EurAsian Journal of BioSciences

Eurasia J Biosci 8, 38-50 (2014)

http://dx.doi.org/10.5053/ejobios.2014.8.0.4

Embryonic and larval development of the

suckermouth sailfin catfish Pterygoplichthys pardalis
from Marikina River, Philippines
Joycelyn Cagatin Jumawan1*, Annabelle Aliga Herrera2, Benjamin Vallejo Jr3
1Biology Department, Caraga State University, Butuan City, Philippines
2Institute of Biology, University of the Philippines-Diliman, Diliman, Quezon City, Philippines
3Institute of Environmental Science and Meteorology, University of the Philippines-Diliman, Diliman,

Quezon City, Philippines

*Corresponding author: joycejumawan@gmail.com

Abstract
Background: There is little information about the early development of this invasive fish species in
order to understand its early life history and developmental strategies towards invasion.
Material and Methods: Female Pterygoplichthys pardalis were induced to spawn using human
chorionic gonadotropin (HCG) so as to study the developmental stages from fertilization until yolk
resorption.
Results: The females subjected to a single dose of HCG responded positively to treatment (97%)
with higher fertilization success (88%) compared to the untreated females (21%). Nonetheless, the
HCG-induced fertilized eggs had a low hatching success (49%), while from the free-living embryos
successfully hatched, a high number (90%) survived to become juveniles. Embryonic development
in P. pardalis was completed 168 h and 30 min after fertilization, with the total yolk resorption
completed on the 8th day post hatching, during which the suckermouth gradually shifted from
rostral to ventral position to commence the loricariid algae-scraping feeding mode.
Conclusions: Pterygoplichthys pardalis does not undergo a true larval metamorphosis between the
free-living embryo and the juvenile stage and a definitive adult phenotype is developed directly.
These results provided basic, yet essential information on the early developmental features of this
invasive species whose spawning and early developmental strategies were difficult to observe in the
field. Implications of some ontogenetic features in this species with regards to invasion are also
discussed.
Keywords: Development, embryogenesis, invasion, larvae, morphology, Pterygoplichthys pardalis.
Jumawan JC, Herrera AA, Vallejo JrB (2014) Embryonic and larval development of the suckermouth
sailfin catfish Pterygoplichthys pardalis from Marikina River, Philippines. Eurasia J Biosci 8: 38-50.
http://dx.doi.org/10.5053/ejobios.2014.8.0.4

EurAsian Journal of BioSciences

Pterygoplichthys pardalis (Castelnau, 1855) is a

native of South America (Weber 2003), but has been
Siluriformes exhibit diverse reproductive introduced to the Philippines through aquarium
strategies and most studies are just focused on late trade. It has invaded many freshwater systems of
embryonic and larval development (Adriaens and the country along with another hypostomine
Vandewalle 2003). Nonetheless, knowledge of the loricariid, Ptergoplichthys disjunctivus. These two
crucial phases of the life history of invasive species is species, popularly known in the country as janitor
vital in order to understand their developmental fish, are not regarded as important commercial
strategies and identify the advantageous fishes because of their hard body armour, very little
ontogenetic
features
for
invasion
and meat, propensity to compete for food resources,
establishment. The management of invasive alien and their potential to bioaccumulate heavy metals in
fish species with known high fecundity potential and polluted environments (Chavez et al. 2006, Lam and
high success rate of establishment in invaded Su 2009, Jumawan et al. 2010a). Nonetheless, the
environments requires knowledge of its early life hardy nature of this genus, its capacity to down
stages, such as the timing of embryogenesis and
organogenesis and the shift from endogenous to
Received: January 2014
exogenous feeding (Godinho et al. 2003).
Received in revised form: April 2014
Accepted: April 2014
The armoured suckermouth sailfin catfish
INTRODUCTION

Printed: May 2014

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EurAsian Journal of BioSciences 8: 38-50 (2014)

regulate metabolism during periods of scarcity of

food (German et al. 2010), its tolerance to poor
water conditions and its ability to breathe air under
hypoxic water conditions (Armbruster 1998) enabled
this fish to invade and successfully establish itself
even in disturbed freshwater systems.
The seasonality of reproduction and the gonad
features of the Pterygoplichthys spp. population in
Marikina River has been previously described,
showing the females to be highly fecund and the
oocytes exhibiting features associated with parental
care (Jumawan et al. 2010b). However, the burrowspawning and nest-guarding nature involved during
the critical period of embryogenesis in these fishes
was found difficult to replicate under laboratory
conditions. To date, there are no available studies
describing the early development of P. pardalis and
P. disjunctivus in their original habitat for comparison
with species thriving in non-native environments.
The present study will serve as a baseline reference
of the early embryogenesis, larval development and
organogenesis of the invasive P. pardalis through invitro fertilization.

