Directory UMM :Data Elmu:jurnal:A:Animal Feed Science and Technology:Vol86.Issue3-4.Aug2000:
Animal Feed Science and Technology
86 (2000) 239±249
Feed intake, sperm output and seminal
characteristics of Ethiopian highland sheep
supplemented with different levels of leucaena
(Leucaena leucocephala) leaf hay
Negussie Danaa,*, Azage Tegegneb, Teshome Shenkorua
b
a
Debre Zeit Agricultural Research Center, P.O. Box 32, Debre Zeit, Ethiopia
International Livestock Research Institute, P.O. Box 5689, Addis Ababa, Ethiopia
Received 7 September 1999; received in revised form 15 February 2000; accepted 2 May 2000
Abstract
The effect of feeding increasing levels of Leucaena leucocephala leaf hay on the potential
fertility and feed intake of male highland sheep maintained on roughage diet was studied. In
treatment 1 (NS), animals received only chickpea haulm. Treatments 2 (L100), 3 (L200) and 4
(L300) consisted of 100, 200 and 300 g per head per day of sun dried leucaena leaf hay while sheep
under treatment 5 (C300) received 300 g per head per day of concentrate supplement. Chickpea
haulm was offered ad-libitum to all groups and water and mineral licks were freely available.
Addition of leucaena improved total dry matter (DM) and crude protein (CP) intake signi®cantly
without depressing the intake of the basal diet. Supplementation increased the percentage of motile
cells (10 vs. 76% for L300 ) and mass activity/motility score (1.5 vs. 3.2 for L300) of spermatozoa
while reducing the incidence of total morphologically defective sperm cells (34 vs. 5% for L200).
Volume of ejaculate (0.36 vs. 1.1 ml for L200), sperm concentration (2.8 vs. 7.1109 mlÿ1 for
L300), and total number of spermatozoa per ejaculate (1.96 vs. 5.92109 per ejaculate for L300),
increased signi®cantly as a result of supplementation. Testicular size showed signi®cant differences
among treatment groups and generally increased with supplementation. It was concluded that
supplementation of up to 300 g per head per day of leucaena resulted in improved feed intake,
testicular growth, sperm production and semen quality of Ethiopian highland sheep offered
chickpea haulm basal diet. # 2000 Elsevier Science B.V. All rights reserved.
Keywords: Highland sheep; DM intake; Leucaena leucocephala; Chickpea haulm; Seminal characteristics
*
Corresponding author. Tel.: 251-1-338555; fax: 251-1-338061.
E-mail address: dzarc@telecom.net.et (N. Dana).
0377-8401/00/$ ± see front matter # 2000 Elsevier Science B.V. All rights reserved.
PII: S 0 3 7 7 - 8 4 0 1 ( 0 0 ) 0 0 1 5 2 - 8
240
N. Dana et al. / Animal Feed Science and Technology 86 (2000) 239±249
1. Introduction
Forage supplements have enormous potential for ruminant production in the tropics
because of their easy availability in the farms; high nutritive value and reduced feeding
cost. Of the forage supplements used, legumes have been particularly advantageous.
Leucaena, a native of Central America, is a tropical legume of mimosa family. It exists
through out the cool subtropics and equatorial subtropics. Of all tropical legumes it has
the widest variety of uses one of which is its use as forage (NAS, 1977). Especially, the
drought tolerance nature of this plant makes it a potential feed source to increase meat
and milk production during the dry season.
In some countries like Ethiopia the use of leucaena as animal feed is limited mainly by its
content of the toxic non-protein amino acid, mimosine (b-(N-(3-hydroxy-4-oxypyridyl))-a
aminopropionic acid) and the rumen degradation product 3-hydroxy-4 (1H)-pyridone
(3,4-DHP) (Hammond, 1995). The aromatic amino acid, mimosine occurs mainly in the
seeds and leaves. It is responsible for toxic effects such as abortion and infertility (Hamilton
et al., 1971), inhibition of deoxyribonucleic acid (DNA) replication, organ damage and
even death (D'Mello, 1992) while DHP is a potent goiterogen (Hammond, 1995).
Information on the effect of feeding leucaena on the reproductive performance of
ruminants is scarce and often inconsistent. For instance, inclusion of up to 75% of
leucaena in the ration resulted in no adverse effect on the reproductive performance of
goats in Philippines (Abilay and Arinto, 1981). On the other hand, a review by Hammond
(1995) revealed that leucaena in unadapted cattle resulted in reduced calving percentage
due to early embryonic mortality. The in¯uence of feeding different levels of leucaena on
sheep reproduction is not well known. Hence, this study was conducted to assess the
effect of feeding increasing levels of leucaena on the feed intake and quality and output of
semen of highland sheep maintained on chickpea haulm as basal diet.
