In-vivo Evaluation of Anti-Coccidial Efficacy of Aqueous Extract of Allium sativum and Carica papaya in Experimentally Infected Broiler Chicks
Udo EJ*, Abba AM, Abdulhamid Y and Mudassir I
Department of Biological Sciences, Nigerian Defence Academy, Nigeria
Submission: August 13, 2018;Published: October 03, 2018
*Corresponding author: Udo EJ, Department of Biological Sciences, Nigerian Defence Academy, P. M. B 2109, Kaduna, Nigeria.
How to cite this article: Udo EJ, Abba AM, Abdulhamid Y, Mudassir I. In-vivo Evaluation of Anti-Coccidial Efficacy of Aqueous Extract of Allium sativumand Carica papaya in Experimentally Infected Broiler Chicks. Curr Trends Biomedical Eng & Biosci. 2018; 16(5): 555946. DOI:10.19080/CTBEB.2018.16.555946.
A study was carried out to evaluate the efficacy of Allium sativum (garlic) and Carica papaya (pawpaw) with respect to growth and PCV value in Eimeria tenella infected broiler chicks. A total of 36 chicks (21 day old) were randomly divided into 9 groups (Group 1-9) of 4 chicks each. All the groups were infected with 5100 viable sporulated oocysts of Eimeria tenella orally except G7 and G8 which served as control. G1 and G2 were treated with supplement of Allium sativum at 10 and 5 ml/L, respectively, G3 were treated with aqueous garlic + pawpaw extracts at 10ml/litre in drinking water. G4 and G5 were treated with supplement of Carica papaya extracts at 10 and 5 g/L, respectively. G6 was treated with Amprolium at 1.25 g/L (+ve control) while G9 was Eimeria tenella infected chicks and non-treated (isolated positive control). Data collected were analyzed using analysis of variance (ANOVA) at P<0.05 level of significance. The study showed that treatment with Allium sativum and Carica papaya resulted in marked reduction in the number of coccidian oocysts shed by treated chicks.Even though it was not statistically significant (p>0.05), there was an increase in the mean weight and PCV values in the Eimeria tenella infected but treated groups. Thus, the plants extract can be used as alternative of synthetic chemical drugs in combating coccidiosis and improve chicken health.
Broiler chicken (Gallus gallusdomesticus) is a gallinaceous domesticated fowl, bred and raised specifically for meat production. It is a hybrid of the egg-laying chicken and a subspecies of the red jungle fowl (Gallus gallus) and typically, has white feathers and yellowish skin . Worldwide, consumption surveys indicate that broiler chicken is the second largest consumed meat. Broiler meat accounts for approximately 33% of world meat intake, serving as the chief meat in consumer diets in many low to middle income countries. Overall, the outlook for global chicken consumption is optimistic as it is expected to increase along with human population and income growth. The total number of broiler meat consumed in the world was nearly 47 billion in 2004; of these, approximately 19% were consumed in the US, 15% in China, 13% in the Europe 25% and 11% in Brazil; and Worldwide, 86.6 million tonnes of broiler meat were estimated to be consumed by 2014 .Therefore, broiler production as healthy and nutritious when compared to beef and mutton can serve as the keys in bridging the protein in-take gap since broiler chicken meat has lower iron contents .
The profession of bird keeping has assumed great importance in improving the employment opportunity and animal food pro duction in the World. It is also regarded as an important means of sustainable livelihood and a way of achieving a certain level of economic independence. Poultry have a significant effect on both international and national economy. Reports showed that about 10% of Nigerians are engaged in poultry production, mostly on subsistence and small or medium-sized farms . The economic efficiency of poultry production is however challenged with various constraints namely, feed inputs such as protein supplements which are usually expensive, limited access to capital, exploitation of the unconventional ingredients and by-product ; and a number of diseases such as new castle disease, infectious bursal disease and Coccidiosis. Among these constraints, coccidiosis has been reported as one of the major problems of poultry industry in the world .
