Livestock Science 195 (2017) 3844

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Livestock Science
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Increase in dietary arginine level could ameliorate detrimental impacts of MARK

coccidial infection in broiler chickens

M. Laikaa, R. Jahanianb,
a
Department of Animal Sciences, College of Agriculture, Isfahan University of Technology, Isfahan 84156-83111, Iran
b
Poultry Nutrition Research Center, Bioscitech Research Institute, Isfahan 81398-67433, Iran

A R T I C L E I N F O A BS T RAC T

Keywords: The present study was conducted to investigate the eect of dietary supplementation of Arg on growth
Broiler chickens performance, carcass characteristics, and morphological indices of jejunal epithelial cells in coccidia-challenged
Arginine broiler chickens. A total of 288 one-day-old broiler chickens were randomly distributed among 3 experimental
Coccidiosis treatments with 8 replicate pens of 12 broiler chickens each. Experimental treatments consisted of the graded
Carcass characteristics
levels of dietary Arg (100, 105, and 110% of the standard recommendations during dierent growth periods).
Intestinal morphology
From 16 to 20 d of age, half of the replicate pens of each dietary Arg level were orally challenged with a mixture
Growth performance
of Eimeria species (acervulina, tenella, maxima, and necatrix). Results showed that dietary Arg had no marked
eect on average daily feed intake (ADFI) and gain (ADG) during the starter (0 to 14 d of age) and grower (14 to
28 d of age) periods. Although ADFI wasnt aected by coccidial challenge, ADG of Eimeria-challenged broiler
chickens were lower (P < 0.05) than those of uninfected ones during both grower and nisher periods. Dietary
supplementation of 105 and 110% of Arg, however, prevented depressed ADG in coccidia-infected broiler
chickens during the nisher period (28 to 42 d of age) compared with 100% of Arg (Argcoccidiosis, P < 0.05).
Coccidial challenge increased (P < 0.01) feed conversion ratio (FCR), while 110% of Arg supplementation
improved (P < 0.05) FCR values during both grower and nisher periods. Although coccidial challenge increased
FCR value in control broiler chickens (100% of Arg), dietary Arg supplementation of 105 and 110% improved
FCR values during the grower period, resulted in an Argcoccidiosis interaction (P < 0.01). Subjecting the
broiler chickens to coccidiosis reduced liver weight (P < 0.001) and carcass yield (P < 0.05). Dietary Arg
supplementation increased (P < 0.001) villi height (VH) to crypt depth (CD) ratio. On the other hand, both VH
and VH to CD ratio were decreased (P < 0.001) in coccidial-infected broiler chickens. Although subjecting the
broiler chickens to coccidiosis increased (P < 0.001) muscular layer thickness (MLT) of jejunum, supplemental
Arg of 110% resulted in a decrease (P < 0.01) in jejunal MLT. In addition, supplemental Arg reduced (P < 0.05)
fecal oocyst count, with the lowest count assigned to the broiler chickens fed 110% of Arg. Dietary Arg
supplementation of 110% improved morphological indices in coccidia-challenged broiler chickens, while it had
no obvious impact in untreated ones, resulted in the Argcoccidiosis interactions (P < 0.05). The present
ndings showed that supplementing the diets with Arg above the recommended values could ameliorate
negative eects of Eimeria on growth performance and morphological indices of broiler chickens.

1. Introduction
The key target of poultry production for the food chain is to obtain
the optimum growth rate and feed conversion eciency, while main-
taining optimal animal health. However, occurrence of some infectious
and parasitic diseases diminishes poultry productivity, resulting in the
economic losses in poultry industry. Coccidiosis is the most frequently
reported and economically important poultry-related disease world-
wide (Dalloul and Lillehoj, 2006). This parasitic disease is an expensive
disease and is estimated to cost the poultry industry by 3.2 billion USD
annually (Dalloul and Lillehoj, 2006; De Gussem, 2007). The major
part of these costs, around 80%, is due to the losses in growth
performance, and the rest is due to the costs of prophylaxis and
treatment (Vermeulen et al., 2001). Several Eimeria species cause
coccidiosis in chickens, with the most prevalent being E. acervulina, E.
maxima, and E. tenella (Vermeulen et al., 2001). Coccidial infection in
broiler chickens results in damage to epithelial cells, diarrhea, osmotic
stress in the intestine (Kettunen et al., 2001, 2003; Perez-Carbajal

