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. 2022 Sep 1;8(6):2511–2520. doi: 10.1002/vms3.923

The effects of green tea in the diet of broilers challenged with coccidiosis on their performance, carcass characteristics, intestinal mucosal morphology, blood constituents and ceca microflora

Keyvan Jelveh 1, Behrouz Rasouli 1, Isam T Kadim 2, Marina Ivanovna Slozhenkina 3, Ivan Fedorovich Gorlov 3, Alireza Seidavi 1,✉, Clive J C Phillips 4,5
PMCID: PMC9677413  PMID: 36049150

Abstract

Background

Coccidiosis is an endemic protozoal disease of chickens normally controlled by ionophores. However, coccidiostats are also antibiotics, and evidence of resistance in both coccidia and bacteria may develop and reduce antibacterial activity in humans. This has led to a search for natural coccidiostats, such as green tea.

Objectives

To study the effects of supplementing broilers with various levels and types of green tea, in comparison to use of a conventional coccidiostat or a control, unsupplemented diet.

Methods

A total of 360 male, day‐old Ross 308 broilers (days 1–42) were used to evaluate the gut morphology and performance when challenged with coccidiosis and fed varying dietary levels of green tea powder or extract. Treatments were Negative control (NC, unsupplemented control diet); positive control (PC, control diet + commercial coccidiostat); control diets with 0.2, 0.3 or 0.4 g/kg green tea extract (GTE 0.2, 0.3 and 0.4); and control diets with 1, 2 or 3 g/kg green tea powder (GTP 1, 2 and 3).

Results

Compared with NC, PC and all green tea treatments, but particularly GTE0.4, increased feed intake and growth rate, with the best feed conversion ratio at GTE0.4. As a proportion of carcase weight, higher inclusion rates increased intestine weight and decreased abdominal fat. The duodenum, jejunum and ileum of birds fed green tea, and particularly GTE0.4, had longer, wider villi, and shallower crypts. Epithelium thickness was reduced by green tea and PC, compared to NC. Clostridium perfringens and coliform populations decreased in proportion to green tea inclusion rate and decreased in PC. Lactobacilli increased with green tea and were more for NC than PC. Green tea at the highest concentrations reduced blood glucose and LDL and VLDL cholesterol.

Conclusions

Green tea offers a possible replacement for conventional ionophores to control coccidiosis in broiler chickens. The best inclusion rate was 0.4 g/kg.

Keywords: antibody, blood parameters, broiler, Camellia sinensis, growth parameters, feed conversion ratio, feed intake

To study the effects of supplementing broilers with various levels and types of green tea, in comparison to use of a conventional coccidiostat or a control, unsupplemented diet. In conclusion, green tea offers a possible replacement for conventional ionophores to control coccidiosis in broiler chickens. The best inclusion rate was 0.4 g/kg.

graphic file with name VMS3-8-2511-g001.jpg

1. INTRODUCTION

Poultry production is growing in many regions, which contributes to increased protein requirements for humans (Dieye et al., 2010). However, the expanding poultry sector faces many challenges, including diseases. Antibiotics have widely been used for many decades, both as growth promoters and to prevent diseases (Phillips et al., 2004). Despite improving feed utilisation efficiency, there may be residue problems, and resistant bacteria, when consumed by humans in poultry products may lead to antibiotics being ineffective in poultry and humans (Gould, 2008; Van de Bogaard & Stobberingh, 2000). Therefore, many countries, including the EU, have made the inclusion of antibiotics in poultry feed illegal (Butaye et al., 2000; Catalá‐Gregori et al., 2002). There is now a widespread search for alternatives to antibiotics.

Herbal products in poultry diets can enhance performance (Teteh et al., 2013) and reduce mortality (Kumar et al., 2003). Green tee (Camellia sinensis) is a natural, non‐toxic product containing many bioactive compounds: alkaloids, polyphenols, polysaccharides, volatile oils, vitamin C and minerals (Karori et al., 2007; Khan, 2014). The leaves contain approximately 92.2% dry matter, 82.4% organic matter, 19.3% crude fibre, 8.7% ether extract, 9.8% ash, 18.1% crude protein, 36.2% nitrogen free extract and 3002 kcal/kg (Abdo et al., 2010). They have therapeutic properties: antimicrobial, antioxidant, immune modulatory (Erener et al., 2011; Ko & Yang, 2008; Nishida et al., 2006), and, most importantly for poultry, anticoccidial (Jang et al., 2007). Coccidiosis is an endemic protozoal disease of the intestine transmitted between broilers through faeces. It causes diarrhoea unless controlled with an ionophore coccidiostat in the feed, which also controls some bacteria. These antimicrobials are classified as feed additives in the EU, as farmers want to retain use of ionophores for controlling coccidiosis, despite resistance in coccidia, but also resistance to bacitracine and vacomycine, both antibiotics used in human medicine [Norwegian Scientific Committee for Food Safety (VKM) 2015]. When birds are fed coccidostats, the risk of exposure to coccidiostat‐resistant bacteria is high in people handling manure and medium in people handling carcases and raw meat. In the United States, coccidiostats are classified as antibiotics and therefore not allowed in the rapidly developing antibiotic‐free production systems (Swormink, 2019).