Coccidiosis is a disease caused by a single celled obligate parasite belonging to the phylum: Apicomplexa, subclass: Coccidia, and the genus: Eimeria [7,8]. In chickens, nine Eimeria spp. have been described and the commonest species are Eimeria tenella, which causes caecal coccidiosis and E. acervulinaas well as E. maxima which cause chronic intestinal coccidiosis . Coccidiosis has still remained one of the most important and common parasitic disease to the broiler industry . Infection is caused by different species of Eimeria which invades the mucosal epithelium of
the intestines and causes enormous economic losses, particularly
by reducing growth rate. Eimeria sporozoites invade the intestinal
epithelium and results in inflammation leading to the initiation
of the immune response to the production of nitric oxide (NO)
and other powerful pro-oxidants that belong to reactive oxygen
species (ROS). These compounds are not only toxic for sporulated
oocysts but also have negative side effects on the host if not protected
by an antioxidant system . To prevent economic losses
due to negative effects on production, anticoccidial drugs are used
In many parts of the world, the use of coccidiostats such as
amproliumsulphonamides, quinoline, polyether ionophorous and
clopidol in treatment of coccidiosis is very common. However, the
increasing resistance of coccidia to currently used anticoccidial
drugs , coupled with the increasing costs of developing new
drugs and the public’s distrust of drug-treated meat demonstrate
the urgent need to explore alternative methods of controlling this
disease. Drug or antibiotic residues in poultry products may inflict
toxic effects on the animal health. Therefore, nutritional methods
mainly based on natural feed additives are considered to be one of
the most promising alternatives to anticoccidials .
Several herbal and medicinal plants have been used as coccidiostat
in traditional medicine in different parts of the world. These
plants include Anona senegalensis, Khaya senegalensis and Butyrospermumparadoxum,
Allium sativum and Carica papaya[13-15].
Garlic (Allium sativum) and pawpaw (Carica papaya) have been
reported to cure and prevent various diseases in animals (Delaha
and Garagusi, 1985). Informations on their efficacy in the treatment
of coccidiosis in broiler birds are scanty. Hence this study
was designed with the view to evaluate the effects of Allium sativum
(garlic) and Carica papaya (pawpaw) extracts on the viability
of coccidia oocysts in vivo and its effects on weight and PCV
values of coccidia infected broiler chicks.
Fresh Allium sativum and unripe fruits of Carica papaya were
bought from an urban market (Chechenia) in Kaduna, Kaduna
State. Plant samples were transported in polythene bags to the
Department of Biological Sciences Laboratory, Nigeria Defence
Academy for identification. In the laboratory, the bulb and fruit
of sampled plants were identified by a Botanist and assigned a
Fresh faecal samples from chickens naturally infected with
Emeria species were collected from the college of Agriculture
poultry farm Mando, Kaduna, Kaduna State, Nigeria, using a wide
mouth sample bottle. Samples collected were placed on ice in
thermox box and transported to the Department of Biological Sciences
Laboratory NDA Kaduna. In the Laboratory, faecal sample
was transferred to a 50ml test tube containing 2% Pottssium Dichromate.
Oocysts were isolated using sieving and sedimentation
techniques as described by Soulby.
Isolated oocysts were suspended in 2.5% (w/v) potassium dichromate
in petri dishes and incubated at room temperature for
7 days in Biological Sciences Laboratory, NDA. After 7 days sporulation,
oocysts were recovered by centrifugation at 3,000rpm for
5min in saturated sodium chloride solution followed by washing
in distilled water. Sporulated oocysts were then quantified using
Mc Master counting chamber and diluted in distilled water to
make a suspension of 5100 oocysts per ml.This was kept at room
temperature until needed.
Thirty-six (36) 2-weeks old broiler chicks were purchased
and were kept for 1 week to acclimatize during which each bird
was observed carefully for signs of infection. Faecal samples were
also collected from birds daily and examined microscopically for
signs of cocciodiosis. The thirty-six (36) broiler chicks were randomly
divided at 21st day of age into 9 groups (Group 1-9) of four
(4) chicks each. Detailed grouping of chicks were as follows:
Group 1:Eimeria tenellainfected chicks were treated with
aqueous garlic extracts at 10ml/litre in drinking water.
Group 2:Eimeria tenellainfected chicks were treated with
aqueous garlic extracts at 5ml/litre in drinking water.
Group 3:Eimeria tenellainfected chicks were treated with
aqueous garlic + pawpaw extracts at 10ml/litre in drinking water.
Group 4:Eimeria tenellainfected chicks were treated with
aqueous pawpaw extracts at 10ml/litre in drinking water.
Group 5:Eimeria tenellainfected chicks were treated with
aqueous pawpaw extracts at 5ml/litre in drinking water.
Group 6:Eimeria tenellainfected chicks were treated with
Amprolium powder at 1.5g / liter drinking water.
Group 7: Non-infected and non-treated (negative control).
