EFFECTS OF POST-HATCH NUTRITION OF ARGININE, LYSINE, METHIONINE AND THREONINE ON PERFORMANCE AND INTESTINAL MORPHOLOGY IN BROILER CHICKENS

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1 Bulgrin Journl of Veterinry Medicine, 2015, 18, No 4, ISSN ; DOI: /bjvm.861 Originl rticle EFFECTS OF POST-HATCH NUTRITION OF ARGININE, LYSINE, METHIONINE AND THREONINE ON PERFORMANCE AND INTESTINAL MORPHOLOGY IN BROILER CHICKENS A. SADRZADEH 1, H. HASSANPOUR 2, M. H. MEHRAEI HAMZE- KOLAEI 3, P. FARHADNIYA 1, M. R. EHSAN 4 & S. S. TOHIDI FAR 3 1 Deprtment of Clinicl Sciences, Fculty of Veterinry Medicine, Islmic Azd University, Grmsr Brnch, Grmsr, Irn; 2 Deprtment of Bsic Sciences (Physiology division), Fculty of Veterinry Medicine, Shhrekord University, Shhrekord, Irn; 3 Deprtment of Clinicl Sciences, Fculty of Veterinry Medicine, Shhrekord University, Shhrekord, Irn; 4 Deprtment of Clinicl Science, Fculty of Veterinry Medicine, Tehrn University, Tehrn, Irn Summry Sdrzdeh, A., H. Hssnpour, M. H. Mehrei Hmzekolei, P. Frhdniy, M. R. Ehsn & S. S. Tohidi Fr, Effects of post-htch nutrition of rginine, lysine, methionine nd threonine on performnce nd intestinl morphology in broiler chickens. Bulg. J. Vet. Med., 18, No. 4, To investigte the effects of post-htch nutrition of rginine, lysine, methionine nd threonine on performnce nd intestinl morphology of broilers, 180 dy-old Ross-308 were ssigned to 6 tretments. Chickens were provided post-htch (48 h) feeding of ech mino-cid in 4 groups. Chickens in other 2 treted groups received no feed nd wter (fsting group) or just wter (wtering group). On dy 14, performnce prmeters were determined, nd then ll chickens from ech tretment were killed. After weighing body orgns, the length of segments of smll intestine were mesured nd prepred for evlution of intestinl morphology. The men body weight nd FCR between wtering, fsting, methionine nd threonine groups did not differ while they ll were greter thn lysine fedchickens (P<0.05). In comprison with other groups, the lmin propri thickness of duodenum nd ileum were greter in methionine nd threonine groups, nd in jejunum it ws more in methionine group (P<0.05). The sum of mesured villus surfce re ws higher nd lower in threonine nd lysine groups respectively (P<0.05). In conclusion, mong the 4 mino cids, post-htch feeding of threonine hs beneficil effects especilly on the growth of intestine. In contrst, lysine hs dverse effects on the body performnce nd intestinl morphology. Key words: mino cid, broiler, intestine, post-htch feeding

