Use of Organic Acids and Competitive Exclusion Product As an Alternative to Antibiotic As a Growth Promoter in the Raising of Commercial Turkeys

PRODUCTION, MODELING, AND EDUCATION

Use of organic acids and competitive exclusion product as an alternative to antibiotic as a growth promoter in the raising of commercial turkeys

E. L. Milbradt ,* 1 A. S. Okamoto ,* J. C. Z. Rodrigues ,* E. A. Garcia ,† C. Sanfelice ,† L. P. Centenaro ,† and R. L. Andreatti Filho *

* Department of Veterinary Clinics, and † Department of Animal Production, School of Veterinary Medicine and Animal Husbandry (FMVZ), São Paulo State University (UNESP), Botucatu, São Paulo, Brazil 18618-970

ABSTRACT A study was conducted to investigate the In the fattening phase (d 28–70), the feed intake of the effects of organic acids (OA) and competitive exclu- OA-treated group was lower than compared with the sion product (CE) on growth performance, intestinal control. The birds that received diet with OA and CE morphology, and concentration of volatile fatty acids product presented higher concentrations of propionic in the cecal content. The experiment lasted for 10 wk. acid, at 14 d, and butyric acid in cecal content at 28, Four hundred twenty 1-d-old female commercial cross 56, and 70 d, compared with the control. Dietary in- turkey poults (British United Turkeys, BUT Big 9) clusion of additives had no significant effects on intes- were distributed into 4 treatments with 5 replicates/ tinal villus height, crypt depth, and villus:crypt ratio. pen of 21 birds each. The birds were fed a basal diet Organic acids had negative effects either on early gain without growth promoter (control), diet with lincomy- or feed intake throughout the study. Because the test cin (44 mg/kg), diet with organic acids (2 g/kg), and was conducted under controlled experimental condi- diet with product of CE (109 cfu/kg). Dietary levels tions, the additives that showed results similar to those of other nutrients, housing, and general management found by using antibiotics should be studied further in practices were similar for all treatments. On the first commercial farms to obtain results that can be incor- week (d 0–7), the BW and BW gain of the birds that porated into practice. fed diets with OA were lower than in the control group. Key words: antimicrobial drug, organic acid, performance , turkey 2014 Poultry Science 93 :1855–1861 http://dx.doi.org/10.3382/ps.2013-03593

INTRODUCTION pacity to inhibit the inflammatory response of the host and that the modification of the microbiota is due to The supplementation of an animal diet with antibiot- the action of the immune system and not to the direct ics with the aim of controlling pathogenic agents during action of antibiotic growth promoter because these are the digestive process and promoting improvement in added to the ration at levels lower than the minimum animal-husbandry indices, maximizing production, has inhibitory concentration for pathogens. been a common practice in the poultry industry (Gas- Independent of the action mechanism, it is known kins et al., 2002). that the vast majority of the drugs employed for eco- Some authors report that the action mechanism of nomic and disease prevention uses in poultry are of lim- antibiotics as growth promoters is related to interac- ited or no importance in human medicine. In addition, tions with intestinal microbial population and that the removal of antibiotic growth promoters from ani- this practice can lead to development of resistance by mal feed can increase production costs due to losses in pathogenic bacteria, which can have an impact on pub- animal-husbandry performance and elevated incidence lic health (Dibner and Richards, 2005). However, ac- of enteric diseases, such as the necrotic enteritis caused cording to Niewold (2007), the action mechanism of by Clostridium perfringens (Smith, 2011). antibiotics as growth promoters is linked to their ca- Strategies to reduce or eliminate the use of antibiotics in the poultry chain include improvement of biosafe- ty programs, use of vaccines, genetic selection, qual- ity control of the raw material used in the ration, and © 2014 Poultry Science Association Inc. the use of other additives such as organic acids (OA) Received September 2, 2013. Accepted March 17, 2014. and products of competitive exclusion (CE; Sun et al., 1 Corresponding author: [email protected] 2005). Like antibiotics, the OA can also control the in-

