ULTRASTRUCTURAL MEASUREMENTS OF THE EGGSHELL AS A TOOL TO DETECT THE CHICKEN BREED EFFECT ON EGGSHELL STRENGTH TRAIT

Transcription

1 ULTRASTRUCTURAL MEASUREMENTS OF THE EGGSHELL AS A TOOL TO DETECT THE CHICKEN BREED EFFECT ON EGGSHELL STRENGTH TRAIT El-Safty, S.A., **Fathi, M.M., **Zein El-Dein, A., and **Lamiaa, M. Radwan ABSTRACT This trial was carried out to detect and investigate the influence of chicken breed (Fayoumi and Dandarawi as Egyptian native breeds) on the ultrastructural properties of eggshells by using scanning electron microscope (SEM) technique. At 26 weeks of age, twelve samples of eggshells were randomly taken from the eggs collecting from the two breeds (6 each). The eggshell samples were prepared according to the scanning electron microscopy procedures to investigate ultrastructural variants. The main results obtained could be summarized as follows: The eggshell of Fayoumi breed had a significantly higher effective thickness (palisade layer %) compared to Dandarawi counterpart. Per contra, both absolute and relative mammillary layer length was significantly lower in eggshells of Fayoumi breed compared to those of Dandarawi ones. With regard to total structural score, the results revealed that Fayoumi breed had a significant better structural score (lower value) to overall ultrastructural traits. Where, the corresponding figures were 22.5 and 27.5 for Fayoumi and Dandarawi breeds, respectively. It can be concluded that the eggs of Fayoumi breed were required high force to break its eggshell, consequently, increased shell strength compared to Dandarawi breed. The previous result was attributed to Fayoumi breed was a better ultrastructural of eggshell and shell quality compared to the eggshells of Dandarawi breed. Key words: Ultrastructural traits, eggshell, Fayoumi and Dandarawi breeds, shell strength *Animal Prod. Dept., Fac. of Agric., Sebha Univ., Sebha, Libya - **PoultryProd.Dept., Fac. of Agric., Ain Shams Univ., Cairo, Egypt Journal of Sebha University-(Pure and Applied Sciences)-Vol.11 No.1 (2012)

2 ULTRASTRUCTURAL MEASUREMENTS OF INTRODUCTION: Ultrastructural traits of eggshell has greatly enhanced our understanding of the eggshell architecture and has reinforced the view that the mechanical - properties of eggshell cannot be defined by a simple eggshell thickness measurement. Extensive studies were conducted to evaluate the ultrastructural variations in eggshell quality among breeds and species of poultry (Bain, 1990; Solomon, 1991; Solomon, 1999; Ruiz and Lunam, 2000; Fathi, 2001 and Afifi et al., 2007). In this context, Solomon (1991 and 1999) stated that the organization of the palisade column is a major determinant of shell stiffness and thereby of shell strength. Shell strength is directly related to shell thickness, therefore, it is likely that alteration in the thickness of the palisade layer, independent of structural reorganization of the palisade columns could affect shell strength (Ruiz and Lunam, 2000). Genetic differences in eggshell formation characteristics exist between breeds, strains and families within the species (Fathi, 2001; Anderson et al., 2004; El-Safty, 2004; Bain, 2005; Bain et al., 2006a; Afifi et al., 2007). According to Dunn and Waddington, (2006), up to 10% of eggs laid are lost through cracking during routine egg handling procedures. From an economic stand point of view, the broken eggs lose sustained by the UK poultry industry in 2005 could have amounted to as much as 16.7 million (Bain et al., 2006b). Many researches was conducted to evaluate the laying performance of all Egyptian local breeds, but a few data were published on the new techniques to measure ultrastructural measurements of eggshell. So that, this paper aimed to test the chicken breed (Fayoumi and Dandarawi) impact on ultrastructural traits of eggshell, and consequently eggshell strength using scanning electron microscopy (SEM). MATERIALS AND METHODS: Experimental procedures: Layers management: Two Egyptian pure lines chicken (Fayoumi and Dandarawi) were raised at the Poultry Breeding Farm, Ain Shams University. At the beginning of the experiment (16 weeks of age), one hundred and sixty two chickens (97 Fayoumi and 65 Dandarawi) were randomly taken Journal of Sebha University-(Pure and Applied Sciences)-Vol.11 No.1 (2012)

