Protective effect of Aloe vera on growth performance, leucocyte count and intestinal injury in broiler chicken infected with coccidiosis

ABSTRACT

Groups were designated as a positive control (+C), negative control (−C), Amprolium (AM1), Aloe vera, 2.5 g/l water (AV2.5) and Aloe vera 5 g/l water (AV5) during the finisher phase. The experiment was conducted using 360 broiler chicks, which were divided into five different groups. Overall feed intake was significantly (P<.01) lower in Aloe vera supplemented groups. The overall weight gain was significantly (P<.01) higher in AV5 and AMI compared to –C. No significant difference was found in the feed conversion ratio between AMI and Aloe vera-treated birds. The caecal lesion, oocysts count and intestinal lesion were significantly (P<.01) reduced in Aloe vera-treated birds. The number of heterophils and total leucocyte count significantly (P<.01) decreased, while the number of monocytes and eosinophils increased (P<.01) in the treatment groups. From the results, it is also clear that the infection increased the monocytes and eosinophils and decreased the heterophils and TLC. In conclusion, the use of Aloe vera at the rate of 5 g/l of water produced encouraging results in broilers in comparison with Amprolium.

KEYWORDS:

• Amprolium
• aloe vera
• oocyst
• lesions

Introduction

Among important poultry disease, avian coccidiosis is one of the economically most dangerous diseases caused by several species of genus Emeria. Typically, the protozoan enters into the intestinal tissues, leading to the pathological changes (Chand et al. 2016; Ali et al. 2019). This disease is typically characterized by low feed consumption; poor feed efficiency and high morbidity and mortality in the flock. Usually, the disease is controlled by using antibiotics (Tanweer et al. 2014).

Anticoccidial drugs have shown great efficacy against the coccidial pathogens; however, there are many negative implications of the prolonged use of these drugs (Hafeez et al. 2020). A number of negative effects on health have been reported; however, the use of antibiotics in controlling coccidiosis is still continuing. The European Union (EU), therefore, banned the use of antibiotics to justify them as animal feed additives (Khan et al. 2012; Abudabos et al. 2018; Safiullah Chand et al. 2019; Nasir et al. 2020). There is an instant need to discover novel feed additives for the effective controlling of avian coccidiosis.

More than 400 different species belong to genus Aloe have been used for different pharmacological activities for the last more than 2000 years. This plant grows well in a hot and humid environment and available in different parts of Asia and other continents (Boudreau and Beland 2006). Most part of this plant comprises water and the remaining small solid portion contains approximately 75 biologically active compounds having broad range pharmacological activities (Yim et al. 2011). The objective of this study was to investigate the beneficial effect of Aloe vera on the production performance, intestinal lesion score and leucocytes count in experimentally induced coccidiosis in the broiler.

Materials and methods

All experimental protocols were carried out according to the protocol of Faculty of Animal Husbandry and Veterinary Sciences, The University of Agriculture, Peshawar.

Birds and experimental design

A total of 360-day-old chicks (Cobb 500) were distributed into five groups, which were further subdivided into six replicates. The study comprised the starter and finisher phase of total 42 days. Water and feed were offered ad libitum. The composition of the feed is given in Table 1. The groups were positive control (+C, infected by coccidial oocysts with no treatment), negative control (−C, neither infected by coccidial oocyst nor treated), AM1 (infected by coccidial oocyst and treated with Amprolium at the rate of 1 g/l water), AV2.5 (infected and treated with Aloe vera at 2.5 g/l water) and AV5 (infected and treated with Aloe vera at 5 g/l water). Mixed sporulated oocysts of different Eimeria species (Eimeria tanella, Eimeria nacatrix and Eimeria acervulina) were collected from diverse poultry farms and slaughter houses. On day 22, about 30,000 sporulated oocysts were mixed in drinking water of all groups except negative control.

Table 1. Basal composition of feed during the starter and finisher phase (%).