1855 1856 Milbradt et al. testinal microbiota because they can exert bactericidal the NRC (1994; Table 1). The amino acids in the diets and bacteriostatic action, depending on the physiologi- were formulated on a total amino acid basis and were cal status of the organism and on the physical-chemi- kept similar among the diets. Biosecurity procedures cal characteristics of the external environment (Ricke, were maintained between treatment groups. The pens 2003). were separated laterally to prevent cross-contamination According to Schneitz (2005), the most efficacious of feed or water. Separate boots and laboratory coats and inoffensive method for controlling avian microbiota were used for each treatment, and bird care was sched- is by means of CE because it consists of a biological uled such that birds without CE product were serviced treatment, which does not leave residue. Furthermore, first followed by those on CE product diet. studies demonstrate that CE products can positively The experimental protocol was approved by the Com- affect the performance of birds (Goren et al., 1984; mission for Ethics in the Use of Animals (n° 177/2012 Schneitz et al., 1998). CEUA) of the Universidade Estadual Paulista “Júlio de The majority of research studies related to the use Mesquita Filho,” Campus of Botucatu. OA and CE products have been conducted on broiler chickens, often with inconsistent results (Gunal et al., 2006; Hernández et al., 2006; Isabel and Santos, 2009; Experimental Diets and Additives Houshmand et al., 2011). The present study aimed to The experimental treatments received a corn-soybean evaluate the effect of continuous supplementation of meal basal diet and were as follows: (control) basal diet OA and CE product in the diet on growth performance, without additive supplementation; diet with lincomycin height of intestinal villi, crypt depth, villus:crypt ratios, (44 mg/kg of feed); diet with OA (2 g/kg of feed); and and the concentration of volatile fatty acids (VFA ) in diet with CE product (109 cfu/kg of feed). the cecal content of commercial turkeys, as an alterna- Feed additives used in the experimental diets were tive to the use of antibiotics as a growth promoter. lincomycin 44% as the antibiotic growth promoter, in- cluded at 44 mg of active principle per kg of feed. MATERIALS AND METHODS Organic acids: commercial product in powdered form, containing a mixture of short- and medium-chain fatty Birds and Experimental Treatments acids, with the following composition: acetic acid, formic acid, propionic acid, sorbic acid, and vegetal fatty Four hundred twenty 1-d-old female commercial cross acids. The guaranteed levels are acetic acid (minimum turkey poults (British United Turkeys, BUT Big 9) 37.4 g/kg), formic acid (minimum 73.6 g/kg), and veg- were obtained from a commercial hatchery. The poults etal fatty acids (minimum 377.1 g/kg). The inclusion were vaccinated, in a hatchery, against Newcastle dis- was 2 g of product per kg of feed. ease and infectious rhinotracheitis. The procedures of The CE product was a commercial product in liquid housing, feed management and daily care were similar form, composed of nonspecific intestinal microbiota of for all the birds, as recommended by British United specific-pathogen-free turkeys, at least 109 cfu/mL. For Turkeys (BUT Big 9, 2012). inclusion in the feed, the CE product was incorporated The experiment was initiated with 420 birds, which by aspersion, after a pelleting process at a dosage of 1 were divided into lots of 21 birds, to obtain 20 ho- mL of the product per kilogram of feed. Feed samples mogenous experimental units (BW). Starting from the (2 per treatment, 8 total/phase) were collected and homogenization, the birds were allocated in 4 experi- sent, labeled but unidentified, to the laboratory for de- mental treatments in a completely randomized man- tection of lactic acid bacteria. ner; each treatment had 105 poults arranged in 5 repli- For the substitution of organic acids into the other cates of 21 birds each. The poults were housed in floor diets, rice husk meal (inert) was used. Starter diet was pens (5 m2) with new wood shaving litter. Poults were offered as 2.0-mm pellets, grower and developer diets initially maintained at 32°C and the temperature was as 6.0-mm pellets, and ﬁnisher diet as 7.0-mm pellets. gradually reduced by 3°C per week to reach a tempera- None of the diets contained coccidiostats. ture of 21°C by the end of wk 4. This temperature was maintained for the duration of the experiment. The following lighting program was adopted: 24 h of light with Experimental Parameters Measured an intensity of 100 lx in the first 24 h, followed by 18 Performance h of light per day until d 14, and 16 h of light per day until the end of the growing period. Birds and feed were weighed by pen at d 7, 28, and Birds were fed corn and soybean meal-based starter 70 to determine average BW, BW gain, feed intake (0 to 3 wk), grower (4 to 6 wk), developer (7 to 9 wk), (FI), and feed conversion ratio (FCR). In addition, and ﬁnisher (9 to 10 wk). The trial lasted for 70 d. Wa- overall BW gain, FI, and FCR were calculated for the ter and feed were supplied ad libitum. The experimental whole duration of the experiment (d 0–70). Mortalities diets were formulated to meet or exceed the minimum were recorded daily, and BW gain, FI, and FCR were nutrient requirements of turkeys of turkeys according to corrected for mortality. ORGANIC ACIDS AND COMPETITIVE EXCLUSION PRODUCT 1857