Jumawan et al.

(Merck, Germany) bath for 4 min. The female P.

pardalis received an intramuscular injection of
human chorionic gonadotrophin (HCG) (Argent
Chemicals, Philippines) at 4 IU/g body weight
(injection volume: 1L/g BW). The hormone-injected
females were then placed separately in 1 m x 0.5 m
plastic tanks containing de-chlorinated tap water to
a depth of 0.4 m each. Females untreated with HCG
served as controls and were placed in a separate
tank.
Approximately 14-18 h after HCG administration,
the females were checked for ovulation by applying
pressure to the abdomen to confirm ovulation. Eggs
from ovulated females were then stripped in a dry
plastic basin. At about the same time, males were
anesthetized, sacrificed by a sharp blow to the head
and had their testes removed. Milt were collected
after maceration of the testes and then immediately
diluted with 0.9% NaCl to obtain milt solution. The
milt solution was poured into a bowl containing the
stripped eggs and mixed for 30-60 sec using a
feather. Approximately 5 mL of tap water was added
to the bowl to ensure fertilization. After 2 min of
gentle stirring, the fertilized eggs were transferred
to a plastic strainer and rinsed with running water
MATERIALS AND METHODS
for about 1 min to remove excess milt. Fertilized
eggs were immediately transferred to a 45x30x30
Strip method for artificial fertilization
The artificial breeding protocol of P. pardalis cm aerated plastic aquarium for incubation. The
adapted the procedure by Tan-Fermin et al. (2008) aquaria were provided with partial shade by using a
with some modifications. All experiments were 1x1 m black, plastic polyethylene bag to simulate
carried out in triplicates. Collection and the darkened burrows in the field. The eggs were
experimentation were performed during the peak examined 10-15 min after gamete mixing to check
spawning months (July to September) of the for blastodisc formation. Unfertilized eggs were
Pterygoplichthys spp. population in Marikina River. carefully removed from the aquaria using fine
Because of the absence of defined sexual forceps.
dimorphism in this fish, large and mature P. pardalis
Initial observations showed that the fertilized
weighing 350-500 g were collected. Females were eggs had a low hatching success rate when
selected based on their full and heavy body, gravid incubated in de-chlorinated tap water (10-15%) or
abdomen and reddish, swollen vent. To identify the rain water (10-20%), but had an improved hatching
sex of the fish, pressure on the abdomen to extrude rate when using a mix of natural river and tap water.
oocytes and the use of cannula were also attempted
Hence, the subsequent trials used a mix of aerated
for most samples. Males were selected based on a
Marikina River water and tap water as an incubation
streamlined body and flat abdomen. A total of 18
medium at 1:1 ratio. Prior to mixing the water
females and 18 males were utilized for this study.
medium, river water and tap water were analyzed
Prior to hormonal induction, all females were
anesthetized in a 200 ppm 2-phenoxyethanol for hardness (CaCO3/L), chloride (mg/L), calcium

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EurAsian Journal of BioSciences 8: 38-50 (2014)