2. Materials and methods
2.1. Description of the study area
The study was conducted at Debre Zeit Agricultural Research Center (DZARC)
located 45 km East of Addis Ababa at an altitude of 1900 m a.s.l. Based on 22 years data,
the average annual rainfall of the area is 851 mm and the average minimum and
maximum temperatures range from 8.9 to 24.38C, respectively, the mean average being
16.68C. According to the weather report of DZARC (1996), annual rainfall of 950 mm,
average minimum, maximum and mean temperatures of 11.5, 26.5 and 198C,
respectively, and relative humidity of 56.4% were recorded during the study period.
2.2. Animals and management
Forty intact male Arsi lambs of similar age and body weight (16±17.2 kg) were
purchased from Arsi region (ca. 100 km South of Debre Zeit). The testes, seminal
vesicles and scrotum of each lamb was palpated and carefully inspected before purchase.
241
N. Dana et al. / Animal Feed Science and Technology 86 (2000) 239±249
Table 1
Chemical composition of chickpea haulm, Leucaena leucocephala leaves and concentrate mixture (% of DM)
Feed
DM
OM
CP
NDF
ADF
Ca
P
Chickpea haulm
Leucaena
Concentrate
94
92
93
82.3
78.9
72
5.6
24.4
21.9
57.3
23.9
20.9
41
11.6
10.7
0.72
1.48
0.47
0.11
0.19
0.60
Sheep were quarantined upon arrival at the station and treated against endo- and ectoparasites. The animals were randomly divided in 10 groups housed in group pens and
each group randomly allocated to one of the ®ve treatments that was replicated
twice.Data on feed intake and seminal characteristics were taken after the animals were
given adaptation period to the environment and experimental diets for 30±40 days. The
experiment lasted for 15 weeks.
In treatment 1 (NS) animals were offered only chickpea haulm, while those on
treatments 2 (L100), 3 (L200) and 4 (L300) were supplemented with 100, 200 and 300 g
per head per day of sun dried leucaena leaf hay. Animals on treatment 5 (C300) were
supplemented with 300 g per head per day of concentrate mixture. The concentrate was
composed of wheat bran, noug (Guizotia abyssinica) cake, molasses and salt and was
formulated to supply about 200 g CP/kg DM. Table 1 presents the chemical composition
of feeds used in the experiment.
The basal diet, chickpea haulm, was fed ad-libitum to allow 25% feed refusal (as-fed
basis) based on intake of the previous day. Mineral licks and water were freely available
at all times. Chickpea haulm was offered once and the supplements twice daily (10:00
and 16:00 h).
2.3. Data and sample collection and analysis
2.3.1. Feed intake and composition
Amount of feed offered and refused was recorded daily to determine feed intake. Daily
samples of feeds offered and refused were collected, bulked weekly and subsamples taken
after thorough mixing. The samples were analyzed for dry matter (DM), organic matter
(OM) and crude protein (CP) according to AOAC (1985). Neutral detergent ®ber (NDF)
and acid detergent ®ber (ADF) were determined using the procedures of Goering and Van
Soest (1970).
2.3.2. Seminal measurements
Semen was collected fortnightly by using an electro-ejaculator (probe for sheep and
goats, Lane Manufacturing Inc., USA) after training the sheep for few weeks. Ejaculate
volume was determined by collecting semen into graduated tubes. The percentage of
motile spermatozoa (motility percent, MOP) and wave motion (motility score, scored
from 1 to 5) were estimated immediately after ejaculation by microscopic examination
(10) of a drop of semen placed between a slide and 2232 mm cover slip in a warm
(34±378C) stage (Salamon, 1976). The wave motion (motility score) was de®ned as good
when the sperm cells showed a vigorous straight forward movement and poor when a
242
N. Dana et al. / Animal Feed Science and Technology 86 (2000) 239±249
weak, slow and spasmodic movement of the sperm cells was noted (Kemp et al., 1989).
Fifty microliter of semen samples were dropped into tubes containing 2.9% PBSglutaraldehyde solution for subsequent determination of sperm concentration and
evaluation of sperm morphology. Sperm cell counts were made in duplicate using
Neubar haemocytometer and a phase contrast microscope (600) as described by Moss
et al. (1979). The number of sperm cells produced per ejaculate was calculated for each
sheep by multiplying the volume of ejaculate with the sperm cell concentration.
The percentage of morphologically abnormal spermatozoa was determined adopting
the classi®cation scheme proposed by Garner and Hafez (1980). Spermatozoa were
examined microscopically on unstained smear of each semen sample, and the percentage
of defective sperm cells was determined from a count of 300 cells per smear.