Group 8: Non-infected but supplemented with garlic extracts
+ pawpaw extract.
Group 9:Eimeria tenellainfected and non-treated (isolated
All groups except group 7 and 8 (unifected control) were infected
with 5100 sporulated oocystsorallythroughdrinking waterby
gavage at 21 day old age. Treatment of chicks in all groups commenced
on the 3rd day of post experimental infection following
the detection of occyst in the faeces and presentations of clinical
manifestations after 3 days post infection, and subsequently therafteron
a daily basis until the experiment was terminated on the
11th day.Treatment with crude extracts was achieved in birds after
thirst of 1hr.
Fresh faecal droppings were collected from the litter spread
on the ground from each groups for oocysts count daily from the
day of commencement of treatment till the 11th day when the experiment
was terminated. The mean number of oocyst per gram
of faeces for each group was determined using a modified Mc Master
Birds in all the groups were weighed in grams (g) on a daily
basis using weighing balance (spring balance) scale. Mean weights
from each replication were also recorded to establish the growth
rates against the expected normal growth curve of a broiler.
Packed cell volume (PCV) of birds in all the groups was determined
as described by Slaoui&Fietteusing duplicate capillary
tubes. Blood samples were collected from each chicken in
all the groups through their wings with the help of 5ml syringe.
The blood were filled in capillary tubes at three quarter level and
then sealed at one end with wax. The sealed capillary tubes were
then centrifuged at 12,000 rpm at 10min in a microhaematocrit
centrifuge (Hawksley MHC) read using the Hawksley haematocrit
reader and expressed in percentage.
Data generated from the study was analyzed using Statistical
Package for Social Scientist (SPSS) version II (2011). Analysis of
variance was used to determine significance between groups;
simple percentage was used to determine percentage reduction
while least significant difference (LSD) was calculated at
Birds in all infected groups had ruffled feathers, decrease
appetite, depression and paralysis, in some cases by day 3 post
infections. Bloody diarrhea (stool) was passed by infected birds
on the 4th day post infection. Maximum presentations of these
clinical signs were observed at day 7 and 9 post infection. However,
there was regression and disappearance of bloody diarrhoea
afterwards till the end of the experiment on day 12. In groups 1-6
(G1, G2, G3, G4, G5 and G6) treated with different concentrations
of A. sativum and C. papaya either as aqueous extract or powder
preparation, manifestations of symptoms of coccidiosis became
milder as treatment progressed. However, birds in G5 treated with
5ml/l of C. papaya showed evidence of depression, decrease feed
intake, and persistence of bloody diarrhoea and death of 25% of
the birds at day 3 through 7 of treatment period. Birds in Gr 9 that
formed the infected non-treated group were observed to have ruffled
feathers, decrease appetite, depression, bloody diarrhea and
paralysis which persist all through the experimental periodresulting
in the death ofbirds in this group.
TD: Treatment Days, Values are means ± standard deviation. Mean values with different letters at the same row differ significantly
The Effects of Allium sativum and Carica papaya crude aqueous
extracts on E. tenellaoocyst count in infected and treated birds
is presented in Table 1. Despite the treatments given to birds in all
treatment groups, oocyst counts increased steadily from day 1-3,
including control group treated with Amprolium. However, there
was a significant drop in oocysts count in all treated groups (G1- G5) on the 4th day of treatment (P≤0.05). A significant increase
(5920±00) was also observed in total oocysts count in G9 (infected
non-treated group) when compared to other groups. Moreover,
a significant decrease in total oocysts count in G1 (1100±216) and
G6 (678.8±150.9) compared with other supplemented groups (G2,
G3, G4 &G5) and G9 (infected, non-treated group) (1812.3±473.2,
1275±359, 1556.3±347.6 & 2480±537 and 4380±00 respectively)
at the first day of post treatment. In comparing various groups
treated with different concentrations of the extracts, there a was
a significant difference when comparing the two groups treated
with garlic extract (G1&G2), and the two different concentrations
of papaya extract (G4&G5) they also differed significantly at
(P≤0.05), there was no significant difference in comparing the two
extracts at (P>0.05). Garlic supplemented at 10ml/liter and Amprolium
induced high reduction in total oocyts count.
TD: Treatment Days, Values are means ± standard deviation. Mean values with different letters at the same row differ significantly
The percentage Reduction in oocysts count of infected birds
treated with crude extracts of Allium sativum and Carica papaya
is presented in Table 2. Group 1 (treated with Allium sativum)
showed a progressive higher percentage oocyst reduction counts
from the 4th day of treatment with 70.5% and a complete reduction
of (100%) was achieved at the 11th day of post treatment.
