The effects of incubation temperature on the sex of Japanese quail

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The effects of incubation temperature on the sex of Japanese quail
Research Note
The effects of incubation temperature on the sex of Japanese quail chicks
A. Yılmaz,1 C. Tepeli, M. Garip, and T. Çağlayan
Department of Animal Science, Faculty of Veterinary Medicine, University of Selcuk, 42003,
Selçuklu, Konya, Turkey
ABSTRACT The effects of incubation temperature on
the sex of Japanese quail chicks were investigated in
this study. The study was conducted on Japanese quail.
In all, 4500 eggs obtained from 2 generations were used.
At the beginning of the study, a new flock was formed
from available hatching eggs. Hatching eggs were gathered at 3 different ages (8 to 10 weeks, 16 to 18 weeks
and 22 to 24 weeks of age) from the laying period in this
flock. These eggs were exposed to 5 different incubation
temperatures (36.7, 37.2, 37.7, 38.2, and 38.7°C). The
hatching results were evaluated for each group. Chicks
obtained from these temperature groups were reared
separately to obtain quail for breeding. Eggs for incubation were gathered from these breeding quail when
they were between 15 and 18 weeks of age. These eggs
were placed in an incubator at a standard (37.7°C)
temperature, separated by F1-generation temperature
groups. The chicks in all groups were reared separately,
and the sex of the chicks was determined at maturity.
Statistical differences (P < 0.05) were found for the sex
of the chicks in the third group (22 to 24 weeks) of the
F1 generation, compared with other groups. This result confirmed the hypothesis that different incubation
temperatures for the first generation (at the embryo
stage) might influence the sex of the next generation
of chicks. Further studies are needed to investigate the
effects of incubation temperature on chicks from different perspectives.
Key words: incubation, temperature, sex determination, chick
2011 Poultry Science 90:2402–2406
doi:10.3382/ps.2011-01471
INTRODUCTION
Some investigators (Fleming and Crews, 2001; Blumberg et al., 2002; Pace and Brenner, 2003; Juliana et
al., 2004) have reported that the sex of turtles, birds,
and some lizards in tropical areas is determined strictly
according to genotype. Accordingly, they have concluded that incubation temperature, especially in birds and
tropical reptiles, does not have a phenotypic effect on
sex differentiation. In this respect, tropical reptiles differ from other reptiles. However, additional reports have
suggested that enzyme and molecular factors related to
incubation temperature may affect sex determination in
poultry, reptiles, and especially in snakes (Graves and
Shetty, 2001; Crews, 2003; Göth and Booth, 2005). For
instance, it has been reported that different temperatures applied with different levels of the histidine triad
(HIT), P450arom, and P45017α affected gene expression,
and that expression of these genes influenced the development of different sexes (specifically, the rates of production of male and female embryos) (Dawson, 1998;
O’Neill et al., 2000; Pace and Brenner, 2003).
©2011 Poultry Science Association Inc.
Received March 9, 2011.
Accepted June 19, 2011.
1 Corresponding author: [email protected]
Several studies of sex determination in poultry have
been published in recent years (Ishimaru et al., 2008;
Mattsson et al., 2008a,b; Alekseevich et al., 2009;
Brunström et al., 2009; Lee et al., 2009; Schoenmakers
et al., 2009; Smith et al., 2009). In studies of poultry,
quail are commonly used because these birds are considered to represent appropriate experimental models;
therefore, they serve as models for commercially produced poultry such as chicken and turkey (Takada et
al., 2006; Mattsson et al., 2008a,b; Brunström et al.,
2009). Conway and Martin (2000) have stated that embryonic development occurs without any defect as long
as the air that surrounds the eggs during the incubation period remains within a critical temperature range
(26°C to 40.5°C). These investigators also stated that
any increase or decrease of the temperatures outside
the critical range increases the death rate of the embryos. Appropriate incubation temperatures for many
poultry species are reported to be from 37.0 to 37.8°C
(Ensminger, 1992; Harvey, 1993; Woodard et al., 1993;
Robbins, 1998). In some studies of poultry (Van Krey,
1993; Dawson, 1998; Kraak and Pen, 2002; Sundström,
2003; Miller et al., 2004), the data collected suggest
that the sex of the individual becomes distinct in the
first days of incubation, yet oogonia of the individuals
differentiating as females develop during the last third
of the incubation period. New data have been obtained
2402
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RESEARCH NOTE
on the mechanism of sex determination of egg-laying
vertebrates, especially as the result of molecular studies
(Crews et al., 1996; Willingham et al., 2000; Broderick
et al., 2001; Gabriel et al., 2001; Milnes et al., 2002).