Ingredients	Starter	Finisher
Corn	53.21	60.75
Soybean meal	37.92	25.00
Corn gluten meal	2.00	7.10
Corn oil	2.20	2.80
Dicalcium phosphate	2.30	2.05
Limestone	0.83	0.68
Salt	0.45	0.50
VM Mix^a	0.50	0.50
DL-Methionine	0.20	0.10
Lysine-HCl	0.22	0.37
Threonine	0.11	0.10
Choline chloride	0.05	0.05
Chemical composition
ME, Kcal/kg	3000	3150
Crude protein, %	23.5	21.30
Methionine, %	0.55	0.44
Lysine, %	1.42	1.23
Sulphur amino acids, %	0.96	0.80
Threonine, %	0.95	0.85
Calcium, %	1.05	0.90
Phosphorus, %	0.50	0.45

^aVitamin–mineral premix contains in the following per kilogram: vitamin A, 2,400,000 IU; vitamin D, 1,000,000 IU; vitamin E, 16,000 IU; vitamin K, 800 mg; vitamin B1, 600 mg; vitamin B₂, 1600 mg; vitamin B₆, 1000 mg; vitamin B₁₂, 6 mg; niacin, 8000 mg; folic acid, 400 mg; pantothenic acid, 3000 mg; biotin 40 mg; antioxidant, 3000 mg; cobalt, 80 mg; copper, 2000mg; iodine, 400; iron, 1200 mg; manganese, 18,000 mg; selenium, 60 mg, and zinc, 14,000 mg.

Performance of birds

Feed intake was taken on a daily basis. Weight gain was measured after every week. The feed conversion ratio (FCR) was calculated on the given data of feed intake and weight gain.

Blood collection

At the end of the experimental period, two birds per replicate were slaughtered and 3 ml blood was collected from each bird with EDTA. The blood samples were subjected to the determination of the number of total lymphocyte count (TLC) and differential leucocyte count (heterophil, monocyte, eosinophil and lymphocyte) as described by the method of Sultan et al. (2018).

Lesion score of the intestines

Two birds from each group were defeathered and intestines were aseptically removed. Intestines along with caeca were removed from the infected birds for the lesion scores (0 to + 4 counting system) as described by Tanweer et al. (2014). 0 score for no lesion, +1 (small lesions), +2 (moderate lesions), +3 (lesions with severity) and  + 4 (more severe lesions).

Number of oocysts per gram of faeces

The load of oocysts was determined by collecting faecal samples on day 42. The number of oocysts was determined by the method described by Tanweer et al. (2014) using the McMaster technique.

Statistical analysis

Data were analysed using completely randomized design with the statistical package SPSS 21.0 using one-way analysis of variance (ANOVA) and the Tukey test at 5% probability level.

Results

Table 2 shows the comparative efficacy of commercial coccidiostat and Aloe vera on feed intake in broilers. At the end of first, second, third week and overall starter phase, feed intake was not significantly different (P>.05) between the groups. At the end of the fourth week, feed intake was significantly higher (P<.01) in the group −C in comparison with all other groups. Feed intake in the group AM1 and group AV5 was significantly higher (P<.01) than the group  +C with no difference with the group AV2.5. At the end of the fifth week, feed intake was significantly higher (P<.01) in the group −C when compared with group AV5 followed by the group AV2.5 and group +C. After the sixth week, feed intake was significantly higher (P<.01) in the group −C than the group AV5 and AV2.5 followed by the group +C. In the finisher phase, feed intake was significantly higher (P<.01) in the group −C in comparison with the group AM1 and group AV5 followed by the group AV2.5 and group + C. Feed intake for the overall period was significantly higher (P<.01) in the group −C than group AV5 and AV2.5 followed by the group + C.

Table 2. Comparative efficacy of Amprolium and Aloe vera against coccidiosis challenge on feed intake (g) in broilers.

Weeks	+ C	−C	AM1	AV2.5	AV5	P-value
1	131 ± 3.1	132.50 ± 7.3	133 ± 7.5	129.75 ± 2.2	131.25 ± 6.8	.93
2	282 ± 6.6	285.25 ± 4.9	282.75 ± 8.2	284.25 ± 4.9	282.75 ± 6.2	.94
3	509.75 ± 13.2	516.75 ± 14.4	518 ± 14.0	525 ± 10.2	518.25 ± 18	.68
Starter phase	922.75 ± 15.6	934.50 ± 17.5	933.75 ± 22.9	939 ± 14.2	932.25 ± 27.3	.83
4	755.25^c±7.9	809.50^a±13.9	783.50^b±5.9	768.50^bc±6.1	785.25^b±4.3	<.01
5	821^d±6.4	884.75^a±10.4	877.25^ab±6.2	845^c±3.9	864.75^b±7.3	<.01
6	856^d±4.7	936^a±8.2	927.25^ab±5.7	907^c±9.9	915.50^bc±7.2	<.01
Finisher phase	2432.25^d±13.4	2630.25^a±24.2	2588^b±8.04	2520.50^c±12	2565.50^b±13.7	<.01
Overall (d1–42)	3355^d±20	3564.75^a±19.8	3521.75^ab ±25	3459.50^c±25	3497.75^bc±27	<.01

Means in the same row with different superscripts are significantly different (P<.01).