Table 1. Composition of basal diets

Starter Grower Developer Finisher Item (1 to 21 d) (22 to 35 d) (36 to 62 d) (63 to 70 d) Ingredient (%) Soybean meal 51.30 47.86 42.43 38.67 Corn 35.91 39.77 45.76 49.66 Soybean oil 5.77 5.80 5.66 6.25 Dicalcium phosphate 2.85 2.59 2.17 1.63 Limestone 1.37 1.28 1.33 1.27 Rice bulk meal (inert) 1.00 1.00 1.00 1.00 Sodium bicarbonate 0.60 0.56 0.53 0.50 dl-Methionine 0.39 0.35 0.35 0.33 Mycotoxin adsorbent 0.30 0.30 0.30 0.30 Mineral premix1 0.15 0.15 0.15 0.15 Threonine 0.13 0.11 0.11 0.06 l-Lysine HCl 0.13 0.13 0.13 0.10 Vitamin premix2 0.10 0.10 0.08 0.08 Total 100 100 100 100 Calculated nutrient composition ME (kcal/kg) 3.000 3.050 3.100 3.200 CP (%) 28.00 26.00 24.00 21.00 Calcium (%) 1.40 1.30 1.20 1.05 Available phosphorus (%) 0.75 0.70 0.60 0.50 Lysine (%) 1.82 1.70 1.50 1.30 Methionine (%) 0.73 0.68 0.63 0.57 Methionine + cysteine (%) 1.18 1.11 0.96 0.91 Arginine (%) 1.95 1.82 1.65 1.43 Threonine (%) 1.16 1.09 0.96 0.85 1Mineral enrichment per kilogram of feed: Cu: 15 mg; Fe: 65 mg; Mn: 110 mg; Zn: 100 mg; I: 1.5 mg; Se: 0.3 mg. 2 Vitamin enrichment per kilogram of feed: vitamin A: 11,250 IU, vitamin D3: 4,000 IU; vitamin E: 50 IU; vitamin K3: 4.5 mg; biotin: 0.25 mg; thia- mine: 4.5 mg; riboflavin B2: 8 mg; pyridoxine: 7 mg; vitamin B12: 0.020 mg; niacin: 75 mg; pantothenic acid: 23 mg; folic acid: 2 mg; antioxidant: 15 mg.

Quantification of VFA distal part of the duodenal loop to the yolk sac diverticulum; the ileum, from the diverticulum yolk to the At 14, 28, 42, 56, and 70 d of age, 10 turkeys per ceca; and the ceca, from the insert to its extremities. treatment (i.e., 2 bird/pen) were randomly selected and Fragments (2 cm) were removed from the medial por- euthanized by the intramuscular administration of 2 tion of all above-mentioned segments. The fragments mg/kg of xylazine (pectoralis major), followed by the were immersed in 10% neutral buffered formalin for 48 intravenous administration of 15 mg/kg of sodium thio- h, dehydrated with increasing concentrations of etha- pental. After the unconsciousness of bird was detected, nol, cleared with 2 passages through xylol, and placed 3 mL of potassium chloride (KCl) was administered to into paraffin plastic. induce death. The histological sections (5 µm) were stained with The ceca were removed for the collection of 1 g of hematoxylin and eosin. Measurements of villus height their content, which was diluted in 2 mL of formic acid (villi) and crypt depth were made, and villus:crypt ra- (17%). Cecal digesta VFA concentrations were deter- tios were calculated. The measurements were obtained mined in duplicate in the supernatants of homogenates through the analysis of images of histological sections after centrifugation at 12,000 × g for 10 min at 4°C. made with the aid of a computerized image-capture The supernatant (2 mL) was injected onto an auto- system (Leica, Qwin Lite 3.0, Leica Microsystems, matic sampler (AS3000, Thermo Scientific, Waltham, Wetzlar, Germany). Measurements for villus height MA) coupled with a gas chromatograph (Focus GC, were taken from the tip of the villi to the valley be- Thermo Scientific). The determination of acetic, propi- tween individual villus and crypt depth was determined onic, and butyric fatty acids was carried out according from the base of the crypt to the level of the opening. to the description of Erwin et al. (1961). The results This yielded 15 measurements per intestinal region per were expressed in mM/L. treatment group.