These molecular studies suggest both similarities and
differences in the sex determination mechanisms of
egg-laying nonmammalian vertebrates and of mammals
(Kraak and Pen, 2002; Miller et al., 2004). A recent
report has produced the first evidence that incubation
temperature has an effect on sex composition in the
Australian brush turkey (Alectura lathami) (Göth and
Booth, 2005). We are not aware of any other studies
examining incubation temperature and its effect on sex
determination in poultry.
The purpose of this study was to investigate the possible effects on sex determination of different temperatures during the incubation of quail eggs.
MATERIALS AND METHODS
Experimental Materials
In all, 4500 hatching eggs obtained from the quail
were used in the study. To feed the birds, an appropriate ratio of chick growing feed to laying quail feed was
determined according to the age and production period
of the birds in the experiment. The feed used in the
study was obtained from a commercial firm. Hatching
eggs were obtained from a quail farm and were placed
into an incubator. The chicks that hatched from the
eggs were reared to 6 wk of age, after which a total of
160 female and 40 male quail were selected from this
stock. These birds constituted the first maternal group
(grandparents, F0) in the study. The chicks in the parental (F1) generation were kept in a brooder for 3 wk
and then in rearing cages for 3 wk. This research was
approved by The Ethics Committee of the Faculty of
Veterinary Medicine, University of Selcuk (report no:
2009/73).
Feeds Used and Feed Content
During the first 4 wk, chick growing feed (24 to 28%
crude protein, 2800 to 3000 kcal/kg metabolic energy)
was used. Beginning with the 5th wk, laying quail feed
(20% crude protein, 2800 kcal/kg metabolic energy)
was used. Subjects were fed ad libitum, and the birds
were always supplied with fresh water.
Cages and Cabinets
Twenty laying cages (45 × 93 × 27 cm) (width ×
depth × height), multideck incubator brooders (70 ×
180 × 27 cm) (width × depth × height), a lower/upper
deck battery (90 × 201 × 52 cm), a fumigation room,
and 5 incubators, each having a capacity of 768 quail
eggs, were used. Eggs were gathered from standard
cages, which were categorized according to groups, every morning at 07:00 and 08:00 and every afternoon at
16:00 and 17:00. All of the eggs were placed in a disinfection chamber and fumigated for 20 min by mixing 20
g potassium permanganate into 40 mL of 10% formalin
solution per cubic meter of chamber. All eggs were then
placed in the storage room at 15 to 16°C stable temperature and 65 to 70% relative humidity. Before the
incubation period, eggs were gathered from breeding
groups for a week (1 to 8 d) and stored.
Experimental Design
The general plan of the study is given in Table 1.
