Effects of ginger (Zingiber officinale) and garlic (Allium sativum) essential oils on growth performance and gut microbial population of broiler chickens

F E Dieumou, A Teguia, J R Kuiate*, J D Tamokou*, N B Fonge and M C Dongmo

Department of Animal Production, Faculty of Agronomy and Agricultural Sciences, University of Dschang, P O Box 222, Dschang-Cameroon
feldieu@yahoo.com   ,   alexisteguia@justice.com   ,   brikatte@hotmail.com   and   mcfrdongmo@yahoo.fr
Laboratory of Microbiology and Antimicrobial Substances, Faculty of Sciences, University of Dschang, P O Box 67, Dschang-Cameroon
jrkuiate@yahoo.com   ,   jtamokou@yahoo.fr

Abstract

An experiment was conducted at the University of  Dschang poultry research farm to evaluate the effect of ginger and garlic essential oils on some blood parameters, growth performance and gut microbial population of broiler chickens. Forty two male and female day old chicks of Arbor acres line were arranged in a fractional factorial experiment of an unbalanced completely randomised design and allotted to three treatments given by stomach tube except for the control in three doses 0 (Control),10mg/kg/day, 20mg/kg/day,and 40mg/kg/day. All diets were iso-nitrogenous, containing 22% and 19% crude protein in the starter and the finisher periods, respectively. The trial lasted for seven weeks and there were no differences in feed intake, body weight gain and the feed conversion ratio among the birds.

 
All organ weights and carcass characteristics were not affected by the treatments, except for a decrease (P< 0.05) in relative liver weight of birds on garlic oil treatment compared with those given ginger oil and control. Similarly, a lower (P< 0.001) proportion of the head weight of birds given essential oils was observed compared to the control. Dosages effects showed a decrease in relative weight of organs only for the head  (P<0.001) and the gizzard (P<0.05) compared to the control. Male broilers deposited less (P<0.001) than the females. There were no significant differences observed in the activities of the serum transaminases (AST & ALT) and blood creatinine level, indicating that none of the three dosages of essential oils given to birds was toxic. However, Escherichia coli, and other Enterobacteria counts in the ileo-cæcal digesta numerically decreased (P<0.05) compared to the control as the doses of essential oils given increased. The same observation was made for the Salmonella and Shigella species  (P< 0.001).The colony forming units (CFU) of Staphylococci spp were statistically similar between the two oil-treated groups, but were significantly (P< 0.01) reduced compared with the control group. Yeast and mold fungi were found in the ileo-cæcal digesta of all the groups.

Key words: broiler chickens, essential oils, growth performance, microbial population

Influence des huiles essentielles du gingembre (Zingiber officinale) et d’ail (Allium sativum)  sur les performances de croissance et la flore intestinale des poulets de chair

Résumé

Un essai a été réalisé à la Ferme d’Application et de Recherche de l’Université de Dschang pour déterminer l’influence des huiles essentielles du gingembre et d’ail sur quelques parametres sanguins, performances de croissance et la flore intestinale des poulets de chair. Quarante deux poussins d’un jour de deux sexes de la souche Arbor acres étaient arrangés dans un schéma factoriel fractionnel du dispositif expérimental complètement aléatoire. Ils étaient soumis à trois traitements administrés chacun par gavage à l’exception du témoin en trois doses, (0) témoin ,10mg/kg/jour, 20mg/kg/jour et 40mg/ kg/jour. Tous les aliments distribués aux poulets étaient iso protéiniques avec 22 %  et 19 % au démarrage et à la finition respectivement. L’essai a duré sept semaines.

L’analyse des resultats obtenus indique qu’il n’y a pas eu de différence significative pour l’ingéré alimentaire, le gain de poids corporel,et l’indice de consommation entre les différents groupes de poulets. A l’exception d’une baisse (P<0.05) de poids relatif du foie au poids vif des poulets soumis à l’huile essentielle d’ail par rapport à ceux sous traitement d’huile de gingembre et control, puis d’une faible (P<0.001) proportion de la tête de poulets sous traitement aux huiles essentielles par rapport au témoin,tous les organes et les caractéristiques de la carcasse n’étaient pas affectés par les traitements. Les doses d’huiles essentielles administrées ont réduit seulement la proportion de la tête (P<0.001) et du gésier (P<0.05) des poulets comparativement au lot témoin. Les poulets mâles ont déposé moins (P<0.05) de graisse que les femelles. Il n’y a pas eu de différence significative entre les traitements pour l’activité des transaminases sériques (AST & ALT) ainsi que pour la teneur en créatinine sanguine indiquant qu’aucune des trois doses d’huile administrées aux poulets était toxique. Toutefois, le nombre d’Escherichia coli, et d’autres entérobactéries dans le digesta iléo-cæcal diminuaient (P< 0.05) par rapport au control au fur et à mesure que les doses d’huiles essentielles administrées aux poulets augmentaient. La même observation était faite pour Salmonella et Shigella spp (P<0.001). Le nombre d’unités formant une colonie de Staphylococci spp était statistiquement semblable pour les deux groupes de poulets soumis aux huiles essentielles mais décroissait (P<0.01) comparativement au témoin. Les  levures et moisissures étaient également présents dans le digesta iléo-cæcal de tous les groupes.