Corresponding author.
E-mail address: r.jahanian@gmail.com (R. Jahanian).

http://dx.doi.org/10.1016/j.livsci.2016.11.002
Received 26 March 2016; Received in revised form 16 September 2016; Accepted 4 November 2016
1871-1413/ 2016 Elsevier B.V. All rights reserved.
M. Laika, R. Jahanian Livestock Science 195 (2017) 3844

et al., 2010), and subsequent nutrient malabsorption (Persia et al., Table 1

2006; Metzler-Zebeli et al., 2009). Feed ingredients and nutrient composition of experimental diets during different growth
periods.a
Historically, poultry industry has prevented and controlled cocci-
dial infections with the inclusion of anticoccidial feed additives. Item Starter (0 to 14 Grower (14 to 28 Finisher (28 to 42
However, concerns over the future use of anticoccidial agents have d of age) d of age) d of age)
increased the needs for development of alternative strategies (both
Ingredients (g/kg)
management and nutritional) to prevent or overcome coccidial infec-
Corn, yellow 534.2 590.0 675.1
tions in poultry ocks. One of the main reasons for elimination of Soybean meal 371.8 289.1 218.2
anticoccidials is development and establishment of drug-resistant Corn gluten meal 30.0 50.0 50.0
strains, which continue to emerge across the world (Williams, 2002). Soybean oil 14.5 22.9 10.6
One potential strategy to control coccidial infections is through Limestone 11.8 11.6 11.0
Dicalcium 19.7 18.8 18.4
improving host immune defenses by nutritional modications.
phosphate
Arginine is one of the most important amino acids involving in immune Common salt 2.3 2.3 2.3
functions (Jahanian, 2009; Tan et al., 2009; Ren et al., 2014). Similar Na bicarbonate 1.5 1.5 1.5
to other amino acids, Arg is basically known for its role in protein Vitamin premixb 2.5 2.5 2.5
Mineral premixc 2.5 2.5 2.5
biosynthesis. Identication of the Arg pathway that produces NO,
DL-Met 2.0 1.5 1.1
however, led the researchers to demonstrate that Arg is needed for L-LysHCl 1.7 2.3 1.8
optimal immune responses (Kidd et al., 2001; Jahanian, 2009). L-Thr 0.5
Production of NO as a result of infection can be considered as a part Inert llerd 5.0 5.0 5.0
of the inammatory responses, because it is most often triggered by
Composition
inammatory cytokines (Allen, 1999). Obviously, it has been known
MEn (MJ/kg) 12.14 12.76 12.76
that Arg is the sole substrate for NO synthesis in biological systems and CP (g/kg) 225.0 205.0 180.0
NO has been implicated as a key detector factor in disease situations Ca (g/kg) 10.0 9.5 9.0
(Billiar, 1995; Rodeberg et al., 1995). Several studies have indicated NPP (g/kg) 4.8 4.5 4.3
Na (g/kg) 1.5 1.5 1.5
that supplemental Arg could ameliorate growth depression and repro-
Met (g/kg) 5.7 5.1 4.4
ductive failure in infected animals (Ren et al., 2012; Yin et al., 2014). Met+Cys (g/kg) 9.4 8.6 7.6
Furthermore, Ren et al. (2014) reported that dietary Arg supplementa- Lys (g/kg) 13.0 11.6 9.5
tion favored jejunal bacterial populations and activated intestinal Thr (g/kg) 9.0 7.7 6.7
innate immunity through dierent signaling pathways. Therefore, Trp (g/kg) 3.3 2.8 2.3
Arg (g/kg) 14.2 12.2 10.3
dietary Arg supplementation above the recommended values seems
to promote immune system and its response against dierent infec- a
MEn=metabolizable energy corrected for zero nitrogen retention; CP=crude protein;
tious invasions, including coccidiosis. Because there isnt enough and NPP=non-phytate phosphorus.
b
information regarding the benecial impacts of supplemental Arg Vitamin premix provided per kilogram of diet: vitamin A (retinyl acetate), 13,000 IU;
during coccidial infections, the present study was designed to investi- cholecalciferol, 3,800 IU; vitamin E (DL--tocopheryl acetate), 60 IU; menadione
(menadione dimethyl-pyrimidinol), 4.5 mg; thiamine, 3.2 mg; riboavin, 6.5 mg; nicotin
gate the eect of dietary Arg overdose on growth performance and
amide, 55 mg; calcium pantothenate, 30 mg; pyridoxine, 7.6 mg; folic acid, 1.2 mg;
morphological indices of jejunal epithelial cells during an experimen- biotin, 0.25 mg; vitamin B12, 0.03 mg; choline (choline chloride, 60%), 450 mg; and
tally-induced coccidiosis. ethoxyquin, 80 mg.
c
Mineral premix provided per kilogram of diet: Mn (MnSO4H2O), 95 mg; Zn (ZnO),
90 mg; Fe (FeSO47H2O), 55 mg; Cu (CuSO45H2O), 10 mg; I (Ca (IO3)2H2O), 1.9 mg;
2. Materials and methods
and Se (Na selenite), 0.35 mg.
d
Inert ller (washed builder sand) was replaced with equal amounts of L-ArgHCl
2.1. Broiler chickens, treatments, and experimental procedure (Ajinomoto Co., Inc., Kawasaki, Japan) to achieve dierent experimental Arg levels.

