**4. Discussion**

This present review used a methodological approach to conduct a comprehensive literature search, which enabled a logical interpretation of the recent results obtained from broiler chicken incubation published studies. Thus, the effects of thermal manipulation on incubation performance, hatchability and hatching quality of broiler chicks could be examined.

#### **4.1 Thermal manipulation and thermotolerance acquistion**

The hatchery industry is expected to change dramatically with increasing demand for quality chicks and production efficacy. It is well established that incubation conditions significantly influence incubation and post-hatch performance besides, hatching quality in chickens [4]. During perinatal stage (critical period), incubation conditions may result in persistent variations in the epigenetic programming of different body systems and their roles in chickens [56]. One condition of interest is incubation temperature, which when manipulated by short or long-term may induce epigenetic adaptation thus enhancing development and maturation of particular body systems and their functions, which begins during the early periods of embryonic development [10].

TM is well known for inducing improved thermotolerance acquisition (thermoregulatory functions) in chickens, which is evidenced by reduced body temperature at hatch and during the first days post-hatch. Intermittent manipulation in incubation temperature between different embryonic ages resulted in thermoregulatory functions being boosted; 3 h of 39.5°C/d from E16 to E18 [21], 39.5°C for 12 h/d at E7–E16 [15, 16, 24], and 60 minutes exposure to 15°C at E18–E19 [57]. In our review, the similar effect was confirmed by [49, 50] at 38.5–39.5°C for 18 h/d from E10 to E18 and, David et al. [18] at 39.5°C for 18 h/d from E7 to E16. It's clear that thermotolerance acquisition in broiler chickens can be enhanced by application of TM between E7 to E18, a period that is termed the critical stage and, the ideal embryonic age for TM.

It is scientifically proven that successful TM should be between E7 and E18, a period which enables efficient alternation in threshold stimulus of the regulatory systems during the development and maturing of the thermoregulatory mechanism (hypothalamus-hypophysis-thyroid axis) and the stress control (hypothalamushypophysis-adrenal axis) [9, 11, 12, 58]. It is clearly reported that thermotolerance acquisition is improved via reduced plasma triiodothyronine (T3) concentrations and basal metabolism, accompanied with lowered body temperature [15, 16, 24]. In addition, T3 is the thyroid hormone of interest in the last week of incubation because it is vital for increasing extra energy requirements during hatching [14].

However, some recent studies have reported that TM increased [30, 51] and had no influence on hatch body temperature [48, 52]. These differences may be associated with the possible elevation or similarity in hormones that regulate metabolism (T3) and growth (GH) leading to elevated or similar metabolic rate and heat production, accompanied with elevated and similar body temperature in thermal manipulated chickens and both thermal manipulated and control treatments, respectively [36, 39].

## **4.2 Thermal manipulation and embryo, hatch, or chick weight**

Short-term (intermittent) alteration in incubation temperature during varying age of embryogenesis can boost muscle growth and development at hatch and in the first weeks post-hatch (early period (E0–E5) [22]; mid-term (E16–E18) [59]; long-term (E12–E18 and E10–E21) [60]). In the current review, the similar effect was identified with short and long-term TM (36.8°C for 24 h/d from E7 to E13 [32]; 39°C for 9–18 h/d from E10 to E18 [37, 50]; 36.°C for 6 h/d from E10 to E5 [2]), which was indicated by increased embryo, hatch and 1-day-old chick weight. In addition, it is evidenced that TM has significant effect on proliferation and differentiation of satellite cells, and thus growth and development of embryonic and chick muscles [61].

TM at 39.5°C for 12 or 24 h/d from E7 to E16 result in accelerated myoblast proliferation and cell differentiation, which is evidenced by increased myoblast number (25–48%) in the pectoral muscle and increased expression of myogenin in embryonic muscles, respectively [17]. Similarly, Al-Zghoul et al. [37] and Al-Zghoul and El-Bahr [44] found upregulation of MyoD, myogenin, insulin-like growth factor 1 (IGF-1), and growth hormone (GH) after TM at 38.5–39°C for 9–18 h from E12 to E18 in embryos and 1-d-old chicks. Furthermore, a linear increase in embryo breast muscle weight with embryonic age was observed but significantly elevated in the TM-treated embryos compared with controls during the second quarter of embryogenesis. The interpretation of the above findings explains the possible reasons for elevated embryo, hatch and 1-day-old chick weight after TM. However, the ability of myoblasts to proliferate declined in the embryos after TM compared with embryos incubated at 37.8°C in the last quarter of incubation [17]. Furthermore, Piestun et al. [59] reported increased muscle hypertrophy in thermal manipulated embryos at 39.5°C for 3 or 6 h/d from E16 to E18. This was evidenced by upregulation of myogenin, and IGF-1 mRNA expressions in TM embryos compared with control treatment.

Studies by Zaboli et al. [41], Al-Zghoul et al. [45], and Dalab and Ali [46] reported depressed embryo, hatch, or chick weight due to intermittent high-temperature TM, which partially agrees with Piestun et al. [14]. However, Piestun et al. [14] reported that only continuous (24 h) elevation in incubation temperature (39.5°C) from E7 to E16 negatively affected embryo growth and development and hatch weight. Although the above variation could have resulted from differences in factors such as breed or strain, flock age, incubation layout, and embryo age at the time of TM, the contradicting results due to TM length may suggest a strong gap for continuous studies on the length of TM.

In the current review, TM did not influence embryo, hatch, or chick weight in 67% of the intermittent TM studies that reported the above parameter. This result has been attributed to a possible similarity in plasma T3, T4, and GH leading to similar metabolic growth rate and heat production, which result in incubation duration and chick body weight being similar in both thermal manipulated and control treatments [36, 39].

