Behavioural Processes 64 (2003) 23–29
Type of intruder and reproductive phase influence male
territorial defence in wild-caught Siamese fighting fish
Mullica Jaroensutasinee∗ , Krisanadej Jaroensutasinee
Institute of Science, Walailak University, 222 Thasala District, Nakhon Si Thammarat 80160, Thailand
Received 1 October 2002; received in revised form 17 February 2003; accepted 14 April 2003
Abstract
This study investigated how parental care increases with offspring age by examining the level of male aggressiveness toward
potential intruders during egg guarding in a natural population of Siamese fighting fish (Betta splendens Regan). The degree
of aggressiveness was measured at two reproductive phases in response to three types of intruders: male, female and females
that have laid eggs. The nest-holding males became more aggressive after their eggs hatched than after the eggs were laid.
Nest-holding males displayed gill cover erection, biting, tail beating and attacking at the highest rate towards male intruders,
intermediate towards female intruders and the least aggressive towards females, which have laid eggs.
© 2003 Elsevier B.V. All rights reserved.
Keywords: Territorial defence; Egg guarding; Aggressive behaviour; Siamese fighting fish; Betta splendens
1. Introduction
To understand the evolution of mating systems, it
is necessary to determine how parental care increases
offspring survival. Many theoretical studies have examined the parental effort and the optimisation of
fecundity with respect to adult mortality (Williams,
1966; Pianka, 1976; Pressley, 1976, 1981; Ricklefs,
1977). These models predict that during the period of
parental care, increases in the cost of parental effort
will be favoured when (1) the average reproductive
value of the young increases, (2) the number of young
under parental care increases, and (3) the parent’s reproductive value decreases.
In an animal with a restricted reproductive season,
the probability of an animal to breed again decreases
∗ Corresponding author. Tel.: +66-075-672005;
fax: +66-075-672004.
E-mail address: jmullica@wu.ac.th (M. Jaroensutasinee).
with the time spent caring for a particular brood.
Therefore, investments should increase with offspring
age (Weatherhead, 1982; Sargent and Gross, 1985;
Curio, 1987). There are many studies of changes
in parental care over the breeding cycle and in response to different intruders in birds (e.g. reviewed
in Montgomerie and Weatherhead, 1988; Andersson
et al., 1980; Brunton, 1990; Rytkönen et al., 1990;
Pavel and Bureš, 2001). However, a few studies have
been investigated in fishes including three-spined
stickleback (Gasterosteus aculeatus; van Iersel, 1953),
smallmouth bass (Micropterus dolomieui; Ridgway,
1988, 1989), bluegill sunfish (Lepomis macrochirus;
Coleman et al., 1985), cichlid (Cichlasoma nigrofasciatum; Lavery and Keenleyside, 1990), freshwater goby (Padogobius martensii; Torricelli et al.,
1985), common goby (Pomatoschistus microps;
Magnhagen and Vestergaard, 1993) and sand goby
(Pomatoschistus minutus; Lindström and Wennström,
1994). These studies showed that parents increase the
0376-6357/$ – see front matter © 2003 Elsevier B.V. All rights reserved.
doi:10.1016/S0376-6357(03)00106-2
24
M. Jaroensutasinee, K. Jaroensutasinee / Behavioural Processes 64 (2003) 23–29
intensity of aggressive behaviours as their offspring
get older.
Parental care is common among teleost fish and can
be costly in term of reduced survival, breeding rate
and fecundity to the parents (Townshend and Wootton,
1985; Smith and Wootton, 1995, 1999). Parents spend
substantial amounts of time and energy on care including egg guarding, egg ventilating to provide a
flow of oxygenated water and dead egg removal from
the nest (reviewed in Clutton-Brock, 1991; Smith and
Wootton, 1995). The wild Siamese fighting fish (Betta
splendens, Regan) conforms to this pattern in its care
behaviour. Males in the breeding season establish and
defend territories in rice paddies. They construct their
bubble nests, entice females to spawn in them and
care for fertilised eggs and fry (Gordon and Axelrod,
1968; Jaroensutasinee and Jaroensutasinee, 2001a).
Therefore, the ability to compete for territories is
critical to the reproductive success of male Siamese
fighting fish. It could be due to male–male competition and possible direct benefits of female mate
choice (Jaroensutasinee and Jaroensutasinee, 2001b).
We investigated the level of male aggressiveness
towards potential intruder during egg guarding in
wild-caught Siamese fighting fish. The degree of aggressiveness was measured at two reproductive phases
in response to the territorial intrusion of a male, a female and a female that has laid eggs. By quantifying
territorial behaviour during encounters, the following
hypotheses were investigated.