Table 3 shows the comparative efficacy of commercial coccidiostat and Aloe vera on weight gain in broilers. At the end of the first, second, third week and starter phase, weight gain was not significantly different (P>.05) between the groups. At the end of the fourth week, weight gain was significantly higher (P<.01) in the group −C when compared with the group AM1, group AV2.5 and group AV5 followed by  +C. At the end of the fifth week, weight gain was significantly higher (P<.01) in the group −C and group AM1 in comparison with the group AV2.5 followed by the group +C. After the sixth week, results showed that weight gain was significantly higher (P<.01) in the group −C, group AM1 and group AV5 than group AV2.5 followed by the group +C. At the finisher phase, weight gain was significantly higher (P<.01) in the group -C than the group AM1 and group AV5 followed by the group AV2.5 and group +C. For an overall period, weight gain was significantly higher (P<.01) in the group −C when compared with the group AM1 and AV5.

Table 3. Comparative efficacy of Amprolium and Aloe vera against coccidiosis challenge on weight gain (g) in broilers.

Weeks	+ C	−C	AM1	AV2.5	AV5	P-value
1	113 ± 4.5	123 ± 7.1	121 ± 4.2	124 ± 4.6	120 ± 5	.08
2	237 ± 4.3	238 ± 3.7	240 ± 3.3	236 ± 6.02	240 ± 5.4	.57
3	329 ± 7.4	330 ± 7.8	331 ± 5.1	328 ± 5.4	330 ± 5.9	.95
Starter phase	680 ± 3.2	692 ± 4.2	693 ± 10.4	689 ± 12.2	691 ± 6.5	.18
4	333^c ± 4.8	420^a ± 11.5	389^b ± 2.7	376^b ± 7.2	389^b ± 5.3	<.01
5	284^c ± 7.1	368^a ± 2.1	363^a ± 5.4	347^b ± 10.6	354^ab ± 7.4	<.01
6	323^c ± 3.6	375.75^a ± 8.2	364.50^a ± 5.5	341.25^b ± 6.8	365.25^a ± 5.9	<.01
Finisher phase	941^d ± 11.2	1164.75^a ± 17.1	1117.25^b ± 10.6	1065.75^c ± 16.3	1109.25^b ± 15	<.01
Overall (d1–42)	1621^c ± 9.8	1856.75^a ± 21.2	1810.75^b ± 8.1	1755.00^c ± 20.3	1801.00^b ± 18.2	<.01

Means in the same row with different superscripts are significantly different (P<.01).

The average daily feed intake and the average daily gain of the infected broiler supplemented with Aloe vera are given in Figure 1. The results showed that there was no significant difference between the control and treatment groups.

Figure 1. Average daily feed intake and average daily gain in coccidiosis-infected broiler fed with Aloe vera.

The comparative efficacy of Amprolium and Aloe vera against coccidiosis challenge on FCR has been described in Table 4. No significant difference (P>.05) was observed for FCR in the first, second and third week as well as at the end of the starter phase. At the end of the fourth week, significantly better (P<.01) FCR was recorded for the group −C when compared with the group AM1, group AV2.5 and group AV5 followed by the group +C. At the end of the fifth week of the trial, FCR was significantly higher (P<.01) in the group +C in comparison with the group −C, group AM1, group AV2.5 and group AV5. At the end of the sixth week, FCR was significantly higher (P<.01) in the group AV2.5 when compared with group AV5 followed by the group AM1 and group –C. At the finisher phase, significantly higher (P<.01) FCR was recorded for the group +C than group AV2.5 followed by group AM1, AV5 and group –C. The same trend was followed for the overall period of trial where significantly higher (P<.01) FCR was recorded for group  +C than group AV2.5, followed by the group AM1, AV5 and group –C.

Table 4. Comparative efficacy of Amprolium and Aloe vera against coccidiosis challenge on FCR in broilers.

