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Effect of feeding different ratios of concentrate roughage during induced estrus on hormonal and reproductive performances of ewes

W Laghrour, B Safsaf, N Alleg1, M Tlidjane and A Ouarest

ESPA Laboratory, Department of Veterinary Sciences, Institute of Veterinary and Agricultural Sciences, University of Batna1, Batna 05000, Algeria
laghrourwafa@yahoo.fr
1 Private veterinarian, Timgad-Batna 05000, Algeria

Abstract

The aim of this study was to examine the effects of feeding different concentrate roughage ratio on serum concentrations of some metabolites and progesterone as well as on the reproductive performance of Ouled Djellal ewes subjected to induction of estrus (progesterone plus 500 IU of PMSG) and natural mating during the winter. The study design involved the use of 60 Ouled Djellal ewes aged between 1.5 and 4 years, assigned to three groups consisting of three ratios of concentrate (C) to roughage (R): group1 (C80%; R 20%), group 2 (C70%; R 30%) and group3 (C 60%; R 40%). The blood samples were collected three times for metabolite analysis; data on pregnancy were recorded and serum levels of progesterone(P 4), urea (URE), total protein (TP), albumin (AB), triglyceride (TR), cholesterol (CH) and beta-hydroxy-butyrate (BHB) were determined. The blood metabolites (except triglyceride p>0.05) were affected by an increasing concentrate roughage ratio: the group 1 had a higher concentration of urea and cholesterol (p<0.05) and a lower total protein than the groups 2and 3. Also, higher BHB and albumin ( p<0.05) in serum were observed in the group 2 of ewes than in ewes of the other groups. The progesterone concentrations (P4) were similar (p≥0.39) among treatments until sample2, and sample3 were higher ( p≤0.05) in group 1 than in the other groups. Fertility, and twin rate were higher in group1 than the other groups. The results from this study suggest that increasing the level of concentrate from 60% to 80% exerted a positive effect on reproductive performance and blood metabolites in ewes.

Keys words: Algeria, BHB, metabolite, Ouled Djellal, pregnancy, progesterone


Introduction

The nutritive value of poor pasture during winter season cannot maintain the livestock. An incorporation of concentrate in animal diets is therefore needed to overcome the nutrients requirements such as protein, mineral and vitamins and to optimize the efficiency of feed utilization for growth, gestation, or milk production (Reddy et al 2005). Moreover, concentrate diets are generally provided because the dry matter intake (DMI) and the volatile fatty acids (VFA) are higher than with diets based only on roughage (Suarez et al 2007). To maximize reproductive performances, roughage and concentrate should be associated to reduce the feed cost and concentrate wastage.

Recently, it has been reported that the role of nutrition during the 1 st month of pregnancy depends largely on the degree of maturity of ewes, as adolescent and mature ewes differ in their response to different level of nutrition in early pregnancy (Annett and Carson, 2006). Energy intake can serve as a significant limitation to sheep production rates among animals raised on pasture. An insufficient energy results in decreased growth, and reproductive performance (Sobrinho 1996). The adoption of a good concentrate roughage ratio before and during the mating period results in a significant increase in ovulation rates (Molle et al 1997), decrease of follicular atresia (Sobrinho 1996), better body condition at mating (Jardim 1983), and higher incidences of twin births (Mukasa-Mugerwa and Lahlou-Kassib 1995).

Estrus synchronization is generally applied for reproductive management in sheep worldwide (Boscos et al 2002) and several methods have been performed with varying degrees of success (Twagramungu et al 1995). The main hormonal method for estrus synchronization in ewes is intra vaginal devices impregnated with progesterone or progestogen such as Fluorogestone Acetate (FAG). When progestogens were maintained during 10 to 14 days and were associated with or without pregnancy mare serum gonadotropin (PMSG) (Ataman et al 2005) reduced fertility rates have been attributed to many factors including breed, seasonality, age, environment quality, female reproductive status, hormonal treatment, and nutrition (Yavuzer 2005).