2.4. Statistical analysis
The data were subjected to variance analysis techniques according to the general linear
model (GLM) procedures of SAS (SAS, 1994). Treatment means were separated using
Tukey's studentized range test protected by a signi®cant F-test for treatment effects. Data
that were found to be deviating from the assumptions of the analysis of variance were
transformed before running the analysis. Accordingly, the data on concentration and
number of spermatozoa were log transformed, while square root transformation was
applied to morphological data. Relationships among parameters were analyzed using
correlation analysis.The statistical model used in the analysis was:
Yij m ai eij
where m is the overall mean, ai the treatment effect and eij the random error term.
3. Results
3.1. Dry matter and crude protein intakes, and feed conversion ratio
Daily DM intake and animal performance are shown in Table 2. Leucaena
supplementation increased signi®cantly total DM intake but did not depress intake of
the basal diet. However, differences in total DM intake between levels of leucaena were
not signi®cant.
The CP intake differed signi®cantly (p0.05).
b
NS: not supplemented.
c
AL-MOP: anti-logarithm of motility percentage (%).
d
AL-CONCT: anti-logarithm of sperm cell concentration (109 mlÿ1).
e
AL-TSPZ: anti-logarithm of total spermatozoal output (109 per ejaculate).
f
SC: scrotal circumference (cm).
g
Morph. defects: morphological defects.
244
N. Dana et al. / Animal Feed Science and Technology 86 (2000) 239±249
signi®cantly increased the volume of ejaculates, sperm concentration and the total
number of spermatozoa per ejaculate.
The quantity of morphologically abnormal spermatozoa in the ejaculates was
signi®cantly reduced as a result of supplementation. The percentage of normal
spermatozoa for treatments L100 to C300 ranged from 88.5 to 95%. These treatments
did not differ from each other in the extent of sperm cells with major (primary) sperm cell
defects while a signi®cant difference was noted in the proportion of spermatozoa having
minor morphological abnormalities between L100 and L200.
3.3. Scrotal circumference
A highly signi®cant difference was observed in scrotal circumference (SC) of rams
receiving the different treatment rations (Table 3). Addition of 100, 200 and 300 g per
head day of leucaena in the diet increased testicular size by 20, 24 and 21%, respectively,
while a 25% increment was achieved with the concentrate supplement compared to the
control basal diet.
4. Discussion
4.1. Dry matter and crude protein intakes, and feed conversion ratio
The CP content of chickpea haulm was below the critical threshold of 7±8% to
avoid that voluntary intake and digestibility could be affected (Van Soest, 1982).
Values obtained for leucaena were comparable to those reported earlier by NAS
(1977).
Inclusion of up to 300 g per head per day of leucaena in the diet did not have any
signi®cant effect on the daily dry matter intake of the basal diet, although a positive effect
was noted. Results on the effect of fodder legume supplementation on intake of basal
diets were inconsistent. Topps (1995) suggested that supplementation of fodder legumes
at levels less than 30±40% increases intake of basal diets. However, leucaena
supplementation was reported to reduce the voluntary intake of hay in Tanzanian goats
(Mtenga and Shoo, 1990). A study by Tomkins et al. (1991) also showed that
supplementation (of about 30% of the ration) of dried L. leucocephala leaves to rams
reduced intake of pangola grass basal diet by 8%.
Total daily DM intake, on the other hand, was signi®cantly improved by leucaena
supplementation con®rming earlier reports by Mtenga and Shoo (1990) and Tomkins et al.
(1991). This would be expected since nitrogen supplementation to ruminants increases
feed intake by increasing the rate of fermentation in the rumen (Van Soest, 1982) and
thereby resulting in improved digestibility and faster rates of passage of digesta through
the gastro-intestinal tract (Preston and Leng, 1987).
The amount of CP supplied by chickpea haulm (33.2 g/day) was slightly greater than
the protein supplied from average quality roughages in Ethiopia (28 g/day) (Seyoum and
Zinash, 1989). Animals receiving chickpea haulm alone required a larger quantity of feed
per unit gain due to lower intake and digestibility of this diet.
N. Dana et al. / Animal Feed Science and Technology 86 (2000) 239±249
245
4.2. Seminal characteristics
4.2.1. Volume of ejaculate
The minimum ejaculate volume, recorded with the control diet (chickpea haulm lone),
was below the standard range of 0.8±1.2 ml suggested by Foote (1980). This low semen
volume was probably due to the lower level of protein in the diet resulting in retarded
growth and maturity of reproductive organs. This situation, however, is not to be rated
adverse since the values were still with in the normal range (0.3±1.6 ml) suggested by
Moss et al. (1979). But adverse effects could be noted when protein de®ciency becomes
severe enough to cause a live weight loss of about 32% (Parker and Thwaites, 1972).