Green tea may enhance growth in poultry (Cross et al., 2007; Demir et al., 2003) and in humans, it has many health benefits: reduced obesity and blood pressure, blood glucose control, anti‐hypercholesterolaemia (Liao et al., 2001). With these benefits, green tea may enhance production and replace antibiotic growth promoters in broilers (Cao et al., 2005; Kaneko et al., 2001; Khan, 2014). However, the physiological effects are largely unknown; hence, we studied these in broilers supplemented with green tea products, a conventional coccidiostat or no supplement.

2. MATERIALS AND METHODS

This study was conducted at a commercial poultry farm.

2.1. Birds, diets and experimental groups

A total of 360 male Ross 308 broiler day‐old chicks were purchased from a local hatchery and randomly distributed into 8 treatments, with 3 replicates of 15 birds in eight treatments, in a completely randomised design:

  • Group 1 (NC) (negative control): no supplement

  • Group 2 (PC) (positive control): commercial coccidiostat (Salinomycin; C42H70O11) at 0.5 g/kg (Rooyan Darou Co)

  • Group 3 (GTE0.2): PC and 0.2 g/kg green tea extract

  • Group 4 (GTE0.3): PC and 0.3 g/kg green tea extract

  • Group 5 (GTE0.4): PC and 0.4 g/kg green tea extract

  • Group 6 (GTP1): PC and 1 g/kg green tea powder

  • Group 7 (GTP2): PC and 2 g/kg green tea powder

  • Group 8 (GTP3): PC and 3 g/kg green tea powder

Green tea was obtained from an autumn harvest of Chinese green tea (Camellia sinensis, Langeroud's Nooshineh Co., http://www.nooshinehco.com). Green tea extract was prepared by heating 100 g dry tea leaves with 200 ml distilled water at 80°C for 10 min (Bombik et al., 2012). Green tea powder was obtained by grinding and passing through a 0.5 mm sieve. Chicks were reared from days 1 to 42, in starter (1–10 days), grower (11–24 days) and finisher (25–42 days) periods. Dietary ingredients and nutrient composition of experimental diets, fed as mash, are in Table 1. Feed and water were available ad libitum.

TABLE 1.

Feed ingredients and nutrient analysis of the diets used during the starter (days 1–10), grower (days 11–24) and finisher (days 25–42) periods

Ingredient (%) Days 1–10 Days 11–24 Days 25–42
Maize (Zea mays) 57.8 58.6 61.6
Soybean meal (44% P) 36.71 35.47 31.66
Soybean oil 1.60 2.20 3.17
Limestone 1.26 1.24 1.09
Dicalcium phosphate 1.56 1.30 1.15
NaCl 0.20 0.25 0.32
Mineral and vitamin mixture* 0.50 0.50 0.50
DL‐Methionine 0.20 0.25 0.30
L‐Lysine hydrochloride 0.04 0.05 0.07
Total 100 100 100
Calculated nutrients
Energy (ME) (kcal/kg) 2900 2950 3000
Crude protein 21.0 20.5 18.75
Crude fat (%) 2.42 4.64 2.86
Linoleic acid (%) 2.81 2.13 1.46
Calcium (%) 0.94 0.87 0.78
Available phosphorus (%) 0.42 0.38 0.35
Sodium (%) 0.19 0.17 0.15
Lysine (%) 1.30 1.10 1.10
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*

Containing: calcium pantothenate: 4 mg/g; niacin: 15 mg/g; vitamin B6: 13 mg/g; Cu: 3 mg/g; Zn: 15 mg/g; Mn: 20 mg/g; Fe: 10 mg/g; K: 0.3 mg/g; vitamin A: 5000 IU/g; vitamin D3: 500 IU/g; vitamin E: 3 mg/g; vitamin K3: 1.5 mg/g; vitamin B2: 1 mg/g.

The animals in each group were kept in 1.2 ×1.2 m pens, bedded with wood shavings. A standard thermo‐neutral ambient temperature was maintained, in accordance with standard rearing practices (Aviagen 2007). Lighting was provided for 23 h on days 1–7 and 40–42, and for 20 h/day on days 8–39.