Intestinal Morphology Statistical Analysis From the same bird group cited previously, at 28 The pen served as the experimental unit for perfor- and 70 d, intestinal segments were collected for mea- mance and individual bird for the VFA, villus height, surement of intestinal mucosal villus height and crypt and crypt depth measures. The results for performance depth. The segments were divided according to Dyce et and length of the intestinal villi were submitted to al. (1996) into the duodenum, the portion from the py- ANOVA complemented with Tukey’s test of multiple lorus to the distal duodenal loop; the jejunum, from the comparisons (0.05%). To analyze the concentration of 1858 Milbradt et al. VFA, the nonparametric ANOVA with model of 2 fac- VFA Concentration tors (treatment and age) was used, complemented with Dunn’s test of multiple comparisons (0.05%; Zar, 2010). There was no difference in the concentration of acetic acid among the different experimental groups (Table 3). All the groups presented elevation in the concentra- RESULTS tion of VFA between d 14 and 56. The birds that re- 5 ceived OA (2.83 mM/L) and CE product (2.32 mM/L) The CE product feed samples contained 10 cfu of presented a higher concentration of propionic acid on lactic acid bacteria per kilogram of pelleted feed. No the 14 d of age, when compared with the group that lactic acid bacteria were detected in the feed samples received lincomycin (1.80 mM/L) and the control group to which no CE product was added. (1.26 mM/L). At 28 and 70 d of age, turkeys fed diets containing OA and CE product had a higher concen- Performance tration of butyric acid in the cecum compared with control. Poult BW did not differ between the experimental treatments on d 1 (i.e., experimental start; Table 2). Intestinal Morphology However, at 7 d of age, the diet supplemented with organic acids exerted a negative effect on BW and BW The treatments did not influence the height of intes- gain of the poults. At 28 d of life, the end of the grower tinal villi, crypt depth, and villus:crypt ratios in any phase, no dietary effect was observed on the perfor- phase of life. mance. During the finisher phase (d 28–70), the group that received OA presented lower FI, but the BW, BW DISCUSSION gain, and FCR were similar to those that consumed the basal diet and diet with lincomycin. During the The birds presented excellent performance, obtaining whole experimental period (d 0–70), only the FI was results superior to those recommended by the British influenced by the diets. The birds that received OA United Turkeys (BUT Big 9, 2012). During wk 1 (d presented lower FI, when compared with birds that re- 0–7), the group that received OA presented decreased ceived CE product and lincomycin. These birds also numerically in FI and lower BW in relation to the other presented BW and BW gain numerically inferior to the groups. The lower FI was the only dietary effect on per- birds of the other treatments. formance at the total rearing period (d 0–70).

Table 2. Mean BW, BW gain, feed intake (FI), feed conversion ratio (FCR), and livability of commercial turkeys fed with basal diet and diet with lincomycin, organic acid, and competitive exclusion (CE) product