2 A. Sdrzdeh, H. Hssnpour, M. H. Mehrei Hmzekolei, P. Frhdniy, M. R. Ehsn... INTRODUCTION Severl studies hve determined the criticl role of post-htch nutrition of fst growing broiler chicks on their performnce (Mdsen et l., 2004; Noy & Uni, 2010; Mhmoudi et l., 2012). Erly nutrition influences development nd growth of chickens over severl dys (Bigot et l., 2003; Quentin et l., 2005). Industril chicks re usully htched for two dys; then they re sexed, vccinted, nd finlly trnsported to industril frms. During this time (2 3 dys of ge), newly htched chicks do not feed or drink nything. Mdsen et l. (2004) showed tht this condition could dely growth of chicks. Pinchsov (1991) nd Noy & Skln (1999) hd lso confirmed tht inccessibility of chicks to feed nd wter in the first dys of life, decresed body weight nd other body performnce. In contrst, shortening the time of deprivtion in post htched chicks, improved body growth nd developed intestine, pncres nd immune system (Jin et l., 1998; Mdsen et l., 2004). It hs been suggested tht n ccumultion of nutrients in crop of chicks my contribute in the development of digestive system even before rriving to industril frm (Bhnj et l., 2010). Severl experiments hve shown tht mino cids in chickens re n importnt requirement for development of different orgns of body, nd the reltion between mino cid/protein requirements nd fctors such s sex, ge, het stress, energy concentrtion (Grci et l., 2006; Bstinelli et l., 2007; Fischer d Silv et l., 2007). Pesti & Miller (1997) identified tht mino cids in optiml levels were not suitble for the mximistion of body performnce e.g., weight gin, feed conversion rte nd growing of brest muscle. Recent studies determined the positive effects of mny essentil mino cids on body weight fter feeding to post-htched chickens (Noy & Skln, 2002; Stringhini et l., 2009). The present reserch ws imed t evluting the effects of the first mino cids (lysine, rginine, threonine nd methionine) eten post htching (from 0 to 48 h) on subsequent (14 dys of ge) body performnce nd intestinl morphology. This study investigted whether the intke of single mino cid in the erly life could hve positive effects on the mentioned prmeters during the further growth period until dy 14. MATERIALS AND METHODS Animls, mngement nd tretments One hundred nd eighty broiler chicks of either sex from Ross 308 breed were used in this study. Ech bird ws weighed in the htchery nd then trnsported immeditely to the neighbour reserch frm without ny loss of time (less thn 1 h). The chicks were housed in pens 1 1 m of size nd wood shvings flooring. They were rndomly distributed to 6 tretments with 3 equl replictes of 10 chicks per pen. Equl numbes of both sexes were settled in ech tretment groups. The treted groups were ssigned s: 1) No feed nd wter in 48 h of post-htch (fsting group); 2) No feed but free ccess to wter in 48 h of post-htch (wtering group); 3) Intubtion of rginine (120% of NRC; Kidd et l., 2001) in 48 h of post-htch (rginine group); 4) Intubtion of lysine (110% of NRC; Kidd et l., 1997) in 48 h of post-htch (lysine group); 5) Intubtion of methionine (130% of NRC; Ahmed & Abbs, 2011) in 48 h of post-htch ( methionine group); nd 6) Intubtion of threonine (100% of NRC; Kidd et l., 1997) in 48 h of post-htch (threonine group). BJVM, 18, No 4 295

3 Effects of post-htch nutrition of rginine, lysine, methionine nd threonine on performnce. Hndling nd intubtion were done in humne wy ccording to Irnin niml welfre regultions. After 48 h of rering, ll treted groups were fed until 14 dys of ge strter diet bsed on corn/soyben (13 MJ metbolisble energy/kg of diet, 230 g/kg crude protein) formulted to meet requirements for broilers ccording to NRC (1994). Performnce prmeters nd crcss trits Body weights of broilers were recorded post-htch nd fter 14 th dy of ge. Feed intke nd weight gin were recorded t the end of the 14 th dy. The rtio between feed intke nd weight gin ws clculted to offer the feed conversion efficiency (FCR). At the end of the experiment (14 dys of ge), ll chickens were weighed nd killed by decpittion, then the internl orgns (i.e., burs of Fbricius, liver, gizzrd, proventriculus, spleen nd duodenum+pncres) were weighed nd the per cents of orgn weight/body weight (BW) were clculted. Moreover, the length of intestine (duodenum from gizzrd's end to the end of the pncres; jejunum from the end of the pncres to the vitelline diverticulum; ileum from the vitelline diverticulum to the junction of cec; nd rectum from the junction of cec to cloc) were mesured in millimeters using ruler. Morphometric nlysis of the intestine The intestinl morphometric vribles i.e., villus sizes (length, width, surfce re nd lmin propri thickness) were evluted in the duodenum, jejunum nd ileum of killed birds t 14 dys of ge ccording to Skmoto et l. (2000). Briefly, fter fixtion of the mentioned segments of intestine in Clrk solution, ech segment ws divided into 3 sections long its length. Sections were stined by periodic cid Schiff (PAS) regent, then rows of villi were seprted in thickness of the sections, trnsferred over glss slides nd covered with cover-slip. Eyepiece grticules (10 ) nd mgnifiction of 100 were used in microscope to mesure villus sizes in these smples. The villus length nd width were used to clculte villus surfce re ccording to formul π (villus width) (villus length), s described by Skmoto et l. (2000). The lmin propri thickness ws mesured t the spce between bse of the villus nd top of the musculris mucos. This experiment ws done for 30 chickens/tretment. Sttisticl nlysis All dt re offered s men ± SE. Intestinl nd performnce prmeters were compred between chicken groups using one-wy ANOVA with Tukey s post hoc test (SigmStt 3.1. Ink). The P vlues less thn 0.05 were sttisticlly significnt. RESULTS Growth performnce nd crcss trits The men initil body weight ws the sme between different groups of chickens (dt not shown). At the end of the experiment (dy 14), the men body weight of chickens between wtering, fsting, methionine nd threonine groups did not differ while they ll were greter thn lysine group (P<0.05). The feed conversion rtio ws higher in lysine group thn in other groups (P<0.05). The per cent of burs/bw in the fsting group ws higher thn in rginine, lysine, methionine nd threonine groups (P<0.05). This prmeter in lysine group ws lower thn in ll 296 BJVM, 18, No 4

