Low Temperature Effects on Embryonic Development and Hatch Time 1

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1 Low Temperature Effects on Embryonic Development and Hatch Time M. E. SUAREZ/ H. R. WILSON,^ B. N. MCPHERSON,* F. B. MATHER,+ and C. J. WILCOXt *Programa de Ganaderia, Colegio de Postgraduados, Montecillo, Estado de Mexico, 6, Mexico, f Dairy and Poultry Sciences Department, University of Florida, Gainesville, Florida 6, and tarbor Acres Farm, Inc., Glastonbury, Connecticut 6 ABSTRACT A series of experiments was conducted to estimate phenotypic correlations between incubation characteristics, and to evaluate the effects of cold stress and genotype during incubation on chick, egg loss, hatching, and embryonic mortality. Eggs were cooled at 8 or 4 C, for, 4, 6, 48, or 7 h beginning on Day 8,,4,6, or 8 of incubation. Other eggs were cooled intermittently for 6 h every 48 h or h every 96 h. A control group in each experiment was not cold stressed. Results indicated a low and negative correlation between hatching and chick, and a low and positive correlation between hatching and loss from transfer to hatching when variability due to egg was removed. Chick s at hatching were lower in chicks from cooled eggs than those of chicks from eggs incubated under normal temperature. The chicks from cooled eggs were more susceptible to dehydration during holding in the hatcher. Incubation s were delayed approximately as long as the s of embryonic cooling. Embryonic mortality was significantly increased under continuous (single period) cold stress, but not under intermittent cooling (6 h every 48 h). Significant genotype by environment interactions were found in the response of embryos of various strains to cold stress. Exposure for 6 h or longer had detrimental effects on chick and embryo viability, but these effects were modified by interactions among the factors involved. The results indicated that embryos from cooled eggs lose more during incubation and that the neonatal chicks are more susceptible to dehydration during holding, and have a longer incubation period, and a greater embryonic mortality. (Key words: cold stress, embryo, embryonic mortality, incubation, genotype) 996 Poultry Science 7:94-9 INTRODUCTION Effects of temperature on embryonic development and duration of the incubation period depends on the temperature (low or high), age of the embryo, duration of, and their interactions, as well as humidity and type of incubator (Wilson, 99a). A large body of information is available pertinent to effects of temperature on hatchability, including reviews by Barott (97), Kosin (964), Landauer (967), Lundy (969), and Wilson (99b). However, few studies have reported the influence of low temperature on loss of the embryo and length of the incubation period. Sarpong and Reinhart (98) found no significant difference in body of -d-old cooled chicks and controls. Taylor et al. (9) indicated that chilling eggs by interruption of the incubator power supply for h resulted in hatching periods of 8 to h longer than the control group. However, this was not true for eggs Received for publication August 7, 9. Accepted for publication March 7, 996. Florida Agricultural Experiment Station Journal Series Number R-46. To whom correspondence should be addressed. chilled on Day of incubation. Similar results were reported by Booth and Rahn (99), who indicated that intermittently cooling eggs for a total of 6 h (6 h every day) at 4 C delayed hatching 7.4 h. Decuypere and Michels (99) reported a delay of only 4 h in hatching of eggs cooled for 4 h (6 h/d from Day to of incubation). Reduction in hatchability from long- or short-term deviations from the normal artificial incubation temperature (7. C) have been reported, probably due to the inability of avian embryos to regulate their own temperature (Oppenheim and Levin, 97). One of the effects of cooling eggs during incubation is a greater mortality, and consequently, a lower hatchability (Booth and Rahn, 99; Taylor et al, 9; Lancaster and Jones, 988; Decuypere and Michels, 99; and others). Sarpong and Reinhart (98) indicated significantly higher hatchability rates for broiler eggs cooled for 4 h at Day 6 of incubation. Similar results were obtained by Callebaut (99) with consecutive daily 8-h interruptions of incubation of quail eggs, but when the period of cooling was extended to h, hatchability decreased by about 7%. Discrepancies found in the literature concerning the extent of hatching delays due to specific periods of 94 Downloaded from on 7 December 7