Weeks	+ C	−C	AM1	AV2.5	AV5	P-value
1	1.15 ± 0.04	1.08 ± 0.04	1.09 ± 0.09	1.04 ± 0.03	1.10 ± 0.06	.21
2	1.18 ± 0.01	1.19 ± 0.01	1.18 ± 0.04	1.20 ± 0.03	1.17 ± 0.02	.55
3	1.54 ± 0.07	1.56 ± 0.07	1.57 ± 0.06	1.59 ± 0.03	1.56 ± 0.04	.82
Starter phase	1.35 ± 0.02	1.35 ± 0.02	1.34 ± 0.04	1.36 ± 0.01	1.34 ± 0.04	.95
4	2.26^a ± 0.05	1.93^c ± 0.02	2.01^b ± 0.03	2.04^b ± 0.04	2.01^b ± 0.01	<.01
5	2.88^a ± 0.08	2.40^b ± 0.04	2.44^b ± 0.03	2.43^b ± 0.08	2.41^b ± 0.04	<.01
6	2.64^ab ± 0.03	2.49^c ± 0.06	2.50^c ± 0.04	2.65^a ± 0.05	2.54^bc ± 0.04	<.01
Finisher phase	2.58^a ± 0.04	2.26^c ± 0.02	2.31^bc ± 0.02	2.37^b ± 0.03	2.31^bc ± 0.02	<.01
Overall (d1–42)	2.07^a ± 0.02	1.92^c ± 0.01	1.94^bc ± 0.01	1.97^b ± 0.01	1.94^bc ± 0.02	<.01

Means in the same row with different superscripts are significantly different (P<.01).

The comparative efficacy of Amprolium and Aloe vera against coccidiosis challenge on microscopic lesion scoring and oocyst count has been described in Table 5. The results revealed that significantly higher (P<.01) caecal lesion was observed in the group  +C than the group −C and group AV5. Caecal oocysts were significantly higher (P<.01) in the group +C than group AV2.5 and group AV5 followed by the group −C. The same trend was followed for intestinal lesions in the group +C and AV5 followed by the group –C. Intestinal oocysts were found significantly higher (P<.01) in the group +C than group AM1 and group AV2.5 followed by the group AV5 and group –C.

Table 5. Comparative efficacy of Amprolium and Aloe vera on the histopathology of the caeca and intestine against Coccidiosis in broilers.

Parameters	+ C	−C	AM1	AV2.5	AV5	P-value
Caeca lesions	3.00^a ± 0.8	0.25^c ± 0.5	2.00^ab ± 0.8	1.75^abc ± 0.9	1.25^bc ± 0.5	<.01
Caeca oocysts	2.25^a ± 0.5	0.25^c ± 0.5	1.75^ab ± 0.5	1.00^bc ± 0.0	1.00^bc ± 0.0	<.01
Intestinal lesions	2.50^a ± 1.0	0.00^c ± 0.0	1.75^ab ± 0.9	1.25^abc ± 0.5	1.00^bc ± 0.0	<.01
Intestinal oocysts	3.25^a ± 0.5	0.00^c ± 0.0	1.25^b ± 0.5	1.25^b ± 0.5	0.75^bc ± 0.5	<.01

Means in the same row with different superscripts are significantly different (P<.01).

Table 6 shows the comparative efficacy of Amprolium and Aloe vera on TLC and DLC against coccidiosis in broilers. Heterophil count was significantly lower (P<.01) in the group −C and group AM1. Significantly, higher (P<.01) monocyte count was observed in the group −C than group AV2.5 and AV5 followed by group +C. Eosinophil count was significantly higher (P<.01) in the treatment groups compared to the control. No significant difference (P>.05) was recorded for lymphocyte count. Total leucocytes count was significantly higher (P<.01) in the group + C than group AM1, group AV2.5, group AV5 and group −C.

Table 6. Comparative efficacy of Amprolium and Aloe vera on total leucocytes and differential leucocytes count against coccidiosis in broilers.

