ABSTRACT
A study was conducted using 231 chicks (101 Normal feather, 56 frizzle and 74 Naked neck) to evaluate growth traits and semen quality characteristics using biochemical and IGF-1 as molecular marker on three genotypes (normal, frizzle and naked neck) of Nigeria indigenous chickens. Each of the genotypes comprised of twenty five hens and five roosters mated in the ratio of 1 rooster: 5 hens (1:5). Body weight (BW) and measurements of breast girth (BG), shank length (SL), body length (BL), wattle length (WL), wattle width (WW), comb length (CL), comb height (CH), wing span (WS) and shank diameter (SD) were taken at an interval of 4 weeks starting from 8 weeks of age. Semen quality characteristics (volume, colour, pH, concentration and live and dead ratio) and morphological defects (acrosoma swelling, detached head, mid piece drop, bent tail, curled tail and full tail) were evaluated thrice at 24, 28 and 32 weeks of age, respectively. Blood samples for the haematological, biochemical and molecular studies (IGF-1) were collected at 20 weeks of age. The data collected were subjected to General Linear Model procedure of SAS. The result showed that mean BW between 8 and 20 weeks ranged from 310.46g to 1117.80g and between 1096.80g and 1285.80g from 24 to 32 weeks with significant (P<0.05) differences with the normal feathered birds having the superior BW between 20 to 32 weeks of age. However, apart from BW and BL which were significant at all ages, other body linear measurements were not significantly (P>0.05) different at all the ages (8 to 32 weeks). The semen quality characteristics for the parameters were significantly (P<0.05) different except for semen colour and semen pH which were not significant (P>0.05) affected by the age of the birds. The morphological
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defects showed that most of them were significantly (P<0.05) different for all the genotypes, but the mid piece was not significantly (P>0.05) different at 32 weeks. The results of the haematological and biochemical profile show that only Chloride in the biochemical parameter was significantly (P<0.05) different. The phenotypic correlation coefficients between traits show that some traits had positive correlation but not statistically significant (P>0.05) while others traits showed between moderate and high correlations amongst themselves. Two alleles; A and B and three genotypes: AA, AB and BB were observed with the A allele (0.72) being predominant and conforms to Hardy-Weinberg equilibrium. The relationship among IGF-1 marker, BW and semen quality was significant (P<0.05). The AA genotype was superior in body weight over the AB and BB genotypes at 28 and 32 weeks. Therefore, it is concluded that the normal feather genotype did better in terms of BW and body linear measurements (BLM). The semen characteristics revealed that as age increased the quality improved up to 28 weeks, while the IGF-1 gene had a positive relationship with BW and semen characteristics making it a good genetic marker for selection for growth traits (BW) and semen characteristics in the Nigerian indigenous chickens.
TABLE OF CONTENTS
TITLE PAGE .……………………………………………………………………………….i FLY PAGE ………………………………………………………………………………….ii
COVER PAGE……………………………………………………………………………..iii
DECLARATION ………………………………………………………………………………………………. iii
CHAPTER ONE ……………………………………………………………………………………………….. 1
1.0 INTRODUCTION ……………………………………………………………………………………….. 1
1.1 JUSTIFICATION ………………………………………………………………………………………… 6
1.2 HYPOTHESES ……………………………………………………………………………………………. 8
1.3 OBJECTIVES……………………………………………………………………………………………… 9
CHAPTER TWO…………………………………………………………………………..10
2.0 LITERATURE REVIEW …………………………………………………………………………….. 10
2.1 HISTORICAL ORIGIN OF THE INDIGENOUS CHICKENS…………………………… 10
2.2 GROWTH PERFORMANCE OF THE INDIGENOUS CHICKENS …………………… 10