2 EMBRYONIC DEVELOPMENT AFTER LOW TEMPERATURE EXPOSURE 9 cooling during incubation, and its effects on other variables of incubation, suggest that interactions between age of the embryo when cold stress is applied, hours of, and temperature to which the embryo is exposed, as well as its genotype, play important roles. Five experiments were conducted to determine: ) correlations between incubation, as affected by cold stress, and other variables of incubation; ) the effects on chick, egg loss, and incubation resulting from of embryos to cold stress at different ages, different temperatures, different lengths of, and their interactions during the incubation period; and ) effects on embryonic mortality due to these factors. MATERIALS AND METHODS Eggs obtained from a broiler breeder (Arbor Acres) or White Leghorn (Hy-Line W-6) breeder 4 flocks were distributed randomly in groups of, identified individually, weighed to nearest. g, and placed in setting trays to be incubated. Each tray was considered a replicate and trays distributed at random in the incubator. The the incubator was turned on was recorded as zero hour of the experiment. During the incubation period, according to the treatments, some trays were removed from the incubator and placed in chambers at a low temperature (8 and 4 C in Experiment ; 4 C in Experiments,, and ), or on a table at environmental temperature (hatchery room = 4 C in Experiment 4) for 6 or h. A temperature of 4 C was selected because it is the approximate temperature in commercial hatchery rooms, and it is just below the physiological zero temperature. A group of eggs was incubated under normal incubation (7. C) throughout the entire incubation period in each experiment and was designated as the control group. After to a lower temperature for the selected on each treatment, trays were returned to the incubator. At 46 h of incubation, all eggs were weighed and candled to identify infertile eggs and dead embryos. Those eggs containing viable embryos were transferred to hatching baskets provided with dividers to identify the chicks individually, and then returned to the incubator. At 48 h of incubation, and every 4 h thereafter, the incubator was opened, hatched chicks were removed and weighed, and of hatching recorded. Then, chicks were returned to the incubator and held until servicing (termination of incubation). At this, each chick was removed from the incubator and weighed. Servicing for the control group Arbor Acres Farm, Inc., Glastonbury, CT 6. 4 Poultry Science Department, University of Florida, Gainesville, FL 6. Jamesway, James Incubator Co., Monroe, NC 869; or Robbins IHA, Robbins Incubator Co., Denver, CO 8. was at or 6 h of incubation. In groups exposed to cold stress, servicing was delayed by the same number of hours that each group of eggs was cooled, or was determined by the when the frequency of hatching chicks declined to near zero per treatment for the period. After chicks were removed from the incubators, and servicing was recorded, unhatched eggs were returned to the incubator for 4 h to assure that all eggs were given full opportunity to hatch. All unhatched eggs (except for Experiment ) were examined macroscopically for characterization of embryonic development. The incubator temperatures (dry and wet bulb) were recorded daily at 8 h during the incubation period; means values were calculated for each incubator. The effects of opening the door were not assessed in any experiment. Response variables considered in all experiments were: hatching and servicing, loss, incubation (recorded in hours) and embryonic mortality. Egg s at set and transfer were recorded to calculate loss. Experiment This experiment was designed to explore the effects of cooling the embryo below the physiological zero temperature. Broiler eggs were cooled at 8 or 4 C on 4,6, or 8 d of incubation for s of,4, or 6 h. Cold stress was applied as described previously. Eggs (n =,4) were set in three adjacent Robbins