Effect of Probiotic on Gut Development of Domestic FowlsKh. Amber, M. H. El-Deep, and M. A. M. Sayed
Animal Prod. Inst. Agric. Res..Center , Ministry of Agric, Egypt. Poultry Production Dept, Faculty of Agric, Kafr El-Sheikh Univ, Egypt*.
This experiment was conducted to study the effects of effective microorganisms (EM) and Zinc bacitracin on gut development, digestibility coefficient of nutrients and intestinal histology of Inshas chicken (a local Egyptian chicken strain) . Five hundredand fourty chicks were randomly assigned to 1 to 6 dietary treatments for 41 wk. The dietary treatments were 1) control; 2) Basal diet + EM (2.5 ml/kg diet ); T3) Basal diet + EM (5.0 ml/kg diet ); T4) Basal diet + EM (7.5 ml/kg diet ); T5) Basal diet + EM (10.0 ml/kg diet ) and T6) Basal diet + Zinc bactracin (500 mg/kg). The obtained results showed that, villi height , villi thickness and villi surface area were significantly increased in birds fed EM with different levels and Zinc bactracin diets . The data on the digestibility coefficient of nutrients revealed that, all nutrients of EM diets were more efficiently digested than that of Zinc bactracin diet (p≤0.01) . While, digestibility coefficient of OM, DM, CP, EE, CF and NFE was significantly increased as compared with chicks kept on the control diet. Moreover, It was generally noticed that intestinal histology was almost following the same trend observed with gut development and digestibility coefficient .
The impact of biotechnology in poultry nutrition is of significant importance. Biotechnology plays a vital role in the poultry feed industry. Nutritionists are continually putting their efforts into producing better and more economical feed. Good feed alone will not serve the purpose but its better utilization is also essential. Dietary changes as well as lack of a healthy diet can influence the balance of the microflora in the gut thus predisposing to digestion upsets. A well-balanced ration sufficient in energy and nutrients is also of great importance in maintaining a healthy gut. A great deal of attention has recently been received from nutritionists and veterinary experts for proper utilization of nutrients and the use of probiotics for growth promotion of poultry.
In poultry nutrition, probiotic species belonging to Lactobacillus, Streptococcus, Bacillus, Bifidobacterium, Enterococcus, Aspergillus, Candida, and Saccharomyces have a beneficial effect on poultry performance , modulation of intestinal microflora and pathogen inhibition , intestinal histological changes , immunomodulation , certain haematobiochemical parameters , improving sensory characteristics of dressed broiler meat  and promoting microbiological meat quality of broilers.
Effective Microorganisms (EM) is a microbial preparation developed by Professor T. Higa of University Of The Ryukyus in Japan. The EM is composed of different microbes that include bacteria, yeasts and/or fungi. Some of the benefits claimed to accrue from the use of EM include improved meat and manure quality, improved animal health, reduction of foul smells and absence of toxic effects on bird growth . Use of EM in Africa is a new innovation and novel idea. There is no available literature regarding use of microbial preparations in broiler production.
Therefore, this experiments was designed to investigate the possibility of using probiotic namely, (EM) effective microorganism ( instead of using antibiotics) to Inshas chickens (Egyptian local strain ) , and to evaluate its effects on gut development .
A total number of 540 unsexed vaccinated Inshas (local Egyptian chicken strain) one day-old-chicks were weighed , wing banded and randomly divided into six experimental groups ( three replicates each group ) . The birds were placed in a room (floor pens) maintained at a constant temperature of 28+/-3 oC and a relative humidity of 70+/-3% .Food and water were always available ad libtum. The basal diet was formulated to meet the nutrient needs suggested by the NRC, 1994. For enteric morphometric analysis, birds on the designated evaluation day were euthanized, and small intestines were collected. A 1-cm segment of the midpoint of the lower ileum were removed and fixed in 10% buffered formalin for 72 h. Each segment was then embedded in paraffin, and a 2μm section of each sample was placed on a glass slide and stained with hematoxylin and eosin for examination with a light microscope . The parameters evaluated were villus height, villus and thickness, villus surface area. Morphological parameters were measured using the Image Pro Plus v 4.5 software package. Fourteen measurements were taken per bird per parameter. Villus height was measured from the top of the villus to the top of the lamina propria. Villus surface area was calculated using the formula (2π)(VW/2)(VL), where VW = villus width, and VL = villus length , and we used anther way villus surface area index = perimeter length × mucosal thickness .The experimental design consisted of six dietary treatments as follows; (T1) Basal diet ( control ), (T2) Basal diet + EM (2.5 ml/kg diet ), (T3) Basal diet + EM (5.0 ml/kg diet ), (T4) Basal diet + EM
MATERIAL AND METHODS
(7.5 ml/kg diet ), (T5) Basal diet + EM (10.0 ml/kg diet ) Multiple Range Test . Statements of statistical and (T6) Basal diet + Zinc bactracin (500 mg/kg).. The significance are based on P<0.05. results obtained were statistically analyzed using Duncan’s
RESULTS AND DISCUSSION
Morphometric Analysis of the Gut
Results obtained showed that villi height was significantly ( p≤0.05) increased in birds fed EM with different levels and Zinc bactracin diets in Fig (1 ). The villi length was longer (P < 0.05) by about 12.6 to 15.2% for birds fed EM and Zinc bacitracin diet as compared with the control. There are no significant differences among chicks fed diets with different levels of EM.