Routine vaccination and deworming were applied according to a programme certified by the regional veterinary authority. At day 9 of age, birds were vaccinated against infectious bronchitis and on days 9, 18 and 31 against Newcastle disease, all vaccinations being administered via the drinking water.

2.2. Coccidiosis challenge

All chickens were challenged with coccidiosis according to standard protocols (Ott et al., 2018). At day 21, all treatments except for NC were orally gavaged with 1.5 ml/chick suspension containing 200,000 live spored oocysts, including Eimeria tenella, E. negatrix, E. maxima and E. acervulina.

2.3. Broiler measurements

Feed intake and weight gain were recorded over two periods (days 1–21 and 22–42). Feed conversion ratio was calculated as feed intake/weight gain, and measurements were made, based on standard methods (Giannenas et al., 2011; Shabani et al., 2015). In brief, at day 42 after 4 h of fasting to evacuate the gastrointestinal tract, one bird with a representative body weight for each replicate was slaughtered for measuring carcass yield and distribution of meat and gastrointestinal tract characteristics. Feathers were removed by dry plucking. Neck, wingtips and feet at the tibio‐tarsal joint were separated from the carcass. The gastrointestinal tract (from oesophagus to rectum), heart, kidney and liver were removed, and the empty or edible carcass weighed.

The different parts and the digestive tract of the carcasses were dissected, and the economically relevant parts weighed: first, breast muscle, including skin and sternum, then legs (thighs and drumsticks, dissected by disarticulation of the hip joint and dissecting tissue from the iliac bone). Cecum content was collected to evaluate microflora. All dimensions were rounded to whole integers. The weights of all dissected parts were related to the totally eviscerated carcass [(weight of component(s)/eviscerated carcass weight) × 100]. Three samples of 1 cm length from the centre of three intestinal segments were taken: first, the middle of the duodenum from gizzard outlet to the end of the pancreatic loop, second, middle of the jejunum from the pancreatic loop to Meckel's diverticulum, and third, 5 cm section from Meckel's diverticulum to the ileocaecocolic junction. After washing with soluble phosphate‐buffered saline (PBS), samples were fixed in 6 ml 10% buffered formalin for 2 days, embedded in paraffin, sectioned at 3 μm and stained the haematoxylin‐eosin and periodic acid‐Schiff methods. One cm long intestine segments were taken from the centre of each sample for morphometric studies under light microscopy. Three researchers independently measured villus length and width, crypt depth, epithelium thickness and number of goblet cells in the epithelium in each sample. Using ten villi and the corresponding crypts per section, villous length was estimated as the vertical distance from the tip to the crypt junction level, crypt depth as the vertical distance from villous‐crypt junction to crypt base (Nahavandinejad et al., 2014).

2.3.1. Blood samples

Measurements were based on methods described previously (Nahavandinejad et al., 2014). Briefly, at the end of the 42‐day experiment, one bird from each replicate was randomly selected for morning blood sampling – 3 birds per group. Prior to blood collection and slaughter, feed was removed for 4 h to stabilise plasma constituents. Samples (∼5 ml/bird) were collected from the wing vein (Vena cutanea ulnaris) into tubes with 10 mg of the anticoagulant ethylenediaminetetraacetic acid (EDTA) for plasma separation, and immediately transferred to the laboratory. Plasma was harvested after centrifugation (3000 rpm, for 10 min at room temperature) and stored at –20°C until analysed.

2.3.2. Determination of blood parameters

Blood parameters analysed were glucose (Gl), total cholesterol (Chol), triglycerides (TG), high‐density lipoprotein (HDL), low‐density lipoprotein (LDL), very‐low‐density lipoprotein (VLDL), albumin (Alb), total protein (TP) and alkaline phosphatase (AP). Blood plasma parameters were analysed using a Roche Cobas Integra autoanalyzer (Roche Diagnostics, GmbH, Mannheim, Germany), based on standard protocols prescribed by the manufacturer of commercial kits (Pars Azmoon Co., Tehran, Iran) and described previously (Nahavandinejad et al., 2014; Shabani et al., 2015).

2.3.3. Composition of cecum microbiota

Measurements were based on methods described previously (Dibaji et al., 2014). Briefly, agar plates were streaked with ceca contents for determination of bacterial growth from colony counts. Collecting tubes were weighed, wrapped in aluminium foil and autoclaved for 10 min. Culture mediums were prepared 24 h before collecting samples. Tryptose‐sulphite Cycloserin Egg Yolk agar‐TSC (CM 587 oxoid) was used for Clostridium perfringens, MacConkey agar (105465.0500) to culture coliforms, and MRS agar (Man Rogosa Sharpe agar, 1.10660.500) to culture Lactobacilli. Samples were transferred to the laboratory and weighed, with sample weight the difference between these two values. Tubes for bacteria isolated from gastrointestinal contents and preparation of suspension were shaken for 30 minutes. One millilitre was removed from the suspension and added to 9 ml buffer phosphate saline (PBS). The suspension was prepared from 10−1 dilutions and serial dilutions to 10−2, 10−3, 10−4, 10−5 and 10−6. A 100 μl sample was then removed from 10−4, 10−5 and 10−6 dilutions and poured into the Petri dish previously prepared with the medium and completely distributed. Lactobacilli bacteria were incubated at 37°C in anaerobic conditions for 72 h, then bacteria were counted by a colony counter (Digital Magnificent Colony Counter, P118, S.ELC).