Treatment1

Item Control Lincomycin Organic acid CE P-value BW (g) Start 62 ± 0.7 61 ± 0.5 62 ± 0.5 61 ± 0.5 0.78 7 d 188 ± 2.7b 183 ± 1.9ab 175 ± 3.2a 181 ± 1.6ab 0.01 28 d 1,268 ± 30.5 1,283 ± 16.3 1,177 ± 49.4 1,273 ± 56.3 0.24 70 d 6,772 ± 171.9 6,908 ± 75.8 6,432 ± 194.8 6,777 ± 111.3 0.16 BW gain (g) 0 to 7 d 127 ± 3.1a 122 ± 1.7ab 113 ± 3.0b 119 ± 1.4ab 0.01 0 to 28 d 1,207 ± 30 1,222 ± 16 1,115 ± 49 1,211 ± 56 0.23 28 to 70 d 5,503 ± 146 5,624 ± 61 5,254 ± 159 5,504 ± 82 0.20 0 to 70 d 6,710 ± 171 6,841 ± 313 6,396 ± 194 6,715 ± 135 0.08 FI (g) 0 to 7 d 150 ± 8.7 149 ± 2.4 134 ± 2.3 141 ± 1.3 0.08 0 to 28 d 1,693 ± 46 1,778 ± 37 1,732 ± 50 1,780 ± 64 0.55 28 to 70 d 10,304 ± 170b 10,492 ± 129b 9,034 ± 72a 10,313 ± 150b 0.001 0 to 70 d 5,998 ± 450ab 6,135 ± 348b 5,383 ± 379a 6,520 ± 361b 0.023 FCR (g of feed/g of BW) 0 to 7 d 1.18 ± 0.06 1.22 ± 0.02 1.18 ± 0.03 1.18 ± 0.01 0.823 0 to 28 d 1.43 ± 0.02 1.45 ± 0.04 1.56 ± 0.05 1.48 ± 0.08 0.239 28 to 70 d 1.88 ± 0.06a 1.87 ± 0.03a 1.71 ± 0.04b 1.89 ± 0.03a 0.039 0 to 70 d 1.64 ± 0.09 1.66 ± 0.05 1.64 ± 0.05 1.67 ± 0.05 0.75 Livability (%) 7 d 99.1 ± 0.31 100 ± 0 98 ± 0.21 99.1 ± 0.29 1 28 d 99.1 ± 0.31 100 ± 0 97.2 ± 0.42 99.1 ± 0.33 1 70 d 98.2 ± 0.28 99.1 ± 0.36 98.1 ± 0.25 97.1 ± 0.11 1 a,bMeans within a row with different superscripts differ (P < 0.05). 1Control: diet without additive growth promoter; lincomycin: diet with lincomycin (44 mg/kg of feed); organic acid: diet with organic acids (2 g/kg of feed); CE: diet with product of CE (109 cfu/kg of feed). ORGANIC ACIDS AND COMPETITIVE EXCLUSION PRODUCT 1859

Table 3. Median, minimum, and maximum concentration values of acetic, propionic, and butyric acids in cecal content

Treatment1 VFA Age (mM/L) (d) Control Lincomycin Organic acid CE Acetic 14 9.6 (4.5; 16.1)a 7.4 (6.4; 12.9)a 7.0 (3.9; 27.7)a 9.4 (5.9; 23.4)a 28 28.9 (24.2; 40.9)bc 25.1 (16.2; 28.4)b 32.3 (29.0; 35.3)c 25.3 (17.1; 31.5)b 42 36.4 (30.9; 41.2)c 34.8 (22.2; 37.7)c 31.5 (29.1; 39.6)c 31.7 (21.6; 35.1)bc 56 36.3 (30.9; 41.2)c 33.4 (28.9; 44.1)c 37.1 (34.8; 44.3)c 35.2 (30.7; 36.2)c 70 22.3 (18.1; 23.6)b 23.9 (18.8; 27.7)b 20.6 (17.1; 25.0)b 24.0 (17.8; 24.8)b Propionic 14 1.8 (1.1; 4.4)a,A 1.3 (0.5; 1.8)a,A 2.8 (0.9; 3.6)a,B 2.3 (0.8; 5.8)a,B 28 11.7 (5.7; 15.5)b 12.0 (6.5; 18.4)b 15.5 (13.5; 19.8)c 13.2 (5.6; 19.2)b 42 16.7 (11.6; 24.4)c 15.9 (11.6; 21.3)c 15.1 (9.3; 21.6)c 14.1 (4.4; 20.8)b 56 14.4 (11.8; 26.7)c 16.1 (11.7; 18.8)c 18.2 (12.9; 26.5)c 12.4 (9.9; 16.4)b 70 10.2 (8.2; 11.6)b 8.8 (5.7; 11.8)b 9.2 (5.6; 10.6)b 8.1 (7.1; 10.0)b Butyric 14 1.4 (0.4; 3.0)a 1.3 (0.4; 2.6)a 1.9 (0.2; 5.7)a 1.8 (0.4; 2.3)a 28 7.1 (3.8; 9.2)b,A 9.5 (4.5; 11.5)b,AB 11.9 (3.6; 17.3)b,B 12.3 (3.9; 19.1)b,B 42 7.9 (5.2; 11.1)bc 7.8 (5.1; 10.6)bc 9.6 (5.6; 16.7)bc 10.3 (6.3; 9.1)b 56 8.7 (8.2; 12.1)b,A 8.6 (7.7; 10.2)b,A 10.6 (6.4; 16.1)b,AB 12.2 (6.8; 15.7)b,B 70 4.9 (4.3; 6.1)c,A 5.1 (3.8; 6.3)c,AB 7.2 (3.5; 9.6)c,B 8.9 (4.2; 10.7)c,B a–c;A,BMedian followed by different lowercase letters in a column and means followed by different uppercase letters in a row differ significantly by Dunn’s test of multiple comparisons (5%). 1Control: diet without additive growth promoter; lincomycin: diet with lincomycin (44 mg/kg of feed); organic acid: diet with organic acids (2 g/kg of feed); CE: diet with product of competitive exclusion (CE; 109 cfu/kg of feed).