IHA incubators. Average temperatures were 7., 7., and 7. C for dry bulb and.6,.9, and 9.6 C for wet bulb in Incubators,, and. Incubator was considered as a source of variation in the statistical analyses. Experiment This experiment was designed to test the upper limit of to low temperature that can be used to delay hatch with no detrimental effects on embryonic development. White Leghorn eggs (n = 6) were divided into groups of eggs each and set in the top, center and bottom of a Robbins IHA incubator. Treatments were not replicated within location in the incubator. Embryos were cooled at 4 C on,4, or 6 d of incubation for 4,48, or 7 h. Unhatched eggs were destroyed inadvertently; therefore, embryonic mortality before and after treatment could not be determined. Fertility was determined by candling the eggs at transfer and total mortality was determined by differences between numbers of fertile eggs set and those that hatched. Experiment Effects of low temperature on younger embryos were examined in this experiment. Broiler eggs (n = 94) were incubated in three adjacent incubators (see Experiment ). Three replicates ( eggs each) per treatment were distributed randomly in each incubator. Embryos were cooled at 4 C on 8,, or 6 d of incubation for 4 or 6 h. Downloaded from on 7 December 7

3 96 SUAREZ ET AL. TABLE. Coefficients for orthogonal contrasts by treatment, Experiment Treatment Contrast -4 () -48 () 4-4 () 4-48 (4) 6-4 () 6-48 (6) (7) vs all others + vs vs vs * ! Age by hours of. Age linear. Age quadratic. Comparison between 4 and 48 h. Incubator was considered as a source of variation in the statistical analyses. Experiment 4 Effects of intermittent cooling of embryos were tested. Broiler eggs (n = ) were divided into three groups (five replicates of eggs each) and incubated in a Robbins IHA incubator. Group was the control (not cooled); Group was cooled for 6 h on alternate days (every 48 h) starting on Day ; and Group was cooled for h every 4 d (every 96 h) starting on Day 4. Therefore, eggs of Groups and were cooled a total of 48 h at 4 C. Experiment Responses of embryos to a single period of low temperature, as affected by genotype of the breeders, were evaluated. One hundred and twenty-five eggs from each of commercial broiler crosses (five female lines crossed with two different male lines and identified as A, IB, A, B, A, C, 4A, 4D, A, and E) were divided into groups. Eight groups were cooled at or 6 d of incubation for 4 or 48 h. The other two groups were used as controls (not cooled). Two Jamesway incubators were used. The average temperatures were 7., 6.9 C dry bulb, and 9.8,9. C wet bulb in Incubators and, respectively. Incubator was considered as a source of variation in the statistical analyses. Statistical Analyses Eggs were not grouped by. To eliminate bias due to variability in egg s, least squares analyses of variances were performed using set as a covariable, except for mortality. Data were analyzed as a factorial considering age, temperature,, incubator, and genotype as main effects, according with each experiment. To obtain estimates of main effects and their interactions, data were analyzed without the control group because solutions could not be obtained if the control group were included. Comparisons of main effects least squares means were performed by orthogonal contrasts (Steel and Torrie, 98). Then, additional analyses were performed considering treatment (including the control group) and incubator (or location) as source of variation (Littell et cd., 99). Comparisons of least squares mean of the control group vs other treatments were by orthogonal contrasts (coefficients for Experiment are shown in Table ). All effects in the model, except error (residual), were considered fixed. Variable Set Transfer Hatch Service Hatch Weight loss Weight loss Weight loss Weight loss total TABLE. Simple and partial correlations between characteristics of broiler breeder eggs exposed to low temperature during incubation, Experiment Set Hatch Weight loss = set to transfer; loss Used as covariate for other responses. Simple correlations above diagonal. 4 Partial correlations below diagonal Hatch Weight loss transfer to hatch; loss = hatch to service, loss total = set to service. Downloaded from on 7 December 7