2.4. Statistical analysis

Results are presented as means ± standard error of the mean. The Shapiro–Wilk test confirmed normal distribution of the data, except bacterial counts, which were transformed to logarithm10 no. bacteria/g. Significance of treatment differences was analysed using ANOVA, followed by a Tukey's post hoc test to separate means, using IBM SPSS Statistics 21 software for Windows® (SPSS 1997). p Values ≤ 0.05 were regarded as statistically significant.

3. RESULTS

All birds remained healthy throughout the study, with no evidence of diarrhoea and no mortality.

3.1. Productive performance

In the first half of the study, feed intake was least in NC, increased in PC and further increased by GTE0.4 (Table 2). In green tea powder treatments, however, feed intake was intermediate between NC and PC. Weight gain was least in NC, increased in PC and green tea treatments and was highest in GTE0.4. FCR was least in GTE0.4, and reduced for all green tea levels and PC, compared with NC.

TABLE 2.

Performance of Ross 308 male broilers (n = 360) fed different levels of green tea extract (GTE) or powder (GTP), a neutral control (NC) or a positive control (PC)

Treatment Feed intake (g/chick) Weight gain (g/chick) Feed conversion ratio
First half of study (days 1–21)
NC 978c 641d 1.52a
PC 1031b 740b 1.39bc
GTE0.2 1031b 736b 1.40bc
GTE0.3 1036ab 743b 1.40bc
GTE0.4 1047a 762a 1.37c
GTP1 1030b 726bc 1.42bc
GTP2 1027b 715c 1.44b
GTP3 1025b 710c 1.44b
p Value 0.0001 0.0001 0.0001
SEM 4.13 7.33 0.0097
Second half of study (days 22–42)
NC 3792b 2009d 1.89
PC 3937a 2145ab 1.84
GTE0.2 3841b 2098bc 1.83
GTE0.3 3934a 2147ab 1.83
GTE0.4 3943a 2192a 1.80
GTP1 3798b 2074cd 1.83
GTP2 3823b 2070cd 1.85
GTP3 3833b 2065cd 1.86
p Value 0.0001 0.0007 0.1219
SEM 13.755 13.097 0.007
Total study
NC 4770c 2650d 1.80a
PC 4968a 2885b 1.72bc
GTE0.2 4872b 2833bc 1.72bc
GTE0.3 4970a 2890b 1.72bc
GTE0.4 4990a 2953a 1.69c
GTP1 4828b 2800c 1.72bc
GTP2 4850b 2785c 1.74b
GTP3 4858b 2775c 1.75b
p Value 0.0001 0.0001 0.0001
SEM 16.2950 18.8301 0.0071
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*

Means within each column with common superscripts do not differ significantly (p < 0.05).

NC: negative control; PC: positive control; GTE: green tea extract; GTP: green tea powder; SEM: standard error of mean.

In the second half of the study, feed intake was increased in GTE0.3 and 0.4 and PC, compared with other treatments. Weight gain was highest in these treatments, and least at the higher powder inclusion rates and NC. FCR was not affected by treatment.

Over the entire study, feed intake was greatest for in GTE0.3 and 0.4 and PC, least for NC and intermediate in other treatments. Weight gain was greatest in the GTE0.4, least in NC and was greater in PC than the 1–3 g/kg powder treatments. FCR was least in GTE0.04 and greatest in NC.

3.2. Carcase composition

There were no significant treatment effects on the carcase weight, or on breast, drumstick or proventriculus weights, as % of carcase weight (Table 3). However, ventriculus weight was increased at the three highest green tea inclusion rates, as powder, compared with the other treatments. Intestine weight was greatest at the highest powder inclusion rate and was also greater at the second highest inclusion rate than the controls and lower inclusion rates. Abdominal fat was greatest in the two control treatments and decreased with inclusion rate of green tea.

TABLE 3.