(mg/L) and magnesium (mg/L) levels. The physical

and chemical parameters of the mixed water in the
aquaria, such as temperature (C), pH, salinity and
dissolved oxygen (D.O.), were recorded throughout
the experiment.
Embryogenesis, larval ontogenesis and
biometry
For each of the 3 replicate plastic aquaria, twenty
developing eggs were observed at 10-30 min
intervals until completion of cleavage, 3 times per
day until hatching. Hatchlings were documented
twice daily, until the yolk was fully resorbed. The
developmental stages were divided into embryo and
free-living embryo stages. The embryonic stage
occurs inside the chorion and ends in hatching. The
free-living embryo stage is characterized by the
nutritive contribution of the yolk sac and the stage
ends when the free-living embryo becomes capable
of exogenous feeding after the yolk has been
consumed (Geerinckx et al. 2008).
Three subsamples of fertilized eggs were
collected daily until yolk resorption and were fixed
in Bouins fluid for histological purposes.
Additionally, three more fertilized eggs and
hatchlings were collected until yolk resorption and
were directly fixed in 4% neutral formaldehyde for
biometry using a digital caliper (accuracy 0.001 mm;
Control Company, USA), following the parameters
described in the study of Guimaraes-Cruz et al.
(2009) (Fig. 1).
The fertilization rate (number of eggs with
blastodisc/total oocytes x 100), hatching rate
(number of hatched eggs/number of fertilized eggs
x 100) and survival rate (number of surviving
juveniles/total number of hatched embryos x 100)
were recorded. Three replicate runs from
fertilization to hatching were conducted.
Analysis of variance (ANOVA) was used to
compare the mean values of the morphometric
variables according to each stage of larval
development, and between oocyte mean diameter
values for HCG-induced and non-HCG-induced
samples. All tests were conducted in a 0.05
significance level using Graphpad Prism 5.

RESULTS
Description of fertilized eggs
The adult female P. pardalis were administered
HCG a day after samples were obtained from the
field. Difficulty in distinguishing between males and
females was noted due to lack of defined sexual
dimorphism as well as difficulty in extruding oocytes
and milt from the vent after the application of
considerable abdominal pressure and cannulation.
Table 1 shows the physico-chemical parameters of
the water used for embryo incubation. Oocyte
extrusion was performed 18 h after the exposure to
HCG. Approximately 200-250 oocytes were extruded
from the ovaries of a single female exposed to HCG
due to difficulty of handling and applying pressure in
the abdomen of the fish. The females injected once
with HCG responded positively to the treatment
(97% success rate) and hydrated, producing nearly
uniform sized (2-3 mm), transparent-yellow oocytes
after the ovary was stripped. In contrast, the oocytes
from females not exposed to HCG were mostly
opaque yellow with occasional pre-vitellogenic
oocytes (<1 mm) along with larger vitellogenic
oocytes. Adhesion of oocytes was observed
immediately after any excess milt was washed off.
The HCG-injected females had a higher fertilization
success (88.3%) compared to the untreated females
(20.9%). The fertilized eggs from HCG-injected
females had low hatching success (48.6%), with
mortalities during early stage somitogenesis.
From the successfully hatched embryos, a high
number survived to become free-living embryos up
to the termination of experiments on the 8th day post
hatching (8 dph) (Table 2). The diameter of the eggs
did not differ significantly between the two
treatment groups. No change in the diameter of the
fertilized oocytes (3.20.23 mm) was also observed
from the onset of fertilization until the pre-hatching
stage, although a very small perivitelline space was
formed surrounding P. pardalis eggs a few minutes
after fertilization.
Embryonic development
The observations of P. pardalis were divided into
two periods: (1) before hatching (embryo), and (2)
between hatching and yolk sac depletion (free-living

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Jumawan et al.

Fig. 1. Lateral view of a P. pardalis free-living embryo (4 dph;

14.2 mm TL).
TL: Total length; SL: Standard length; PAD: Pre-anal distance; HL:
Head length; HH: Head height; SNL: Snout length; ED: Eye diameter;
YSL: Yolk sac length; YSH: Yolk sac height, BH: Body height.

Fig. 3. Stages of embryonic and larval development in P.

pardalis.
(a) Late gastrula: formation of the embryonic shield (es); (b)
Somites development (black arrow); (c) Late Neurula stage; f:
Forebrain; m: Midbrain; h: Hindbrain, oc: Otic capsule; (d-e) Rostral
location of suckermouth (*), Note egg sac removed; (f) P. pardalis at
6 dpf; (g) Newly hatched embryo (7 pf); (h) Note the ventral
position of the suckermouth in (g).
Scale bar: 1 mm.

Fig. 2. Stages of embryonic development in P. pardalis.