The present study was performed in the Poultry Research Station of
Isfahan University of Technology (Isfahan, Iran). All protocols used
were approved by the Isfahan University of Technology Animal Care
and Use Committee. A total of 288 one-day-old broiler chickens (Ross
308; Navid Morgh Guilan Co., Guilan, Iran) were randomly distributed
among 3 dietary treatments with 8 replicate pens of 12 broiler chickens
each. Dietary treatments were dierent levels (100, 105, and 110% of
the standard recommended values; Aviagen, 2009) of dietary Arg. After
the starter period, 4 replicate pens of each dietary Arg level were
randomly selected and challenged with a mixture of Eimeria species (E.
acervulina, E. tenella, E. maxima, and E. necatrix) from 16 to 20 d of
age. Each broiler chicken was administered with 1 mL oral dose of
sporulated Eimeria oocysts per day (each inoculum was containing
195,000 oocysts of E. acervulina, E. tenella, E. maxima, and E.
necatrix at the proportions of 10:1:1:1, respectively).
Dietary Arg levels of 100, 105, and 110% of recommendations were
14.2, 14.9, and 15.6 g/kg during the starter period. The respective
values were 12.2, 12.8, and 13.4 g/kg for the grower, and 10.3, 10.8,
and 11.3 g/kg for the nisher period. Basal experimental diets
(Table 1) were formulated to meet the nutritional requirements of
broiler chickens as provided by management manual (Aviagen, 2009)
during dierent periods. Broiler chickens were maintained on a 23
light:1 dark lighting regimen in the oor pens (11 m) in a thermo-
statically-controlled room, and had free access to water and experi-
mental diets throughout the duration of study. Temperature was set on
34 C during the rst week of age; then, reduced by 3 C/week until the
broiler chickens were 5-wk-old and xed at 22 C for the remaining
trial period.
2.2. Chemical analysis
Before formulating the diets, the main feed ingredients (i.e., corn,
soybean meal, and corn gluten meal) were analyzed for crude protein
(Method 976.06), ether extract (Method 954.02), crude ber (Method
978.10), and ash (Method 942.05) according to the standard proce-
dures of AOAC (2002). The metabolizable energy values of ingredients
were then calculated using NRC (1994) recommended formula. In
addition to basic chemical composition, the feed ingredients were
analyzed for amino acid composition according to Jahanian and
Rasouli (2014). Briey, all feed samples were hydrolyzed with HCl
(containing 0.1% phenol) for 24 h at 110 C in glass tubes sealed under
vacuum. Amino acid contents of feed samples were detected using an
ion exchange HPLC (Biochrom 20 Amino Acid Analyzer; Biotronik
GmbH, Maintal, Germany), through post-column ninhydrin derivatiza-
tion and uorescence detection. The resultant chromatograms were
integrated using the instrument software. The basal experimental diets
(during dierent growth periods) were formulated according to the

39
M. Laika, R. Jahanian Livestock Science 195 (2017) 3844

Table 2
Effect of dietary Arg supplementation on growth performance of coccidial-challenged broiler chickens.a