Group 4 (treated with Carica papaya) showed a progressive percentage
oocyst reduction count from the second day of treatment with 68.9% and achieved a reduction with 99.8% at the 12th day
of post treatment. The difference was no t statistically significant
at p≤ 0.05. Amprolium group achieved a significant (p≤0.05) oocyst
reduction count with 100% at the 9th day post treatment.
The daily mean live weight of coccidia infected chicks treated
with different concentrations of aqueous extract of A. sativum and
C. papaya is presented in Table 3. Birds in groups 1-8 exhibited
significant increase in weight ranging from 425±26.8g -547±69g
(p≤0.05) from day 1 through day 11 when treated with different
extract concentrations at 2.5ml, 5.0ml and 10ml or Amprolium
given at 1.5ml. However, birds in group 9 which served as infected
and untreated control showed a significant decrease in live weight
from day 5 till the experiment was terminated on day 11(p≤0.05).
The loss in mean live weight of birds in Group 9 when compared
to other birds in either of the treated groups (1-8) was in range of
112±59g - 154 ±72g.
Values are means ± standard deviation. Mean values with different letters at the same row differ significantly
The haematological parameters for Eimeria-infected chickens
treated with A. sativum and C. papaya crude extracts is presented
in Table 4. Values of PCV increased in chicks of groups 1-7 during
the first three (3) days of the experiment and then decreased from
day 4 through to the 7th day, and then increased again from day 8
till the experiment was terminated on day 11. In group 6 treated
with Amprolium at 1.5g, there was progressive increase in PCV
from day 1 through today 12. However, chicks in Gr9 that were
infected but not treated (control) exhibited a steady decrease in
PCV from 36.06% ±00.23 to 16.66%±00. Although the variation in
PCV values within each group did not show any significant difference,
the variation in PCV values of chicks in Gr 9 was found to be
Values are means ± standard deviation. Mean values with different letters at the same row differ significantly.
The clinical signs of coccidiosis in chicks observed during the
In vivo study include weakness, reduced appetite, ruffled feathers
and bloody diarrhea. These are signs normally associated with
coccidiosis in infected chickens. Some of these manifestations are
evidence of cumulative tissue damage associated with second or
third generation schizogony of the parasite and depends on many
factors such as parasite species, viability, infectivity, virulence,
tropism, host age, nutritional status and immunological competence.
El bana et al.  reported the association of the detrimental
effect of coccidiosis and manifestation of clinical signs which
resulted in mortality of severely affected bird.
Allium sativum and Carica papaya extracts induced anticoccidial
effect as evident in the oocyst output in treated groups. In
this study, despite the treatments given to birds in all treatment
groups, oocyst counts increased steadily from day 1-3, including
control group treated with Amprolium. However, there was a significant
drop in oocysts count in all treated groups on the 4th day
of treatment. This is line with the finding of Biu et al.  who
reported consistent decreases in faecal oocyst count over 4 days
of treatment and at all doses of neem (Azadirachtaindica) in broiler
chickens infected with Eimeria species. Garlic supplemented
group at 10ml/liter showed high reduction in total oocyts count.
This suggests that the effect of the plant extract on coccidian oocyst
output was dose dependent. This cannot be unconnected with
the work of El-Khtam et al. &Biu et al.  who reported similar
dose related responses in faecal oocyst count and mortality
from coccidian infection in broiler chickens. There was observed
differential oocyst output between extract treated groups and the
control. Amprolium group achieved a significant oocyst reduction
count with 100% at the 9th day post treatment. This confirms the
work of El-Khtam et al. , Nghonjuyi et al. &Tan and Long
, who reported lowest range of faecal oocyst output in the
group treated with amprolium.
Percentage reduction in oocysts count could also probably
be due to inhibition or impairment of invasion and or replication
and development of Eimeria parasites species in the gut tissues of
chicks treated with plant extracts leading to relatively non-viable
oocysts formation and release. In other words, the effect of the extracts
may be on the intracellular stages of the parasite especially
at the 2nd schizogony stage (late stages of asexual cycle) as well as
those of the sexual stage of parasite in the lumen of the intestine. It
could also be attributed to the effect of antioxidant present in both
the plants evaluated in this study. Antioxidants are reported to be
responsible for the control of Eimeria infections during oxidative
stress and lipid peroxidation of the intestinal mucosa tissues.
