• Ramadan S Shewita
  • Karima El-Naggar
  • Walaa S Abd El Naby



This study was conducted to evaluate the effects of dietary supplementation of different levels of vitamin C (VC) in broiler chicken reared under high stocking density on the growth performance, blood biochemical parameters and the expression of heat shock protein genes. A total of 150, one day old mixed sex broiler chicks (Cobb 500) were randomly distributed in five equal groups. Group1 was reared in normal stocking density (10.6 birds/m2) and fed on the basal diet (BD) without VC (control). While, chicks in other groups (2, 3, 4 and 5) reared in high stocking density (15.6 birds/m2) and fed on BD supplemented with VC at 0, 200, 400 and 600 mg / kg diet, respectively. Birds reared in high stocking density showed a reduction of the final body weight and total feed intake, with high mortality (6.6%). Moreover, they revealed a significant up-regulation of HSP70 mRNA and elevated HSP90 and HSF1 mRNA expression in heart and liver tissue. Graded dietary levels of VC provided variable protection against the hazard of high density through improved final body weight and total feed intake, decreased the mortality % and downregulated liver HSP70 expression level. However, the best performance was observed in birds supplemented with 200 mg/kg VC (group 3).

Key words: chicks' growth performance; gene expression; stocking density; vitamin C


Thaxton JP, Dozier WA, 3rd, Branton SL, Morgan GW, Miles DW, Roush WB, et al.Stocking density and physiological adaptive responses of broilers. Poult Sci 2006; 85(5): 819–24.

Pandurang LT, Kulkarni GB, Gangane GR, More PR, Ravikanth K, Maini S, et al.Overcrowding Stress Management in Broiler Chicken with Herbal Antistressor. IJAS 2011; 1(1): 49–55.

Whitehead CC, Keller T.An update on ascorbic acid in poultry. World's Poult Sci J 2003; 59(2): 161–84.

Zhang AW, Lee BD, Lee SK, Lee KW, An GH, Song KB, et al.Effects of yeast (Saccharo-myces cerevisiae) cell components on growth performance, meat quality, and ileal mucosa development of broiler chicks. Poult Sci 2005; 84(7): 1015–21.

Campo JL, Dávila S.Changes in heterophil to lymphocyte ratios of heat-stressed chickens in response to dietary supplementation of sev-eral related stress agents. Arch Fur Geflugelk 2002; 66: 80–4.

Ipek H, Avci M, Yerturk M, Iriadam M, Aydilek N.Effects of ascorbic acid and vitamin E on performance and hematological parame-ters of Japanese quails under high ambient tem-perature in Sanliurfa. Arch Fur Geflugelk 2007; 71: 130–4.

Lin H, Jiao HC, Buyse J, Decuypere E.Strategies for preventing heat stress in poultry. World's Poult Sci J 2007; 62(1): 71–86.

Al-Aqil A, Zulkifli I.Changes in heat shock protein 70 expression and blood charac-teristics in transported broiler chickens as af-fected by housing and early age feed restriction. Poult Sci 2009; 88(7): 1358–64.

Zulkifli I, Liew PK, Israf DA, Omar AR, Hair-Bejo M.Effects of early age feed re-striction and heat conditioning on hetero-phil/lymphocyte ratios, heat shock protein 70 expression and body temperature of heat-stressed broiler chickens. J Therm Biol 2003; 28(3): 217–22.

Jacquier-Sarlin MR, Fuller K, Dinh-Xuan AT, Richard MJ, Polla BS.Protective effects of hsp70 in inflammation. Experientia 1994; 50(11-12): 1031–8.

Jackson SE.Hsp90: structure and func-tion. Top Curr Chem 2013; 328: 155–240.

NRC, Nutrient requirements of poultry. 9th Ed. Washington, DC:National Academic Press,1994.

Houshmand M, Azhar K, Zulkifli I, Bejo MH, Kamyab A.Effects of prebiotic, protein level, and stocking density on performance, immunity, and stress indicators of broilers. Poult Sci 2012; 91(2): 393–401.

Rao X, Huang X, Zhou Z, Lin X.An im-provement of the 2^(-delta delta CT) method for quantitative real-time polymerase chain re-action data analysis. Biostat, Bioinforma Bi-omath 2013; 3(3): 71–85.