Values recorded in the supplemented groups were slightly larger compared to those
(0.7±0.8 ml) reported by Chiboka (1980) for West African dwarf rams electro-ejaculated
weekly but smaller compared to that (1.25 ml) reported by Ihukwumere and Okere
(1990) for Nigerian Yankassa rams. These variations could possibly be attributed to either
breed, age (Moss et al., 1979) or other environmental factors.
Absence of signi®cant response to increasing levels of leucaena (Fig. 1B) most likely
indicates that inclusion up to 300 g per head per day of leucaena in the diet would not
have adverse effect on the accessory sex glands of rams.
Fig. 1. Changes in mean values of (A) scrotal circumference; (B) seminal volume (volume); (C) motility percentage;
and (D) motility score of spermatozoa of rams offered chickpea haulm basal diet supplemented with 100 (100 g l),
200 (200 g l) and 300 g per head per day (300 g l) of leucaena leaf hay and 300 g per head per day of concentrate.
246
N. Dana et al. / Animal Feed Science and Technology 86 (2000) 239±249
4.2.2. Spermatozoal motility
Acquisition of progressive motility is considered to be one of the strong evidences for
sperm maturation and a useful determinant of conception. The range in the proportions of
actively motile spermatozoa recorded in this study was wide (10±76%). Supplementation
generally improved the nutritional condition of the animals consequently improving
maturation and motility of spermatozoa. This is in agreement with the results of Chiboka
(1980), who reported motility of 40% in West African dwarf rams grazing on dry
roughage which later increased to 59% with improved grazing conditions. Oldham et al.
(1978) also noted signi®cant improvements in spermatogenesis of rams with protein
supplementation. In this study, except for rams maintained on chickpea haulm alone, the
values were within the standard range (60±80%) set for mature rams (Foote, 1980).
4.2.3. Concentration and number of sperm cells
Despite the variations between treatments, sperm cell concentrations recorded here
appeared to be slightly higher compared to the normal range of 1.6±6109 and 2±
3109 mlÿ1 suggested by Moss et al. (1979) and Foote (1980), respectively, for mature
rams. While comparative data in these aspects are lacking in other sheep breeds of
Ethiopia, Chiboka (1980) recorded only 0.9109 mlÿ1 in the semen of West African
dwarf rams under grazing conditions in Nigeria. However, concentration of sperm cells is
a highly variable determinant of semen quality (Garner and Hafez, 1980); and thus such
disparities in the values reported are supposed to be normal.
Increasing sperm concentration is generally considered to be bene®cial because it
allows insemination of a larger number of females (Foote, 1980). During natural mating
higher sperm concentration ensures entry of more spermatozoa into the cervical reservoir
and then into the oviduct, consequently increasing the chance of fertilization (Garner and
Hafez, 1980).
Both leucaena and concentrate supplementation increased the total number of
spermatozoa per ejaculate in a manner similar to the seminal volume and sperm cell
concentration.
4.2.4. Motility score
Leucaena supplementation has signi®cantly although not consistently improved the
mass activity (motility score) of spermatozoa (Fig. 1D). Spermatozoa from rams kept on
the roughage diet showed a very slow and spasmodic motion. This is in line with the work
of Salamon (1976) who reported signi®cantly higher scores for spermatozoa of rams
receiving high level of supplementary feeding (600 g TDN, 165 g CP) compared to those
on a lower level of feeding (310 g TDN, 45 g CP). Similarly, Parker and Thwaites (1972)
noted the adverse effect of poor nutrition on mass activity of sperm cells in ram semen.
The motility scores for spermatozoa of supplemented sheep (2.8±3.3) were comparable to
the values reported by Hamani et al. (1996) for White Fulani and Tuareg rams in Niger.
4.2.5. Morphology of sperm cells
Morphological abnormalities varied from 5 to 34%, which were larger compared to the
®gures (4.5±11%) reported by Hamani et al. (1996). A signi®cantly greater proportion of
abnormalities (34%) was observed in spermatozoa of rams fed chickpea haulm alone.
N. Dana et al. / Animal Feed Science and Technology 86 (2000) 239±249
247
According to Moss et al. (1979), this ®gure appears to be out of the normal range (5±
20%). However, there is no concrete evidence to conclude that the semen of these rams is
infertile. Certain types of abnormalities may not be associated with infertility, even when
the proportions exceed 20% (Foote, 1980). Out of the total defects observed in
spermatozoa of non-supplemented rams, only 10% were major defects while the
remaining 24% were minor. The latter defects were mainly due to bent or terminally
coiled tails, which usually occur during the passage of spermatozoa through the
epididymis (Jainudeen and Hafez, 1980). A higher proportion of these abnormalities
might be attributed to disturbances in the function of the epididymis probably due to
subnormal levels of testosterone (Hainonen, 1989). Production of this hormone was stated
to be seriously suppressed as a result of feeding poor quality diets for a prolonged period
(Parker and Thwaites, 1972).