Interestingly, Janisch et al. [32] and Rocha et al. [2] observed increased hatch weight at low-temperature TM compared with high-temperatures. This result may be associated with the variations based on factors such embryo age at TM, strain, and incubation temperature profile. However, it is well established that yolk weight is a critical factor that accounts for 20% of hatch weight [62]. At low incubation temperatures, nutrient metabolic rate, and the embryo's ability to draw liquids from the yolk sac are reduced, which result in increased yolk weights at hatch [63], and consequently, elevated hatch weight.

*Thermal Manipulation: Embryonic Development, Hatchability, and Hatching Quality… DOI: http://dx.doi.org/10.5772/intechopen.101894*

#### **4.3 Thermal manipulation and hatchability**

The effect of TM on hatchability in the present review is contradictive, with 65% of 20 studies that reported hatchability found no significant effect, 30% being reduced, and a comparative study by Dalab and Ali [46] reported increased and decreased hatchability with intermittent TM at different embryonic age. Also, earlier studies regarding TM and hatchability have shown contradicting results, for instance, Yahav et al. [11] and Piestun et al. [22] reported significantly increased hatchability with TM at 39.5°C for 3 h/d from E8 to E10 and 38.1°C for 24 h/d from E0 to E5, respectively. Yahav et al. [29] identified no effect on hatchability at 38.5°C for 3 h/d from E8 to E10. Piestun et al. [24] found decreased hatchability with TM at 39.5°C for 12 h/d from E7 to E16.

It is well documented that hatchability is depressed by overheating embryos however, length, strength, and embryo age at the time of high-temperature TM determine the effects of the application on hatchability [25]. Reduced hatchability has been associated with reduction in corticosterone concentrations at internal pipping after TM at 39°C for 2 h/d from E13 to E17 [64]. Continuous TM at 39.5°C from E7 to E16 depressed embryonic growth and development, which was accompanied by lower hatchability compared with intermittent and control treatments [14]. Low hatchability was associated with reduced development of pipping muscle (musculus complexus) on E18 and E19 day, which muscle is stated to have a significant role during hatching [14].

Meanwhile, embryo mortality rate and incubation duration or hatching time have been associated with hatchability. Brannan et al. [54], for instance, revealed increased embryonic mortalities (mid and late) after TM, which periods of development overlap with the plateau in eggshell temperature during TM at 39.5°C from E7 to E16, consequently, reduced hatchability. In addition, the above authors stated that fluctuating effect of TM on hatchability is associated with harmful levels of incubator temperature on embryo development besides, flock age, genotype, incubation design, etc. Almeida et al. [36] reported longer incubation period at low-temperature TM, which was followed by reduced hatchability compared with standard incubation and high TM.

Furthermore, reduced hatchability is linked to decreased chick quality, which is a well-known indicator for incubation challenges and investigated for assessment of incubation conditions [65]. While Elmehdawi et al. [35] identified no negative effect of high-temperature (38.4°C) TM from E18 to E20 on hatchability and chick quality, Dalab and Ali [46] observed lower hatchability and chick quality after TM at 39°C for 18/h from E15 E7 to E18. Similar to hatchability, the effect of exposure of embryos to low or high temperatures on chick quality is thought to depend on length and level of TM besides the stage of embryo development at the timing of TM [25].

Tzschentke [10] reported that slight increase in incubation temperature is expected to yield no depressing effects of TM in the last stages of embryogenesis, a period in which the development of mechanisms that regulates temperature in peripheral and central nervous systems, besides other body systems and their roles are completed. This could be the possible reason for no significant effect of TM on hatchability in most studies in the current review.

#### **4.4 Thermal manipulation and eggshell temperature**

Studies by Morita et al. [39] and Amjadian and Shahir [48] identified that exposure of embryos to high temperatures increased eggshell temperature in comparison to standard incubation temperature. The eggshell temperature reflects embryo body temperature [66]. The air temperature and heat transfer between the egg and the

incubator affect embryo body temperature, however, the correlation between heat production by the embryo and heat loss by the incubator determine the embryo temperature [67, 68]. It is established that the rate of chicken embryo heat production is proportional to increase in embryo development thus embryo body temperature reflects embryo development [69]. This could explain the longer incubation duration for low-temperature TM compared with control and high-treatments observed by Almeida et al. [36] and Morita et al. [39]. Earlier, Willemsen et al. [70] found significantly higher eggshell temperature (41.1°C) in high-temperature (40.6°C) compared with 35.5°C in low-temperature (34.6°C) thermal manipulated embryos, which was significantly reduced in comparison to 38.3°C of control temperature (37.6°C) from E17 to E18. Similarly, Piestun et al. [24] found that eggshell temperature was higher in thermal manipulated eggs at 39.5°C compared with standard incubation (37.8°C), accompanied by elevated hatching process of 6 h earlier. However, between E19 and E21, the eggshell temperature decreased although both the thermal treated and untreated eggs were placed in the same hatcher. Delay in hatching period has been linked to depressed metabolism in embryos after exposure to lower incubation temperatures than the standard [25].

In the current review, 65% of studies used 37.8°C as the standard incubation temperature, which also acted as the control treatment. During TM, any elevation in incubation temperature (above 37.8°C), RH is adjusted to 65% to eliminate excessive water loss from the eggs [14]. In addition, setting RH at 60% from E0 to E21 was thought to reduce the influence of RH on embryogenesis and embryonic mortality [36].