Parameters	+C	−C	AM1	AV2.5	AV5	P-value
Heterophils (%)	75.00^a ± 2.98	65.12^c ± 298	65.34^c ± 369	70.56^b ± 230	73.98^ab ± 208	<.01
Monocytes (%)	3.5^c ± 0.21	4.95^a ± 0.12	3.6^ab ± 0.60	3.48^bc ± 0.26	3.52^bc ± 0.1	<.01
Eosinophil (%)	2.32^b ± 0.27	3.00^a ± 0.81	2.82^a ± 0.12	2.67^a ± 0.95	2.91^a ± 0.81	<.01
Lymphocytes (%)	16.95 ± 1.5	14.75 ± 1.2	16.00 ± 0.81	15.17 ± 0.56	15.75 ± 0.95	.091
TLC (×10^3)	3.4^a ± 9.57	2.67^b ± 6.29	2.9^b ± 8.16	2.9^b ± 8.16	2.8^b ± 9.32	<.01

Means in the same row with different superscripts are significantly different (P<.01).

Discussion

Eimeria is a highly infectious disease and can easily be transferred by insects, contaminated feed, equipments and water. Coccidiosis causes serious economic losses and can be controlled by antibiotics and vaccination. However, there is a need to develop alternatives due to the rising drug-resistance against the parasites (Chand et al. 2016). The phytochemicals can interfere with disease propagation through a number of the signalling pathway; therefore, herbal remedies are emerging as new alternative and attractive methods to control coccidiosis (Kaingu et al. 2017). Aloe vera is among the earliest known plants for different pharmacological uses. Aloe vera and A. secundiflora have been shown to inhibit the invasion and replication of different species of coccidia in the broiler (Yim et al. 2011; Kaingu et al. 2017).

The findings of the current study suggested that the overall performance was significantly higher in –ve control group when compared with groups treated with Amprolium and Aloe vera at 5 g/l water and remained the lowest in the +ve control group. Conflicting results have been published in the literature for treating birds with Aloe vera with and without coccidial infection (Yim et al. 2011; Shokraneh et al. 2016; Islam et al. 2017; Singh 2017). Aloe gel has several medicinal uses including the correction of digestive disorders. In addition, it has positive effects on microflora and reduces inflammation. Surjushe et al. (2008) concluded that Aloe vera contains growth hormones (auxins and gibberellins) and polysaccharide (glucomannan) which reduced the inflammation and haemorrhages in the intestines caused by Eimeria species that led to better digestion and enhanced weight gain.

The findings of the present study regarding intestinal lesions indicated that caecal lesions, caeca oocyst, intestinal lesions and intestinal oocysts were observed less in the negative control group and more frequently observed in the positive control group when compared with birds treated with Amprolium. Positive effects of Aloe vera supplementation in broiler production infected with coccidia have been reported in the previous studies (Yim et al. 2011; Akhtar et al. 2012; Kaingu et al. 2017). Aloe vera may interfere with critical stages of Emeria development and reduces damages to the intestinal wall of the chicken by reducing oocysts count, lesion score and haemorrhages. The positive effect of Aloe vera may be associated with its antioxidant activity which may reduce the severity of the infection (Kaingu et al. 2017). Researchers in the recent past are in agreement with the benefits of Aloe vera against different diseases since Aloe vera has active compounds including beta-carotene, vitamin C and E, vitamin B12, folic acid, choline, glucomannans, acemannans, proteins and minerals, which effectively control multiplication of oocysts in the intestines and caeca of the birds (Surjushe et al. 2008). Due to the multiple beneficial compounds, we may infer that the pathological lesions caused by coccidiosis infection may have been reduced.

In the current study, the number of heterophils and TLC significantly decreased while the number of monocytes and eosinophils increased in the treatment groups. The number of lymphocytes was similar among all groups. From the results, it is also clear that the infection increased the monocytes and eosinophils and decreased the heterophils and TLC. On the other hand, the treatment of Aloe vera ameliorated these alterations. Normal TLC and DLC counts help in the proper functioning of the body and enabling the birds to fight the infection and restore the deteriorating feed efficiency in birds (Ajayi and James 2015). Singh et al. (2013) demonstrated that Aloe vera contains different active compounds (glucomannans, acemannans, carotene, vitamin B12) which help in boosting the immunity by increasing total leucocytes count. Healing and anti-inflammatory properties of Aloe vera may have helped the birds in ceasing Eimeria oocysts multiplication in the intestines and caeca leading to the normal leucocytes count (Surjushe et al. 2008).

Conclusion

The findings of the present study indicated that supplementation of 5 g/l Aloe vera gel in drinking water produced encouraging results in broiler during the coccidial challenge.

Disclosure statement

No potential conflict of interest was reported by the author(s).