2.2.1 Body Weight and Body Linear Measurements ………………………………………………. 11
2.3 THE TESTIS ……………………………………………………………………………………………… 13
2.3.1 The Male Anatomy …………………………………………………………………………………… 13
2.3.2 Spermatogenesis ………………………………………………………………………………………. 13
2.3.3 Production of Semen in Cocks ……………………………………………………………………. 14
2.3.4 Semen Composition ………………………………………………………………………………….. 15
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2.4 SEMEN QUALITY CHARACTERISTICS …………………………………………………….. 16
2.4.1 Semen Volume and Concentration ………………………………………………………………. 16
2.4.2 Semen Motility, pH and Colour ………………………………………………………………….. 17
2.4.3 Semen Quantity and Testis Size ………………………………………………………………….. 19
2.5 MORPHOLOGICAL DEFECTS ASSOCIATED WITH SEMEN QUALITY ………. 20
2.6 CORRELATION OF BIOCHEMICAL AND HAEMATOLOGICAL WITH SEMEN CHARACTERISTICS IN THE NIGERIAN INDIGENOUS COCKERELS …………. 21
2.7 PHENOTYPIC CORRELATION AMONG BODY WEIGHT, BODY LINEAR MEASUREMENTS AND SEMEN CHARACTERISTICS ……………………………….. 22
2.8 SELECTION FOR SEMEN QUALITY ………………………………………………………….. 23
2.9 SEMEN QUALITY ……………………………………………………………………………………… 24
2.10 FACTORS AFFECTING SEMEN QUALITY ………………………………………………… 25
2.10.1 Age and Strain …………………………………………………………………………………………. 25
2.10.2 Feed Nutrition ………………………………………………………………………………………….. 27
2.10.3 Ambient Temperature………………………………………………………………..28
2.10.4 Photoperiod …………………………………………………………………………………………….. 29
2.10.5 Semen pH…………………………………………………………………………….30
2.11 BLOOD CHEMISTRY………………………………………………………………………………… 30
2.12 HAEMATOLOGICAL PARAMETERS…………………………………………………………. 31
2.12.1 Packed Cell Volume (PCV)…………………………………………………………..31
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2.12.2 Haemoglobin (Hb) ……………………………………………………………………………………. 32
2.12.3 Mean Corpuscular Volume (MCV), Mean Corpuscular Haemoglobin (MCH) and Mean Corpuscular Haemoglobin Concentration (MCHC) ……………………………….. 33
2.13 BIOCHEMICAL PARAMETERS …………………………………………………………………. 34
2.14 INSULIN GROWTH FACTOR ……………………………………………………………………. 35
2.14.1 The IGF ligands ……………………………………………………………………………………….. 35
2.14.2 Insulin Growth-like Factor-1 (IGF-1) ………………………………………………………….. 36
2.14.3 Allele Frequency and Genotypic Frequency in Different Chicken Populations …… 37
2.14.4 Associations of IGF-1 Genes in Chicken with Growth Traits and Reproduction …. 38
CHAPTER THREE …………………………………………………………………………………………. 39
3.0 MATERIALS AND METHODS ………………………………………………………………… 39
3.1 LOCATION OF THE STUDY AREA …………………………………………………………….. 39
3.2 EXPERIMENTAL BIRDS …………………………………………………………………………….. 39
3.2.1 Base Population (Foundation Stock) ……………………………………………………………… 39
3.2.2 Housing and Management …………………………………………………………………………… 40
3.2.3 Vaccination and de-worming ……………………………………………………………………….. 40
3.2.4 Mating and Egg Collection ………………………………………………………………………….. 40
3.2.5 Incubation Process …………………………………………………………………………………….. 42
3.2.6 Brooding and Rearing of the Progeny ……………………………………………………………. 42
3.2.7 Management of F1Generation ……………………………………………………………………… 42
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3.3 BODY WEIGHT AND LINEAR BODY MEASUREMENTS…………………………46
3.4 SEMEN COLLECTION AND EVALUATION ……………………………………………….. 48
3.4.1 Ejaculate Volume …………………………………………………………………………………….. 48
3.4.2 Ejaculate Colour and Consistency ……………………………………………………………….. 49
3.4.3 Sperm Motility…………………………………………………………………………………………. 49
3.4.4 Sperm Concentration ………………………………………………………………………………… 51
3.4.5 Semen pH ……………………………………………………………………………………………….. 52
3.4.6 Live and Dead Ratio …………………………………………………………………………………. 52
3.4.7 Sperm Morphological Defects …………………………………………………………………….. 52
3.5 BLOOD COLLECTION AND EVALUATION ………………………………………………… 53
3.5.1 Evaluation of Haematological Parameter ……………………………………………………… 54
3.5.2 Evaluation of Biochemical Parameter ………………………………………………………….. 54
3.5.3 Insulin-like Growth Factor-1 (IGF-1) DNA Isolation ……………………………………… 54