4 A. Sdrzdeh, H. Hssnpour, M. H. Mehrei Hmzekolei, P. Frhdniy, M. R. Ehsn... Tble 1. Effects of post-htch nutrition of different mino cids on body performnce nd reltive orgn weights (%) in broiler chickens (men± SEM, n=30) Fsting group 1 Wtering group 2 Arginine group Lysine group Methionine group Threonine group Body weight (g) 386± b 396± ± b 248± c 380± b 378± b Burs (%) 0.23± ± ± ± ± ±0.010 Pncres ± ± ±0.044 b 1.75± ± ±0.096 duodenum (%) Gizzrd (%) 2.42± ±0.062 b 2.24±0.053 b 2.31± ±0.086 b 2.69±0.059 b Liver (%) 4.46± ±0.229 b 3.95±0.203 b 4.2±0.133 b 4.11±0.157 b 4.3±0.062 Spleen (%) 0.08± ± ± ± ± ±0.007 Proventriculus (%) 0.73± ± ± ± ± ±0.026 Feed conversion rtio,b Mens with different superscripts within the sme row differ (P<0.05); 1 No feed, no wter; 2 No feed but free ccess to wter. Tble 2. Effects of post-htch nutrition of different mino cids on intestinl prmeters in broiler chickens (men± SEM; n=30) Fsting group 1 Wtering group 2 Arginine group Lysine group Methionine group Threonine group Duodenl length (cm) 20.2± ± ± ±1.79 b 19.7± ±0.767 Jejunl length (cm) 47.5± ± ± ± ± ±1.077 Ilel length (cm) 47.1± ± ± ± ± ±1.016 Rectl length (cm) 6.1± ± ± ± ± ±0.199 Duodenl lmin 0.11± ±0.007 b 0.12± ± ±0.08 b 0.21±0.013 b propri (mm) Jejunl lmin propri (mm) Ilel lmin propri (mm) 0.09± ±0.009 b 0.10± ± ±0.017 b 0.14±0.013 b 0.06± ±0.012 b 0.06± ±0.006 b 0.11±0.007 b 0.10±0.011 b,b Mens with different superscripts within the sme row differ (P<0.05); 1 No feed, no wter; 2 No feed but free ccess to wter. BJVM, 18, No 4 297