Mots clés: flore intestinale, huiles essentielles, performances de croissance, poulets de chair

Introduction

The fast growing nature of broilers and their short generation interval has been associated  over the years with the use of antibiotic growth promoters used at sub-therapeutic doses in animal feeds in order to improve the quality of the product, with a lower percentage of fat containing a relatively high concentration of polyunsaturated fatty acids and a higher protein content of the meat ( Kinsella et al 1990; Nettleton 1991). Other benefits of these compounds include control of zoonotic pathogens such as Salmonella,Campylobacter, Escherichia coli and Enterococci species in the gut (NOAH  2001).Although birds raised with these feed additives achieved good performance, their potential side effects became a real public health problem worldwide (Bager 1998; Donoghue  2003) and led to the ban of  these products by the European Union in January 2006. This decision has therefore stimulated a search for alternatives. Essential oils have been proven to control pathogens due to their antimicrobial activity (Dorman and Deans 2000), to have antioxidative potential (Hui 1996) by delaying lipid oxidation in broiler meat, and to enhance digestion (Brugalli 2003) by stimulating the endogenous enzymes. It is in this respect that essential oils from garlic (Allium sativum) and ginger (Zingiber officinale) supplements were evaluated as natural alternatives to feed antibiotics in broiler diets. Thus, the objective of this study was to investigate the effects of different dosages of essential oils extracted from these two plants on some blood parameters,  intestinal microflora population and  growth performance of broiler chickens.

Materials and methods

Experimental birds and management

Forty two day old Arbor Acres chicks, with equal numbers of males and females were kept in a brooding house for 21 days under a decreasing room temperature regime, that was reduced from 32°C during the first week of life to 26°C at three weeks. The chicks were given an anti-stress agent from the first to the third day of age and before, during and after the vaccinations. In order to boost their immunity they were vaccinated against Newcastle disease and infectious bronchitis on the seventh and 25^th days of age, while Gumboro vaccine was administered on the tenth day of the experiment. Birds received feed and water ad libitum every day. The experiment lasted for seven weeks during which feed intake, weekly weight gain and feed conversion ratio were monitored.

The entire flock was affected by coccidiosis from the fourth week of the experiment and was therefore subject to four (04) days of cure with Amprolium coupled with anti-stress administration from the 29^th day.

Feed preparation and feeding

The ingredient and calculated chemical composition of the basal diets are shown in Table1 for the starter and finisher growth periods respectively. The diets were formulated to meet the nutrient requirements of growing broilers.

Essential oil extraction and chemical analyses

Ginger roots were harvested from the agricultural zone of Santchou in the Menoua division, in the West Province of Cameroon, while the garlic used came from the northern part of the country. The plant materials were ground in a mortar and pestle in order to liberate the tissues. The plant tissues were mixed with water and were then packed into the cooking unit of a Clevenger-type hydrodistillator. The essential oils collected were then dehydrated using sodium sulphate anhydride before storage at 4°C. Unlike ginger oil, the steam extraction  (Oetting  2005) of garlic oil produces two fractions: a hydro-soluble whitish fraction and a hydrophobic yellowish fraction. Essential oils in the hydro-soluble fraction was partitioned with hexane using a separating funnel. The upper hexanic phase was then evaporated to dryness at 40°C under reduced pressure using a rotary evaporator. The dried hydro-soluble garlic oil obtained was mixed with its corresponding hydrophobic fraction and stored at +4°C until use. One kg of garlic yielded 1.9ml of oil weighing 2.0g (0.2%), while 1kg of ginger produced 2.0ml of essential  oil weighing 1.7g (0.17%) after 6 hours of  distillation.