Carcass weight as a % of total weight, and the weight of breast, drumstick, ventriculus, proventriculus, intestine and body fat, as % of carcase weight, at day 42 of age in Ross 308 male broilers (n = 24) fed different levels of green tea extract (GTE) or powder (GTP), a neutral control (NC) or a positive control (PC)

Treatment Eviscerated carcass (% total weight) Breast (% CW) Drumstick (% CW) Ventriculus (% CW) Proventriculus (% CW) Intestine (% CW) Abdominal fat (% CW)
NC 64.86 28.68 24.90 0.94b 0.49 5.84cd 1.36a
PC 65.51 28.08 24.26 0.94b 0.43 5.46d 1.42a
GTE0.2 65.77 29.11 24.62 0.94b 0.47 5.78cd 1.21b
GTE0.3 66.55 29.24 24.91 0.94b 0.48 5.77cd 1.21b
GTE0.4 66.75 29.90 25.43 0.88b 0.45 5.49d 1.03c
GTP1 65.75 28.46 24.39 1.17a 0.43 7.02bc 0.95cd
GTP2 65.35 29.51 25.27 1.17a 0.43 7.42b 0.88d
GTP3 65.55 28.57 25.18 1.18a 0.48 8.92a 0.89d
p Value 0.37 0.07 0.25 0.0003 0.63 0.0002 0.0001
SEM (standard error of mean) 0.224 0.164 0.132 0.0267 0.0099 0.268 0.423
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*

Means within each column with common superscript do not differ significantly at p < 0.05.

NC: negative control (diets not supplemented); PC: positive control (diets supplemented commercial coccidiostat); GTE: green tea extract; GTP: green tea powder.

3.3. Gut mucosa morphology

3.3.1. Duodenum

Villus length was longest for GTE0.04% extract treatment and least in the two control treatments (Table 4); it declined at higher inclusion rates of powder. Villus width increased, and epithelium thickness decreased, in the green tea treatments compared with NC, and intermediate in PC. Crypt depth was decreased at all levels of green tea inclusion, compared with NC, with PC intermediate. The ratio of villus length to crypt depth was greatest in the GTE0.4, then the lower extract concentrations, then the higher, powder, concentrations and, finally, least in NC and PC. Treatment did not affect the number of goblet cells.

TABLE 4.

Duodenal morphology at day 42 of age in Ross 308 male broilers (n = 24) fed different levels of green tea extract (GTE) or powder (GTP), neutral control (NC) or positive control (PC)

Treatment Villus length (μm) Villus width (μm) Crypt depth (μm) Villus length to crypt depth ratio Thickness of epithelium (μm) Number of goblet cells in epithelium
NC 1700d 148c 321a 5.31c 41.8a 5
PC 1910d 170bc 270b 7.08c 30.1c 8
GTE0.2 2393abc 197a 231bc 10.42ab 35.3b 10
GTE0.3 2390ab 199a 213c 11.36ab 34.2b 13
GTE0.4 2443a 213a 200c 12.31a 33.3b 15
GTP1 2310bc 193ab 233bc 10.01b 34.3b 11
GTP2 2320bc 191ab 239bc 7.79b 34.1b 11
GTP3 2290c 188ab 240bc 9.56b 33.3b 11
p Value 0.0001 0.001 0.0002 0.0001 0.002 0.35
SEM (standard error of mean) 53.35 4.57 8.24 0.486 0.69 0.93
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*

Means within each column with common superscript do not differ significantly at p < 0.05.

NC: negative control; PC: positive control; GTE: green tea extract; GTP: green tea powder.

3.3.2. Jejunum

Villus length and width were increased, and crypt depth was decreased, by green tea treatments, compared with NC and intermediate in PC (Table 5). Villus length to crypt depth ratio increased in all green tea treatments, compared with NC, and increased in all green tea treatments except the highest powder concentration, compared with PC. Epithelial thickness was greatest in NC and least in PC and green tea treatments up to 1 g/kg, and intermediate in GTP2 and 3. Goblet cells were least in NC and PC and increased for green tea treatments, with a peak for GTE 0.4.

TABLE 5.

Jejunum morphology at day 42 of age in Ross 308 male broilers (n = 24) fed different levels of green tea extract (GTE) or powder (GTP), a neutral control (NC) or a positive control (PC)

Treatment Villus length (μm) Villus width (μm) Crypt depth (μm) Villus length to crypt depth ratio Thickness of epithelium (μm) Number of goblet cells in epithelium
NC 990d 141c 217a 4.61c 43.2a 5c
PC 1180cd 170bc 190ab 6.22bc 30.2c 10c
GTE0.2 1400ab 188ab 155c 9.04a 33.1bc 18ab
GTE0.3 1461ab 195ab 153c 9.57a 32.0bc 19ab
GTE0.4 1483a 209a 148c 10.01a 30.2c 24a
GTP1 1383abc 189ab 158bc 8.83a 34.3bc 19ab
GTP2 1378bc 190ab 163bc 8.65a 34.7b 17b
GTP3 1381abc 190ab 163bc 8.47ab 35.0b 17b
p Value 0.0009 0.03 0.003 0.001 0.002 0.0001
SEM (standard error of mean) 34.85 5.13 5.41 0.388 0.91 1.26
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*

Means within each column with common superscript do not differ significantly at p < 0.05.