Argb Coccidial ADFI (g/d per chicken) ADG (g/d per chicken) FCR (g feed/ g gain)
challengec
Starter (0 to Grower (14 to Finisher (28 to Starter (0 to Grower (14 to Finisher (28 to Starter (0 to Grower (14 to Finisher (28 to
14 d) 28 d) 42 d) 14 d) 28 d) 42 d) 14 d) 28 d) 42 d)

100 35.99 101.4 173.2 24.63 60.88 79.80 1.46 1.66 2.17
+ 103.0 175.8 59.12 76.09 1.74 2.31

105 36.40 103.8 174.7 24.92 61.07 77.85 1.46 1.70 2.24
+ 102.6 178.7 60.20 77.06 1.70 2.32

110 36.21 102.5 172.3 24.57 61.27 78.42 1.47 1.67 2.20
+ 101.4 172.6 60.19 77.79 1.68 2.22

SEM 0.24 1.1 1.3 0.21 0.62 0.86 0.01 0.01 0.02

P-values

Arg
100 vs. 105+110% 0.292 0.762 0.977 0.659 0.210 0.914 0.510 0.219 0.941
105 vs. 110% 0.464 0.148 0.011 0.176 0.969 0.508 0.229 0.016 0.026
Coccidiosis 0.794 0.055 0.035 0.049 0.001 0.002
Arg coccidiosis 0.414 0.390 0.782 0.041 0.002 0.026

a
ADFI=average daily feed intake; ADG=average daily gain; FCR = feed conversion ratio; and SEM=standard error of the means. n=8 for performance criteria during the starter
period and n=4 for performance during other experimental periods.
b
Dietary Arg levels of 100, 105, and 110% were 14.2, 14.9, and 15.6 g/kg during the starter period. The respective values were 12.2, 12.8, and 13.4 g/kg for the grower stage, and
10.3, 10.8, and 11.3 g/kg for the nisher period.
c
Each broiler chicken was administered with 1 mL oral dose of sporulated Eimeria oocysts per day (each inoculum containing 195,000 oocysts of E. acervulina, E. tenella, E. maxima,
and E. necatrix at the proportions of 10:1:1:1) from 16 to 20 d of age.

analyzed values to be containing dietary Arg level of 100% during each
period. Additional Arg levels were obtained by stepwise addition of L-
ArgHCl (Ajinomoto Co., Inc., Kawasaki, Japan) to the basal diets.
2.3. Growth performance
Average daily feed intake (ADFI) and average daily gain (ADG) were
recorded periodically as the starter (0 to 14 d of age), grower (14 to 28
d of age), and nisher (28 to 42 d of age) periods. Weight gains were
measured by pen basis after 3 h of feed withdrawal. Feed conversion
ratio (FCR) was calculated for 3 dierent periods. Mortality was
recorded to adjust feed intake, weight gain, and resultant FCR values.
times during xation process), and then were dehydrated through
consecutive embedding in graded ethanol solutions. Thereafter, the
xed samples were embedded in paran. Transverse and longitudinal
sections with 5 m in thickness were prepared using microtome,
stained with Hematoxyline-Eosin, and examined under the light
microscope (Olympus CH-2; Olympus Corp., Tokyo, Japan) for mea-
suring villi height (VH), villi width, crypt depth (CD), villi surface area
(VSA), submucosal layer thickness, and muscular layer thickness
(MLT). The examinations were made by a veterinary technician under
the lens 40 x and each slide was examined 2 times. All measurements
were obtained using 15 villi per slide, and the mean values of 2 slides
per pen were used for statistical analysis.

2.4. Carcass characteristics and internal organ weights 2.6. Oocyst shedding

At the end of study (d 42 of age), 2 broiler chickens per pen, nearest
to the average weight of the same pen (small or heavy chickens were
ignored from the selection), were selected to evaluate internal organs
weight and carcass yield. Feed was removed 3 h before slaughter. Each
broiler chicken was exsanguinated by cutting the jugular vein, and
allowed to bleed for approximately 2 min. Viscera were removed
immediately; thereafter, the weights of liver, pancreas, heart, abdom-
inal fat, and carcass were obtained using a sensitive (0.01 g) digital
scale (UX2200H; Shimadzu Corp., Kyoto, Japan). Carcass yield and
relative weights of internal organs were calculated as the percentages of
live body weight and average values of 2 broiler chickens per pen were
used for analysis of variance, as described by Jahanian et al. (2008).
Fecal samples were collected from each pen on d 26 and 27 of age
(6 and 7 d post infection). Random collection of the samples from each
pen is necessary to obtain an accurate estimate of oocyst excretion. All
areas of each pen oor were sampled, and a representative amount of
excreta was collected from each pen. Samples were kept in the separate
airtight plastic bags. After thoroughly homogenization, samples were
stored at 4 C (refrigerator) until oocyst counting could be performed.
Homogenized samples were diluted 10-fold with distilled water and
further diluted 10-fold with a saturated saline solution. Oocyst count
was determined via microscope using McMaster counting chamber (JA
Whitlock & Co., Eastwood, NSW, Australia), and expressed as the
number (104) of oocysts per gram of excreta (Christaki et al., 2004).