The essential oils of Zingiber officinale and Allium sativum were subjected to gas chromatography–mass spectra (GC/MS) analysis on an argilent apparatus consisting of a model 6890N Network GC system/ 5975 InertXL mass selective detector at 70 eV and 20°C. The capillary column was a CP-Sil 8 CB LB with a 30 m long fused silica, an interior diameter of 0.25mm and a film thickness of 0.25 µm. The carrier gas was helium at a flow rate of 1.2ml/min.The oven temperature was programmed from 50 to 300°C with an initial increase of 10°C/ 1mn .One microlitre of each essential oil sample prepared in acetone (1% concentration) was injected into the apparatus. Identification of oil components was based on their retention indices, which were determined with references to a series of normal alkanes, and by comparison of their mass spectral fragmentation pattern with Nist 89 database and Wiley237 Library (Adams 2001).

Microbial counts in the intestinal content

Fecal samples collected from the ileo-cecum of the forty eviscerated birds on the farm were put into sterilised vials and conveyed immediately to the laboratory. Then 1g of digesta taken from each sample was added to 9 ml cooked meat broth and mixed for 1minute. A tenfold serial dilution  was made by transferring 1ml from each mixture to the same broth volumes. Finally 1ml was pipetted from the 1/1000 dilution test tube of each sample and inoculated on the solid culture medium prepared in Petri dishes the previous day. Dispersion was done using a sterile spreader sterilised after each step over a bunsen flame.

Yeast and mold fungi were cultured on Sabouraud agar medium mixed with 250mg chloramphenicol in order to inhibit any bacterial growth and were incubated at room temperature up to two weeks for identification. Bacterial counts were performed using Salmonella/Shigella agar medium for Salmonella and Shigella  species, MacConkey agar for Escherichia coli and other enterobacters, then Chapman Stone medium for Staphylococci spp respectively after aerobic incubation for 24 hours at room temperature.

Determination of the trans aminase (AST and ALT) activities and creatinine level  in the serum

Blood samples were collected from the jugular vein of birds stunned before slaughter using non-heparinized tubes and  immediately subjected to  centrifugation at 2000 revolutions per minute for 15 minutes to obtain blood serum. Alanine amino transferase and Aspartate amino transferase activities and creatinine level in samples of blood serum obtained were determined  using commercial kits ( Biosystem Reagents and Instruments).

Experimental design, data collection and statistical analysis

The gavages of essential oil to birds started from the fourth day right to the end of the experiment. The forty two chicks  were given three treatments (control, ginger and garlic) having three dosages each (10ppm, 20ppm, 40ppm) except for the control and consisting of six ,eighteen and eighteen birds respectively. They were arranged in a fractional factorial experiment of an unbalanced completely randomised design. Each dosage consisted of three replicates with a male and a female chick per replicate in deep litter pens for the starter period and in battery cages during the finisher period. The daily dosages of essential oil given to birds by oral intubation in proportion to their mean weight gain are shown in Table 2.    

The quantity of oil administered was calculated taking into account the proportion of the major oil components and the chicken live weights.

Significance of treatment effects and the interactions were assessed by the General Linear Model (GLM) and Compare Means options of Statistical Package for Social Sciences software (version 11.0) as described by Steel and Torrie (1980) and differences in treatments means were compared as per the standard method of Duncan (1955).
 

Results and discussion 

The results of gas chromatography coupled with mass spectrometry (GM/MS) analyses of the oils are summarised in Table 3 and 4.

The main constituents identified in Allium sativum  essential oil were trisulfide-di-2-propenyl, disulfide, di-2-propenyl and trisulfide-methyl-2-propenyl. In the ginger oil, 26 constituents were identified, with zingiberene, sabinene, camphene, geranial, z-citral and 1,8-cineole as major components.

These results agree with the findings of Suad Khamis et al (2004) in terms of the major constituents in garlic and ginger essential oils but are respectively higher and lower with regards to the highest percentage content of major components.

Performance of broiler chickens

The overall growth performance of broilers  during the experiment is shown in Table 5.

There were no significant differences among treatments for daily feed intake (DFI), daily weight gain (DWG) and feed conversion ratio (FCR), or in the interaction between treatments and dosages, indicating that the two factors acted independently on the growth performance. Similarly, all offals and organs expressed as a proportion of body weight and the carcass characteristics of birds were not affected by treatment and dosage (Table 6).