3.6 Statistical Analysis ……………………………………………………………………………………….. 58
CHAPTER FOUR ……………………………………………………………………………………………. 61
4.0 RESULTS ……………………………………………………………………………………………… 61
4.1 GROWTH PERFORMANCE AND SEMEN QUALITY CHARACTERISTICS OF THE THREE GENOTYPES …………………………………………………………………………. 61
4.1.1 Body Weight and Growth Traits …………………………………………………………………. 61
4.1.2 Semen Characteristics of the Three Genotypes ………………………………………………. 65
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4.1.3 Morphological Defects ………………………………………………………………………………. 66
4.1.4 Interaction between Age and Genotype on the Semen Quality Characteristics ……… 66
4.2 LEAST SQUARE MEANS (±SE) FOR BIOCHEMICAL AND HAEMATOLOGICAL PARAMETERS OF THE COCKERELS ………………………………………………………… 70
4.3CORRELATION BETWEEN BIOCHEMICAL, HAEMATOLOGICAL PARAMETERS AND SEMEN QUALITY OF THE NIGERIAN INDIGENOUS COCKERELS…………………………………………………………………………..74
4.4 PHENOTYPIC CORRELATION BETWEEN GROWTH TRAITS AND SEMEN QUALITY IN THE NIGERIAN INDIGENOUS COCKERELS AT 24, 28 AND 32 WEEKS OF AGE ……………………………………………………………………………………….. 77
4.5 INSULIN GROWTH FACTOR-1 (IGF-1) POLYMORPHISM AND ASSOCIATION WITH BODY WEIGHT AND SEMEN QUALITY ………………………………………….. 82
4.5.1 Insulin-like Growth Factor-1 (IGF-1) Polymorphic Forms ………………………………… 82
4.5.2 Association of the different Alleles on IGF-1 Gene, Body Weight and Semen Characteristics of the Indigenous Cockerels ………………………………………………….. 84
CHAPTER FIVE ……………………………………………………………………………………………… 86
5.0 DISCUSSION…………………………………………………………………………………………. 86
5.1 MEAN PERFORMANCE ……………………………………………………………………………… 86
5.1.1 Body weight and growth traits ……………………………………………………………………… 86
5.1.2 Semen characteristics of the three genotypes …………………………………………………. 86
5.1.3 Morphological defects related to cockerels spermatozoa …………………………………. 88
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5.1.4 Interaction between age and genotypes of the Nigerian indigenous cockerels ……… 89
5.2 BIOCHEMICAL PARAMETERS OF THE THREE GENOTYPES …………………….. 90
5.3 HAEMATOLOGICAL PARAMETERS OF THE COCKERELS ………………………… 92
5.4 CORRELATION BETWEEN BIOCHEMICAL, HEAMATOLOGICAL PARAMETERS WITH SEMEN CHARACTERISTICS OF THE NIGERIAN INDIGENOUS COCKERELS ………………………………………………………………………. 93
5.5 PHENOTYPIC CORRELATION BETWEEN BODY WEIGHT, LINEAR BODY TRAITS AND SEMEN QUALITY OF THE NIGERIAN INDIGENOUS COCKRELS AT VARIOUS AGES ………………………………………………………………………………….. 95
5.6 INSULIN-LIKE GROWTH FACTOR 1 (IGF-1) POLYMORPHISM …………………… 96
5.7 VARIATION IN BODY WEIGHT AND SEMEN QUALITY WITH IGF-1 POLYMORPHIC FORMS AT DIFFERENT AGES …………………………………………. 96
CHAPTER SIX ……………………………………………………………………………………………….. 97
6.0 SUMMARY, CONCLUSIONS AND RECOMMENDATIONS ………………………… 97
6.1 SUMMARY ………………………………………………………………………………………………… 97
6.2 CONCLUSIONS………………………………………………………………………………………….. 99
6.3 RECOMMENDATIONS ………………………………………………………………………………. 99
REFERENCES ………………………………………………………………………………………………. 100
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CHAPTER ONE
1.0 INTRODUCTION
The poultry industry is a versatile business in Nigeria but the development has been to the detriment of the indigenous birds which have been neglected and not properly managed (Adebambo et al., 1999). The indigenous chickens (Gallus domesticus) have been kept in Africa for many generations in subsistence systems. The Nigerian indigenous chicken constitutes between 80 and 90 percent of the local population of chickens in Nigeria. Three other major genes have been identified in these local chickens apart from the normal feathered type. These are: Frizzle, naked neck and sex-linked dwarfism. Each of these genes plays a significant role in the productive adaptability of the Nigerian local chicken (Ozoje and Ikeobi, 1995). Despite being in Nigeria for long, the indigenous chickens consist of various unimproved sub-populations of heterogeneous characteristics, not yet classified into breeds and varieties since they do not breed true to type and have no clear plumage colours (Obioha, 1992; Ibe, 2001).