(a) Early stage fertilized egg with distinct invagination at the animal
pole before blastodisc formation; (b) Blastodisc stage; (c) Cleavage
at 2-cell stage; (d) Cleavage at 4-cell stage; (e) Cleavage at 8-cell
stage; (f) Cleavage at 64 blastomeres; (g) High blastula stage; (h)
Low blastula; (i) Early gastrula.
Scale bar: 1 mm.

embryo). The main events in embryogenesis and

their respective times of observation are
summarized in Table 3 and Figs. 2-4. The fertilized
eggs of P. pardalis show meroblastic cleavage in
which the blastoderm is restricted to a small area at
the animal pole. The first segmentation that divided
the blastodisc into two blastomeres occurred within
41

5 to 15 min after fertilization. Subsequent

successive cleavage until 64 blastomeres was
observed until it was completed after 5 h with
blastomeres very much decreased in size.
Blastomeres very fine in appearance eventually
flattened at the animal pole at 14-15 h post
fertilization. The spread of the blastoderm was
evident, covering the yolk with the embryo body
becoming more elongated, while the head and tail
ends of the embryo can be clearly seen at 20 h.
Finally, full yolk invasion and closure of the
blastopore was observed at 24 h.
Differentiation of the embryo
Pre-hatching organogenesis in P. pardalis
commenced with the formation of the notochord
and observations of the cranial-caudal portions of
the embryo (29 h), as well as the first somites and
the optic vesicles (36 h). Somitogenesis was
observed starting 2 days post fertilization (2 dpf)

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EurAsian Journal of BioSciences 8: 38-50 (2014)

Table 1. Some physico-chemical features of the water medium used in this study.

*Data analyzed only once, through the Research and Analyti cal Servi ces Laboratory (RASL), Natural Sciences Research Institute (NSRI);
Remaining data monitored once in every 3 days for 15 days. Values are listed as meansS.E.

Table 2. Percent (%) response of P. pardalis to HCG-injection at 24C

*P<0.001; Values are listed as meansS.E.

and subsequent somite formation enabled tail

movement, although the embryo was largely
confined within the perivitelline space. No
corresponding somite formation for each hour
during the onset of embryogenesis in P. pardalis was
noted because of the difficulty in turning the
strongly adhesive eggs to locate the somites
without causing mechanical injury to the developing
embryos. The time when the pericardial cavity was
formed was not determined; however, the pulsating
heart and blood circulation was visible as a very faint
stream of capillary network in the pericardial cavity
at 36 h after fertilization. Ectodermal thickening to
form the lens of the eyes was observed starting at
36 h, with the eye lens fully formed at 49 to 50 h post
fertilization. The suckermouth was rostrally
positioned inside the membrane (Fig. 3d-e, Fig. 4a).
Tail movement (72 h post fertilization) was
observed before the formation of the vitelline
circulatory system. Movement in the gill cavity and
blood circulation in the gill arches was noted
beginning at 5 dpf. Hatching was observed 167-168

h post fertilization. The caudal tail region was

observed detaching from the yolk mass with a
subsequent increase in body movement, causing the
rupture of the chorion and the emergence of the
embryo from the capsule.
Free-living embryo development
Details of development and average body
measurements are reflected in Tables 4 and 5.
Histologic sections of larvae at different days post
hatching are shown in Figs. 5-7.
Hatchling
Newly hatched, free-living embryos or eleutherembryos (Balon 1986) have a mean total length of
7.860.12 mm, still containing a large amount of yolk
(Fig. 5a). The newly hatched larvae had already
ventrally located the suckermouth and maxillary
barbels and were able to attach to the sides of the
glass substrata with their suckermouth, with water
inflow for the sucking action passing through the
furrows of the maxillary barbels, and attachment
sometimes assisted by body and tail movements
(Fig. 3h). Embryos initially had a rostrally located

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Jumawan et al.