NC: negative control (diets not supplemented); PC: positive control (diets supplemented commercial coccidiostat); GTE: green tea extract; GTP: green tea powder.

3.3.3. Ileum

Villi were taller in all the green tea treatments compared with the two control treatments, which did not differ (Table 6). GTE0.4 had taller villi than other green tea treatments. Villus width increased and crypt depth decreased in the green tea treatments, compared with NC, with PC intermediate. Villus length to crypt depth ratio was increased in all green tea treatments, compared with NC and PC, and it increased in GTE0.4 compared with other green tea treatments. Epithelial thickness was less in the green tea treatments and PC, compared with NC. Goblet cells were greatest in GTE0.4, then other green tea treatments, then PC and least in NC.

TABLE 6.

Ileal morphology at day 42 of age in Ross 308 male broilers (n = 24) fed different levels of green tea extract (GTE) or powder (GTP), a neutral control (NC) or a positive control (PC)

Treatment Villus length (μm) Villus width (μm) Crypt depth (μm) Villus length to crypt depth ratio Thickness of epithelium (μm) Number of goblet cells in epithelium
NC 880c 120c 256a 3.43c 41.2a 5d
PC 990c 143bc 185bc 5.61c 30.0b 10c
GTE0.2 1150b 175ab 173bc 6.78b 33.2b 18b
GTE0.3 1175b 180a 168bc 7.02b 33.1b 19b
GTE0.4 1300a 195a 143c 9.11a 30.1b 24a
GTP1 1145b 168ab 175bc 6.57b 34.2b 19b
GTP2 1130b 171ab 183ab 6.21b 34.1b 18b
GTP3 1130b 173ab 180bc 6.45b 33.4b 17b
p Value 0.0001 0.005 0.03 0.0006 0.004 0.0001
SEM (standard error of mean) 27.2 5.55 7.81 0.354 0.816 1.21
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*

Means within each column with common superscript do not differ significantly at p < 0.05.

NC: negative control; PC: positive control; GTE: green tea extract; GTP: green tea powder.

3.4. Blood composition

Blood glucose was reduced in GTP2 and 3, compared with the two control treatments, with other green tea treatments intermediate (Table 7). Total cholesterol was reduced in all green tea treatments and PC compared with NC, and further decreased at the highest green tea inclusion rate, GTP3. Triglycerides were reduced at this highest rate compared to NC and PC and the lowest inclusion rate, GTE0.2. HDL cholesterol was not affected by treatment, but LDL cholesterol was reduced by all the green tea treatments, and particularly the highest level, compared with NC. PC was intermediate and greater than all green tea treatments except GTE0.2. VLDL was reduced by most green tea inclusion rates, but particularly the highest level, compared with NC and PC.

TABLE 7.

Blood parameters at day 42 of age in Ross 308 male broilers (n = 24) fed different levels of green tea extract (GTE) or powder (GTP), a neutral control (NC) or a positive control (PC)

Treatment Glucose (mg/dl) Total cholesterol (mg/dl) Triglycerides (mg/dl) HDL Cholesterol (high‐density lipoproteins) (mg/dl) LDL Cholesterol (low‐density lipoproteins) (mg/dl) VLDL (mg/dl)
NC 238ab * 201a 119a 61 117a 23.8a
PC 240a 173b 117a 58 91ab 23.3a
GTE0.2 219abc 168b 108ab 67 79bc 21.6ab
GTE0.3 213abc 169b 85bcd 75 78c 17.0bcd
GTE0.4 203abc 160b 75cd 74 71c 15.0cd
GTP1 198bc 164b 103abc 66 78c 20.6abc
GTP2 193c 165b 93abcd 67 79c 18.6acd
GTP3 183c 133c 74d 70 49d 14.8d
p Value 0.04 0.0001 0.008 0.18 0.0001 0.008
SEM (standard error of mean) 5.39 4.05 4.30 1.74 3.83 0.859
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*

Means within each column with common superscript do not differ significantly at p < 0.05.

NC: negative control; PC: positive control; GTE: green tea extract; GTP: green tea powder.

3.5. Cecum bacterial content

Populations of Clostridium perfringens were reduced in proportion to green tea inclusion rate, compared with NC, with PC the same as all green tea inclusion rates except GTP3 (Table 8). Coliforms were decreased in approximate proportion to green tea inclusion rate, compared with NC, but PC was the same as higher levels of green tea. Lactobacilli were increased by green tea inclusion, more at GTP2 and 3, compared with NC and particularly PC.