Proximal protoplasmic droplets were the major abnormalities encountered in common
to all the treatment groups. The greatest (p
86 (2000) 239±249
Feed intake, sperm output and seminal
characteristics of Ethiopian highland sheep
supplemented with different levels of leucaena
(Leucaena leucocephala) leaf hay
Negussie Danaa,*, Azage Tegegneb, Teshome Shenkorua
b
a
Debre Zeit Agricultural Research Center, P.O. Box 32, Debre Zeit, Ethiopia
International Livestock Research Institute, P.O. Box 5689, Addis Ababa, Ethiopia
Received 7 September 1999; received in revised form 15 February 2000; accepted 2 May 2000
Abstract
The effect of feeding increasing levels of Leucaena leucocephala leaf hay on the potential
fertility and feed intake of male highland sheep maintained on roughage diet was studied. In
treatment 1 (NS), animals received only chickpea haulm. Treatments 2 (L100), 3 (L200) and 4
(L300) consisted of 100, 200 and 300 g per head per day of sun dried leucaena leaf hay while sheep
under treatment 5 (C300) received 300 g per head per day of concentrate supplement. Chickpea
haulm was offered ad-libitum to all groups and water and mineral licks were freely available.
Addition of leucaena improved total dry matter (DM) and crude protein (CP) intake signi®cantly
without depressing the intake of the basal diet. Supplementation increased the percentage of motile
cells (10 vs. 76% for L300 ) and mass activity/motility score (1.5 vs. 3.2 for L300) of spermatozoa
while reducing the incidence of total morphologically defective sperm cells (34 vs. 5% for L200).
Volume of ejaculate (0.36 vs. 1.1 ml for L200), sperm concentration (2.8 vs. 7.1109 mlÿ1 for
L300), and total number of spermatozoa per ejaculate (1.96 vs. 5.92109 per ejaculate for L300),
increased signi®cantly as a result of supplementation. Testicular size showed signi®cant differences
among treatment groups and generally increased with supplementation. It was concluded that
supplementation of up to 300 g per head per day of leucaena resulted in improved feed intake,
testicular growth, sperm production and semen quality of Ethiopian highland sheep offered
chickpea haulm basal diet. # 2000 Elsevier Science B.V. All rights reserved.
Keywords: Highland sheep; DM intake; Leucaena leucocephala; Chickpea haulm; Seminal characteristics
*
Corresponding author. Tel.: 251-1-338555; fax: 251-1-338061.
E-mail address: dzarc@telecom.net.et (N. Dana).
0377-8401/00/$ ± see front matter # 2000 Elsevier Science B.V. All rights reserved.
PII: S 0 3 7 7 - 8 4 0 1 ( 0 0 ) 0 0 1 5 2 - 8
240
N. Dana et al. / Animal Feed Science and Technology 86 (2000) 239±249
1. Introduction
Forage supplements have enormous potential for ruminant production in the tropics
because of their easy availability in the farms; high nutritive value and reduced feeding
cost. Of the forage supplements used, legumes have been particularly advantageous.
Leucaena, a native of Central America, is a tropical legume of mimosa family. It exists
through out the cool subtropics and equatorial subtropics. Of all tropical legumes it has
the widest variety of uses one of which is its use as forage (NAS, 1977). Especially, the
drought tolerance nature of this plant makes it a potential feed source to increase meat
and milk production during the dry season.
In some countries like Ethiopia the use of leucaena as animal feed is limited mainly by its
content of the toxic non-protein amino acid, mimosine (b-(N-(3-hydroxy-4-oxypyridyl))-a
aminopropionic acid) and the rumen degradation product 3-hydroxy-4 (1H)-pyridone
(3,4-DHP) (Hammond, 1995). The aromatic amino acid, mimosine occurs mainly in the
seeds and leaves. It is responsible for toxic effects such as abortion and infertility (Hamilton
et al., 1971), inhibition of deoxyribonucleic acid (DNA) replication, organ damage and
even death (D'Mello, 1992) while DHP is a potent goiterogen (Hammond, 1995).
Information on the effect of feeding leucaena on the reproductive performance of
ruminants is scarce and often inconsistent. For instance, inclusion of up to 75% of
leucaena in the ration resulted in no adverse effect on the reproductive performance of
goats in Philippines (Abilay and Arinto, 1981). On the other hand, a review by Hammond
(1995) revealed that leucaena in unadapted cattle resulted in reduced calving percentage
due to early embryonic mortality. The in¯uence of feeding different levels of leucaena on
sheep reproduction is not well known. Hence, this study was conducted to assess the
effect of feeding increasing levels of leucaena on the feed intake and quality and output of
semen of highland sheep maintained on chickpea haulm as basal diet.