Table 3. Main events of the embryonic development of P. pardalis and their respective times (mean) after HCG-induced
fertilization at 24C. n: 192 developing eggs/stage.

mouth, which gradually shifted into the ventral

location during hatching. Dorsal and caudal fins were
observed. The bodies of the newly hatched freeliving embryos were transparent, except for the
onset of dendritic pigmentations initially found
interspersed finely on the head and on the edges of
the snout. The eye diameter was small (0.510.02
mm). The dorsal and caudal fins were well
recognizable. Serial sections of the digestive system
during the day of hatching showed that the gut is
tubular and closed at both ends, while the intestine
is composed of simple cubic epithelium. The first
outlines of the gill arches supported by blood
vessels were observed in the gill cavity. A tubular
heart was observed.
3-d old free- living embryo
A continued reduction of yolk sac and an increase
in body length (TL 12.410.05 mm) were observed
along with increasing pigmentation in the retina and
increasing complexity of the gills. Increasing
dendritic chromatophore pigmentation was
observed in the outlines of the head and dorsal side
43

of the body. The digestive system was characterized

by a simple striated border in the intestine (Fig. 5f).
The cranial kidney was now well developed with
readily recognizable glomeruli tufts within the
network of reticular fibers (Fig. 5e). The spleen could
be observed on the left lateral-dorsal-right lateral
part of the borderline between the gut and the
kidney. The liver could be observed ventrally located
in the cranial region with the hepatic parenchyma
very homogenous in appearance.
5-d old free-living embryo
The average length of a 5-day old free-living
embryo was about 14.150.04 mm TL with the yolk
sac length becoming gradually reduced (3.220.03
mm). Increasing pigmentation all throughout the
body was observed with dendritic chromatophores
reaching the lateral region caudally to the pectoral
fin (Fig. 4c-d). The yolk was visible, although less
compact near the digestive system. The intestines
became looped and lengthened, with the intestinal
epithelium exhibiting a complex columnar
arrangement of mucous and goblet cells. Intestinal

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EurAsian Journal of BioSciences 8: 38-50 (2014)

Fig. 4. Free-swimming embryo development in P. pardalis.

(a) Embryo with yolk sac removed. Note the rostral position of the
mouth; (b) Development at 3 dph; (c-d) Development at 5 dph; Note
the capacity of the ventrally positioned mouth for attachment in
the glass substrata in; (c) Body pigmentation in (d); (e)
Pigmentation in the ventral position of the 7 dph free-living
embryo.

content (residue) was observed in this stage.

Nephric ducts in the cranial kidney were very visible;
the encephalon was compact with associated cells
(Fig. 6e). The heart was compartmentalized with a
very visible atrium (Fig. 6f).
7-d old free-living embryo
The average length of 7-day old free-living
embryo was about 14.840.34 mm TL with the yolk
sac length becoming gradually reduced (2.320.09
mm). Increased pigmentation all throughout the
body including the ventral part previously occupied
by the yolk sac (Fig. 4e). Full pigmentation of the
eyes observed (Fig. 7c). Increased use of the
suckermouth to anchor body in the glass aquaria was
observed in larvae. The gills were more developed
with elongated filaments and gill lamellae with
intense vascularization. Gas bladder was observed
with simple squamous epithelium (Fig. 7d). The heart
presented two compartments. Small intestine
mucosa can be seen, including microvilli, columnar
epithelium and muscularis mucosa. The head kidney

Fig. 5. Organogenesis in P. pardalis.

(a) Newly hatched free-living embryo; medial portion; (b) 2dph,
cranial portion; (c) 3 dph, Retina (*) and its layers; (d) 3 dph, cranial
portion; (e) 3 dph, Digestive system; (f) 3 dph, intestine with simple
striated border. N: Nostrils; E: Encephalon; H: Heart; B: Barbells; CK:
Cranial kidney; GB: Gas bladder; I: intestine; y: Yolk; G: Ganglionar
cell; IN: Inner nuclear layer; ON: Outer nuclear layer; YG: ocular
globe; GA: gill arches.
Scale bars: a,d,e: 200 m; c,f: 20 m; b: 40 m.

had visible renal tubules and extensive hematopoetic tissue.

8-d old free-living embryo
The average total length of free-living embryo
was 6.950.20 mm TL and the yolk was fully
resorbed. The body had become opaque with the
accumulation of pigments all throughout its surface
as chromatophores became more numerous and
darker, maintaining the same distribution pattern.
Undifferentiated gonad with primordial germ cells
was observed (Fig. 7e), while a true larval stage
(Balon 1986, 1999) was not observed. P. pardalis
underwent a direct transition from a free-living
embryo with large yolk into a juvenile without
undergoing a true larval stage when the yolk was
fully consumed at 8 dph. Except for the absence of
hardened armour covering the body and the
abdominal pattern distinct for P. pardalis, an adultlike appearance was observed at the moment the
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Jumawan et al.