4 EMBRYONIC DEVELOPMENT AFTER LOW TEMPERATURE EXPOSURE 97 TABLE. Least squares means of main effects and probability values of orthogonal contrasts for characteristics of broiler breeder eggs exposed to low temperature during incubation, vs a group incubated under normal conditions, Experiment Effect* Age 4 d 6 d 8 d Temperature 8 C 4 C Exposure h 4 h 6 h Orthogonal contrasts vs all others Age linear Age quadratic Temperature Hour linear Hour quadratic Hatching Chick Servicing M \&> Weight loss (h) Weight loss = set to transfer; loss = transfer to hatching; loss = hatching to servicing, loss total = set to servicing. Adjusted by set (9. g). P <...6. Thus, the residuals were used as error terms for all effects. Data for s and hatching s were analyzed using the SAS General Linear Models (GLM) and Correlation procedures (SAS Institute, 989). Partial correlations were obtained from multivariate analyses of variance (MANO- VA). Data concerning mortality were analyzed using the SAS Categorical Model (CATMOD) procedure. Only least squares means for main effects and orthogonal contrasts by treatments are reported. RESULTS AND DISCUSSION The coefficients for total and partial correlation followed the same pattern in all the experiments. Only coefficients estimated for Experiment are reported (Table ). Incubation correlations with all variables measured before hatching occurred (above diagonal) were positive and highly significant. When partial correlations were estimated using set as a covariate (below diagonal), levels of significance, as well TABLE 4. Least squares means of main effects and probability values of orthogonal contrasts for characteristics of White Leghorn breeder eggs exposed to low temperature during incubation, vs a group incubated under normal conditions. Experiment Chick Weight : loss Effect* Hatching Servicing Age d 4 d 6 d Exposure 4 h 48 h Orthogonal contrasts vs all others Age linear Age quadratic Time 4 vs 48 h Weight loss = set to transfer; loss : Adjusted by set (6.7 g). P < (g) (h) transfer to hatching; loss = hatching to servicing, loss total = set to servicing. Downloaded from on 7 December 7

5 98 SUAREZ ET AL. TABLE. Least squares means of main effects and probability values of orthogonal contrasts for characteristics of broiler breeder eggs exposed to low temperature during incubation, vs a group incubated under normal conditions, Experiment Effect Age 8 d d 6 d Exposure 4 h 6 h Orthogonal contrasts vs all others Age linear Age quadratic Time 4 vs 6 Incubator vs Incubator vs + Hatching Chick Servicing Ol \S> Weigh! t loss Weight loss = set to transfer; loss = transfer to hatching; loss = hatching to servicing, loss total Adjusted by set (7.8 g). P < (h) set to servicing. as magnitudes and signs of the coefficients between incubation and the other variables, differed from total correlation coefficients. The variation in chick at hatching is influenced primarily by egg and egg loss during incubation (Tullet and Burton, 98). Reports indicate that large chicken eggs usually require longer incubation s than small ones (McNally and Byerly, 96; Williams et ah, 9; Wilson, 99b). From eggs of the same, the variability of chick at hatching is influenced by the water lost from the chick and by size of the residual yolk sac (Tullet and Burton, 98). This might explain differences between our total and partial correlation coefficients, because variability due to egg was removed. The response of the chick embryo to low temperature during the incubation period depended upon the age of the embryo at which it was cooled, hours of, temperature, as well as the incubator and some interactions among the factors involved. Results are shown in Tables through 6. Hatching was lower for chicks from cooled eggs, but this reduction was significant (P =.) only for White Leghorn cooled eggs (Experiment, Table 4), This result may indicate that Leghorn eggs are more susceptible to dehydration during embryonic development than broiler eggs, but no statistical comparison with broiler eggs was possible. A similar effect was suggested by Lancaster and Jones (988), but the opposite was concluded by Decuypere and Michels (99) in a comparison of different strains. Within cooled treatments there was an inverse relationship between hatching and age of the embryo when it was TABLE 6. Least squares means and probability values for orthogonal contrasts for characteristics of broiler breeder eggs exposed to low temperature during incubation vs a group incubated under normal conditions, Experiment 4 Chick Weight loss Treatment ' Hatching Servicing 6 h every 48 h h every 96 h Orthogonal contra sts vs all oth ers Cold 6 h vs cold h (g) * (h) height loss = set to transfer; loss = transfer to hatching; loss = hatching to servicing, loss total = set to servicing. Adjusted by set (7.7 g). Room temperature (4 C). 