(1) Males should become more aggressive after the
eggs have hatched.
(2) Males should be the most aggressive towards male
intruders, intermediate towards female intruders
and the least aggressive towards females that have
laid eggs. This is because nest-holding males may
perceive male intruders as an additional threat of
territorial take-over and perceive female intruders
as mates. Therefore, the nest-holding males should
perform specific amounts of aggressive acts towards different types of intruders.
1.1. Fish biology
The Siamese fighting fish is an Anabantid native
to Southeast Asia. Typical fighting fish habitats in
Thailand are quiet fresh water ponds with muddy bot-
toms or flooded rice paddies (Gordon and Axelrod,
1968). Unlike domesticated fighting fish, wild fighting fish are small, inconspicuous, and dull brown or
green in colour (Jaroensutasinee and Jaroensutasinee,
2001a,b). They hide beneath water plants, presumably
to minimise predation from fish-eating egrets, herons
and kingfishers.
Males in this species build one bubble nest, court
females, and care for a single brood of the developing
eggs and newly hatched larval fish at a time. However, the successful males may have 2–3 successive
broods per one breeding season. Each male defends
a territory in the water column near the surface. His
territory is centred on a bubble nest built by the male
(Forselius, 1957). Fertilised eggs need to be aerated
by being attached to bubble nests. Males retrieve eggs
or larval fish that fall out of the nest or stray and spew
them back into the bubble nests (Gordon and Axelrod,
1968; Jaroensutasinee and Jaroensutasinee, 2001a,b).
Fertilised eggs hatch approximately 36 h after eggs
were laid. Males exhibit parental care behaviour both
in the field and laboratory during when the young
have hatched up to 5–7 days (personal observation).
They do not cannibalise their eggs and larval fish
during the period of parental care. They have very
aggressive social displays including gill cover erection, fin spreading, biting, and tail beating (Clayton
and Hinde, 1968; Simpson, 1968). Fighting usually
involves physical damage and can result in death.
Females are duller in colour and usually smaller than
males. After the females finish laying eggs, the males
chase the females out of the bubble nest areas and
solely provide parental care for the developing eggs
and larval fish. Wild fighting fish males are heavier
and bigger than females. They build their nests in aggregations in dense emergent vegetation. Dense male
nesting aggregations may help males to attract more
mates. Their territory size would be less than half of
the distance to the nearest nest.
2. Materials and methods
The test subjects were wild-caught fish collected
from the same rice field from July–September 2001 in
Nakhon Si Thammarat, Thailand (8◦ 38 N 99◦ 53 E).
The fish were maintained in the laboratory with
natural light (i.e. approximately 12 h-light/12 h-dark
M. Jaroensutasinee, K. Jaroensutasinee / Behavioural Processes 64 (2003) 23–29
cycle) and fed daily with mosquito larvae. Males and
females were housed in separated 1-l bottles that were
wrapped around with a piece of paper to prevent visual
contact and fighting. Prior to the test, the nest-holding
male was placed in his 1-l bottle next to a female in
her 1-l bottle until the female became gravid.
To control for confounding effect of size differences
between males, the fishes were measured prior to the
experiments. The following procedure was followed
in measuring the fish body length (Jaroensutasinee and
Jaroensutasinee, 2001a,b). First, each fish was placed
in an aquarium (20.0 cm × 12.0 cm × 16.0 cm high)
filled with water to the depth of 3 cm. The fish were
not anaesthetised because most types of anaesthetic
alter fish appearance (Kodric-Brown, 1989). Secondly,
a piece of Plexiglas with a ruler was placed in the
aquarium to provide a standard calibration. Finally,
after a 1-min acclimatisation period, the fish were photographed with a digital camera. The digital pictures
were used to estimate the fish’s standard body length
from the tip of the upper jaw to the caudal peduncle.
The nest-holding male was placed in a 37-l aquarium, measuring 0.50 m × 0.25 m × 0.30 m high,
densely planted with aquatic vegetations. Males built
their bubble nests within 24 h after being placed in the
aquarium. A gravid female was placed in the aquarium
with the nest-holding male in the evening at approximately 15:30 h of the second day after the nest-holding
male had been placed in the aquarium. After the female was placed in the aquarium, the nest-holding
male chased the female and tried to entice her to come
and spawn under his bubble nest. The female usually
spawned between 07:00 and 10:00 h the morning after she had been placed with the nest-holding male.