The indigenous chicken, also known as the local chicken is the general term given to those chickens kept on an extensive system, scavenging free-range, having no identified description, and being dual-purpose and unimproved (Horst,1989; Pedersen, 2002). They appear to be the cheapest and sustainable means of producing high quality animal protein for the expanding populations of developing countries like Nigeria. They are self reliant and hardy birds with the capacity to withstand harsh weather conditions and adaptation to adverse environments. They are valued for their ability to scavenge, disease tolerance,
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meat quality and general hardiness (Ssewayana et al., 2003); they also have the ability to hatch and brood. This is clear evidence of the positive attributes of the indigenous chickens. Their products are preferred by the majority of Nigerians because of the pigmentation, taste, leanness and suitability for special dishes (Horst, 1989). However, the Nigerian indigenous chickens are inferior in growth and egg production characteristics compared to the exotic chickens (Ajayi, 2010). Despite these drawbacks, the indigenous chicken plays a vital role in the socio-economic life of those who keep them. Apart from being a source of meat and egg, they are also a source of income for many resource-poor people particularly those living in the rural areas (Ssewayana et al., 2003). Growth is affected by genetic and non genetic factors (Singh and Singh, 1983; Gupta et al., 1988). Growth is a fundamental property of biological systems and it can be defined as an increase in the number of cells in the body size per unit of time (Schulze et al., 2002; Lawrence and Fowler, 2002). Growth of fowl is analogous to growth of mammalian; consisting of three or four cycles (Grossman, 1988), two of these cycles, however, occur after hatching.
Body weight and body conformations are two important parameters for measuring growth in the domestic chicken (Udeh and Ogbu, 2011). The price of chicken largely depends on body weight. The indigenous chicken, like the improved breed, has a sigmoid growth pattern with differences in growth rate and feed efficiency (Nwosu, 1979), although the indigenous fowl seemed to complete the rapid growth phase earlier than
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improved breeds (Oluyemi, 1980). However, there is generally no weighing machine or scale available in poultry markets in villages. It is important that the farmer monitors the growth performance of his birds on regular basis to know when they have attained the desired market weight. In places where scales are not available, as is the case in most rural African communities (Nesamvuni et al., 2000), linear body measurements such as shank length, drum stick length and wing length can be used in a predictive equation to predict body weight in broilers (Akanno et al., 2007). For instance, the reports of Kabir et al. (2006) showed that there was a positive interrelationship between body weight and shank length in Rhode Island chickens and similar observations were reported by Malik et al. (1997). Reproductive performance is critical for efficient production in poultry and suitable selection criteria for males based on semen characteristics in roosters (McDaniel et al., 1998). The male chicken is known to be responsible for fertilizing the eggs of a number of females and most studies concerning the reproductive efficiency of breeder birds have centered on the cocks and artificial insemination (AI) of large breeder hens for the production of hatching eggs. The artificial insemination is one method showing promise in improving reproductive efficiency of cocks (McDaniel, 1994; McCartney, 1996).