In Timgad semi-arid area at Eastern Algeria, ewe breeding is one of the most important activities of these rural areas; the predominant breed was and is still Ouled Djellal ewe, breed exclusively for meat. Ewes were exposed to the feedlot housing management and feeding as confined animals (zero pasture) using different concentrate roughage ratio during winter season.

This study was planned to compare the effect of three concentrate roughage ratio on the metabolic profile and P4 level during induced estrus, in order to define appropriate concentrate roughage ratio that ensure high reproductive performances. The synchronization of estrus response and mating, and consequently of lambing, may be a great advantage conferred by hormonal treatments because it could allow the production of homogeneous lots of lambs, which could eventually support the demand of the sheep meat market.


Materiel and methods

All procedures have been approved by the Ethical Committee of the Institute of Veterinary and Agronomic Sciences of Batna-1University (Algeria).

Experimental location

The study was performed in three private farms belonging to Timgad area in Batna Province (Eastern Algeria), a semi-arid region located at 1027 m of altitude (35°29′03″N 6°28′07″E), characterized by cold winters and hot and dry summers.

Animals and experimental design

Sixty Ouled Djellal ewes, weighting 45-48 kg and aged between 1.5-4 years have been used in this study. They were identified by ear tags and regularly vaccinated and treated against ecto-and endoparasites.

The experiment was conducted in winter 2018 (January) where there is a relative reduction in temperature and feed and continued to September 2018 coinciding in lambing season. The ewes were exposed to the feedlot housing management and feeding as confined animals (zero pasture). As mentioned in Table 1, the animals were assigned into three similar groups (20 each):

Group C80: concentrate mixture 80 % roughage 20%

Group C70: concentrate mixture 70 % roughage 30%

Group C60: concentrate mixture60 % roughage 40%

Table 1. Feed composition

(%)

C60

C70

C80

Roughage

Wheat straw A

40%

30%

20%

Concentrate B
Cereal mix#

20%

27%

38.5%

Wheat bran

37%

40%

38.5%

Mineral mix
A+B

3%
100%

3%
100%

3%
100%

#Cereal mixture of barley and wheat

The ewes were allowed mineral salt licks and water was available ad libitum during all the experiment period. The animals were fed twice daily; between 6.00 and 7.00 am and between 5.00 and 6.00pm.

Feed measurement

The feed samples were analyzed for dry matter (DM), ash, crude protein, crude fiber, starch and fat using near-infrared reflectance spectrophotometer (NIRS) instrument (Model Burker Tango FT-NIR spectrometer, Burker Optics, Germany). The instrument was specifically calibrated for forage formula (mhaygpfe. eqa) and concentrate mixed rations formula (tmrgpfe. eqa) of NIRS procedure as described by Aufrere and Michalet-Doreau (1988) and Berglund et al(1990), respectively. NIRS is a proven technique wildly used in Algerian animal feed milling. It provides simultaneous results of moisture and protein starch and ash in less than a minute (Garnsworthy et al 2000).

Body condition scoring (BCS)

The scoring of body condition of ewes was performed using a five-point scale (1-5) at 0.5 intervals between 2 and 4 (1, 2, 3, 3.5,4 and 5). The animals were examined by palpation of the loin region after last rib by the finger (López-Gatius et al 2002). The condition was scored always by the same person. Body weight and body condition score of animals were recorded at the beginning and at the end of the experiment.

Estrus synchronization

The ewes were kept isolated from rams for at least 2 months and were synchronized with one intra-vaginal progestagen-impregnated sponge (40 mg fluorogestone acetate, FGA, Chronogest®, Intervet, International B.V., Angers, France) inserted on day 0 and left for 14 days. The sponges were withdrawn (day 14) followed by an intramuscular injection of 500 IU PMSG (Chronogest®,Intervet International B.V., Angers, France) (Cognie 1988). Estrus detection was done by visual inspection. The onset of estrus was defined as the time when the ewe first stood to be mounted by the ram. The duration of estrus was defined as the interval between the onset and the end of estrus signs. Two days after sponge removal, the ewes were submitted to one controlled natural mating by introduction of one fertile ram /5 ewes (Figure 1).