3 El-Safty, S.A., Fathi, M.M., Zein El-Dein, A., and Lamiaa, M. Radwan They were housed in suitable individual cages from 16 weeks of age up to the end of the experiment (50 weeks of age). They were reared under similar environmental, managerial and hygienic conditions. They were fed a laying diet containing 16.3% CP and 2900 kcal ME/kg diet and water were offered ad libitum. They were exposed to16h light per day throughout the experiment. Preparation of Samples for ultrastructural analysis using scanning electron microscopy (SEM): At 26 weeks of age, twelve samples of eggshell were randomly taken from Fayoumi and Dandarawi breeds (6 each) to detect ultrastructural variations. The specimens were prepared by cutting a piece (1 cm 2 ) of shell from the equatorial region of each egg. The shell membranes were carefully removed by first soaking in water. The loosely adhering membranes were then gently peeled from the edge of the sample inwards. To remove the remaining tightly bound membrane fibers, each sample was then immersed overnight in 6 % sodium hypochlorite, 4.12% sodium chloride and 0.15% sodium hydroxide. Thereafter, the specimen was rinsed with water and left to dry at room temperature. Following these preparative treatments, two samples from each egg were mounted in inner side uppermost and in vertically manner on aluminum stubs, coated with gold for 3 min in an Emscope Sputter Coater. These samples were examined using JEOL JSM-T330A scanning electron microscopy at 15 Kv. The incidence of ultrastructural variants (Table 1) at the level of the mammillary layer was assessed according to the methodology and terminology developed by the Poultry Research Unit, University of Glasgow (Bain, 1990, 1992 and Solomon, 1991). The cross-sectional lengths of palisade and mammillary layers were directly measured in µm using scaling software provided with the SEM at a magnification of x200. The total thickness of each specimen was measured as the distance from its' outermost surface to the point where the basal caps inserted into the shell membranes. The thickness of the mammillary layer was also assessed, this being the distance from the basal caps to the point at which the palisade columns first fused. Subtraction of these two measures provided a length of the palisade thickness or effective thickness (Bain, 1990 and Solomon, 1991). Triplicate measures were performed in each case and the mean Journal of Sebha University-(Pure and Applied Sciences)-Vol.11 No.1 (2012)

4 ULTRASTRUCTURAL MEASUREMENTS OF values were used in the statistical analysis. Structural variation in the eggshell which alter resistance to breakage: two categories; firstly the structural variants which decrease resistance to breakage, and secondly the structural variants which increase resistance to Ultrastructural measurements of breakage. mammillary layer were divided into Table (1): The structural variations which occur at the mammillary layer. Decrease resistance to crack Increase resistance to crack 1- Late fusion 1-Early fusion 2- Type As 2- Good cap formation (i.e., close binding between 3- Type Bs organic and inorganic fraction of shell). 4- Aragonite 3-Cuffing 5- Pitting : depression, erosion,pin holes 4-Confluent mammillae 6- Alignment 5-A high mammillary density 7- Cubics 8- Changed membrane 9- A low mammillary Statistical analysis: Data were subjected to a one-way ANOVA using GLM (SAS Institute, 2001) with breed as fixed effect. Data of ultrastructural traits are presented as means and the pooled SEM. RESULTS AND DISCUSSION: Ultrastructural examination of eggshells has greatly enhanced our understanding of the eggshell architecture and has reinforced the view that the mechanical properties of eggshells cannot be defined by a simple thickness measurement. As can seen from Table (2), Fayoumi breed had satisfied confluence and early fusion traits (the main factors affecting stiffness of eggshells) as compared to Dandarawi breed. The terms of fusion, alignment, erosion and good caps measurements were significantly differed between the two breeds. Where, Fayoumi breed had satisfied good caps and early fusion as compared to Dandarawi one, so that the effective thickness of the shell and shell thickness without membranes significantly increased in Fayoumi eggs compared to Dandarawi counterparts. As depicted in Fig (1) the mammilary caps were make good attachment with membranes (extensive confluence) and good cap formation for Fayoumi breed, while poor Journal of Sebha University-(Pure and Applied Sciences)-Vol.11 No.1 (2012)