Semen quality of cocks determines the fertility of the male chicken while the female contributes the eggs (Liu et al., 2008). Body weight, length of shank, comb and wattle lengths have been reported to be good predictors of semen attributes in chicken (Galal, 2007). For good results in the artificial insemination of chickens, the quality of semen
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should be ensured (Tabatabaei et al., 2009). The importance of semen evaluation in poultry breeding for selecting breeding males or for routine monitoring their reproductive performance is well recognized (Cheng et al., 2002). The assessment of semen quality characteristics of poultry gives an excellent indicator of their reproductive potential and has been reported to be a major determinant of fertility and subsequent hatchability of eggs (Peters et al., 2004). Also, sperm morphology is used as one of the most important criteria in the evaluation of semen quality (Den Daas, 1992). Semen of the domestic fowl, according to Hafez (1990), varies from dense opaque suspension to a watery fluid with a relative high density. The author further stated that the differences in volumes and sperm concentration of the domestic fowl depends largely on the relative contribution of the various reproductive glands, the number of spermatozoa that could be obtained from a breed/strain and the extent to which the genetic potentials can be exploited. The use of blood examination as a way of assessing the health status of animals has been documented (Owoyele et al., 2003; Muhammad et al., 2004). This is because it plays a vital role in physiological, nutritional and pathological status of organisms (Muhammad et al., 2000; Muhammad et al., 2004) and it also acts in regulatory, protective and homeostatic tissue of an organism (Nasyrova et al., 2006; Eze et al., 2010).
Haematological parameters provide valuable information on the immune status of animals (Kral and Suchy, 2000). This information ranges from giving the level of the
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blood to detecting ailment or disorders through them. Such information, apart from being useful for diagnostic and management purposes, could equally be incorporated into breeding programmes for the genetic improvement of indigenous chickens. Glucose, cholesterol, calcium, total protein, alkaline phosphatase, uric acid, sodium, potassium and chloride levels are diagnostic values for diabetes mellitus, liver disease, hypoparathyroidism, chronic hepatopathy and liver disease, gout, kidney disease, chronic diarrhea and dehydration (Simaraks et al., 2004). Besides, haematological values are important to clinicopathological diagnosis such as traumatic injury, parasitism, organic disease, bacterial septicemia and nutritional deficiencies. Moreover, managing abnormalities in birds requires an understanding of how diseases change the biochemical function and electrolyte of the bodies. Because the clinical signs of illness in birds are frequently subtle, clinical chemistry is necessary to evaluate cellular changes (Ritchie et al., 1994).
The insulin-like growth factors (IGFs) gene is a candidate gene for growth, body composition and metabolism, skeletal characteristics and growth of adipose tissue and fat deposition in chickens (Zhou et al., 2005). Insulin-like growth factors (IGFs) belong to the family of polypeptide hormones; they are structural homologues of insulin and also have a similar function (Li et al., 2003). The IGF-1 and IGF-2 stimulate the proliferation, differentiation, and metabolism of myogenic cell lines from different species (Florini et al., 1996 and Siddle et al., 2001). The IGFs have been shown to regulate body and muscle growth in chickens (Duclos et al., 1999). Similarly, the metabolic effects of avian
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IGFs include increase in amino acid and glucose uptake and the regulation of DNA and protein synthesis (McMurtry, 1998). This molecular characterization of the chicken IGF-1 and IGF-2 has provided valuable clues for understanding how they are regulated and expressed. Most of the functions of the growth hormone in chickens are mediated by insulin-like growth factors (IGF) (Lei et al., 2005) which stimulate amino acid uptake, glucose metabolism, DNA synthesis (McMurtry 1998), protein synthesis, proliferation of different cell types and regulation of growth (McMurtry et al., 1997). Growth rate stimulation by IGF is known in many species of animals but studies have shown that there is no direct relationship between the levels of growth hormone (GH) and the growth rate in chickens. It might be useful therefore to study insulin-like growth factors (Beccavin et al., 2001), as mediators of the functions of the growth hormone (Lei et al., 2005). Surprisingly, higher GH levels in chickens are consistently associated with low growth rates, whether in unrelated genotypes (such as broiler vs. layer) (Goddard et al., 1988) or in divergently selected genotypes (Nir et al., 1987). This suggests that GH levels do not directly explain growth rate in the chicken and implies the need for studies on the factors acting downstream of GH, namely the insulin-like growth factors (IGFs).