Pregnancy diagnosis

The pregnancy diagnosis during early pregnancy (day 24 post mating) was assessed for the first time on all animals by progesterone concentration [P 4] >2.5 ng/l (Boscos et al 2003) and for the second time at 60day post mating by transabdominal ultrasonography using a portable wed 3000 with a R40/ 3.5 MHz curvilinear transducer (China). During the procedure, the sheep were restrained in a crate, in an inverted dorsal position. All ultrasonographic examinations were performed by the same operator. Still images of the fetus were transferred to a PC.

Reproductive performance

The following measurements were calculated for all ewe groups:

Fertility rate

Fertility rate is one of the most important parameters of the productivity of the sheep, where the number of lambs obtained at lambing is a good indicator (Petrovic et al 2012).

Prolificacy rate

The prolificacy represents the ability of a female to give a number of lambs characterizing the size of the litter (Dudouet 1997). Moreover, it represents the best criterion for the qualification of a sheep and is the basic element of the reproductive selection in terms of prolificacy.

Fecundity rate

The fecundity is the parameter representing the reproduction process; it characterizes the reproductive ability of a sheep or herd (Casamitjina 1996).

Frequency of twinning



Figure 1. Experimental protocol
Blood samples and analysis

The blood samples were withdrawn between 8-11am for three times:

S1: sample done at insertion of sponge;

S2: sample done at 48hours post withdrawal of sponge of estrus;

S3: sample done at day 24post-mating.

Beta-hydroxybutyrate level (BHB)

The samples were obtained in the morning, 4h after feeding, through a puncture of the jugular vein using sterile needles. The blood concentrations were determined in situ using strips of a hand held ketometer (Free Style Optimum from Abbott Diabetes Care) (Oetzel 2010).

Metabolic profile

For cholesterol (CHOl), total proteins (TP), Albumin (ALB), urea (URE) and triglycerides (TG), the blood samples were collected into evacuated tubes without anti-coagulant and kept in a cool place and centrifuged at 3000r/10min. The tubes were labeled and frozen then stored at -20 °C until the time of analysis. The serum concentrations of the mentioned biochemical parameters were determined using available commercial kits:

· Total proteins were estimated by colorimetric method (Roch-Hitachi Cobas 6000, Roche Diagnostics, France),

· Albumin by color. BCG PF method (Cobas6000, Roche Diagnostics, France),

· Urea by Cinét. Method (Cobas 6000, Roche Diagnostics, France),

· Cholesterol by color- enzymatic method (Cobas 6000, Roche Diagnostics, France),

· Triglycerides by GPO/PAP method (Roche-Cobas c 501, Roche Diagnostics, France).

Progesterone profile

The serum concentrations of progesterone were determined using chemiluminescence immunoassay by an automated system ECLIA (Cobas 6000, Roche). The measurement of ovarian activity was performed from the analysis of serum levels of progesterone, effectively declaring a ewe in sexual activity from a concentration ≥ 0.5 ng/ml and pregnant from a concentration>2.5 ng/ml (Boscos et al 2003).

Statistical analysis

Statistical analysis of data was performed using GraphPad Prism 7.00 software. The effect of fixed factors, diet (C60, C70 andC80) and time of sampling (sample1, sample2 andsample3) of the study and also their interaction on the various studied parameters were analyzed using a two-way analysis of variance. Tukey’s multiple comparisons test was conducted to test the significance between means of the different sub­groups. Differences were considered significant when p <0.05. Results were presented as the mean ±SD for metabolites and mean ±SEM for progesterone concentration.


Results and discussion

Chemical analysis of diets

The experimental diets (Table2) were formulated to meet the requirements for ewes that were 45-48kg of body weight (NRC 2007) and to be isolipidic. This has been achieved by chemical analysis as indicated in Table 2. Total dietary fat was under 4%, and this falls within the desired range of<5% total dietary fat. Dietary fat levels exceeding 6 to 7% reduce fiber digestion and, consequently, lower DM intake in dairy sheep (NRC 2007). The DM content of the concentrate mixtures C80, C60 andC70 was 881, 887 and 890 g/kg DM, respectively (Table 2). The crude fiber content was higher in C70 and lower in C80.