2. Materials and methods
2.1. Description of the study area
The study was conducted at Debre Zeit Agricultural Research Center (DZARC)
located 45 km East of Addis Ababa at an altitude of 1900 m a.s.l. Based on 22 years data,
the average annual rainfall of the area is 851 mm and the average minimum and
maximum temperatures range from 8.9 to 24.38C, respectively, the mean average being
16.68C. According to the weather report of DZARC (1996), annual rainfall of 950 mm,
average minimum, maximum and mean temperatures of 11.5, 26.5 and 198C,
respectively, and relative humidity of 56.4% were recorded during the study period.
2.2. Animals and management
Forty intact male Arsi lambs of similar age and body weight (16±17.2 kg) were
purchased from Arsi region (ca. 100 km South of Debre Zeit). The testes, seminal
vesicles and scrotum of each lamb was palpated and carefully inspected before purchase.
241
N. Dana et al. / Animal Feed Science and Technology 86 (2000) 239±249
Table 1
Chemical composition of chickpea haulm, Leucaena leucocephala leaves and concentrate mixture (% of DM)
Feed
DM
OM
CP
NDF
ADF
Ca
P
Chickpea haulm
Leucaena
Concentrate
94
92
93
82.3
78.9
72
5.6
24.4
21.9
57.3
23.9
20.9
41
11.6
10.7
0.72
1.48
0.47
0.11
0.19
0.60
Sheep were quarantined upon arrival at the station and treated against endo- and ectoparasites. The animals were randomly divided in 10 groups housed in group pens and
each group randomly allocated to one of the ®ve treatments that was replicated
twice.Data on feed intake and seminal characteristics were taken after the animals were
given adaptation period to the environment and experimental diets for 30±40 days. The
experiment lasted for 15 weeks.
In treatment 1 (NS) animals were offered only chickpea haulm, while those on
treatments 2 (L100), 3 (L200) and 4 (L300) were supplemented with 100, 200 and 300 g
per head per day of sun dried leucaena leaf hay. Animals on treatment 5 (C300) were
supplemented with 300 g per head per day of concentrate mixture. The concentrate was
composed of wheat bran, noug (Guizotia abyssinica) cake, molasses and salt and was
formulated to supply about 200 g CP/kg DM. Table 1 presents the chemical composition
of feeds used in the experiment.
The basal diet, chickpea haulm, was fed ad-libitum to allow 25% feed refusal (as-fed
basis) based on intake of the previous day. Mineral licks and water were freely available
at all times. Chickpea haulm was offered once and the supplements twice daily (10:00
and 16:00 h).
2.3. Data and sample collection and analysis
2.3.1. Feed intake and composition
Amount of feed offered and refused was recorded daily to determine feed intake. Daily
samples of feeds offered and refused were collected, bulked weekly and subsamples taken
after thorough mixing. The samples were analyzed for dry matter (DM), organic matter
(OM) and crude protein (CP) according to AOAC (1985). Neutral detergent ®ber (NDF)
and acid detergent ®ber (ADF) were determined using the procedures of Goering and Van
Soest (1970).
2.3.2. Seminal measurements
Semen was collected fortnightly by using an electro-ejaculator (probe for sheep and
goats, Lane Manufacturing Inc., USA) after training the sheep for few weeks. Ejaculate
volume was determined by collecting semen into graduated tubes. The percentage of
motile spermatozoa (motility percent, MOP) and wave motion (motility score, scored
from 1 to 5) were estimated immediately after ejaculation by microscopic examination
(10) of a drop of semen placed between a slide and 2232 mm cover slip in a warm
(34±378C) stage (Salamon, 1976). The wave motion (motility score) was de®ned as good
when the sperm cells showed a vigorous straight forward movement and poor when a
242
N. Dana et al. / Animal Feed Science and Technology 86 (2000) 239±249
weak, slow and spasmodic movement of the sperm cells was noted (Kemp et al., 1989).
Fifty microliter of semen samples were dropped into tubes containing 2.9% PBSglutaraldehyde solution for subsequent determination of sperm concentration and
evaluation of sperm morphology. Sperm cell counts were made in duplicate using
Neubar haemocytometer and a phase contrast microscope (600) as described by Moss
et al. (1979). The number of sperm cells produced per ejaculate was calculated for each
sheep by multiplying the volume of ejaculate with the sperm cell concentration.
The percentage of morphologically abnormal spermatozoa was determined adopting
the classi®cation scheme proposed by Garner and Hafez (1980). Spermatozoa were
examined microscopically on unstained smear of each semen sample, and the percentage
of defective sperm cells was determined from a count of 300 cells per smear.
