Influence of incubation temperature on offspring phenotype

Differences in temperature during incubation have been shown to affect offspring phenotype and potentially influence the fitness of young birds. A series of recent laboratory studies have pinpointed incubation temperature as a major contributor to such phenotypic variation. These studies demonstrated that small differences in incubation temperature, reflecting variation in the temperature found in naturally incubated nests, drastically affected hatchling immunocompetence, locomotor performance, stress endocrinology and growth, and body condition. A previous study in wood ducks revealed that at one to three days prior to hatching, ducklings from eggs incubated at lower temperatures had lower protein content than those incubated at higher temperatures (Hepp and Kennamer, Reference HEPP and KENNAMER2012). Additionally, ducklings incubated at lower temperatures expended more energy during incubation, as noted by higher oxygen consumption, than those from higher incubation temperatures (Hopkins et al., Reference HOPKINS, DURANT, HEPP and HOPKINS2011). These studies suggested that ducklings incubated at lower temperatures have lower energy reserves after hatching to meet the demands of thermoregulatory challenges than those incubated under higher temperatures. Avian ecologists have long appreciated the role incubation temperature plays in hatching success and the length of incubation (Vasudeva et al., Reference VASUDEVA, DEEMING and EADY2014), and current findings have revealed that incubation temperature is an important determinant of avian offspring phenotype, not only affecting the early development of the body, but also having a continued effect on body condition, including final body weight and meat quality (DuRant et al., Reference DURANT, HOPKINS and HEPP2011; Reference DURANT, HOPKINS, HAWLEY and HEPP2012; Reference DURANT, HOPKINS, HEPP and WALTERS2013).

Influence of incubation temperature on offspring sex ratio

The gonads in the early stage of vertebrate embryos have a bisexual structure, meaning they are able to develop into a male or female bird. Environmental conditions can affect the direction of sexual development in animals, which is mainly determined by the combined effects of hormones and temperature. It has been reported that different temperatures applied with hormones influenced the development of different sexes (specifically, the rates of production of male and female embryos). It was stated that embryonic development occurred without any defect as long as the air that surrounded the eggs during the incubation period remained within a critical temperature range (26°C to 40.5°C) (Conway and Martin, Reference CONWAY and MARTIN2000).

Some studies of poultry 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 (Yilmaz et al., Reference YILMAZ, TEPELI, GARIP and CAGLAYAN2011). The different incubation temperatures for the first generation (at the embryo stage) may influence the sex of the next generation of chicks, and males were significantly more common during the embryonic period in parent quail eggs exposed to 36.7°C and 37.2°C.

It has been reported that different incubation temperatures have significant effects on the sex ratio of Muscovy ducklings. The numbers of male ducklings increased with temperature, and the females declined. When pre-incubation temperature was between 36.3 and 37.3°C, the number of hatched male Muscovy ducklings was less than the number of females. The sex ratio (male:female) ranged from 0.24 to 0.95. When pre-incubation temperature was between 37.8 and 39.3°C, the number of hatched male Muscovy ducklings was more than females with a sex ratio ranging from 1.39 to 5.38. Only when the incubation temperature was between 37.3 and 37.8°C, the secondary sex ratio and genetic sex ratio (1:1) were not different. However, the reasons for the sex ratio changing at different incubation temperatures needs further study. Controlling sex ratio using incubation temperature could be used to meet market needs.

Influence of incubation temperature on immunity function

The embryonic development of oviparous vertebrates is completed outside of the mother's body. Thus, immunity is determined by both genetic and environmental factors, with the latter representing an important influence. Incubation temperature is one of the most obvious exogenous environmental factors which impacts poultry embryo development.