Table 2. Ingredients and chimique composition of experimental concentrate

I

C60

C70

C80

Dry matter (g /kg) a

887

890

881

Moisture (% of DM)

11.3

11.0

11.9

Crude protein (% of DM)

11.2

12.2

11.3

Crude fat (% of DM)

3.5

3.5

2.9

Crude fiber (% of DM)

9.6

10.1

6.3

Starch (% of DM)

23.7

18.8

17.0

Ash (% of DM)

7.2

7.1

5.6

Nutritional value#

UF (/kg CM)

0.94

0.94

1.03

PDIN (g/kg CM)

73.6

71.6

77.3

PDIE (g/kg CM)

85.7

84.3

93.7

ME (kcal/kg)

2610

2612

2822

DE (kcal/kg)

3129

3185

3441

DOM
FOM (g/kg)

0.749
631

0.740
624

0.812
709

DM = Dry Matter. # Estimated from INRA Tables (2007) CM= Crude Matter . UF = Feed Unit PDIE = true protein absorbable in the small intestine when rumen fermentable energy (organic matter) is limiting microbial protein synthesis in the rumen . PDIN = true protein absorbable in the small intestine when degradable N is limiting microbial protein synthesis in the rumen ME=Metabolizable energy (Kcal\kg) and DE digestible energy were calculated according to INRA tables 2007, DOM : Digestible organic matter FOM: Fermenticible organic matter

The ME content and protein digestible in intestine (PDIN) for the concentrate mixture C80 were higher than for C70 and C60. The study revealed that the C60 ration contained lower digestible energy (DE) than C70 and C80 rations (Table 2). The increased DE and ME values for C80, compared to C70 and C60 were expected due to the increase in the proportion of concentrate in the complete rations. Our results are in agreement with those of Mcleod and Baldwin (2000), who reported higher ME in the high-concentrate diets than in low-concentrate diets.

The decrease in estimated microbial N production (PDIN) found in low-concentrate diets C60 and C70 compared with high-concentrate dietC80 could be partially explained by the decreased intakes of energy (PDIE) and N, as the yield of microbial biomass is related to the amount of available energy and N. The increase in the proportion of concentrate in the diet augmented significantly the N digestibility. On high-concentrate diet, more fermentable energy is available, which helps rumen microbes to capture N leading to its increased digestibility. The Digestibility of Organic Matter (DOM) and Fermentable Organic Matter (FOM) varied among diet groups; the highest averages were recorded in C80 as the fiber digestibility decreases with increase in proportion of starch in the diet.

Effect of concentrate-roughage ratio on metabolite concentrations

The mean concentrations of biochemical parameters (Table3) were within the limits of most published literature for sheep (Dubreuil et al 2005; Kaneko et al 2008). There were no differences in values of all parameters between S1, S2 and S3 samples, except for concentrations of BHB and total protein. The values for BHB were higher in C70 than in C60 and C80 groups, while albumin and total protein were lower in C80 than in C70 and C60 (Table 3).

The group C70 ewes had lower level of urea (7.79±1.01 mmol/l) than in C60 and C80 ewes (7.94±1.06 mmol/l and 8.18±0.30 mmol/L, respectively, and higher level of albumin (32.0±0.2 g/l vs 1.9±0.4.

BHBA levels were higher in C70 ewes (0.87±0.48mmol/L) than in C 60 and C80 (0.87±0.48 mmol/l), and also this was found in levels of total protein in C60 (73.9±2.0 g/l vs. 70.4±1.1 g/l in C80). There were no differences in protein, triglycerides, cholesterol and albumin between groups during j0, j16 and j24 post mating samplings. Increasing concentrate in the ration increased urea and cholesterol level and had no negative effect on other blood metabolites.