Table 4. Main morphologic events occurring during the development of P. pardalis from hatching until yolk resorption (TL,
mm).

Table 5. Body measurements* (mm) of P. pardalis from hatching until yolk resorption.

TL: Total length; SL: Standard length; PAD: Pre-anal distance; HL: Head length; HH: Head height; SNL: Snout length; ED: Eye diameter; YSH:
Yolk sac height; YSL: Yolk sac length; BH: Body height. Values are listed as meansS.E.; n= 212

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EurAsian Journal of BioSciences 8: 38-50 (2014)

Fig. 7. Organogenesis in P. pardalis.

Fig. 6. Organogenesis in P. pardalis.
(a) 3 dph, cranial portion; (b) 3 dph, spleen and kidney; (c) 4 dph,
dorsal nervous tube; (d) 4 dph, digestive system; (e) 5 dph,
encephalon; (f) 5 dph, heart. E: Encephalon; H: Heart; B: Barbells; K:
Kidney; I: Intestine; y: Yolk; G: Ganglionar cell; IN: Inner nuclear
layer; ON: Outer nuclear layer; YG: Ocular globe; GA: Gill arches; S:
Stomach; SP: Spleen, N: Notochord; M: Myomeres; NC:
Neurocranium.
Scale bars: a: 200 m; b,d: 40 m; c,e,f: 40 m.

(a) 6 dph; gas bladder and kidney; (b) 6dph, intestine; (c) 7 dph,
cranial portion; (d) 7 dph, digestive system; (e) 8 dph,
undifferentiated gonad; (f) 8 dph, segmentation of the notochord.
B: Barbels; GB: Gas bladder; CK: Cranial kidney; I: Intestine; Y: Yolk.
G: Ganglionar cell; IN: Inner nuclear layer. ON: Outer nuclear layer;
YG: Ocular globe; GA: Gill arches; H: Heart; PGC: Primordial germ
cell; N: Notochord; DF: Dorsal fin; M: Muscle.
Scale bars: c,d,f: 200 m; b,e: 20 m.

stages development of P. pardalis, a highly invasive

loricariid, whose early life history has not been
last yolk was consumed at 8 dph.
Biometric parameters registered a gradual studied so far.
The reproduction and spawning behavior of the
increase in all values except for the decrease in the
yolk sac length and height as the free-living embryo janitor fish P. pardalis is difficult to observe, as they
neared and completed the yolk resorption are known to spawn in burrows with males guarding
externally (Table 5). A full yolk resorption in the the fertilized clutch. What is surprising in this
juvenile P. pardalis was observed 336 h or 14 dpf at experiment, however, is the tendency of the eggs to
24C. All the oocytes observed for fertilization until hatch well in river water characterized to be hardy,
yolk resorption in juveniles developed synchrowith higher calcium and magnesium levels compared
nously.
with tap water.
The water quality of key areas along Marikina
DISCUSSION
River where spawning colonies of P. pardalis are
abundant showed that the river is highly eutrophic
Most invasive loricariids do not reproduce
and turbid, and had overall high conductivity,
spontaneously when reared under laboratory
nitrate, ammonia and phosphate levels. Fertilized
conditions (Alfaro et al. 2008). The present study
clutches from the river also have a higher hatching
provides baseline information regarding the early
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rate if incubated in river water. It is not clear how the