TABLE 8.

Mean (±SEM) cecum microbial populations at day 42 in Ross 308 male broilers (n = 24) fed different levels of green tea extract (GTE) or powder (GTP), a neutral control (NC) or a positive control (PC)

Clostridium perfringens Coliforms Lactobacillus
Treatment (log10 CFU/g) (log10 CFU/g) (log10 CFU/g)
NC 2.59a 7.65a 7.54c
PC 2.25bcd 7.40cd 6.48d
GTE0.2 2.34b 7.56ab 7.65b
GTE0.3 2.31bc 7.54ab 7.73ab
GTE0.4 2.22cd 7.40cd 7.78a
GTP1 2.28bcd 7.48bc 7.67b
GTP2 2.20d 7.39cd 7.81a
GTP3 1.80e 7.34d 7.82a
p Value 0.0001 0.0007 0.0001
SEM (standard error of mean) 0.0437 0.0234 0.0873
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*

Means within each column with common superscript do not differ significantly at p < 0.05.

NC: negative control; PC: positive control; GTE: green tea extract; GTP: green tea powder; CFU: colony forming units.

4. DISCUSSION

Green tea increased length and width of villi in the different intestinal segments, as well as reducing crypt depth, compared with NC. This led to increased growth rates and feed conversion ratio, as well as increased feed intake, particularly at the lower inclusion rates. The improvement in weight gain of experimental birds fed with GTE0.4, compared with PC, may be due to the action of flavonoids, known to have anticoccidial effects due to antioxidants (Chen et al., 2008; Jang et al., 2007).

The increase in feed intake contrasts with decreases reported previously (Alimohammadi‐Saraei et al., 2014; Biswas & Wakita, 2001), but these authors included green tea at much higher doses. The smaller increase in intake at our higher inclusion rates suggests a dose dependent effect. Others (Hrnčár & Bujko, 2017) reported no effect of green tea supplementation on feed intake in broilers. Observed reductions in feed intake (Alimohammadi‐Saraei et al., 2014; Biswas & Wakita, 2001), attributed to suppression of lipid metabolism, reduced broilers’ growth, probably unacceptable to the industry. Yang et al. (Yang et al., 2003) also observed a reduction in weight gain of broilers fed 10 g/kg of their diet as green tea by‐product, high in tannins. Biswas and Wakita (Biswas & Wakita, 2001) found that green tea powder included at 5, 7, 10, and 15 g/kg only tended to decrease weight gain at the highest dose. The optimum inclusion rate for production performance, at which FCR is least, may be 0.4 g/kg. Differing effects may also be due to the type of green tea used, in particular catechin content. Improved FCR could be attributed to the most abundant catechin, epigallocatechin gallate. Other green tea components, alkaloids, carotenoids, minerals, amino acids (especially l‐theanine), and volatiles compounds, decrease pathogens and improve nutrient absorption, leading to better FCR (Engelhardt, 2010).

4.1. Carcass characteristics

We observed no effects on dressing %; neither did Biswas and Wakita (Biswas & Wakita, 2001) when they added green tea to broilers’ diet at 5–15 g/kg. In contrast, Guray et al. (2011) observed increased dressing percentage. A reduction in abdominal fat has also been reported by Yang et al. (2003) and Guray et al. (2011). The increase in intestine weight at our two highest levels probably reflects changes in villi. Contrary to our study, Uuganbayar (2004), when supplementing with 0.5% green tea, recorded lighter small intestines compared to a control diet.

4.2. Intestinal morphology

Similar results were obtained from the three sections of the intestine. Villi length and width were increased by green tea, especially in GTE0.4. In Hassanpour et al.’s (2010) study, jejunal but not duodenal or ileal villus length increased, with no increase in villus width. In our study, the proportional increase in villus length was similar in the three intestinal segments (+55%, 48% and 44% for the jejunum, ileum and duodenum, respectively). Epithelial tissue was thinner and there was a tendency for the number of goblet cells to increase, at least in the jejunum and ileum, and again especially in GTE0.4. Thinner epithelial tissue may facilitate nutrient absorption, also longer villi. Intestinal villi are the main site of nutrient absorption (Ray et al., 2002). There is a positive relationship between between villus length, nutrient absorption and digestibility (Mekbungwan et al., 2002). Thus, it was noticeable that birds in treatment GTE0.4 gained most weight, had biggest villi and most goblet cells. Goblet cells secrete mucin glycoproteins that protect the epithelium and they sense microbiota, sampling bacteria and transferring them to dendritic cells, thereby actively participating in immune responses to pathogens (Shira & Friedman, 2018). Shorter villi and deeper crypts will reduce nutrient absorption. According to Yason et al. (1987), deep crypts indicate fast renewal of the villi in response to tissue inflammation or toxins produced by pathogens. Thus, it is likely that it was the changes in crypt depth and thickness of the epithelium, which caused the slower weight gain observed in NC.