Urea concentrations were influenced by the type of diet. Sheep fed with C80 diet showed an increased serum urea (Table 4). Our results are in concordance with those obtained by Tetaouine (2015), 8.6±1.8 mmol/l and Casamassima et al (2008) 7.9 mmol/l.

Table 3. Effect of concentrate roughage ratio on the biochemical and hormonal responses in Ouled Djellal ewes

Parameters

C60

C70

C80

Sample1

Sample2

Sample3

Urea mmol/l

7.94±1.06

7.79±1.01

8.18±0.30

8.11±0.56

7.20±0.69

7.20±0.69

Total protein g/l

73.9±2.0

71.2±2.3

70.4±1.1

70.0±1.8

72.9±2.7

72.6±1.7

Albumin g/l

30.5±0.5

32.0±0.2

31.9±0.4

31.2±0.9

31.7±0.6

31.4±1.0

Cholesterol mmol/l

1.29±0.09

1.64±0.10

1.71±0.05

1.58±0.24

1.59±0.19

1.47±0.24

Triglyceride mmol/l

0.22±0.02

0.21±0.02

0.22±0.17

0.21±0.01

0.23±0.01

0.21±0.02

BHB mmol/l

0.28±0.01

0.87±0.48

0.45±0.10

0.68±0.48

\

0.39±0.13

P4 ng/ml

1.62±1.2

1.51±1.05

2.04±1.65

0.42±0.02

0.41±0.05

4.37±0.90



Table 4. Effect of concentrate-roughage ratio and sampling and their interaction

Factors

Urea
mmol/l

Total
protein g/l

Albumin
g/l

Cholesterol
mmol/l

Triglycerides
mmol/l

BHB
mmol/l

P4
ng/ml

Diet

***

**

**

***

NS

****

NS

Sample

NS

**

NS

NS

NS

***

***

Diet*sample

**

NS

NS

NS

NS

****

NS

NS : non-significant, * : p <0.05; ** : p <0.01, *** : p <0.001

Increasing concentrate level had no effect on serum triglyceride and total protein, but increased blood urea nitrogen. Urea blood levels may indicate short term metabolic changes and are used together with serum total protein and albumin levels (Zobel et al 2015). As seen in our research, total Protein and Albumin levels were lower in day 0 while urea was higher in day 24 post mating (8.6±0.26 mmol/l). This increase of serum urea is due probably to consumption of young grass during the last sampling coinciding the beginning of spring. Indeed, the nitrogen content is higher for young grass compared to regrowth and is maximal at the heading (Verité 1986). Ribeiro et al (2004) determined plasma urea levels in Border Leicester x Texel ewes reared extensively on natural pastures, finding values of 7.6 ±1.8 mg/dL and 7.1±1.7 mg/dL during the initial period of gestation and in non-pregnancy, respectively. On the other hand cereal grains are rather poor in nitrogenous matter (10 to 15% of the DM), whereas they are rich in cytoplasmic carbohydrates, mainly starch (40 to 55% of the MS).

The values of the total serum protein in all groups were in physiological limits as reported by Kaneko et al (2008) and Radostits et al (2006). Our results are in concordance with those obtained by Safsaf et al (2012) and Deghnouche et al (2011) revealing higher concentrations of total protein in non-pregnant ewes during winter season. In contrast to the observation of Caballero et al (1992) who noted the lowest value (65.0g/l) at the start of the grazing period, our samples showed the higher value 72.8 g/l in day 40.This is in agreement with those obtained by Haddad et al (1981) 72±3g/l and Dekar (1994) 72.2g/l.

The concentration of albumin was higher in group C70 (32.0±0.2g/l) without a significant effect of sampling. Our values are in concordance with those obtained by Safsaf et al (2012) and higher than those obtained by Deghnouche et al (2011) in arid zone of south-east Algeria in OD ewes with 24.6 g/l ±8.5 and Haffaf et al (2012) who noted 26.1 g/l ±0.9. The cholesterol serum levels in the group C80 (1.71±0.05mmol/l) were significantly higher comparatively to groups C60 and C70. These values were in concordance with those found by Antunovic et al (2015) in Dubrovnik sheep (1.82 ±0.32 mmol/l). However, our study showed a lower triglyceridemia than the values described by Karapehlivan et al (2007) 0.85±0.01 mmol/l. Moreover, these data remained within the range of norms cited by Mollereau et al (1995) and Titaouine et al (2015) (0.22±0.16 -0.22±0.07 mmol/l).