Factors such as the inclusion rate and organic acid has caused the birds to develop in an environment of a type may be related to the decrease of FI, and accord- low level of stress and pathogenic challenge. Generally, ing to Cave (1984), diets supplemented with propionic it has been suggested that beneficial effects of most acid (20 g/kg of feed) can cause a reduction in FI, additives are clearer in suboptimal and stressful condi- principally due to its influence on palatability. Little is tions, such as a disease condition, a thermic stress, a known about the utilization of a blend of organic acids high stocking density, and bad management practices in the diet of commercial turkeys as growth promot- (Baurhoo et al., 2007). ers. However, in general, the results obtained in the Volatile fatty acids are produced and used by birds present study are similar to the results obtained from as an energy source, but their nutritional significance is broiler chickens. Hernández et al. (2006) did not obtain limited due to their small quantity, which is less than positive effects on the performance (BW gain, FI, and 0.5% of BW (Annison et al., 1968). According to van FCR) and intestinal histomorphology of broiler chick- Der Wielen et al. (2000), soon after hatching, there ens supplemented with formic acid (5 and 10 g/kg of are no VFA in the cecal content, but the concentra- feed). Houshmand et al. (2011) found no positive ef- tion elevates rapidly until 21 d. In the current study, fects on broiler performance and intestinal villi height we observed that the concentrations of the acids were from the use of a blend of organic acids (formic acid, elevated up to d 56. Commercial turkeys are generally citric acid, malic acid, lactic acid, tartaric acid, and slaughtered at approximately 18 wk of age, in contrast orthophosphoric acid) at the inclusion rate of 1.5 g/ to chickens that are killed at 6 wk. Due to the lon- kg of feed. ger raising period, other factors can influence the VFA According to Schneitz (2005), CE can improve the concentration, besides reduction of protein in the diet. performance and diminish the mortality of birds. In Between 6 and 12 wk, there is an expansion of gut- the present study, CE product was added to the feed, associated lymphoid tissue (Lillehoj and Chung, 1992) similar to the concept of direct-fed microbials (Elam and a drop in the growth hormone levels (Scanes et et al., 2003). In agreement with the findings of Ow- al., 1984). The growth hormone stimulates the thyroid, ings (1992), there was no finding of a positive effect of adrenal glands, and pancreas and affects lipid and sug- CE product on bird performance, but several authors ar metabolism. Some functions presented by the host report positive effects triggered by the use of live mi- could include decreased intestinal pH or increased de- crobiota in the feed (Angel et al., 2005; Grimes et al., fensin production (Scupham, 2007). These factors can 2008). Among the factors that can affect the efficacy of affect the VFA concentration directly or indirectly, but CE products are the use of antibiotics, elevated stress, specific studies on the species in question are necessary. fasting, and the presence of diseases before application Acetic acid is the predominant VFA in the cecal con- of the product, and several factors can interfere in the tent of chickens (Engberg et al., 2002), which can also response of birds to additives, including management be validly affirmed for turkeys, as shown in the study. practices, environmental conditions, the type and dose There was no observable effect of lincomycin on the of an additive, and the particular characteristics of the concentration of the VFA evaluated. According