48 h. 4 P <.. Downloaded from on 7 December 7

6 EMBRYONIC DEVELOPMENT AFTER LOW TEMPERATURE EXPOSURE TABLE 7. Embryonic mortality of broiler breeder eggs exposed to low temperature during incubation, Experiment Effect Embryo age 4 d 6 d 8 d Temperature 8 C 4 C Exposure h 4 h 6 h Before After cooled. Servicing was significantly lower for chicks from cooled eggs in all the experiments, except for some interactions when incubator effects were involved. This effect was enhanced by a greater loss of chicks from cooled eggs from hatching to servicing. A greater loss after hatching for chicks from eggs cooled during incubation has been indicated by Decuypere and Michels (99). Incubation was affected significantly by (P <.); the longer the, the longer the incubation. However, the delay was generally shorter than the respective to low temperature. This effect may be attributed to the fact that it takes some for the internal temperature of the egg to equal the incubator or environmental temperature when this temperature is lower than normal (Moreng and Bryant, 96). In Experiment, crosses had a highly significant effect on incubation (data not shown); but, in other experiments with the same crosses this effect was not significant (P >.). The results in Experiment could be due to differences of experimental protocol or because of genotype by environment interactions. Higher hatching of chicks from eggs cooled during incubation have been reported, possibly because the small and weak embryos had succumbed to the cold (Moreng and Bryant, 94), or due to a lower loss before hatching (Kuhn et ah, 98). The opposite was observed in the present study. Sarpong and Reinhart (98) suggested that embryos in cooled eggs have reduced heat production because of a reduced metabolic rates during the of to lower temperature, which may disrupt embryonic development, depending on age of the embryo and of (Moreng and Bryant, 94). However, when the cooled eggs are returned to normal incubation temperature (7. C), compensatory metabolic rate occurs (Kuhn et al., 98; Geers et al, 98), and consequently, higher heat production. No compensatory metabolic rate is evident when the embryo is cooled around 8 d of incubation (Tazawa et al, 988). Geers et al. (98) reported that embryos of eggs incubated at.8 C during the first d had higher heat production per unit of, and per unit of dry body from Day up to the end of incubation. This higher heat production might increase the temperature inside the eggs, which under normal conditions is about 8 C at d of incubation and increases subsequently up to C above incubator environment just before hatching (Tazawa and Rahn, 987). A higher temperature inside the eggs might increase the gradient of temperature between the embryo's body and the incubator, leading in turn to more heat dissipated by evaporation and, thus, to a greater loss before hatching. It is evident by the incubator effect on loss (data not shown) that the lowest loss occurred in eggs incubated at the highest wet bulb temperature, due to a reduction of the moisture gradient between the egg and the incubator (Tullet and Burton, 98; Swann and Brake, 99; Lundy, 969; Gildersleeve, 984). After hatching, the chicks from cooled eggs continued to lose more than the chicks from eggs not cooled, although the holding was not different from the control. This loss could indicate a higher susceptibility to dehydration of chicks from TABLE 8. Embryonic mortality of White Leghorn eggs exposed to low temperature during incubation, Experiment Effect Embryo d 4 d 6 d age Exposure 4 h 48 h Mortality Age by d x 4 h.64 d x 48 h d x 4 h.4 4 d x 48 h d x 4 h. 6 d x 48 h Downloaded from on 7 December 7