1.1 JUSTIFICATION
Great genetic resources embedded in the indigenous chicken await full exploitation that will provide basis for genetic improvement and diversification to produce breeds that are adapted to local conditions for the benefit of farmers in developing countries (Sonaiya et al., 1999). Basic morphometric data, sperm reserve and quality on the three genotypes
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(frizzle, naked-neck and normal feathered chicken) of indigenous chickens are limited (Oke and Ihemeson 2010). This study will help in improving breeding practices, since a baseline assessment of the growth and reproductive system in relation to the sperm quantity and quality of the genotypes will be established. The biochemical testing being investigated will help to ascertain the blood profile in the three genotypes which will enhance proper management, feeding, breeding, prevention and treatment of disease since it is desirable to know the normal physiological information and the biochemical values which can be obtained under local conditions of rearing since valuable information in literatures towards this line of research is limited. The inclusion of the IGF-1 molecular marker will check out the level of relationship among this genetic marker, IGF-1 gene polymorphism, body weight and semen quality in the cockerels in respect to fertility and chronological age since this gene of interest does not only have influence on growth and body weight, but it is also critical for reproductive neuroendocrine (Wang et al., 2011) and reproduction in chicken (Bian et al., 2008). Therefore, this experiment was designed to evaluate growth traits, semen ejaculate quality, and biochemical and IGF-1 molecular markers in three genotypes (normal feathered, naked neck and frizzle) of the Nigerian indigenous cockerels.
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1.2 HYPOTHESES
H0: There is no difference in growth, morphometric traits and semen quality characteristics of the F1 offspring produced from the three genotypes of the Nigerian indigenous cockerels. HA: There is difference in growth, morphometric traits and semen quality characteristics of the F1 offspring produced from the three genotypes of the Nigerian indigenous cockerels. H0: There is no difference in some haematological and serum biochemical parameters of F1 offspring produced from the three genotypes of the Nigerian indigenous cockerels. HA: There is difference in some haematological and serum biochemical parameters of F1 offspring produced from the three genotypes of the Nigerian indigenous cockerels. H0: There is no relationship among the IGF-1 polymorphism, body weight and semen quality characteristics of the F1 offspring produced from the three genotypes of the Nigerian indigenous cockerels. HA: There is relationship among the IGF-1 polymorphism, body weight and semen quality characteristics of the F1 offspring produced from the three genotypes of the Nigerian indigenous cockerels.
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1.3 OBJECTIVES
The followings are the objectives of the study: 1. To evaluate the growth, morphometric trait and semen quality characteristics of F1 offspring produced from the three genotypes of the Nigerian indigenous cockerels. 2. To evaluate the haematological and serum biochemical parameters of the F1 offspring produced from the three genotypes of the Nigerian indigenous cockerels. 3. To determine the relationship of IGF-1 polymorphism with body weight and semen quality characteristics of the F1 offspring produced from the three genotypes of the Nigerian indigenous cockerels. 4. To evaluate the correlation among body weight, morphometric traits and semen quality characteristics at 24, 28, and 32 weeks of age in the F1 offspring of the indigenous cockerels.
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