2.4. Statistical analysis
The data were subjected to variance analysis techniques according to the general linear
model (GLM) procedures of SAS (SAS, 1994). Treatment means were separated using
Tukey's studentized range test protected by a signi®cant F-test for treatment effects. Data
that were found to be deviating from the assumptions of the analysis of variance were
transformed before running the analysis. Accordingly, the data on concentration and
number of spermatozoa were log transformed, while square root transformation was
applied to morphological data. Relationships among parameters were analyzed using
correlation analysis.The statistical model used in the analysis was:
Yij m ai eij
where m is the overall mean, ai the treatment effect and eij the random error term.
3. Results
3.1. Dry matter and crude protein intakes, and feed conversion ratio
Daily DM intake and animal performance are shown in Table 2. Leucaena
supplementation increased signi®cantly total DM intake but did not depress intake of
the basal diet. However, differences in total DM intake between levels of leucaena were
not signi®cant.
The CP intake differed signi®cantly (p0.05).
b
NS: not supplemented.
c
AL-MOP: anti-logarithm of motility percentage (%).
d
AL-CONCT: anti-logarithm of sperm cell concentration (109 mlÿ1).
e
AL-TSPZ: anti-logarithm of total spermatozoal output (109 per ejaculate).
f
SC: scrotal circumference (cm).
g
Morph. defects: morphological defects.
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N. Dana et al. / Animal Feed Science and Technology 86 (2000) 239±249
signi®cantly increased the volume of ejaculates, sperm concentration and the total
number of spermatozoa per ejaculate.
The quantity of morphologically abnormal spermatozoa in the ejaculates was
signi®cantly reduced as a result of supplementation. The percentage of normal
spermatozoa for treatments L100 to C300 ranged from 88.5 to 95%. These treatments
did not differ from each other in the extent of sperm cells with major (primary) sperm cell
defects while a signi®cant difference was noted in the proportion of spermatozoa having
minor morphological abnormalities between L100 and L200.
3.3. Scrotal circumference
A highly signi®cant difference was observed in scrotal circumference (SC) of rams
receiving the different treatment rations (Table 3). Addition of 100, 200 and 300 g per
head day of leucaena in the diet increased testicular size by 20, 24 and 21%, respectively,
while a 25% increment was achieved with the concentrate supplement compared to the
control basal diet.
4. Discussion
4.1. Dry matter and crude protein intakes, and feed conversion ratio
The CP content of chickpea haulm was below the critical threshold of 7±8% to
avoid that voluntary intake and digestibility could be affected (Van Soest, 1982).
Values obtained for leucaena were comparable to those reported earlier by NAS
(1977).
Inclusion of up to 300 g per head per day of leucaena in the diet did not have any
signi®cant effect on the daily dry matter intake of the basal diet, although a positive effect
was noted. Results on the effect of fodder legume supplementation on intake of basal
diets were inconsistent. Topps (1995) suggested that supplementation of fodder legumes
at levels less than 30±40% increases intake of basal diets. However, leucaena
supplementation was reported to reduce the voluntary intake of hay in Tanzanian goats
(Mtenga and Shoo, 1990). A study by Tomkins et al. (1991) also showed that
supplementation (of about 30% of the ration) of dried L. leucocephala leaves to rams
reduced intake of pangola grass basal diet by 8%.
Total daily DM intake, on the other hand, was signi®cantly improved by leucaena
supplementation con®rming earlier reports by Mtenga and Shoo (1990) and Tomkins et al.
(1991). This would be expected since nitrogen supplementation to ruminants increases
feed intake by increasing the rate of fermentation in the rumen (Van Soest, 1982) and
thereby resulting in improved digestibility and faster rates of passage of digesta through
the gastro-intestinal tract (Preston and Leng, 1987).
The amount of CP supplied by chickpea haulm (33.2 g/day) was slightly greater than
the protein supplied from average quality roughages in Ethiopia (28 g/day) (Seyoum and
Zinash, 1989). Animals receiving chickpea haulm alone required a larger quantity of feed
per unit gain due to lower intake and digestibility of this diet.
N. Dana et al. / Animal Feed Science and Technology 86 (2000) 239±249
245
4.2. Seminal characteristics
4.2.1. Volume of ejaculate
The minimum ejaculate volume, recorded with the control diet (chickpea haulm lone),
was below the standard range of 0.8±1.2 ml suggested by Foote (1980). This low semen
volume was probably due to the lower level of protein in the diet resulting in retarded
growth and maturity of reproductive organs. This situation, however, is not to be rated
adverse since the values were still with in the normal range (0.3±1.6 ml) suggested by
Moss et al. (1979). But adverse effects could be noted when protein de®ciency becomes
severe enough to cause a live weight loss of about 32% (Parker and Thwaites, 1972).
Values recorded in the supplemented groups were slightly larger compared to those
(0.7±0.8 ml) reported by Chiboka (1980) for West African dwarf rams electro-ejaculated
weekly but smaller compared to that (1.25 ml) reported by Ihukwumere and Okere
(1990) for Nigerian Yankassa rams. These variations could possibly be attributed to either
breed, age (Moss et al., 1979) or other environmental factors.