5 Effects of post-htch nutrition of rginine, lysine, methionine nd threonine on performnce. Threonine c Methionine c Lysine b Arginine Wtering Fsting Villus surfce re (mm 2 ) in duodenum Fig. 1. The villus surfce re in duodenum of broilers t 14 dys of ge, post-htch fed with rginine, lysine, methionine nd threonine, deprived from wter nd food (fsting) or with only ccess to wter (wtering). All dt re presented s men ± SEM (n=30). Mens with different indices differ significntly (P<0.05). other groups (P<0.05). The percent of pncres+duodenum/bw, spleen/bw nd proventriculus/bw in lysine group were lower thn other groups of chickens. The per cent of liver/ BW nd gizzrd/bw were sttisticlly different mong experimentl groups (Tble 1). Intestinl morphometric ssessment In lysine group, the duodenum were shorter thn in other groups of chickens (P<0.05) while the length of jejunum, ileum nd rectum did not differ mong experimentl groups. The lmin propri thickness of duodenum in methionine nd threonine groups were greter thn fsting, rginine nd lysine groups (P<0.05). The lmin propri thickness of jejunum in methionine group ws lso greter thn in fsting, rginine nd lysine groups (P<0.05). The ilel lmin propri thickness ws higher in methionine nd threonine groups thn in fsting nd rginine groups (P<0.05) (Tble 2). The villus surfce re ws lower in the duodenum of lysine group t dy 14 compred to other experimentl groups of chickens (P<0.05), wheres this prmeter in threonine group ws higher thn in other groups (P<0.05; Fig. 1). The villus surfce re ws incresed in the jejunum of threonine group compred to other groups of chickens (P<0.05), lthough significnt vritions between other groups were lso observed (Fig. 2). The villus surfce re in the ileum of fsting nd lysine groups ws lower thn in other groups (P<0.05); the villus surfce re of ileum mong the other groups ws not significnt, lthough this prmeter in threonine group ws higher (Fig. 3). As presented on Fig. 4, the sum of mesured villus surfce re in three intestinl prts t the end of experiment ws higher nd lower in threonine nd lysine groups respectively thn in other groups of chickens (P<0.05). 298 BJVM, 18, No 4

6 A. Sdrzdeh, H. Hssnpour, M. H. Mehrei Hmzekolei, P. Frhdniy, M. R. Ehsn... Threonine b Methionine b Lysine Arginine b Wtering b Fsting Villus surfce re (mm 2 ) in ileum Fig. 2. The villus surfce re in ileum of broilers t 14 dys of ge, post-htch fed with rginine, lysine, methionine nd threonine, deprived from wter nd food (fsting) or with only ccess to wter (wtering). All dt re presented s men ± SEM (n=30). Mens with different indices differ significntly (P<0.05). Threonine d Methionine b Lysine c Arginine b Wtering b Fsting DISCUSSION Mny studies exmined the effects of post-htch nutrition on chickens, which indicted tht feeding in this period of Villus surfce re (mm 2 ) in jejunum Fig. 3. The villus surfce re in jejunum of broilers t 14 dys of ge, post-htch fed with rginine, lysine, methionine nd threonine, deprived from wter nd food (fsting) or with only ccess to wter (wtering). All dt re presented s men ± SEM (n=30). Mens with different indices differ significntly (P<0.05). rering, could imply positive effects on chicken performnce in comprison with feed deprivtion (Gonzles et l., 2003; Mdsen et l., 2004; Bhnj et l., 2010; Noy & Uni, 2010; Mhmoudi et l., 2012). Erlier, it seemed tht nutrients of BJVM, 18, No 4 299

7 Effects of post-htch nutrition of rginine, lysine, methionine nd threonine on performnce. Threonine c Methionine Lysine b Arginine Wtering Fsting Overll villus surfce re (mm 2 ) in intestine Fig. 4. Overll villus surfce re in intestine of broilers t 14 dys of ge, post-htch fed with rginine, lysine, methionine nd threonine, deprived from wter nd food (fsting) or with only ccess to wter (wtering). All dt re presented s men ± SEM (n=30). Mens with different indices differ significntly (P<0.05). yolk including proteins nd crbohydrtes were enough for growth nd developments of chickens during post-htch period. Next studies clered tht erly feeding of chickens with different percent of crude protein, strch, soyben oil or csein could enhnce growth of different pprtus of body such s digestive nd immune systems (Mdsen et l., 2004; Pinheiro et l., 2004; Bhnj et l., 2010; Ydv et l., 2011). In the present study, the broiler chickens were fed 4 mino cids for 48 h, nd their doses were suggested by previous studies. In ddition, two groups of chicken received no feed or just wter for 48 h. After 48 h, ll chickens were fed stndrd diet. At the end of experiments (dy 14), comprison of body weight nd FCR between experimentl groups showed tht feeding of chickens with mino cids not only did not hve ny dvntges in those spects but lso feeding with lysine hd negtive effects. Stringhini et l. (2009) lso reported tht supplement of methionine to the chickens during pre-strter period did not influence weight gin, feed intke nd feed to gin rtio; lthough, complex of methionine, lysine, threonine nd tryptophn improved feed to gin rtio. In the present study, mny internl orgns of body were weighed, in which results were different mong experimentl groups; but it could be deduced tht growth of orgns such s burs, spleen, proventriculus, pncres nd intestine (duodenl length) in lysine group ws slower thn those in other chickens. The reson for this result is not cler. Perhps, the used dose of lysine in this study, intercted with other yolk mino cids. Chmruspollert et l. (2002) found n interreltionship mong lysine, methionine nd rginine. They suggested tht excess mount of lysine could increse rginse ctivity, which reduced the mount of vilble rginine. Excess dietry lysine lso decresed trnsmidinse ctivity leding to diminished cretine biosynthesis in the muscles. On the other 300 BJVM, 18, No 4