to specie in question (Yang et al., 2009). In this study, the Chaveerach et al. (2004), most antibiotics have no in- birds were raised under suitable conditions of manage- fluence on the production of VFA. The CE product ment, environment, and stocking density. This situation positively influenced the concentrations of propionic 1860 Milbradt et al. and butyric acid, probably due to the establishment Cave, N. A. G. 1984. Effect of dietary propionic and lactic acids on of VFA-producing microbiota in the cecum (Schneitz, feed intake by chicks. Poult. Sci. 63:131–134. Chaveerach, P., D. A. Keuzenkamp, L. J. A. Lipman, and F. Van 1998). At some moments, the birds that received OA Knapen. 2004. Effect of organic acids in drinking water for young presented a higher concentration of butyric and pro- broilers on Campylobacter infection, volatile fatty acid produc- pionic acid than the control group. Although it is be- tion, gut microﬂora and histological cell changes. Poult. Sci. lieved that organic acids in powdered form are rapidly 83:330–334. Dibner, J. J., and J. D. Richards. 2005. Antibiotic growth promoters absorbed in the intestinal tract, thus losing the ability in agriculture: History and mode of action. Poult. Sci. 84:634– to reach the cecum (Thompson and Hinton, 1997), they 643. can exert an influence on intestinal microbiota, reduc- Dyce, K. M., W. Sack, and C. J. G. Wensing. 1996. Anatomia das aves. Pages 631–650 in Tratado de Anatomia Veterinária. 2nd ed. ing the population of bacteria from the family Entero- Guanabara Koogan, Rio de Janeiro, Brazil. bacteriaceae and favoring the establishment of VFA- Elam, N. A., J. F. Gleghorn, J. D. Rivera, M. L. Galyean, P. J. De- producing bacteria. foor, M. M. Brashears, and S. M. Younts-Dahl. 2003. Effects of Supplementation with CE product and OA can lead live cultures of Lactobacillus acidophilus (strains NP45 and NP51) and Propionibacterium freudenreichii on performance, carcass, to beneficial alterations in the intestinal tract such as and intestinal characteristics, and Escherichia coli strain O157 the reduction of pH and increase in the height of villi shedding of finishing beef steers. J. Anim. Sci. 81:2686–2698. (Yang et al., 2009). The action of antimicrobial agents Engberg, R. M., M. S. Hedemann, and B. B. Jensen. 2002. The influ- can diminish the intestinal microbiota and, consequent- ence of grinding and pelleting of feed on the microbial composition and activity in the digestive tract of broiler chickens. Br. ly, cause reduction of toxins, which can lead to altera- Poult. Sci. 43:569–579. tions in the intestinal morphometry such as shortening Erwin, W. S., G. J. Marco, and E. M. Mery. 1961. Volatile fat acid of villi (Xu et al., 2003). In the current study, there analyses of blood an rumen fluid by gas chromatography. J. was no observable influence of additives on the height Dairy Sci. 44:1768–1771. Garcia, V., P. Catala-Gregori, F. Hernandez, M. D. Megias, and J. of intestinal villi. Similar results have been described Madrid. 2007. Effect of formic acid and plant extracts on growth, by Houshmand et al. (2011), who used organic acids as nutrient digestibility, intestine mucosa morphology, and meat growth promoters in the feed of broiler chickens. These yield of broilers. J. Appl. Poult. Res. 16:555–562. authors found no difference in villus height or the crypt Gaskins, H. R., C. T. Collier, and D. B. Anderson. 2002. Antibiot- ics as growth promotants: Mode of action. Anim. Biotechnol. depth in the duodenum, jejunum, and ileum. However, 13:29–42. Baurhoo et al. (2007) and Garcia et al. (2007) reported Goren, E., W. A. de Jong, P. Doornenbal, J. P. 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