5 El-Safty, S.A., Fathi, M.M., Zein El-Dein, A., and Lamiaa, M. Radwan attachment with membranes (weak confluence) was observed in eggshell of Dandarawi breed (Fig.2). Solomon (1999) found good shell ultrastructural beneficial high confluence reflects good attachment with membranes and caps. A good eggshell quality should have mammillary bodies that are even in size and distribution and rounded so that there can be maximum attachment to the fibers of the outer membrane (El-Dlebshany et al., 2007). Early fusion between mammillary cones increased in Fayoumi breed compared to Dandarawi breed, the difference between breeds was significant. Fig. (3) indicates the cones contact with them was earlier, so the effective thickness increased, whereas the late fusion between cones was extremely exist in the eggshell of Dandarawi breed (Fig.4). The last result confirmed that Fayoumi breed significantly increased the effective thickness (stiffness shell), because increased early fusion compared to Dandarawi breed. Bain (1991), Fathi (2001) and El- Safty (2004) reported that the mammillary layer does not contribute to the stiffness of shell but late fusion of adjacent palisade columns is related to fracture toughness. Regarding alignment term, it could be stated that the alignment of the mammillary bodies was significantly increased in Dandarawi breed compared to Fayoumi counterparts. Thereby, Fayoumi breed required high force to crack due to force required to crack columns it s very strong, so the crack almost path way between alignment (Fig. 5). If an egg is subjected to mechanical trauma then the crack line will tend to follow the path of least resistance and ordered mammillae provide such a pathway (Fathi, 2001 and El-Safty, 2004). As shown in Fig (6), the Dandarawi breed increased the occurrence of erosion compared to Fayoumi one. So that the cone of Fayoumi eggshell was stronger than that of Dandarawi eggshell. Accordingly, the eggshell of Fayoumi was high resistant to crack than Dandarawi eggshell. The total structural score estimate proved to be a reliable measurement to determine the ability of a shell to resist the load (this estimate is ranking from one which is the best to five which is the worst). Because no feature could be isolated as the causative factor of lower resistance and because in every single shell a number of structural modifications will be present, only their overall magnitude, represented by the total structural score, could ultimately determine the probable Journal of Sebha University-(Pure and Applied Sciences)-Vol.11 No.1 (2012)

6 ULTRASTRUCTURAL MEASUREMENTS OF performance of the shell as a barrier to bacterial penetration (Fathi, 2001 and Bain et al., 2006a). The results revealed that Fayoumi breed had a significant better structural score (lower value) to overall ultrastructural traits. Where, the corresponding figures were 22.5 and 27.5 for Fayoumi and Dandarawi breeds, respectively. Consequently, Fayoumi breed was required high force to break its eggshell, consequently, increased shell strength compared to Dandarawi breed. The last result was attributed to Fayoumi breed was a better ultrastructural of eggshell and shell quality compared to Dandarawi breed. As presented in Table (3), the eggshell of Fayoumi breed was significantly higher palisade layer (absolute or relative) compared to Dandarawi breed. On the other hand, the mammillary layer (absolute or relative) was significantly lower in eggshells of Fayoumi hens compared to those of Dandarawi counterpart. Generally, these data revealed that the proportion of mammillary and palisade layers is dependent on shell thickness. Moreover, the proportion of palisade layer could be played an important role in shell stiffness in both breeds. According to Bain (1991) and Ruiz and Lunam (2000), the palisade layer provides the stiffness characteristics of the shell and thereby shell strength. Thus, a reduction in its relative thickness could compromise shell strength leading to a higher incidence of breakage. Additionally, Bain et al. (2006a) reported that the eggshell is consist of several different layers, and proposed that each of these different layers must variously contribute to the eggs performance under load. Furthermore, the term of effective thickness (palisade layer) must conserve as a tool for selection programs in both broiler breeders and parent stock of layers. Finally, it could be recommended to use ultrastructural traits of eggshell to distinguish among breeds or strains for eggshell strength traits, and consequently using those traits as tools in selection programs for eggshell characteristics and better eggshell architecture Journal of Sebha University-(Pure and Applied Sciences)-Vol.11 No.1 (2012)