2.5. Jejunal morphology 2.7. Statistical analysis

At the end of study, 2 broiler chickens from each replicate pen were
slaughtered by cutting the jugular vein and tissue samples (jejunal
sections) were collected and processed according to the method
described by Sun et al. (2005). Briey, the 2 cm sections of jejunal
segment, anterior to Meckel's diverticulum, were excised for light
microscopic observations. The histological sections were immediately
xed in 10% formalin solution for 7 d (formalin solution was replaced 3
All data were subjected to analysis of variance using the general
linear model procedures of SAS (SAS Institute, 1999). The pen was the
experimental unit for all measurements. For growth performance
criteria during the starter period, 3 dierent levels of dietary Arg were
compared. For remaining traits, however, data set was analyzed as a
32 factorial arrangement, including dietary Arg levels and coccidial
challenge as the main eects and respective interaction. The orthogonal

40
M. Laika, R. Jahanian Livestock Science 195 (2017) 3844

contrasts were made among dierent Arg levels (100 vs. 105+110%
and 105 vs. 110%) to evaluate the eects of supplemental Arg levels.
Results were considered statistically signicant when P < 0.05.
3. Results
3.1. Growth performance
As shown in Table 2, dietary Arg had no marked eects on ADFI,
ADG, and FCR during the starter period. After coccidial challenge (at
the grower period), however, increase in dietary Arg up to 110% of
recommended values improved (P < 0.05) FCR values during the
grower and nisher periods. On the other hand, ADG wasnt aected
by dietary Arg in the grower and nisher periods. The lowest (P < 0.05)
ADFI was assigned to the broiler chickens fed diet containing the
greatest dietary Arg level during the nisher period. Although experi-
mental challenge with coccidiosis had no marked eect on ADFI, it
resulted in the losses (P < 0.05) in ADG during both grower and
nisher periods. In addition, coccidial challenge increased (P < 0.01)
FCR values during both experimental periods. Dietary Arg supplemen-
tation prevented depressed ADG caused by Eimeria infection, resulted
in an Argcoccidiosis interaction (P < 0.05) for ADG during the nisher
period. Furthermore, there was an Arg coccidiosis interaction for
FCR values during the grower (P < 0.01) period, so that supplemental
Arg of 105 and 110% improved FCR values in coccidia-infected broiler
chickens compared with 100% of Arg. Such improving eect (P < 0.05)
was seen with 110% of Arg during the nisher period.
3.2. Internal organs weight
Eects of dietary Arg on internal organs weight and carcass yield of
coccidia-challenged broiler chickens are presented in Table 3. As noted,
supplemental Arg up to 110% of recommended values increased (P <
0.01) the relative liver weight. Relative weights of other internal
organs, including heart and pancreas, however, werent aected by
dietary Arg. In addition, dietary Arg supplementation had no marked
eect on abdominal fat percentage and carcass yield. On the other
Table 3
Effect of dietary Arg supplementation on carcass characteristics and internal organs
weight (as the percentages of live body weight) of coccidial-challenged broiler chickens.a
Argb Coccidial Liver Pancreas Heart Abdominal fat
challengec
100 2.29 0.23 0.49 1.57
+ 2.01 0.23 0.48 1.41
hand, subjecting the broiler chickens to coccidial challenge decreased
(P < 0.05) carcass yield. The nding of importance was the considerable
reduction (P < 0.001) of relative liver weight as the result of coccidial
challenge.
3.3. Gut morphology
As shown in Table 4, dietary Arg level didnt aect VH and villi
width. On the other hand, CD was decreased (P < 0.01) as the result of
dietary Arg supplementation. As a consequence, the greater (P0.001)
VH: CD was assigned to the broiler chickens fed diets containing 105
and 110% of Arg. Dietary Arg supplementation of 110% of recom-
mended values decreased (P < 0.01) MLT. Challenging the broiler
chickens with a mixture of Eimeria species resulted in decreases (P <
0.001) in VH and VH: CD. As a result, VSA was markedly (P < 0.001)
decreased in coccidia-infected broiler chickens. Moreover, coccidial
challenge increased (P < 0.001) MLT. Although coccidial infection
increased CD in broiler chickens fed diet containing 100% of Arg,
supplemental Arg of 105 and 110% caused numerical decreases in CD
of coccidia-challenged broiler chickens, resulted in an Arg coccidiosis
interaction (P < 0.05). A similar interaction was seen for VH: CD, so
that 110% of Arg supplementation resulted in a similar VH: CD in
coccidia-challenged and uninfected broiler chickens. Subjecting the
broiler chickens to coccidiosis increased MLT in the control group
(100% of Arg), while it had no marked eect on MLT in broiler
chickens supplemented with 105 and 110% of Arg (Argcoccidiosis, P
< 0.05). Fig. 1 shows microscopic alterations of villi as the result of
coccidial challenge and ameliorating eect of 110% of Arg on histolo-
gical indices of jejunal epithelial cells. As shown, uninfected broiler
chickens (Fig. 1a) had the tallest villi, while coccidial infection
decreased VH and increased CD in broiler chickens receiving 100%
of Arg (Fig. 1b). On the other hand, dietary supplementation with Arg
up to 110% of recommended values increased VH in coccidia-chal-
lenged broiler chickens (Fig. 1c).
3.4. Oocyst count
Eect of dietary Arg on oocyst enumeration in fecal contents of
Eimeria-infected broiler chickens is shown in Table 4. As presented,
the lowest (P < 0.01) oocyst count was allotted to the broiler chickens
fed diet containing 110% of Arg. As expected, challenging the broiler
chickens with a mixture of Eimeria species increased (P < 0.001) oocyst
Carcass enumeration in feces. Interestingly, there was an Arg coccidiosis
(skinless) interaction (P < 0.05) for oocyst count, so that 110% of Arg supple-
mentation ameliorated the severity of oocyst shedding in coccidia-
64.01
63.28 challenged broiler chickens, even though this eect wasnt complete.