4.3. Haematological parameters

As has been demonstrated, green tea is hypocholesterolemic, particularly of LDL cholesterol. Green tea reduces lipoprotein lipase and adipose triglyceride lipase (Mohammadpour et al., 2021). This action of green tea may be link to polyphenols, which enhance reverse‐cholesterol transport: cholesterol is removed from peripheral tissues and delivered to the liver, reducing cholesterol absorption (Tebib et al., 2004). Other researchers have found that 1.5 g/kg of green tea significantly decreased plasma triglyceride and total cholesterol, compared to a control group (El‐Deek et al., 2012).

4.4. Cecal microbial population

Coliforms and Clostridium perfringens populations, respectively 7.65 CFU/g and 2.59 CFU/g, were greater for NC, and the lactobacillus population (7.54 CFU/g) for the same group was less (p < 0.05), compared to the other groups. Coliforms and Clostridium perfringens decreased as green tea level increased. The least value in the GTE group was 2.22 CFU/g while 1.8 CFU/g was the least value in GTP group. Green tea increased lactobacilli (Table 8), thus increasing useful intestinal microflora, and inhibiting growth of enteropathogenic strains of Coliforms and Clostridium perfringens. The better nutrient utilisation and improved growth and development of chickens, particularly in GTE0.4, may be a direct result of this. The stimulation of useful and inhibition of pathogenic microflora leads to improved absorption and utilisation of nutrients. Thus, green tea can inhibit and kill a wide range of pathogenic bacteria (Hamilton‐Miller, 1997).

5. CONCLUSIONS

We found benefits of including green tea in the diet of broilers from increased weight gain, improved feed conversion, reduced LDL and VLDL cholesterol, and reduced pathogenic bacteria in the cecum. At least some of these beneficial effects of green tea may stem from direct effects on intestinal mucosa morphology, since we observed that it supported the growth of villi and goblet cells and reduced the thickness of the epithelium. The results also confirm the beneficial effects of green tea as a feed additive, as it increased production performance and enhanced health status of the birds. The most beneficial effects were observed with green tea included in the diet at a concentration of 0.04 g/kg.

AUTHOR CONTRIBUTIONS

Conceptualisation: K.J., B.R., A.S. Methodology: K.J., B.R., A.S. Formal analysis: K.J., B.R., I.T.K., A.S. Investigation: K.J., B.R., I.T.K., M.S., I.G., A.S., C.J.C.P. Resources: K.J., B.R., I.T.K., M.S., I.G. Data curation: K.J., B.R., I.T.K., M.S., I.G. Writing—original draft preparation: I.T.K., M.S., I.G., A.S. Writing—review and editing: C.J.C.P. Supervision: B.R., A.S. Project administration: K.J., M.S., I.G. Funding acquisition: K.J., M.S., I.G., C.J.C.P.

AUTHOR DECLARATIONS

All authors are either employed by, or associated with, a government agency or university, whose primary function is research and education

ANIMAL ETHICAL STATEMENT

All procedures involving animals were approved by the Animal Ethics Committee of the Islamic Azad University (Approval number 11750103931001) and conducted in according to the EU's International Guidelines for Research involving Animals (Directive 2010/63/EU)

CONFLICT OF INTEREST

The authors declare that there are no conflicts of interest.

PEER REVIEW

The peer review history for this article is available at https://publons.com/publon/10.1002/vms3.923.

ACKNOWLEDGEMENTS

The authors are grateful to staff at the commercial poultry farm at Amirabad, Lahidjan, Iran, and to Rasht Branch, Islamic Azad University, Rasht, Iran (grant number 17.16.3.16303). The research study proposed was conducted under the grant of the RSF No. 22‐16‐00041, 21‐16‐00025, GNU NIIMMP.

Jelveh, K. , Rasouli, B. , Kadim, I. T. , Slozhenkina, M. I. , Gorlov, I. F. , Seidavi, A. , & Phillips, C. J. C. (2022). The effects of green tea in the diet of broilers challenged with coccidiosis on their performance, carcass characteristics, intestinal mucosal morphology, blood constituents and ceca microflora. Veterinary Medicine and Science, 8, 2511–2520. 10.1002/vms3.923

DATA AVAILABILITY STATEMENT

The data that support the findings of this study are available from the corresponding author upon reasonable request.

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Associated Data

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Data Availability Statement

The data that support the findings of this study are available from the corresponding author upon reasonable request.

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