7 El-Safty, S.A., Fathi, M.M., Zein El-Dein, A., and Lamiaa, M. Radwan Table (2): Impact of breed on ultrastructural variants of eggshell mammillae for Fayoumi and Dandarawi breeds. Breed Variant Fayoumi Dandarawi Pooled SEM Prob. Confluence Fusion Cuffing Alignment Type Bs Depression Erosion Cubics Aragonite Caps Type As Changed membrane Total score Table (3): Impact of breed on cross-sectional length (µm) of eggshell mammillary layer. Breed Layer Fayoumi Dandarawi Pooled SEM Prob. Palisade Mammillary Total Palisade % Mammillary % Palisade Mammillary Palisade % = X 100 Mammillary % = X 100 Total Total Journal of Sebha University-(Pure and Applied Sciences)-Vol.11 No.1 (2012)

8 ULTRASTRUCTURAL MEASUREMENTS OF المقاييس البنائية لقشرة البيضة كأداة الكتشاف تأثير ساللة الدجاج على صفة قوة ومتانة قشرة البيضة *صالح الدين عبد الرحمن ألصفتي- **معتز محمد فتحي- **علي زين الدين- **لمياء رضوان *قسم اإلنتاج الحيواني- كلية الزراعة- جامعة سبها- ليبيا ** قسم إنتاج الدواجن- كلية الزراعة- جامعة عين شمس- مصر الم ستخلص أخر ت هذه التخربة لالكتشاف والتحقق من تأث ر ساللة الدخاج )من النوع الف وم والدنداراوي كسالالت محل ة مصر ة أص لة( على الصفات البنائ ة لقشرة الب ضة وذلك باستتددا تقن تة الم كروستكو االلكترونت الماست عند ع مر 12 أسبوع ت خمع 71 ع نتة متن وشتور البت و عشتوائ اك متن كلتتا الستاللت ن )2 متن كتس ستاللة(ت وتت إعتداد تلتك الا نتات للفحت إلبقتاك للختراةات الم تباتة ات تحضت ر ع نتات الفحت بالم كروستكو االلكترونت الماس وذلك للكشف عتن االدتالاتات البنائ تة بقشتور بت و الستاللت ن كانتت أهت النتتائح الم تحصتس عل تا علتى النحو التال : Journal of Sebha University-(Pure and Applied Sciences)-Vol.11 No.1 (2012)