105 2.21 0.23 0.46 1.56 64.86 4. Discussion

+ 2.07 0.19 0.54 1.50 63.24

110 2.37 0.22 0.54 1.61 64.06

As noted in Table 2, dietary Arg had no marked eect on growth
+ 2.23 0.22 0.49 1.65 63.48 performance criteria during the starter period (before coccidial chal-
lenge). During the grower and nisher periods, however, supplemental
SEM 0.05 0.01 0.02 0.06 0.44 Arg of 110% improved FCR values. Some research studies (Kwak et al.,
1999; Jahanian, 2009; Khalifeh-Gholi and Jahanian, 2012) have
P-values
shown that Arg supplementation of decient diets increased weight
Arg gain and improved FCR in broiler chickens. However, in the present
100 vs. 105+110% 0.422 0.328 0.345 0.148 0.541 study we formulated the basal experimental diets to be containing Arg
105 vs. 110% 0.033 0.635 0.648 0.096 0.619 at the sucient quantities during dierent periods (based on the
Coccidiosis < 0.001 0.182 0.736 0.266 0.015
Argcoccidiosis 0.291 0.169 0.041 0.333 0.459
standard recommendations); so, the lack of growth performance
response to supplemental Arg wasnt unexpected. In addition, the lack
a
SEM=standard error of the means. n=4. of marked eect of supplemental Arg on ADG of infected broiler
b
Dietary Arg levels of 100, 105, and 110% were 14.2, 14.9, and 15.6 g/kg during the chickens shows that the availability of this amino acid isnt a rate-
starter period. The respective values were 12.2, 12.8, and 13.4 g/kg for the grower stage, limiting factor for growth in Eimeria-challenged broiler chickens.
and 10.3, 10.8, and 11.3 g/kg for the nisher period.
c
Each broiler chicken was administered with 1 mL oral dose of sporulated Eimeria
Subjecting to coccidiosis reduced ADG and caused considerable
oocysts per day (each inoculum containing 195,000 oocysts of E. acervulina, E. tenella, increases in FCR values during both grower and nisher periods.
E. maxima, and E. necatrix at the proportions of 10:1:1:1) from 16 to 20 d of age. Consistent with our ndings, Watson et al. (2005), Parker et al. (2007),