After fertilisation was completed, the mated female
was immediately removed from the breeding tank.
During two reproductive phases (i.e. 1 h after the
eggs were laid and within 1 h of the eggs hatching), three types of intruders were introduced to the
nest-holding males (i.e. males, gravid females and
females which have laid eggs). Each 15 min trial was
separated by a 10 min interval and conducted in random order to minimise an order effect. An intruder
was placed in the breeding tank for 5 min before the
observation began. The aggressive responses of the
nest-holding male was observed, including gill cover
erection, biting, tail beating, attacking and chasing.
At the end of the observation period, the intruders
25
were removed from the experimental aquarium and
returned to their home tank. Sixty replicates were conducted with new sets of fish in order to avoid pseudoreplication. No males or females had been used in the
experiment more than once to avoid an order effect.
The number of the five aggressive acts (i.e. gill cover
erection, biting, tail beating, attacking and chasing)
was recorded (Simpson, 1968). Gill cover erection was
regarded as males erecting the gill cover while oriented towards or parallel to intruders. Lowering of the
gill cover or swimming away from intruder ended gill
cover erection. Biting was recorded when males used
the mouthpart to bite or tear at intruder. Tail beating
was defined as each separate beat of the tail towards
intruder. Attacking was recorded when the focal male
swam rapidly towards its intruder. Chasing was defined as rapid and continuous following. Male aggressiveness was the sum of each five aggressive act that
the nest-holding male performed during the 15 min
observation period.
2.1. Statistical analyses
All variables were tested for normality using Lilliefors’ test. The equality of variances was evaluated
using Levene’s test. The three-way repeated-measures
ANOVA was used to test for types of intruder (male,
female and female that have laid eggs), times of reproductive phase (after eggs were laid and after eggs
hatched), types of aggressive behaviour (gill cover
erection, biting, tail beating, attacking and chasing)
and the interaction among factors, and then followed
by paired samples t-tests.
3. Results
The nest-holding males, male intruder after egg laid
and male intruders after eggs hatched were not different in size (nest-holding males (x̄ ± S.D. = 3.30 ±
0.26), male intruders after eggs laid (x̄ ± S.D. =
3.23 ± 0.34) and male intruders after eggs hatched
(x̄ ± S.D. = 3.26 ± 0.31) one-way ANOVA: F2,177 =
0.81, ns). Females who laid the eggs were the same
size (x̄ ± S.D. = 3.20 ± 0.22) as female intruder after
egg laid (x̄±S.D. = 3.14±0.25) and female intruders
after eggs hatched (x̄ ± S.D. = 3.13 ± 0.21) (one-way
ANOVA: F2,177 = 1.59, ns).
26
M. Jaroensutasinee, K. Jaroensutasinee / Behavioural Processes 64 (2003) 23–29
M. Jaroensutasinee, K. Jaroensutasinee / Behavioural Processes 64 (2003) 23–29
27
Table 1
Three-way repeated-measures ANOVA (Wilks’) of types of intruders at two reproductive phases with five types of aggressive acts
Effect
Hypothesis d.f.
Error d.f.
Value
F
Intruder
Reproductive phase
Aggressive behaviour
Intruder × reproductive phase
Intruder × aggressive behaviour
Reproductive phase × aggressive behaviour
Intruder × reproductive phase × aggressive behaviour
2
1
4
2
8
4
8
57
58
55
57
51
55
51
0.161
0.913
0.188
0.992
0.214
0.930
0.783
148.253
5.509
59.223
0.234
23.383
1.031
1.770
∗
∗∗
P
∗∗
∗
∗∗
ns
∗∗
ns
ns
P < 0.05.
P < 0.001.
The nest-holding males displayed differing amounts
of aggressive acts per 15 min towards three types of
intruders (Fig. 1a–e, Table 1). The nest-holding males
were the most aggressive toward male intruders, intermediate towards female intruders and the least aggressive towards females that have laid eggs (Fig. 1a–e).
For reproductive phase, the amount of aggressive acts
significantly changed with the reproductive phases,
being more frequent after egg hatching than egg laying (Table 1). For types of intruders and aggressive
acts, these had no major effect since for intruder ×
aggressive behaviour, the interaction was significant
(Table 1). The frequency of each of the five types of
aggressive acts significantly varied in relation to the
type of intruder.
The nest-holding males won all 120 fights with female intruder and 120 fights with females that have
laid eggs (chi-square tests for both female intruders
and females which have laid eggs: χ12 = 60, P <
0.001) and lost 1 of 120 fights with male intruders
(chi-square test: χ12 = 58.02, P < 0.001).