Changing incubation temperature may affect the development of immune organs, thus affecting the immune ability of poultry. It has been reported that there is an effect of incubation temperature on duck immunity (Hopkins et al., Reference HOPKINS, DURANT, HEPP and HOPKINS2011). Ducklings hatched from eggs incubated at lower temperatures had slower growth rates and poorer immune function compared to those incubated at higher temperatures. Recent studies on birds have indicated that temperature affects the immune function of hatchings, with decreased immunocompetence at lower temperatures (e.g. 35°C) compared to higher temperatures (e.g. 37°C) (Ardia et al., Reference ARDIA, PEREZ and CLOTFELTER2010; DuRant et al., Reference DURANT, HOPKINS, HAWLEY and HEPP2012).

Research has shown that the development of the duck embryo's immune organs can be inhibited after incubation temperature changed (Liu et al., Reference LIU, LIU, YAN, LI, ZHAO, WANG, ZHANG, WANG, WANG, LI, YANG, LI, HAN and WANG2015). This is similar to the effect of heat stress on the immune organs of chickens. In the whole, in the high temperature treatment group, the relative weight of the spleen, thymus, Bursa of Fabricius in the duck embryo and hatched ducklings showed a downward trend, which meant that embryo and subsequent hatchling immune organ development was inhibited by increases in incubation temperature. Heat stress caused apoptosis of the immune organs, with thymus apoptosis occurring earlier than the Bursa. The main cause of this damage may be that the heat stress enhanced the effect of corticosterone on the thymus gland, the cytoplasm and nucleus of the bursa, which had a cytotoxic effect, inhibited the activity of natural killer cells, antibody production, lymphocyte activation factor and cell growth factor production, inhibited glucose uptake and protein synthesis and led to thymus and bursa atrophy (Liu et al., Reference LIU, LI, HAN, ZHANG and XU2012).

It has been reported that both cold and heat stress could lead to damage of the immune organs (Soren et al., Reference SOREN, BHANJA, MANDAL, MEHRA, GOEL, SOREN, MANDAL and MEHRA2012; Zhao et al., Reference ZHAO, ZHANG, QU, YAO, LI, LI and XU2014). With extended heat stress, the development of the immune organs of broiler chicks was blocked, and relative weight of the thymus, spleen and bursa was reduced. Under too high or low temperatures, endocrine function of the hypothalamus- pituitary- adrenal axis could be enhanced. The content of glucocorticoid in the blood was significantly increased, and immune suppression led to a reduction in immunity. Due to the long period of high temperature incubation, these effects may extend from the embryonic period to after hatching.

Effects of incubation temperature on immune-related genes and enzymes

Immunity-related genes and enzymes may be critical towards understanding how incubation temperature affects offspring immune function. IgD, IgM and IFN-γ are members of the immunoglobulin classes. These immunoglobulins are considered to provide the first line of defence against microbial infection, together with innate immunity factors. The level of IFN-γ rises, which means that the immune function of the body is improved. In the high temperature incubation group, serum globulin and IFN-γ of newly hatched and two week old ducks were lower than in the control group (Liu et al., Reference LIU, LIU, YAN, LI, ZHAO, WANG, ZHANG, WANG, WANG, LI, YANG, LI, HAN and WANG2015), which suggested that immune function was inhibited. There were similar effects of external temperature change on the level of globulin and IFN-γ (DuRant et al., Reference DURANT, HOPKINS, HAWLEY and HEPP2012). These studies indicated that high temperature affects the development of the immune system during the incubation period, reducing the level of immunoglobulin and IFN-γ, and suppressing immune function.

Effect of incubation temperature on the development of muscle fibre

The number of animal muscle fibres has been determined before birth and does not increase after birth. The growth of muscle fibres mainly refers to its hypertrophy (Hammond et al., Reference HAMMOND, SIMBI and STICKLAND2007), which in turn depends on the increase in myofibrils.