nest guarding nature of the males for this species
actually influences the successful hatching because
of the difficulty of observing this process in the field.
Makeshift and darkened plastic aquaria to replicate
burrows in the field in the preliminaries of this
experiment failed to encourage spawning under
laboratory conditions. The low success rate in
hatching for P. pardalis under artificially-induced
spawning conditions may be attributed to the
absence of parental care during the experiment or
to the preference of eggs for river water
environments.
Eggs of P. pardalis are highly adhesive, allowing
the formation of tight clutches of eggs. Teleost eggs
can be non-adhesive, weakly adhesive or strongly
adhesive. Rizzo et al. (2002) point out that adhesive
eggs are often large, laid in smaller numbers and
associated with the sedentary nature of the species
or with parental care. The zona radiata of
Pterygoplichthys spp. in Marikina River was thin
(mean 4.56 m), while its granulosa layer was thick
(mean 2.81 m) (Jumawan and Herrera 2014). A thick
granulosa layer may provide better adhesion of
eggs, while the thin zona radiata may be
compensated by the nest guarding nature of the
males of this species, as was the nature of some nest
guarding loricariids (Suzuki et al. 2000).
It was apparent in the results that although
artificially fertilized eggs had a low hatching rate, all
hatched P. pardalis free-living embryos successfully
survived and had resorbed their yolk at 8 dph. A
large endogenous supply of yolk nutrients enhances
survival during the period when feeding structures
are still developing (Geerinckx et al. 2008), while also
enabling the free-living embryo to avoid an
intermediate larval stage and the cost of
metamorphosis since a definitive adult phenotype
was developed directly (Balon 1986).
Although hardened armour covering the body
was not observed when the last yolk was consumed
at 8 dph, hardening of the head and dorsal structure
towards development of the armoured covering of
the body was eventually observed 30 days post yolk
resorption in juveniles (data not shown).

47

Jumawan et al.

Types of feeding (exogenous, endogenous),

nutrient supply (altricial, precocial) and life history
models (indirect, direct development) were used as
basis in the description of embryos (Balon 1986,
1999). Embryos with indirect development are a
consequence of poor vitellogenesis and depend
entirely on an endogenous nutrient supply, as eggs
are altricial in nature (Balon 1986). The short embryo
period in this type of development appears to be
extended by the larva period that feeds
exogenously prior to the formation of the definitive
phenotype. Fish taxa characterized by direct
development have a prolonged embryo period due
to a large endogenous supply as eggs are precocial
(large amount of yolk) that ultimately enables the
embryo to develop directly into a definitive
phenotype (Balon 1986). Embryos develop for
longer but directly into juveniles that are able to
compete in the adult habitat (Balon 1999). Direct
development occurs more frequently in the
reproductive guilds of guarders and bearers as these
fish groups exhibit parental care such as site
selection, egg deposition and nest guarding (Balon
1986). A true larva requires some tissues and
structures very different to those in the definitive
organism, and so, has to be remodeled through the
process of metamorphosis (Balon 1999). Fish species
with direct development lack the necessary cost of
forming temporary organs through metamorphosis.
The large yolk (mean 3.3 mm) in P. pardalis is an
advantage in this context, as it requires none or little
external nutrients to develop into a definitive
phenotype. It has been proposed that the larger and
more advanced an individual at the onset of
exogenous feeding, the better its chances of
surviving (Balon 1999). This scenario improves
competitiveness in P. pardalis even in its early stages
and could be an advantage for invasion (Balon 1986).
The absence of a true larval stage in P. pardalis is
similar to most loricariids (Geerinckx et al. 2008). It is
however important to note that a slight variation in
measurements may be caused by the fixative used in
this study (4% neutral formaldehyde), as fixatives
may have some dehydrating effects contrary to the
measurement of fresh samples.
The eggs of P. pardalis contain a large amount of

EurAsian Journal of BioSciences 8: 38-50 (2014)