7 9 SUAREZ ET AL. TABLE 9. Embryonic mortality of broiler breeder eggs exposed to low temperature during incubation, Experiment Effect Before After Embryo age 8 d d 6 d Exposure 4 h 6 h Age by 8 d x 4 h 8 d x 6 h d x 4 h d x 6 h 6 d x 4 h 6 d x 6 h cooled eggs. Cooled groups differed from each other, which is consistent with results reported by Kuhn et al. (98) and might indicate differences in body mass due to age of the embryos when they were cooled (Decuypere, 979). Results showing effects on embryonic mortality are shown in Tables 7 through. A significant curvilinear effect was found in embryonic mortality in Experiments and due to age of the embryos, and in Experiment (Tables 8, 9, and ) due to crosses (genotype). In addition, these results suggested higher susceptibility of younger embryos to longer periods of cooling. Hours of were significant in Experiments,, and (Tables 7, 8, and ). Eggs cooled for 7 h (Experiment ) had % embryonic mortality. A strong, interaction of age at which the embryo is cooled with hours of and temperature on embryonic mortality was established by Webb (987). Moreng and Bryant (94) indicated that older embryos were more susceptible to cooling. Martin and Insko (9) found early and late periods to be critical for turkey embryos. Romanoff and Romanoff (97) noted that older embryos were more susceptible to high temperatures, whereas younger embryos were more affected by low temperatures. Moreng and Bryant (96) indicated a high degree of embryonic susceptibility to cooling on Days and of incubation of eggs exposed to to C for 4 and 48 h at different stages of embryonic development. In our study, embryonic mortality indicated that embryos were more susceptible to cooling on Days 8, 4, and 8, and with s of 6 and 48 h. When was 48 h, unhealed navels and abnormal down feathers were noted (visual observations); a similar situation was reported by Lancaster and Jones (988). Intermittent for a total of 48 h was not detrimental to survival (Table ). Differences from previous results could be due to differences in experimental procedure, genotype, not turning of 8 d embryos during the period, or major physiological changes in the embryos when they were cooled (Moreng and Bryant, 96). Cooling eggs during incubation has been suggested as a tool to manipulate incubation in order to meet conveniences of commercial hatcheries and to improve hatchability by reduction of heat stress on the embryo (Sarpong and Reinhart, 98; Lancaster and Jones, 988; Tullet, 99). Results obtained in the current study indicated that embryos from eggs cooled for an extended : ) lost more during incubation and the chicks were more susceptible to TABLE. Embryonic mortality and orthogonal contrasts of broiler breeder eggs exposed to low temperature during incubation, Experiment 4 Treatment eggs hatched not hatched mortality Exposure every 48 h Exposure every 96 h Orthogonal contrasts vs h Exposure 48 vs 96 h df Chi-square Probability x CATMOD procedure. Downloaded from on 7 December 7

8 EMBRYONIC DEVELOPMENT AFTER LOW TEMPERATURE EXPOSURE 9 TABLE. Embryonic mortality of broiler breeder eggs exposed to low temperature during incubation, Experiment Mortality Effect Before After Cross (female x male) A B A B A C 4 A 4 D A E Age d 6 d Exposure 4 h 48 h dehydration during holding, with a consequent lower of the chicks at hatching and servicing; ) have a longer incubation period; and ) have higher mortality. Egg cooling during incubation should be used only after consideration is given to increased mortality, effects on chick quality, and effectiveness in delaying hatching. Management strategies that must be decided include temperature,, the stage of embryonic development for cold, and use of single or intermittent periods. ACKNOWLEDGMENT Special thanks are extended to Arbor Acres, Inc. for furnishing broiler hatching eggs used in these studies. REFERENCES Barott, H. G., 97. Effect of temperature, humidity, and other factors on hatch of hen's eggs and on energy metabolism of chick embryo. USDA Technical Bulletin Number, Washington, DC. Booth, D. T., and H. Rahn, 99. Effect of periodic egg cooling during incubation. Page in: Avian Incubation. S. G. Tullett, ed. Butterworth-Heinemann Ltd., Surrey, UK. Callebaut, M. E., 