. Genome editing with CRISPR/cas9 has been applied to knock out genes in this fish. Its diverse phenotypes, including colours, colour patterns, fin types, and aggressive behaviour, are complementary to those of other model animals. Therefore, this fish could be the next important model organism for studying phenotypic variation and aggressive behaviour. In this paper, we synthesized knowledge about its aquaculture, biology, genetics, genomic tools, phenotypes and novel insights on phenotypic variation, sex determination and aggression. We hope that the information described in this paper will facilitate genetic studies on phenotypic variations in aquaculture and aggressive behaviour in other species, including humans.
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Paracheirodon axelrodi, ou cardinal tetra, é um peixe ornamental cuja ocorrência natural está restrita í­Â Bacia dos Rios Negro e Orinoco. É encontrado principalmente em igarapés de água preta e ácida com baixa velocidade de fluxo, e sua biologia reprodutiva é ainda pouco estudada. Neste trabalho são apresentados dados relativos a reprodução e desenvolvimento embrionário e larval em laboratório. A manipulação do ní­­vel da água (chuva artificial), pH e condutividade elétrica favoreceram a desova de Paracheirodon axelrodi. A espécie possui desova parcelada e os ovos são adesivos. Análises de oito fêmeas de cardinal tetra revelam fecundidade de 154 a 562 ovócitos, positivamente relacionada com o tamanho do peixe. O desenvolvimento dos ovócitos é do tipo sincrônico em mais de dois grupos, isto é, inúmeros ovócitos em diferentes fases de desenvolvimento são liberados em grupos. í­â‚¬ temperatura média de 26,0 ± 1,0°C, o desenvolvimento do ovo apresentou diferenciação embrionária rápida e perí­­odo larval longo. As larvas eclodiram, aproximadamente, 19 a 20 horas após a fertilização, com 2,9 ± 0,2 mm de comprimento total e corpo não pigmentado. No 5o dia de vida, as larvas apresentavam, em média, 4,1 ± 0,2 mm de comprimento total e nadadeiras em desenvolvimento. No 12 o dia de vida foram observados os primeiros pigmentos vermelhos na base da nadadeira anal, e uma faixa azul metálica lentamente formando-se e expandindo-se até a base da nadadeira adiposa. No 22o dia de vida, todas as nadadeiras estavam formadas, e os juvenis se assemelhavam aos adultos, com média de 11,0 ± 1,0 mm de comprimento total.
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Hoplias malabaricus, ou traíra é um peixe predador com ampla distribuição na América do Sul. Ε encontrado com frequência na Amazônia, porém sua biologia ainda é pouco estudada na região. Nesta nota apresentamos dados relativos a sua reprodução e ao início do seu desenvolvimento larval na várzea do rio Solimões. A traíra se reproduz durante todo o ano. As fêmeas têm fecundidade relativamente baixa, que varia conforme o tamanho do peixe. Os ovócitos maduros são grandes (média = 1,5 mm). Ovos fecundados foram encontrados em posturas nas margens dos lagos de várzea. As larvas eclodiram com 4,7 cm de comprimento e detalhes sobre seu desenvolvimento embrionário são apresentados.
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The embryonic and larval development of black skirt tetra, Gymnocorymbus ternetzi, are described under controlled laboratory conditions. In addition, major histomorphological changes and the allometric growth patterns during larval development have been described. The laboratory-reared broodstock, that is 1 year of age, were spawned. Hatching occurred 20–21 h after spawning at 24 ± 0.5°C. The cleavage was finished in 2 h and the early blastula stage occurred at 2:04 hours after spawning. The gastrulation started at 3:20 hours and 30% epiboly was observed at 3:34 hours after spawning. Eight-somite stage was observed at 08:33 hours. And embryonic developmental stage was completed at 21 h after spawning. The newly hatched larvae were 1442 ± 14.3 μm in mean total length (TL). The mouth opened at 3 days after hatching (DAH). The yolk sac had been totally absorbed and the larvae started to swim actively within 3–4 days. Notochord flexion began at 11 DAH. The metamorphosis was completed and the larvae transformed into juveniles at 32 DAH. In this paper, the full developmental sequence from egg to juvenile of G. ternetzi is described for the first time.
1 Introduction.- 2 The egg.- 3. Yolk (vitellus).- 4. Cortex and its alveoli.- 5. Egg envelope.- 6. Accessory structures of egg envelope.- 7. Micropyle.- 8. Sperm.- 9. Fertilization.- 10 Cleavage and formation of periblast.- 11 Gastrulation.- 12. Neurulation.- 13. Fate-maps.- 14. Kupffer's vesicle.- 15. Ectodermal derivatives.- 16. Hatching.- 17. Development of the eye.- 18. Mesodermal derivatives.- 19. Entoderm and its derivatives.- 20. Viviparity.- 21. Synthesis.- References.- Species Index.
At the end of gastrulation the outer layer of the embryo and yolksac consists of ectoderm1. In teleosts it is often referred to as ‘epiblast’ (to contrast it with the inner layer, the ‘hypoblast’). The ectoderm covering the embryonic shield is greatly thickened compared with the extraembryonic area and it undergoes neurulation as described in Chapter 12. Eventually the whole embryo and the yolksac become covered by the one-layered epidermis topped by a flattened layer, the EVL or periderm (Chapter 11).
Embryology (now called developmental biology) of teleosts was intensively analyzed from the beginning of the 19th century onwards. It is, therefore, most intriguing if not to say incomprehensible, that there are textbooks and even atlases of the development of amphibia, birds and various mammals widely available, but so far none on teleost fish. This was the compelling reason for the writing of this book.
Eggs of 23 Characiformes and eight Siluriformes, belonging to nine families with diverse reproductive behaviour, were ultrastructurally analysed. The migratory species exhibited non-adhesive eggs, whereas, most of the sedentary species presented some degree of egg adhesiveness. Among the Characiformes, non-adhesive eggs showed zona radiata with pore-canals or a fibrillar net at the surface; weakly adhesive eggs presented only zona radiata with pore-canals while adhesive eggs exhibited zona radiata with apparatus like globules, filaments, villi or honeycomb-like pores depending on the systematic group. The ' jelly ' coat is strongly related to the Siluriformes eggs apparently without relationship with adhesiveness. A micropylar disc was present in adhesive eggs of a few species of both Characiformes and Siluriformes. Some patterns were characteristic of the animal pole, others of the vegetal pole, and others were common to both poles. The radial ridges converging to the micropyle in Astyanax bimaculatus lacustris appear to be related to fertilization. In general, egg surface structures in the Characiformes varied according to the genus, whereas all Siluriformes showed a similar egg surface pattern, regardless of the group analysed. Multivariate analysis allowed the identification of eight clusters among the Characiformes and three among the Siluriformes showing relationships between reproductive style and egg characteristics. It is suggested that egg surface and adhesiveness may be related to reproductive patterns and to phylogenesis.