41
M. Laika, R. Jahanian Livestock Science 195 (2017) 3844

Table 4
Effects of dietary Arg supplementation on jejunal histology and fecal oocyst count in coccidial-challenged broiler chickens.a

Argb Coccidial challengec VH (m) VW (m) CD (m) VH: CD VSA (mm2) SLT (m) MLT (m) Oocyst (104/g excreta)

100 1,093 110 148 7.40 0.37 23 90 3.50

+ 770 130 178 4.37 0.31 25 148 6.50

105 1,070 135 130 8.23 0.45 28 105 3.25

+ 883 93 125 7.07 0.26 28 128 5.75

110 1,018 113 145 7.05 0.36 18 83 3.25

+ 913 135 128 7.22 0.38 20 90 4.50

SEM 52 7 8 0.30 0.01 5 9 0.27

P-values

Arg
100 vs. 105+110% 0.550 0.888 0.001 0.006 0.549 0.890 0.159 0.018
105 vs. 110% 0.858 0.392 0.262 0.205 0.726 0.084 0.005 0.030
Coccidiosis < 0.001 0.999 0.713 < 0.001 < 0.001 0.703 < 0.001 < 0.001
Argcoccidiosis 0.134 < 0.001 0.026 < 0.001 < 0.001 0.963 0.030 0.013

a
VH = villi height; VW = villi width; CD = crypt depth; VH: CD = villi height to crypt depth ratio; VSA = villi surface area; SLT = submucosal layer thickness; MLT = muscular layer
thickness; and SEM = standard error of the means. n =4.
b
Dietary Arg levels of 100, 105, and 110% were 14.2, 14.9, and 15.6 g/kg during the starter period. The respective values were 12.2, 12.8, and 13.4 g/kg for the grower stage, and
10.3, 10.8, and 11.3 g/kg for the nisher period.
c
Each broiler chicken was administered with 1 mL oral dose of sporulated Eimeria oocysts per day (each inoculum containing 195,000 oocysts of E. acervulina, E. tenella, E. maxima,
and E. necatrix at the proportions of 10:1:1:1) from 16 to 20 d of age.