Formation of the F1 Generation
From the first maternal flock, a total of 2250 hatching
eggs were gathered, 750 per 3-wk period during weeks
8 to 10, 14 to 16, and 22 to 24. The eggs were used for
the production of the parental (F1) generation. The 750
hatching eggs were divided equally into 5 temperature
Table 1. Experimental design1
Grandparent (F0)
Parent (F1)
Offspring (F2)
Egg-gathering
period
Incubation
temperature (°C)
Egg
number (n)
Group
1st period
(8 to 10 wk) 
36.7
37.2
37.7
38.2
38.7
36.7
37.2
37.7
38.2
38.7
36.7
37.2
37.7
38.2
38.7
150
150
150
150
150
150
150
150
150
150
150
150
150
150
150
2,250
1
2
3
4
5
1
2
3
4
5
1
2
3
4
5
 
2nd period
(16 to 18 wk)
3rd period
(22 to 24 wk)
Total eggs
1Incubation
 
Incubation
temperature (°C)
Egg
number (n)
37.7
37.7
37.7
37.7
37.7
37.7
37.7
37.7
37.7
37.7
37.7
37.7
37.7
37.7
37.7
150
150
150
150
150
150
150
150
150
150
150
150
150
150
150
2,250
 
humidity was adjusted to 65% for the first 14 d and to 75% between the 14th and 17th day.
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Yılmaz et al.
Table 2. Incubation characteristics of hatching eggs and sex ratio and viability (%) of chicks (F2, 3rd period)
Group
Item
Parental incubation temperature (°C)
Incubation characteristic
Hatching, all eggs (%)
Hatching, fertile eggs (%)
Fertility rate (%)
Early embryonic death ratio (%)
Middle embryonic death ratio (%)
Late embryonic death ratio (%)
Total embryonic death ratio (%)
Male ratio (%)
Female ratio (%)
Survival (to age 6 wk) (%)
1
2
3
4
5
36.7
37.2
37.7
38.2
38.7
72.67a
76.76
94.67a
8.45
8.45
4.93
23.24
61.80a
38.20b
81.65b
64.00ab
76.80
83.33b
9.60
7.20
4.00
23.20
63.33a
36.67b
93.75a
60.67ab
67.91
89.33ab
9.70
13.43
5.97
32.09
42.86b
57.14a
92.31a
56.67b
65.89
86.00b
12.40
17.83
1.55
34.11
44.87b
55.13a
91.76a
56.67b
70.25
80.67b
10.74
13.22
4.96
29.75
42.86b
57.14a
90.59ab
Significance
*
—
*
—
—
—
—
*
*
*
a,bDifferences
among groups on the same line identified by different letters are significant (P < 0.05).
*P < 0.05; — = not significant.
groups (36.7, 37.2, 37.7, 38.2, and 38.7°C). These incubation temperature groups were maintained separately
thereafter. The quail chicks hatched from eggs of different temperature groups at different periods (8 to 10,
16 to 18, and 22 to 24 wk old) were raised to form the
breeding members of the parent (F1) generation.
Formation of the F2 Generation
By placing 32 female and 8 male quail from the same
temperature group into each breeding cage, 2250 hatching eggs were obtained from 160 females and 40 males
for each period. A total of 480 females and 120 males
were bred for 3 periods to produce the F2 generation.
From each cage, 150 eggs were gathered from each temperature group, marked, and placed into an incubator.
For the production of the offspring (F2) generation, the
incubation temperature was set at 37.7°C; however, the
eggs produced by different temperature groups during
the previous incubation were marked and used for this
incubation cycle. Quail hatched separately from different parental temperature groups were raised to the age
of 5 to 6 wk. The sex of the individuals in the offspring
(F2) generation was assessed based on the results of
tests involving chest feathers and cloaca, and was recorded according to the parental period and temperature group.
Classification of Incubation Results
The number of chicks hatched from the eggs in the
parental (F1) and offspring (F2) generations, the number of embryo deaths associated with various periods,
the number of infertile eggs, and the number of male
and female chicks (4 to 5 wk old) represented the raw
data for the study.
Statistical Analysis
Data on incubation and sex consisted of numerical
counts and were therefore analyzed using the Chi-
squared test. The SPSS (2006) statistical package was
used for the analysis of the data.
RESULTS AND DISCUSSION
Males were significantly more common (P < 0.05)
during the embryonic period in parent quail eggs exposed to 36.7°C and 37.2°C (Table 2, Figure 1). The
critical temperature is known for incubation in reptiles because both sexes hatch at nearly the same ratio
(Doody et al., 2004; Doody et al., 2006; Lance, 2008).