The reason for the low levels of triglycerides in both pregnant and non-pregnant ewes could also be dependent on consistent seasonal variations but not with reproductive status (Yokus et al 2006). The comparison between sample 1 and sample 3 did not reveal any difference (0.21±0.01 mmol/l for both). Our results ranged within values cited by Dubreuil et al (2005) and Kaneko (2008).

According to Clark et al (2006), the blood BHB concentration increased as the % of concentrate in the diet increased. However, Anderson and Lundstrom (1984) made a relationship between time of feeding and BHB blood concentration. The blood samples for BHB concentrations were taken approximately 3-4 hours after the morning feeding, a time when the effect of diet (concentrate) on ketone bodies in circulation would be greatest (Andersson and Lundstrom 1984). Indeed, higher concentrate intake in C80 did not induce metabolic disorder.

Effect of concentrate roughage ratio on progesterone concentration [P4]

Progesterone concentration was higher (Table 5) (4.34 ±0.90 ng/ml) in S3 than S2 (0.41±0.05 ng/ml). No interaction was found between diet and progesterone concentration; however, the highest level was shown in C80. On the other hand, during sampling at day 24 post mating, a difference was revealed between ewes bearing a single lamb and those bearing twins.

The importance of progesterone during early pregnancy in sheep has been well established. Studies using ovariectomized ewes have revealed three important phases of progesterone secretion after mating (Miller and Moore 1976; Wilmut et al 1985a).

First, it is essential to have a low concentration (probably 0,1-0,2 ng/ml) of progesterone from day 1 after mating. Second, the concentration must increase during the period from day 3 to day 7 after mating to values typical of those observed during the luteal phase (3-6 ng/ml). Third, it is important that this luteal-phase value is adequate for the maintenance of pregnancy (Ashworth et al 1989).

At day 0, all ewes had a basal value of [P4] <0.5ng/ml which indicates that all ewes were in anestrus. Our results are in concordance with those of Madani et al (2009) and Benyounes and Lamrani (2006), who reported that the anestrus season of Ouled Djellal ewes extends from december to march. The progesterone concentrations remained at basal levels ([P4] <1ng/ml) throughout the estrus coinciding 48h after sponge removal. Plasma progesterone concentrations of ewes at mating (day16) were close to zero for all animals. The serum progesterone level increased gradually from the mean basal value of 0.38±0.03, 0.48±0.06, 0.38±0.02 ng/ml on day 16 to 4.05±0.95, 3.62±0.46, 5.36±0.78 ng/ml in C60, C70, C80 respectively on day 24 post mating (Table 5)

Table 5. Mean concentration (±SEM) of progesteronemia in ewes during different samplings

[P4] (ng/ml)

C60

C70

C80

Sample S1(Day 0 insertion sponge)

0.43±0.04*b

0.43±0.07*a

0.39±0.03

Sample S2 (Day 16 after insertion sponge)

0.38±0.03

0.48±0.06

0.38±0.02

Sample S 3 (24 days post mating)

4.05±0.95

3.62±0.46* c

5.36±0.78*d

*p <0.05, a : C70 vs C80, b : sample1 (C60) vs sample3 (C60), c : sample1 (C70) vs sample 3 (C70), d : sample1 (C80) vs sample3 (C80)



Table 6. Serum progesterone levels (mean ±SEM) in single and twin bearing OD ewes

[P4]

Sample1

Sample2

Sample3

Single

0.35±0.05

0.24±0.01

4.25±0.38

Twin

0.43±0.06

0.39±0.03

7.75±0.87

p

NS

NS

*

NS: non-significant, * p <0.05; a: single vs. twin

At day 24 post mating, progesterone concentration showed increased values in more than 50% of ewes in all experimental groups with 75%, 50% and 65% respectively. According to Thimonier (2000) and Titi et al (2008) progesterone level greater than 1 ng/ml may be indicative of a gestational state. Whereas, for Ganaie et al (2009) a progesterone level≥1.75 ng / ml is considered as a gestational sign if kept high at day 18 post mating.