Changes in the external environment can affect muscle development. It was found that the changes in myoblast proliferation, muscle fibre fusion, muscle fibre diameter thickening and increasing cross cutting area was relatively obvious before and after the 21 embryonic ages in the late stage of duck embryo development (Zhang et al., Reference ZHANG, LIU, LI, WANG, LIU, HU, YAN, GOU, LI and WANG2014). Changing incubation temperature has been shown to change gene expression in embryonic muscle of Peking ducks (Liu et al., Reference LIU, LIU, YAN, LI, ZHAO, WANG, ZHANG, WANG, WANG, LI, YANG, LI, HAN and WANG2015). Hammond et al. (Reference HAMMOND, SIMBI and STICKLAND2007) found that the number of muscle nuclei and fibres in chicken embryos increased compared to the control group from 12 days after raising the hatching temperature. At the same time, it was found that the whole gastrocnemius muscle mass of the high temperature treatment group was larger than that of the control group. It was reported that the leg muscle of the high temperature group at 20 embryo age and at two weeks after hatching was larger than that of the control group (Liu et al., Reference LIU, LIU, YAN, LI, ZHAO, WANG, ZHANG, WANG, WANG, LI, YANG, LI, HAN and WANG2015). Additionally, pectoral muscle fibres were larger than in the control group.

The reasons for muscle weight loss after increasing incubation temperature at incubation mid-anaphase have been analysed. Increased incubation temperature may speed up the movement of muscle cells, inducing the production of endoplasmic reticulum stress which causes a large number of muscle cells to die. At the same time, the sequence data of the leg muscle group were analysed, which showed that change in incubation temperature decreased gene expression. In total, 335 differentially expressed genes were analysed, in which 182 genes were up-regulated and 153 genes were down regulated. Signal pathway enrichment results showed that ribosomal metabolic processes and MAPK signalling pathways were important molecular pathways in the development of duck skeletal muscle (Liu et al., Reference LIU, LIU, YAN, LI, ZHAO, WANG, ZHANG, WANG, WANG, LI, YANG, LI, HAN and WANG2015).

Incubation temperature affects muscle growth in turkeys (Piestun et al., Reference PIESTUN, ZIMMERMAN and YAHAV2015). Short-term change in incubation temperature, such as 3 hours at 38.5°C, had no effect on hatching rate (Christensen and Bagley, Reference CHRISTENSEN and BAGLEY1989). However, the effect of temperature during late embryonic development on muscle growth is important, which was confirmed in an experiment using different temperatures and time of exposure. The number of satellite cells increased in the chicken embryo during the 16th and 18th days. However, the treatment of temperature had no effect on muscle growth in the embryo from 8 to 10 days (Piestun et al., Reference PIESTUN, HALEVY and YAHAV2009a).

Studies have indicated that the effect of temperature on late embryo stage has a major and lasting effect on the proliferation and differentiation and hypertrophy of muscle cells. Changes in cell proliferation and hypertrophy indicated that fibroblasts were highly sensitive to small changes in heat during the whole process. This was confirmed in a study using short-term and long-term temperature treatments (Piestun et al., Reference PIESTUN, HAREL, BARAK, YAHAV and HALEVY2009b). When turkey embryos were hatched at 38.5°C from the ninth to the 12th days, more muscle fibres were produced at 16 days after hatching (Maltby et al., Reference MALTBY, SOMAIYA, FRENCH and STICKLAND2004). When turkey embryos were hatched at 38.5°C from the fifth to the eighth day, the expression of muscle cells was delayed, proliferation of muscle cells was increased and differentiation was slower. Compared with 37.5°C in the same period of incubation, when turkey embryos were hatched at 38.5°C from day 5 to 11, the turkeys hatched at the higher temperature had more muscle nuclei at 16 days after hatching, which could lead to muscle proliferation and muscle hypertrophy.

Therefore, in order to improve the lean meat rate of poultry, the effects of changing incubation temperature on muscle yield should be further studied. In general, muscle output could be achieved by taking appropriate measures in the embryonic period in mammals. In the above studies, the muscle fibre diameter and cross-sectional area could be increased by changing the incubation temperature before and after hatching. Therefore, increases in muscle output of adult poultry could be achieved.