9 El-Safty, S.A., Fathi, M.M., Zein El-Dein, A., and Lamiaa, M. Radwan تفوق بت و ستاللة التدخاج الف توم بدرختة مانو تة ات مق تال س تمك القشترة المت ثر الم ابتر عتن النستبة المئو تة للإلبقة الوتد ة بالقشرة وذلك م قارنة بما هو عل ه ا ساللة الدنداراوي على الاكل من ذلك اقد لوحظ اندفتاو النسبة والق مة الم إللقتة لإلتوس الإلبقتة الحلم تة بالقشترة بدرختة مانو تة ات التدخاج الف توم م قارنتة ببت و دختاج الدنداراوي أما بالنسبة لمخموع النقاإل الكل للبن ة الترك ب ة لقشرة الب ضةت اقتد أوضتحت النتتائح تستخ س ستاللة الف تتوم تقتتد راك أاضتتس )مخمتتوع النقتتاإل ا(وتتس( م قارنتتة بستتاللة الدنتتداراويت ح تتم ستتخلت ستتاللت الف تتوم والدنتداراوي 1122 و 1122 درختة علتى الترت ت وبشتكس عتا اتذن نتتائح هتذه الدراستة تدلت إلتى أن البت و الم نتح من ساللة الف وم حتاج لقدر أكبتر متن القتوة ل تت كستره نت ختة لق تادة وتوة ومتانتة وشترة الب ضتة م قارنتة بالب و الم نتح من ساللة الدنتداراويت كمتا أكتدت نتتائح الفحت بالم كروستكو االلكترونت الماست أن النت ختة الستتابقة ت اتتقو لقتتوة الترك تت البنتتائ وبالتتتال ختتودة وشتترة البتت و الم نتتتح متتن ستتاللة الف تتوم م قارنتتة بستتاللة الدنداراوي الكلمات الم فتاحية: مقا ل الترك البنائ ت وشرة الب وت دخاج الف وم والدنداراويت ووة القشرة REFERENCES Afifi, Y.K.; S.A. El-Safty; M.M. Fathi. (2007). Assessment of eggshell quality measurements in some Egyptian local breeds of chicken using scanning electron microscopy.1-early egg production cycle. Proceedings of 4 th World s Poult. Conf. Sharm El-Sheikh, Egypt. Pp Anderson, K.E.; J.B. Tharrington; P.A. Curtis and F.T. Jones (2004). Shell characteristics of eggs from historic strains of single comb White Leghorn chickens and the relation of egg shape to shell strength. Int. J. of Poult. 3: Bain, M.M. (1990). Eggshell strength: a mechanical / ultrastructural evaluation. Ph.D. Thesis, University of Glasgow,UK. Bain, M.M. (1992). Eggshell strength a relationship between the mechanism of failure and ultrastructure organization of the mammillary layer. Br. Poult. Sci. 33: Bain, M.M. (2005). Recent advances in the assessment of eggshell quality and their future application. World s Poult.Sci. J. 61: Bain, M.M.; N. Macleod; Thomson; and J.W. Hancock (2006a). Microcracks in eggs. Poult. Sci. 85: Bain, M.M.; I.C. Dunn; P.W. Wilson; N. Joseph; B. De Ketelaere; J. De Journal of Sebha University-(Pure and Applied Sciences)-Vol.11 No.1 (2012)

10 ULTRASTRUCTURAL MEASUREMENTS OF Baerdemaeker and D. Waddington (2006b). Probability of an egg cracking during packing can be predicted using a simple nondestructive acoustic test. Br. Poult. Sci., 47: Dunn, I. and D. Waddington (2006). Materials science meets genetics to reduce egg losses. An article. Poult. Int. 10: El-Dlebshany, A.E.; S.A. El-Safty; M. Bahie El-Deen; M.M. Fathi. (2007). Ultrastructural evaluation of eggshell in Japanese quail genetically selected for higher egg production. Proceedings of 4 th World s Poult. Conf. Sharm El-Sheikh, Egypt. Pp El-Safty, S.A. (2004). Stepwise regression analysis for ultrastructural measurements of eggshell quality in two local breeds of chicken. Egypt. Poult. Sci.24: Fathi, M.M. (2001). Ultrastructural evaluation of eggshell quality produced from two different genetic groups of chicken. Egypt. Poult. Sci. 21: Ruiz, J. and C.A. Lunam (2000). Ultrastructural analysis of the eggshell: contribution of the individual calcified layers and the cuticle to hatchability and egg viability in broiler breeds. Poult. Sci., 41: SAS Institute (2001). SAS/STAT User s Guide:Statistics. Ver.8.2,SAS Institute Inc.,Cary,NC Solomon, S.E. (1991). Egg and eggshell quality, Wolfe Publishing Ltd. London. Solomon, S.E. (1999). An egg ist ein ei, es un huevo, est un oeuf. Br. Poult. Sci. 40: Journal of Sebha University-(Pure and Applied Sciences)-Vol.11 No.1 (2012)