evenly distributed yolk, hence, is classified as

telolecitic (Ribeiro et al. 1995, Marques et al. 2008).
P. pardalis has a meroblastic cleavage pattern
restricted to the animal pole, which is common in
teleosts. Blastopore closure occurred 24 h after
fertilization, indicating fertilization success. The
embryonic development of P. pardalis lasted 7 days
(168 h, 30 min), which is long compared with the
duration of embryogenesis in other siluriforms: 45 h,
50 min at 24C in R. aspera (Perini et al. 2009), 21 h,
20 min at 23C in Pimelodus maculates (Luz et al.
2001) and 18 h at 27C in P. corruscans (Marques et
al. 2008).
Long embryonic periods are known to be
associated with non-migratory species having large
eggs and with those that display parental care
(Sargent et al. 1987), features that both fit the
natural history of P. pardalis. Invasive species
exhibiting parental care may be considered an
advantage in the context of ensuring egg survival
until the juvenile period. By females precisely
selecting a safe and protected site for egg
deposition and males guarding the nest, the
progress of the early developmental stages in P.
pardalis becomes ensured. This protection makes
the early stages difficult to observe in the wild as
they are well hidden from predators and other
threats, emerging only when post-yolk sac juveniles
are capable of feeding and the definitive phenotype
ensures the adult form.
A pigmentation of the retina observed early
during hatching in P. pardalis could be associated
with the need to develop a functional visual system
before the first feeding, typical in some fishes (Hall
et al. 2004). The undifferentiated gonad located
between the cranial kidney and the digestive tract in
P. pardalis already contains a cluster of primordial
germ cells (PGCs) at 8 dph. PGCs in the
undifferentiated gonad were also observed at 5 dph
in R. aspera and at 13 dpf in Pimephales promelas
(Uguz 2008).
The shift of the suckermouth from rostral to
ventral position long before the hatching stage is a
common feature for most loricariids. This shift is of
more advantage to P. pardalis as newly hatched freeliving embryos already had a ventrally flattened

Jumawan et al.

mouth despite the large yolk sac during hatching. On

the contrary, A. cf triradiatus, a relative loricariid,
was noted to have a rostro-ventrally positioned
suckermouth during hatching, but had to wait for 24 days for the yolk sac to resorb for the transition to
a ventrally flattened mouth (Geerinckx et al. 2008).
The newly hatched P. pardalis in this study was able
to attach immediately to the substrate with their
suckermouth. This observation was also reported for
Sturisoma aureum (Riehl and Patzner 1991) and A. cf.
triradiatus (Geerinckx et al. 2008). This feature may
be essentially an advantage for loricariid species
where hatchlings leave the shelter immediately
(Suzuki et al. 2000). In the case of an invasive species
such as P. pardalis, accidental release of eggs and
juveniles may result in assured higher survival rates
in the wild.
The SL of P. pardalis during the time of hatching
(7.860.12 mm) and the large yolk sac during the
same time fall within the size range for most
loricariids (6-8 mm) (Riehl and Patzner 1991,
Nakatani et al. 2001, Geerinckx et al. 2008, Perini et
al. 2009). The morphometric development in P.
pardalis, which reflected a gradual increase in all
parameters with decreasing yolk size, is also
observed in A. cf triradiatus (Geerinckx et al. 2008), R.
aspera (Perini et al. 2009) and Lophiosilurus alexandri
(Guimaraes-Cruz et al. 2009), all of which exhibited a
high degree of allometric growth. Geerinckx et al.
(2008) hypothesized that loricariid hatchlings often
exhibit rapid allometric growth in the snout and
remarkable lip transformation because it is a
necessity and advantageous for the suckermouth to
attach to the substratum for scraping and sucking
food.
In conclusion, this study pointed out several
baseline features of the early life strategies in P.
pardalis that may be of advantage to its biotic spread
potential: (1). The propensity of the embryo to thrive
in polluted water; (2) the adhesiveness of the eggs
allowing for higher hatching success rate, further
contributing to the nest guarding feature of males;
(3) the already ventrally positioned suckermouth
and sucking capacity of the free living embryo,
allowing for higher survival potential once left out of
parental care due to its substratum scraping capacity
48

Jumawan et al.

EurAsian Journal of BioSciences 8: 38-50 (2014)

for food and attachment; and (4) the absence of true

larval metamorphosis between the free-living
embryo and the juvenile stage due to the large
supply of yolk. Information on the early life history
strategies of P. pardalis from its original habitat
would be essential for comparison with the nonnative counterparts in order to determine possible
developmental plasticity of the fish in invaded water
systems.

ACKNOWLEDGEMENTS
JC Jumawan is grateful to the Commission on
Higher Education- Science and Engineering Grants
(CHED-SEGS) for the dissertation grant and to the
Philippine Kidney Transplant Institute (PKDF)
Histology laboratory for the histological
preparations. The authors thank Drs PJ Denusta and
LMB Garcia for the technical help in the in-vitro
experiment.

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