8 A. Sdrzdeh, H. Hssnpour, M. H. Mehrei Hmzekolei, P. Frhdniy, M. R. Ehsn... hnd, they reported tht n increse in rginine cused growth nd feed intke depression while incresing methionine in the diet improved body performnce. Despite of our dt on the body performnce nd crcss trits, the effect of the utilized mino cids especilly methionine nd threonine on the intestinl morphology ws noticeble. The thickness of lmin propri in 3 prts of intestine ws incresed when feeding these two mino cids, which ws pprently due to more development of Lieberkühn's glnds in the spce of lmin propri. This result my influence intestinl secretion in chickens. Nir et l. (1996), Pinheiro et l. (2004) nd Noy & Uni (2010) reported tht development of digestive trct plys n individul role in the erly growth of chickens becuse nutrients in post-htch stimulted the production of digestive enzymes. Uni et l. (1998) exmined the effects of erly withholding feed on morphometric prmeters of chick intestine. They observed considerble decrese of villus volume in the jejunum nd n irregulr morphology in jejunl crypts. Scnning electron microscopy lso confirmed some ltertions (included clumping of microvilli nd bnorml crypt structure) in the morphology of intestine in the mentioned chickens. Bigot et l. (2003) nd Fischer d Silv et l. (2007) lso reported tht erly deprivtion delyed the growth of intestine while mino cids such s glutmine incresed intestinl surfce re nd improved growth of intestinl villi. These dt re in greement with our results, which demonstrted tht villus surfce re s index of villus sizes in the intestine ws incresed. This increse indictes better growth of intestinl villi. Apprently, the positive effect of threonine in this spect ws predominnt, which hs been shown well in clculting the overll villus surfce re. On the other hnd, lysine hd no effect or dverse effect on villus surfce re. However, mny studies confirmed tht post-htch nutrition improved the intestinl prmeters (Bigot et l., 2003; Tbeidin et l., 2011; Ydv et l., 2011) but s mentioned bove, the interctions between mino cids could increse body requirements to mny of them. This sitution between lysine, methionine nd rginine hs been confirmed (Chmruspollert et l., 2002) which is probbly the reson of dverse effects on the intestinl development. It must be noticed tht in the htching chick, the smll intestine mtures lrgely in the immedite post-htch dys, which occurs with intensive cryptogenesis, enterocyte mturtion to functionl forms (Geyr et l., 2001). On the other hnd, intestinl development needs mino cids (Foye et l., 2007; Wijtten et l., 2010). Then, ny mlnutrition with mino cids my influence the intestinl development t posthtch period, which would be bckground for diminished growth of intestine in the rering period. It is concluded tht mong the 4 mino cids (rginine, lysine, methionine nd threonine), post-htch feeding of threonine hs beneficil effects especilly on the growth of intestine, lthough its effects on the broiler chicken performnce were not considerble. In contrst, lysine hd dverse effects on the body performnce nd intestinl morphology. ACKNOWLEDGMENTS This work ws supported by the funds wrded by the Vice Chncellor for Reserch of Islmic Azd University. BJVM, 18, No 4 301

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