99. Research note: Hatching of Japanese quail chicks (Coturnix japonica) following long daily cyclical interruption of their incubation. Poultry Sci. 69: 4-4. Decuypere, E., 979. Effect of incubation temperature patterns on morphological, physiological and reproduction criteria in Rhode Island Red birds. Agricultura 7:6-68. Decuypere, E., and H. Michels, 99. Incubation temperature as a management tool: A review. World's Poult. Sci. J. 48: 8-8. Geers, R., H. Michels, G. Nackaerts, and F. Konings, 98. Metabolism and growth of chickens before and after hatch in relation to incubation temperature. Poultry Sci. 6: Gildersleeve, R. P., 984. The effect of humidity and broiler strain on egg losses during incubation. Poultry Sci. 6:4-44. Kosin, I. L., 964. Recent research trend in hatchability-related problems of the domestic fowl. World's Poult. Sci. J. : Kuhn, E. R., E. Decuypere, L. M. Colen, and H. Michels, 98. Posthatch growth and development of circadian rhythm for thyroid hormones in chicks incubated at different temperatures. Poultry Sci. 6:4-49. Lancaster, F. M., and D. R. Jones, 988. Cooling of broiler hatching eggs during incubation. Br. Poult. Sci. 9: Landauer, W., 967. The hatchability of chicken eggs as influenced by environment and heredity. Storrs Agr. Exp. Sta. Monograph (Revised). Storrs, CT. Littell, R. C, R. J. Freund, and P. C. Spector, 99. SAS System for Linear Models, Third Edition. SAS Institute Inc., Cary, NC. Lundy, H., 969. A review of the effects of temperature, humidity, turning, and gaseous environment in the incubator on the hatchability of the hen's egg. Pages 4-76 in: The Fertility and Hatchability of the Hen's Eggs. T. C. Carter and B. M. Freeman, ed. Oliver and Boyd, Edinburgh, UK. Martin, J. J., and W. M. Insko, Jr., 9. Incubation experiments with turkey eggs. Kentucky Agric. Exp. Sta. Bull. 9: -77. McNally, H. W., and T. C. Byerly, 96. Variation in development of embryos of hen's eggs. Poultry Sci. : 8-8. Moreng R. E., and R. L. Bryant, 94. Effect of sub-freezing temperature on the chicken embryo.. Hatchability, chick and survival to six weeks. Poultry Sci. : Moreng, R. E., and R. L. Bryant, 96. The resistance of the chicken embryo to low temperature. Poultry Sci. :7-77. Oppenheim, R. W., and H. L. Levin, 97. Short-term changes in incubation temperature: behavioral and physiological effects in the chick embryo from 6 to days. Develop. Psychobiol. 8():-. Downloaded from on 7 December 7

9 9 SUAREZ ET AL. Romanoff, A. L., and A. J. Romanoff, 97. Pathogenesis of the Avian Embryo. John Wiley and Sons, New York, NY. Sarpong, S., and B. S. Reinhart, 98. Broiler hatching stress and subsequent growth performance. Poultry Sci. 64: -4. SAS Institute, 989. SAS User's Guide: Statistics. SAS Institute Inc., Cary, NC. Steel, R.G.D., and J. H. Torrie, 98. Principles and Procedures of Statistics. McGraw Hill Book Co., Inc., New York, NY. Swann, G. S., and J. Brake, 99. Effect of incubation dry-bulb and wet-bulb temperatures on of hatch and chick at hatch. Poultry Sci. 69: Taylor, L. W., C. A. Gunns, and B. D. Moses, 9. The effect of current interruption in electrical incubation. California Agric. Exp. Sta. Bull. :-9. Tazawa, H., and H. Rahn, 987. Temperature and metabolism of chick embryo and hatchling after prolonged cooling. J. Exp. Zool. (Suppl.) :-9. Tazawa, H., H. Wakayama, J. Turner, and C. V. Paganelli, 988. Metabolic compensation for gradual cooling in developing embryo. Comp. Biochem. Physiol. 89A: -9. Tullet, S. G., 99. Science and the art of incubation. Poultry Sci. 69:-. Tullet, S. G., and F. G. Burton, 98. Factors affecting the and water status of the chick at hatch. Br. Poult. Sci. :6-69. Webb, D. R., 987. Thermal tolerance of avian embryos: A review. The Condor 89: Williams, C, G. F. Godfrey, and R. B. Thompson, 9. The effect of rapidity of hatching on growth, egg production, mortality, and sex ratio in the domestic fowl. Poultry Sci. : Wilson, H. R., 99a. Effect of egg size on hatchability, chick size, and posthatching growth. Pages 79-8 in: Avian Incubation. S. G. Tullet, ed. Butterworth-Heinemann Ltd., Surrey, UK. Wilson, H. R., 99b. Interrelationship of egg size, chick size, posthatching growth, and hatchability. World's Poult. Sci. J. 47:-. Downloaded from on 7 December 7