This temperature varies according to the species of reptile considered, and it is equal to 37.7°C in poultry.
The present study found an increase in favor of females
starting at the temperature of 37.7°C at 22 to 24 wk for
the F1 quail offspring (P < 0.05).
In the offspring (F2) generation, no statistical differences in the sex ratio were found for other periods.
This result may be a consequence of the fact that the
eggs in the parental (F1) generation that were exposed
to different temperatures were gathered from different
grandparental (F0) age groups. Offspring (F2) were produced at a standard incubation temperature (37.7°C)
from the eggs obtained from the groups consisting of
these parents (F1). However, parental (F1) incubation
temperatures had no statistically significant effect on
the sex ratio (Table 2, Figure 1). This evidence suggests
that grandparental (F0) age can be a significant factor
in sex determination as well (P < 0.05).
Our research has found that temperature effects on
eggs obtained from older-aged grandparents are reflected in the sex composition of the offspring (F2) generation. These results are in agreement with the findings of
Velando et al., (2002), Young and Badyaev (2004) and
others that the sex composition of offspring in certain
types of birds reflects maternal history. Our findings
are consistent with the statement by Crews and Bull
(2009) that specific chromosomal differences, together
with temperature and male or female dominance, affect
genes that influence sex composition. In addition, the
internal organs of parents (F1) reared for breeding may
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RESEARCH NOTE
Figure 1. Male and female ratios according to maternal incubation temperatures (F2, 3rd period). Incubation temperature: top row, offspring
(F2) generation temperature was 37.7°C; bottom row, parent (F1) generation incubation temperatures. Color version available.
reflect the temperature applied during the embryonic
period.
Recent research on sex composition in poultry (Velando et al., 2002; Young and Badyaev, 2004; Hoshino
et al., 2005; Takada et al., 2006; Endo et al., 2007;
Ishimaru et al., 2008; Mattsson et al., 2008a; Mattsson
et al., 2008b; Alekseevich et al., 2009; Brunström et al.,
2009; Lee et al., 2009; Schoenmakers et al., 2009; Smith
et al., 2009) has suggested the idea that sex composition may reflect the operation of different mechanisms
from those traditionally invoked to explain the phenomenon.
We also found that when the parents (F1) had been
incubated at 38.7°C as eggs, the feathers observed over
most of their body were not of a mature type, and some
regions of the body such as back of the neck, abdominal region and back of the body were not feathered in
maturity.
The research reported here indicates that it is possible
to hatch chicks below or above the standard incubation
temperature (37.7°C) if the age of the grandparents at
egglaying is taken into account (F0). However, because
low and high temperatures can cause embryonic deaths,
commercial firms should include a cost analysis when
considering an application of this sort. In addition, further studies should be conducted to find ways of lowering the embryonic death ratio.
Conclusions
The effects on sex composition of differences in the
temperatures applied to the quail eggs during incubation were apparent in the offspring (F2) generation. In
addition, the age of grandparents (F0) might have indi-
rect effects on the sex composition of the F2-generation
chicks. This research further suggests that interdisciplinary studies on sex differentiation in poultry can improve the level of research on the topic. Furthermore,
evaluation of the incubation results obtained at different temperature applications suggests opportunities for
further detailed studies of the effects of incubation temperature.
ACKNOWLEDGMENTS
This research was supported by TÜBİTAK (The
Scientific and Technological Research Council of Turkey), TOVAG (Agriculture, Forestry & Veterinary Research Grant Committee), Project no: 106 O 048 and
Selcuk University Research Foundation as Project no:
06401068.
A portion of the data in this paper was previously
presented as a poster presentation at the XIIIth European Poultry Conference in Tours, France, August
23–27, 2010. A portion of the study was published in
abstract form in the conference proceedings (Yilmaz et
al., 2010).
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