Absence of signi®cant response to increasing levels of leucaena (Fig. 1B) most likely
indicates that inclusion up to 300 g per head per day of leucaena in the diet would not
have adverse effect on the accessory sex glands of rams.
Fig. 1. Changes in mean values of (A) scrotal circumference; (B) seminal volume (volume); (C) motility percentage;
and (D) motility score of spermatozoa of rams offered chickpea haulm basal diet supplemented with 100 (100 g l),
200 (200 g l) and 300 g per head per day (300 g l) of leucaena leaf hay and 300 g per head per day of concentrate.
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N. Dana et al. / Animal Feed Science and Technology 86 (2000) 239±249
4.2.2. Spermatozoal motility
Acquisition of progressive motility is considered to be one of the strong evidences for
sperm maturation and a useful determinant of conception. The range in the proportions of
actively motile spermatozoa recorded in this study was wide (10±76%). Supplementation
generally improved the nutritional condition of the animals consequently improving
maturation and motility of spermatozoa. This is in agreement with the results of Chiboka
(1980), who reported motility of 40% in West African dwarf rams grazing on dry
roughage which later increased to 59% with improved grazing conditions. Oldham et al.
(1978) also noted signi®cant improvements in spermatogenesis of rams with protein
supplementation. In this study, except for rams maintained on chickpea haulm alone, the
values were within the standard range (60±80%) set for mature rams (Foote, 1980).
4.2.3. Concentration and number of sperm cells
Despite the variations between treatments, sperm cell concentrations recorded here
appeared to be slightly higher compared to the normal range of 1.6±6109 and 2±
3109 mlÿ1 suggested by Moss et al. (1979) and Foote (1980), respectively, for mature
rams. While comparative data in these aspects are lacking in other sheep breeds of
Ethiopia, Chiboka (1980) recorded only 0.9109 mlÿ1 in the semen of West African
dwarf rams under grazing conditions in Nigeria. However, concentration of sperm cells is
a highly variable determinant of semen quality (Garner and Hafez, 1980); and thus such
disparities in the values reported are supposed to be normal.
Increasing sperm concentration is generally considered to be bene®cial because it
allows insemination of a larger number of females (Foote, 1980). During natural mating
higher sperm concentration ensures entry of more spermatozoa into the cervical reservoir
and then into the oviduct, consequently increasing the chance of fertilization (Garner and
Hafez, 1980).
Both leucaena and concentrate supplementation increased the total number of
spermatozoa per ejaculate in a manner similar to the seminal volume and sperm cell
concentration.
4.2.4. Motility score
Leucaena supplementation has signi®cantly although not consistently improved the
mass activity (motility score) of spermatozoa (Fig. 1D). Spermatozoa from rams kept on
the roughage diet showed a very slow and spasmodic motion. This is in line with the work
of Salamon (1976) who reported signi®cantly higher scores for spermatozoa of rams
receiving high level of supplementary feeding (600 g TDN, 165 g CP) compared to those
on a lower level of feeding (310 g TDN, 45 g CP). Similarly, Parker and Thwaites (1972)
noted the adverse effect of poor nutrition on mass activity of sperm cells in ram semen.
The motility scores for spermatozoa of supplemented sheep (2.8±3.3) were comparable to
the values reported by Hamani et al. (1996) for White Fulani and Tuareg rams in Niger.
4.2.5. Morphology of sperm cells
Morphological abnormalities varied from 5 to 34%, which were larger compared to the
®gures (4.5±11%) reported by Hamani et al. (1996). A signi®cantly greater proportion of
abnormalities (34%) was observed in spermatozoa of rams fed chickpea haulm alone.
N. Dana et al. / Animal Feed Science and Technology 86 (2000) 239±249
247
According to Moss et al. (1979), this ®gure appears to be out of the normal range (5±
20%). However, there is no concrete evidence to conclude that the semen of these rams is
infertile. Certain types of abnormalities may not be associated with infertility, even when
the proportions exceed 20% (Foote, 1980). Out of the total defects observed in
spermatozoa of non-supplemented rams, only 10% were major defects while the
remaining 24% were minor. The latter defects were mainly due to bent or terminally
coiled tails, which usually occur during the passage of spermatozoa through the
epididymis (Jainudeen and Hafez, 1980). A higher proportion of these abnormalities
might be attributed to disturbances in the function of the epididymis probably due to
subnormal levels of testosterone (Hainonen, 1989). Production of this hormone was stated
to be seriously suppressed as a result of feeding poor quality diets for a prolonged period
(Parker and Thwaites, 1972).
Proximal protoplasmic droplets were the major abnormalities encountered in common
to all the treatment groups. The greatest (p