Allen et al.  reported that antioxidant-rich plants is lethal to
coccidia parasites due to induced oxidative stress and neutralization
of reactive oxygen species which makes it effective in treating
coccidial infections. According the author, antioxidant compounds
are known to reduce the severity of Eimeria tenellainfections by
ameliorating the degree of intestinal lipid peroxidation. Allen and
Danforth (1998) in a separate study concluded that feed supplementations
with products that have antioxidant properties such
as the pawpaw even at 1% appears effective in reducing upperand
mid-small intestinal infections caused by E. tennela. Similarly,
Allen et al.  reported that drinking water supplemented with
different concentrations of pawpaw extract was suppressive towards
the development of coccidiosis in chickens.
In addition, presence of phenolic compounds in the plant extracts
evaluated in the study could also have contributed to the
lower oocysts count recorded in the infected and treated groups.
Sikkema et al.  confirmed that phenols interact with cytoplasmic
membranes and change their cation permeability leading
to impairment of crucial processes in coccidia cells and finally
leading to their death. According to Allen et al.  C. papaya is
rich in phenolic compound which has been shown to have antioxidative,
anti-inflammatory and immunomodulatory properties
and thus could have responsible for the anticoccidial effects of C.
papaya observed in this study. However, the significant reduction
in oocysts count in the infected groups treated with A. sativum
may be associated with the presence of organosulfur compounds
as earlier suggested in the studies. These compounds are found
abundantly in A. sativum. Organosulfur compounds are the most
important component responsible for most of pharmacological
effects of the plant (garlic) . Nghonjuyi et al.  support that
the anti-inflammatory property of the C. papaya leaves with high
concentrations of vitamin A might have acted in caecal epithelium
cell protection which is detrimental to the coccidial reproductive
The weights of the treated birds reduced and subsequently
increase following the progressive treatment. These variations
could be attributed to the deleterious effect of the infection before
treatment. Birds affected with coccidian parasite when not
treated show progressive signs ill health such as poor weight gain
or weight loss, weakness and emaciation and in severely affected
birds may result in death after the appearance of disease. The
progressive decrease in the weight before treatment occurred
probably as a result of severity of the infection. Conway et al.
,McDougald  &Chapman et al.  confirmed that under
conditions of more severe infection with Eimeria, weight gain is
generally reduced. Conway et al.  reported significant reduction
in body weight of broilers infected with high doses of sporulated
oocysts of E. tenella. The improved growth performance by
the 4th day of treatment may be related to reduce severity of the
infection due to the administration of plant extracts.
The results obtained also showed that infection had no significant
effects on the PCV values between extract treated groups.This
however support the findings of Nghonjuyiet al.  who reported
no significant difference between Eimeria infected chickens
treated with ethanolic extracts of C. papaya leaves. Similarly, Jatau
et al.  also recorded no significant differences in the haematological
parameters of chickens in an experiment they carried out
with experimental fowls. However, the effects of the treatment on infected birds could be best appreciated when weights of chicks in
the treated group is compared with birds in the untreated group
(control). Birds in group 9 progressively lost weight till the end of
the experiment. In other words, growth of birds in this group was
adversely affected by coccidiosis. This is also reflected by the continuous
decrease inPCV values due probably to continuous loss of
blood due to passing out of bloody diarhoea which is characteristic
of coccidia infected birds. This invariably results in reduced
blood count in the birds of group 9. Coccidia parasite such as E.
tenella is reported to undergo series of intestinal invasions during
asexual reproduction resulting in the damage of the intestinal lumen
during the course of penetrating the epithelial cells of the
caecum and severe erosion of the mucosal membranes through
cell lysis which lead to profuse watery-to-bloody diarrhea [13,20].
Bloody diarhoea has been reported to have a detrimental effect on
the blood parameters of birds including PCV which may lead to
anaemia in infected birds .
The results of this study suggest that supplementation with
crude extracts of garlic and pawpaw at concentrations of 10 ml/
litre drinking water alleviates the negative impact of Eimeria
infection in broiler chickens. The beneficial effect of the extract on
the number of oocysts per gram of excreta of the infected chickens
was found to be optimally effective though not as comparable to
that exhibited by Amprolium. Allium sativum exhibits a significant
anticoccidial activity than C. papaya although both plants could
serve as alternative to synthetic chemical drugs used in combating
coccidiosis and improves chicken health.