EL-Gohary (2006) observed that progesterone concentrations were associated with the number of corpora lutea counted on the ovaries of ewes after mating. At 24 days post -mating, [P4] level increased in all experimental groups to above 1 ng/mL, as result of pregnancy incidence. Progesterone is essential for pregnancy maintenance and one of the important functions of the blastocyst is to ensure that uterine luteolytic mechanism is counteracted. Progesterone and estrogen determine the proper function of the uterus in preparation for embryo development and implantation. Therefore, increasing P4 level during early pregnancy reduces embryonic losses and increases pregnancy rate and fertility (Ataman et al 2013). At day 0, ewes offered concentrate roughage ratio C80 tended to have a lower concentration of progesterone than C70 (Table 5). Since there were no differences in progesterone concentrations between C60 and C80 during early gestation, we conclude that the ratio did not differ enough to produce a variation in the P4 profile; however, there was a difference between sampling times of S1 and S3 in all groups. In our study, a difference between ewes carrying one and twin fetuses was found in the three groups in early pregnancy (Table5), which is in agreement with Muller et al (2003) who stated that the fetal number can be determined from the 19th day of pregnancy via [P4] concentration with 78% of accuracy. In contrast, Mukasa-Mugerwa and Viviani (1992) and Manalu and Sumaryadi (1998b), respectively, in Menz and Javanese Thin-Tail sheep, found no differences in the first three weeks after insemination between non pregnant and pregnant ewes bearing one or two fetuses. Indeed, the high twin rate may be due either to greater release of pituitary hormones in the C80 ewes, or an increased sensitivity of the ovary to the hormonal action of PMSG.

In spite of progesterone profile being known as a non-pregnancy test, pregnancy incidence was confirmed by echography at day 60. However, in the present study, concentrate / roughage ratio during early pregnancy had no effect on fertility rate. This may be related to the nutritional status of the ewes at mating. Improvements in embryo survival of ewes offered a high level of nutrition in early pregnancy have been reported when ewes were in poor body condition at mating (2.5) (Robinson 2006). It has been reported that the role of nutrition during the 1st month of pregnancy depends largely on the degree of maturity of the ewes, as adolescent and mature ewes differ in their response to different planes of nutrition in early pregnancy (Annett and Carson 2006). More recently, the same authors suggested that mature ewes were insensitive to an over-supply of nutrients as in our study no effect of over –feeding was found during the first 30 days of gestation on conception rate. The best reproductive parameter was recorded in C80 group in which ewes had good body score at the beginning of experimentation and maintained it during the mating season. The apparent reduced sensitivity to low peripheral progesterone concentrations in mature ewes may be related to the overall mean level of the hormone. Parr et al (1987) proposed a progesterone threshold for satisfactory conception of 2ng/ml on day 12 of gestation, although the highest conception rates were obtained for progesterone concentrations of 4 to 5ng/ml. Therefore, while in the current study over-feeding mature ewes in early pregnancy decreased their progesterone concentrations, the level of the hormone remained higher than the threshold.