4. Discussion
This study aimed to test only the first hypothesis of theoretical models that parents will increase
their parental effort during the period of parental care
when the average reproductive value of the young increases (Williams, 1966; Pianka, 1976; Pressley, 1976,
1981; Ricklefs, 1977). The result supports the hypothesis that the nest-holding males become more aggressive as the reproductive value of the young increases
as they age. This is because the probability of the
survival of offspring to reproduce increases. It has
been shown in many fish species that parental males
put more effort into egg guarding and caring as eggs
become older, such as in three-spined sticklebacks
(van Iersel, 1953), freshwater gobies (Torricelli et al.,
1985), common gobies (Magnhagen and Vestergaard,
1993), and sand gobies (Lindström and Wennström,
1994). These males increase their fanning with increased egg age because larval fish need more oxygen, as they grow older (Reebs et al., 1984). However, Yamamoto et al. (1999) found the opposite result
in that an Amazonian cichlid, Pterophyllum scalare
Lichtenstein, was more aggressive after the eggs were
laid than after the eggs hatched. The opposite finding
between Yamamoto’s study and our study may be because wild-caught male fighting fish do not cannibalise
their eggs during the period of parental care but the
Amazonian cichlid has some tendency to eat his own
eggs. Therefore, Amazonian cichlid eggs are at greater
risk than newly hatched fry because they are immobile compared to newly hatched fry (Yamamoto et al.,
1999).
䉳
Fig. 1. Mean (±S.E.) of the number of aggressive acts per 15 min among three types of intruders at two reproductive phases. (a) Gill
cover erection, (b) biting, (c) tail beating, (d) attacking, and (e) chasing. MFL, mated female intruders after eggs laid; MFH, mated female
intruders after eggs hatched; FL, female intruders after eggs laid; FH, female intruders after eggs hatched; ML, male intruders after eggs
laid and MH, male intruders after eggs hatched. Solid bars link those groups that were found to be significantly different between types of
intruders. Dotted bars link those groups that were found to be significantly different between two reproductive phases. Two-tailed paired
t-test, ×× P < 0.001, × P < 0.05.
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M. Jaroensutasinee, K. Jaroensutasinee / Behavioural Processes 64 (2003) 23–29
Our results showed that the nest-holding males performed specific amounts of aggressive acts towards
different types of intruders. Nest-holding males were
the most aggressive towards male intruders. Male intruders may pose an additional threat of territorial takeover. Owning a territory is crucial for male fighting
fish to obtain mates. This also has been shown in Tanganyikan cichlid (Lamprologus ocellatus; Walter and
Trillmich, 1994) and Amazonian cichlid (P. scalare,
Yamamoto et al., 1999). In addition, male fighting fish
were less aggressive towards female intruders. These
males may perceive opposite sex intruder as their potential mates. The nest-holding males were the least
aggressive toward females that have laid eggs. This
suggested that these males might be able to distinguish
between females that have laid eggs and other female
intruders. Females that have laid eggs might pose less
treat of eating their own eggs than other females.
However, there is an alternative explanation for the
different response to the two types of female intruders.
Courtship and aggressive behaviour of male fighting
fish share the same components of display (i.e. tail
beating, gill cover erection and approach, Robertson
and Sale, 1975; Doulrelant et al., 2001). The difference between these two types of display regards which
behaviours are performed less or more commonly
towards the intruders. During courtship behaviour,
males perform very few bites and perform mostly
tail beating, and gill cover erection, while males also
spend more time at the nest. During an aggressive
interaction male bites more, increases the amount of
tail beats and gill cover erection, and decrease the
time they spend under the nest (Robertson and Sale,
1975; Doulrelant et al., 2001). When we look at the
gravid female treatment and compare between reproductive phases, one finds that males perform more
tail beating and gill cover erections after hatching
than after egg laying, but these males do not change
their biting behaviour. This change in behaviour could
represent and increase of the motivation of males to
court gravid females as the eggs hatch and the males
become more available to take care of new eggs.
Acknowledgements
We thank J. Endler for useful suggestions on experimental design and data analysis. We thank J. Endler,
N. Metcalfe, R. J. Matos, T. na Nagara, and A. Roberts
for comments on previous versions of the manuscript.
Invaluable assistance in the laboratory was provided
by N. Kongthong, P. Suebchana, P. Sirisuk, N. Klanniwat and U. Chavalit. This work was supported by the
TRF/BIOTEC Special Program for Biodiversity Research and Training grant BRT R 144003.
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