Statistical analysis of the results obtained proved that EM and Zinc bacitracin diets had a significant effect on the Villi thickness Fig (1 ). Villi thickness was significantly increased in birds fed EM and Zinc bacitracin diets as compared with control diet ( as average 3.8 vs 1.6 μm; P < 0.001).
Villi Surface Area
Villi surface area as influenced by dietary EM with different levels and Zinc bactracin during the experiment of period is presented in Fig (1 ). Villi surface area was increased (P < 0.001) from 17.11 to 71.3 μm as the level of EM increased from 0 to 10 ml/kg in bird diets. Also, increased (P < 0.001) by 101.3 % in chicks fed T6 diet as compared with those fed control diet . These results agree with . Upon consumption, probiotics deliver many lactic acid bacteria into the gastrointestinal tract. These microorganisms have been reputed to modify the intestinal milieu and to deliver enzymes and other beneficial substances into the intestines .
It is well established that probiotics alter gastrointestinal pH and flora to favor an increased activity of intestinal enzymes and digestibility of nutrients . Mechanisms by which probiotics improve feed conversion
efficiency include alteration in intestinal flora, enhancement of growth of nonpathogenic facultative anaerobic and gram positive bacteria forming lactic acid and hydrogen peroxide, suppression of growth of intestinal pathogens, and enhancement of digestion and utilization of nutrients .
Illuminating work from Gordon’s laboratory provides evidence that manipulating the microbiota with probiotics can influence the host. Germ-free mice were colonized with Bifidobacteria thetaiotaomicron, a prominent component of the adult human gut microbiota, and Bifidobacterium longum,a commonly used probiotic. B. longum repressed B thetaiotaomicron expression of antibacterial proteins that may promote its own survival in the gut, as well as influence the composition, structure, and function of its microbial community.
Data on digestibility coefficient of nutrients as shown in Table ( 1) revealed that all nutrients of EM diets were digested more (p≤0.01) efficiently than of Zinc bactracin diet. While, digestibility coefficient of OM, DM, CP, EE, CF and NFE were significantly increased by 3.7, 3.8, 3.9, 9.6, 4.1 and 4.4 %, respectively, in chicks fed T6 diet. Also, increased (p≤0.01) by 7.3, 7.5, 5.4, 9.0, 53.9 and 7.0 %, respectively, in those fed EM diets ( T2, T3,T4 and T5) as compared with chicks kept on control diet.
 reported that supplementing broiler diets with probiotics tended to increase the digestibility of CP in both fresh maize and 10% moldy maize diets. Also,  showed that, supplementing broiler chick diets with growth promoter significantly improved digestion coefficient of nutrients except CF compared to unsupplemented diet. The increased number of beneficial microbes was confirmed and explained by  who found that the number of anaerobic bacteria and cellulytic bacteria was increased, when the diet was supplemented with yeast, due to enhancing Lactate utilization and moderating pH of the media, therefore, yeast improved the nutrients digestibility coefficients.
In this connection,  reported a positive effect of probiotics on apparent protein digestion and attributed this effect to the proteolytic activity of bacteria. It is worthy to note the absence of significant differences in the data as a result of the combination effect of dietary CP level and tested probiotics. Such observation confirmed the previously mentioned opinion that the tested probiotics had a sparing effect of nearly 2.0 % CP. Similarly, the better (P>0.05) digestibility obtained with probiotics supplementation suggests that such addition improved feed and nutrients utilization, which in turn explain the better growth and FCR values obtained with the probiotics supplemented diets. In general, the improvement (P>0.05) due to adding the probiotics may be attributed to improving intestinal microbial balance. In other words, probiotics help to keep the intestinal tract healthy and when the epithelial tissue is healthy, there is improved and better absorption of all nutrients .
Improvement of nutrient digestibility by supplementing chick diets with either microbial probiotics could be attributed to different stimulators such as change in the enteric flora and reduction of E. coli population, lowering gastric pH, synthesis of catabolic enzymes that help in releasing cell compounds including amino acids, sugar and fatty, acids into the intestinal environment and involving of active bacteria with the digestive processes, protein synthesis and nutrient absorption in gastrointestinal tract 
Fig (1): Morphometric Analysis of the Gut as affected by different levels of EM and Zinc bacitracin.
Table (1): Digestibility coefficient of chicks as affected by the different nutrients of the experimental treatment (Means ± SE).
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