Bozkurt et al. (2014), and Amerah and Ravindran (2015) reported that
coccidial challenge reduced weight gain and feed eciency in broiler
chickens. Reduced nutrient digestibility and increased immune costs
associated with coccidial infection are likely to be responsible for the
reduced growth performance in broiler chickens (Williams, 2005). On
the other hand, dietary Arg fortication improved FCR values in
coccidia-challenged broiler chickens, resulted in the Argcoccidiosis
interactions. Although not measured, the benecial impacts of supple-
mental Arg in Eimeria-infected broiler chickens is largely because of
the involvement of this critical amino acid in production of NO. It has
been shown that production of this eector molecule and plasma levels
of its metabolites (i.e., NO2 and NO3) were signicantly increased
during primary infections with E. acervulina (Allen and Teasdale,
1994), E. tenella (Allen, 1997a), and E. maxima (Allen, 1997b).
Because the chickens can't synthesize Arg de novo via the ornithine
cycle, they must obtain this essential amino acid directly from the diet
(Whittow, 2000; Leeson and Summers, 2001). Therefore, in theory, NO
production in response to infection and its subsequent eects on the
host and infecting organism should be manipulatable with dietary Arg
supplementation.
As presented in Table 3, dietary Arg of 110% caused an increase in
the relative liver weight. This eect can be attributed to the inductive
eect of Arg on endocrine secretions, which stimulates the release of
pituitary and pancreatic hormones, including glucagon, insulin, and
growth hormone (Davila et al., 1987). This hormonal induction could
promote protein biosynthesis and increased liver activity and weight,
with consideration of the fact that liver is the most important metabolic
organ (Whittow, 2000) in avian species. In this regard, it has been
reported that Arg and its family (i.e., glutamine, glutamate, proline,
aspartate, and asparagine) serve important regulatory functions in
nutrient metabolism and immune responses, thereby aect organs
(especially liver and small intestine) metabolism (Wu et al., 2007).
As noticed, coccidial challenge decreased both liver weight and
carcass yield. It seems that reduced liver weight as the result of coccidia
challenge is related to the lower ADG in infected broiler chickens,
which dont need to the higher anabolic activity of liver and extra-
hepatic tissues. On the other hand, reduced nutrient digestibility and
increased immune costs (Williams, 2005) associated with coccidial
infection can be responsible for reduced liver weight in coccidia-
challenged broiler chickens. Consistent with our conclusion,
Matthews and Southern (2000) showed that chronic and acute infec-
tions with E. acervulina decreased plasma total protein in broiler
chickens. In contrast to our ndings, Bozkurt et al. (2014) reported that
coccidial infection resulted in substantial increases in relative weights
of liver (by 24%) and pancreas (by 11%) compared with uninfected
broiler chickens. These researchers attributed the increased liver and
pancreas weights with an attempt to overcome the deciencies in the
production of digestive enzymes and bile salts and to maintain hepatic
turnover under the conditions of parasitic infection (Bozkurt et al.,
2014).
Coccidial infection decreased VH, resulted in the lower VH: CD. As
a result, VSA was lower in coccidia-challenged broiler chickens than
that of uninfected ones. On the other hand, subjecting the broiler
chickens to coccidiosis increased MLT. During coccidiosis, sporozoites
infect the cells of the intestinal lining, causing tissue damage and
trauma to the intestinal mucosa and submucosa (Perez-Carbajal et al.,
2010). Furthermore, coccidial infections negatively aect the morphol-
ogy of the intestine, especially shorten villi (Kettunen et al., 2001) and
reduce the activities of digestive enzymes (Williams, 2005). The overall
eect is reduction of digestion and absorption processes. For example,
the negative eects of coccidial challenge on amino acid digestibility
have been reported by Persia et al. (2006) and Parker et al. (2007).
Supplementation of basal diets with L-ArgHCl increased VH, VH:
CD, and VSA, and these benecial eects were more pronounced in
infected broiler chickens. It seems that the positive impacts of
supplemental Arg on morphological indices of jejunal epithelial cells
are largely because of the production of NO through activity of NO
synthase on Arg (Moncada and Higgs, 1993). Because of its oxidant
properties and ability to react with intracellular iron-containing
compounds, NO could be toxic to coccidia, as well as to host cells
harboring the parasite (Moncada et al., 1991; Ovington and Smith,
1992). If NO is toxic to dierent Eimeria species, dietary Arg
supplementation might be expected to increase plasma NO2 + NO3
levels and decrease lesion scores and oocyst shedding in coccidia-
infected broiler chickens. Consistent with this, supplemental Arg
increased VSA and reduced oocyst count in the present study,
especially in coccidia-challenged broiler chickens. On the other hand,
supplemental Arg has been reported to enhance intestinal growth and
development (Yao et al., 2011) and to prevent gut dysfunction (He
et al., 2011) in piglets. It seems that a part of inducing eects of

42
M. Laika, R. Jahanian
Fig. 1. Eect of dietary Arg supplementation on villi height and crypt depth in coccidial-
challenged broiler chickens. a) uninfected broiler chickens with 100% of dietary Arg
recommendations (i.e., 14.2, 12.2, 10.3 g/kg during the starter, grower, and nisher,
respectively); b) coccidia-infected broiler chickens with 100% of Arg; and c) coccidia-
infected broiler chickens with 110% of Arg (i.e., 15.6, 13.4, and 11.3 g/kg during the
starter, grower, and nisher, respectively).
supplemental Arg on VH and VSA in infected broiler chickens is
through increasing the expression of key regulation factors within the
intestinal cells (Yao et al., 2011).
5. Conclusions
This study draws attention to detrimental impacts of coccidiosis on
growth performance and morphological indices of jejunal epithelial
cells in broiler chickens. Coccidial infection reduced VH, resulted in
decreased absorption surface area in jejunal epithelium. In addition,
subjecting to a mixture of Eimeria species reduced liver weight and
carcass yield. These adverse eects were largely ameliorated with
supplemental Arg up to 110% of recommended values. The benecial
impacts of supplemental Arg at practical conditions, however, need
43
Livestock Science 195 (2017) 3844
further investigations.
Conict of interest statement
The authors declare that they have no conict of interest.
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