Relationship between pregnancy and BCS

The best reproductive performances were found in C80 treatment which had the greater initial body score at the beginning of flushing period (2.6±0.3) and final BCS about3.1±0.3 at the end of flushing. Most authors recommend a BCS of 2.5 to 3.0 either for natural or artificial breeding (Husein and Ababneh 2008; Contreras-Solis et al 2009). In a study carried out in inseminated Rasa Aragonesa ewes ( Bru et al 1995), the lowest pregnancy rates (32,7 %) were obtained in sheep with a BCS<2, the average values (48.3%) with BCS between 3 and 2 and the higher values (58.8%) when BCS was >3.Additionally, ewes having a BCS equal to 2.3±0.3 units in C70 had a markedly lower twin rate than in those having higher values (C80). This was also the case for BCS less than 2.0 units in C60 (1.6±0.4), thus confirming our previous findings. By contrast, one of the ewes having BCS of 3 units (the highest of all) in C 80 group had quadruplets. Ptaszynska (2001) noted that in general ewes respond optimally to increasing concentrates in the diet when in medium BCS (2.5-3.5) rather than when exclusively thin or fat. Hence, BCS of 2.5 units is considered critical (as threshold) to affect reproductive performance in sheep breeding.

Reproductive performance

The highest concentrate/ roughage ratio (C80) resulted in high fertility, fecundity and prolificacy rates compared with C60 and C70 (75, 155 and 206 %, respectively). In the present study, percentage of ewes that lambed twins was higher for ewes in C80, as twin births dominated with 40.0% in C80 against 26% in C60 and 23.5% in C70.

Table 7. Effect of feeding different concentrate roughage ratios on reproductive performance of ewes

Fertility
rate

Fecundity
rate

Prolificity
rate

Single
fetus

Double
fetus

Triple
fetus

Quadrable
fetus

C60

70%
(14/20)

70%
(14/20)

100%
(14/14)

66%
(10 /14)

26%
(4/10)

///

///

C70

60%
(12/20)

85%
(17/20)

142%
(17/12)

76%
(13/17)

24%
(4/17)

///

///

C80

75 %
(15/20)

155%
(31/20)

206%
(31/15)

33%
(5/15)

40%
(6/15)

13%
(2/15)

13%
2/15)

Fertility and fecundity

Prolificacy rate and twin frequency were recorded following the 150 ±5 days of ram mating (Table 7). Difference in number of newborn lambs were found between groups. Fertility rate in C80 was 75%, the highest and the lowest number of ewes that lambed was70% in C60 and 60% inC70, respectively. On the other hand, the rate of double fetuses was lower in C70 and C60 compared with C80.The highest recorded fecundity rate was 155 % in C80 and the lowest 70% in C60. These results indicate that induction of estrus increased the fecundity rate, and showed that fecundity was affected by the synchronization treatment (PMSG) and by nutrition. The fecundity rate increased within the C80 group in comparison to C60 and C70.This may be attributed to the increased number of growing follicles associated with the usage of sponge and the higher number of double ovulations in synchronized ewes. Our findings are in concordance with those of Zarkawi (2001), who detected a twinning rate of 50% in Syrian Awassi ewes treated with sponges plus PMSG compaed to 20% for sponge-treated ewes without PMSG. In the other hand, (Lassoued et al (20 01), showed important interactions between genotype and level of nutrition. In this sense, in highly prolific ewes like D’Man breed, higher levels of nutrition prior to and during mating were associated with improved reproductive performance, but in low prolific breeds such as Queue Fine de l'Ouest, neither ovulation rate nor lambing rate were affected by the dietary treatment. Highest values of reproductive performances registered in C80 can be attributed to level of concentrate and BSC (>2.5). Our results were in agreement with those obtained in a study carried out in inseminated Rasa Aragonesa ewes (Bru et al 1995); the lowest pregnancy rates (32,7 %) were obtained in sheep with a BCS<2, the average values (48.3%) with BCS between 2 and 3 and the higher values (58.8%) when BCS was >3.The importance of BCS in fertility has been also reported in Spanish Manchega breed (Montoro 1995).


Conclusions


Acknowledgments

We thank Dr Abedessamed Halim from Sarl Vietam Setif for his help during the study to achieve NIRS analysis. We would also like to thank Dr. Virginie Decruyenaere form Wallon Agriculural Reaserch center ( CRA-W) for predicting chemical composition of concentrate mixture analyzed by NIRS and formulating the ewe’s requirements.


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Received 5 December 2019; Accepted 18 January 2020; Published 2 March 2020

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