These results agree with the findings of Jayalakshmi et al (2006), who studied the influence of sunflower acid oil usage on production performance, carcass traits and economics of broilers, and reported no significant differences in body weight gain, cumulative feed consumption, cumulative feed conversion ratio and liveability between treatment groups from the first week till the end of experiment. Similarly, Botsoglou  et al (2002 ) observed that supplementation of broiler feed with oregano essential oil for thirty eight days had no growth-promoting effects. Treatments effects of essential oils were only significant for the relative heart and head weights of broilers (Table 7).

The results indicate a lower (P<0.05) relative heart weight  of broiler chickens on the garlic essential oil treatments compared with those in the ginger oil treatment and control. Similarly, a decrease (P<0.001) in the proportion of the head was observed in birds given in the essential oils treatments compared with the control.

These findings agree with the reports of Tubtim and Wasiksiri (2007), who subjected male and female Sprague-Dawley rats to a 28-day repeated dose oral toxicity study with Litsea cubeba essential oil and obtained changes in relative liver and kidney weights in certain treated groups compared with the control.

Dosage effects also exclusively affected the head weights (Table 8).

The relative head weights of birds given different dosages of essential oils were statistically similar but significantly (P<0.001) lower compared to the controls. These results are consistent with those observed by Barreto et al (2008), who assessed the efficacy of different plant extracts as alternatives to antimicrobial growth promoters in broiler diets and reported no effects of experimental treatments on organs morphometrics, except for a lower liver relative weight (P<0.05) of birds fed the diet containing red pepper extract, which showed the lowest change in relative liver weight compared to those on the control diet.

The sex effects of essential oils on the parameters studied are shown in Table 9.

Enzymes activities in the serum, creatinine level in the blood and microflora enumeration

The details of treatments and dosages effects of essential oils on enzymes activities in the serum, creatinine level and on the gut microbial load are shown in Tables 7 and 8. The effect of essential oil treatments on the aspartate amino transferase activitiy, alanine amino transferase activity, and creatinine  level in the blood did not differ from the control. Similarly, Tubtim and Wasiksiri (2007) administered  different doses of Litsea cubeba essential oil to male and female Sprague-Dawley rats by stomach tube for twenty eight days and obtained no significant difference from the control group for these same blood parameters.

There was a significant decrease (P<0.001) in the number of colony forming units (CFU) of Escherichia coli and other enterobacteria in digesta of ileo-cecum, compared with the control as the dosage of essential oils used increased. The same observation was made for the Salmonella and Shigella species (P<0.05).The counts of Staphylococci spp did not differ between the oil treated  groups of birds but were significantly (P<0.001) reduced as compared to the control. These findings agree with the reports of Jang et al (2007), who obtained a significant decrease (P<0.05) in the number of Escherichia coli in ileo-cecal digesta of birds fed diets containing antibiotics when compared to those on a control treatment after feeding ROSS 308 broilers for five weeks with a basal diet and diets supplemented with a blend of commercial essential oils or antibiotics.

Similarly, Hinton and Linton (1988), Izat et al (1990), Oliveira (1996) and Byrd et al (2001) demonstrated that the addition of lactic acid, formic acid and propionic acid to diets and drinking water efficiently controlled Salmonella and Shigella species, Escherichia coli and other Enterobacteria in poultry. Yeast and mold fungi were also found in the ileo-cecal digesta of all the groups.

Female birds put on more (P<0.001) abdominal fat and had more zoonotic pathogens in their gut than the males (Pym and Solvyns 1979; Broadbent et al 1981; Ross Breeders 1996; Rondelli et al 2003). However, the carcasses and the legs of male broilers weighed more than those of females. There were no significant differences in the activities of  blood parameters and organs such as the gizzard, pancreas, liver and the heart between male and female birds.

These results agree with the reports of Souza et al (1995), who evaluated the carcass traits of four commercial broiler lines, Arbor Acres, Hubbard, Cobb and Ross, under the same nutritional management and found higher leg and thigh yield in Hubbard males and higher breast yield in the Ross line.           

Conclusions

• The forty five consecutive days of ginger and garlic oils administration to broilers in this experiment did not exert any growth-promoting effects.

  
  

• The treatment effects of these essential oils had a positive impact on the gut microflora as they reduced their loads as compared to the control, contributing to some extent to health maintenance.

  
  

• However, the essential oil-treated groups could not draw much growth  benefits from the decrease in counts of zoonotic pathogens present in their intestine, because unlike rabbits and ruminants, they have very little nutritional advantage from this situation.

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Received 15 April 2009; Accepted 5 May 2009; Published 5 August 2009

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