Xie J, Tang L, Lu L, Zhang L, Xi L, Liu HC, et al.Differential expression of heat shock transcription factors and heat shock proteins after acute and chronic heat stress in laying chickens (Gallus gallus). PloS one 2014; 9(7): e102204.

Nerren JR, Swaggerty CL, MacKinnon KM, Genovese KJ, He H, Pevzner I, et al.Differential mRNA expression of the avian-specific toll-like receptor 15 between hetero-phils from Salmonella-susceptible and -resistant chickens. Immunogenetics 2009; 61(1): 71–77.

SAS, Statistical Analysis System North Carolina:Users Guide Statistics. As. Institute Cary,1996.

Dozier WA, 3rd, Thaxton JP, Branton SL, Morgan GW, Miles DM, Roush WB, et al.Stocking density effects on growth perfor-mance and processing yields of heavy broilers. Poult Sci 2005; 84(8): 1332–8.

Elagib-Hind AA, Omer HM.Effect of Di-etary Ascorbic Acid on Performance and Im-mune Response of Heat Stressed Broiler Chicks. Pakistan J Nutr 2012; 11(3): 216–20.

SabahElkheir MK, Mohammed AMM, Abdel-Gadir SM.Effect of Feed Restriction and Ascorbic Acid Supplementation on Perfor-mance of Broiler Chicks Reared under Heat Stress. Res J Ani & Vet Sci 2008; 3: 1–8.

Sahin K, Kucuk O.Effects of vitamin E and selenium on performance, digestibility of nutrients, and carcass characteristics of Japa-nese quails reared under heat stress (34 degrees C). J anim Physiol Anim Nutr 2001; 85(11-12): 342–8.

Lohakare JD, Ryu MH, Hahn TW, Lee JK, Chae BJ.Effects of Supplemental Ascorbic Acid on the Performance and Immunity of Commercial Broilers. J Appl Poult Res 2005; 14(1): 10–19.

Blaha J, Kroesna K.Effect of vitamin and electrolytes supplements on broiler perfor-mance, slaughter value and chemical composi-tion of meat during the heat stress. Universitäs Agriculturäe Praga Press 1997; 30: 103–13.

Jaffar GH, Blaha J.Effect of ascorbic ac-id supplementation in drinking water on growth rate, feed consumption and feed efficiency of broiler chickens maintained under acute heat stress conditions. Universitas Agriculturae Praga Press 1996; 41: 485–90.

McKee JS, Harrison PC.Effects of sup-plemental ascorbic acid on the performance of broiler chickens exposed to multiple concurrent stressors. Poult Sci 1995; 74(11): 1772–85.

Amady V, Tayb I, Yokhana J. Effects of supplemental ascorbic acid on humeral immune response in broilers reared under heat stress conditions. Res Opinions Anim Veter Sci. 2011; 1: 459–62.

Amakye-Anim J, Lin TL, Hester PY, Thiagarajan D, Watkins BA, Wu CC.Ascorbic acid supplementation improved antibody re-sponse to infectious bursal disease vaccination in chickens. Poult Sci 2000; 79(5): 680–8.

Simsek UG, Dalkilic B, Ciftci M, Yuce A.The Influences of Different Stocking Densi-ties on Some Welfare Indicators, Lipid Peroxi-dation (MDA) and Antioxidant Enzyme Activi-ties (GSH, GSH-Px, CAT) in Broiler Chickens. J Anim Vet Adv 2009; 8(8): 1568–72.

Al-Darajji H, Al-Mashhadanp E, Al-Atharp A.Effect of ascorbic acid supplementa-tion in the diets on haematological traits of Fawbro broiler breeders reared under hot cli-mate Indian J Anim Sci. 2001; 71(9): 857–9.

Kucuk O, Sahin K, Sahin N, Gursu M, Gulcu F, Ozcelik M, et al. Egg production, egg quality, and lipid peroxidation status in laying hens maintained at a low ambient temperature (6 deg C) and fed a vitamin C and vitamin E-supplemented diet. Vet Med 2003; 48: 33–40.

Najafi P, Zulkifli I, Jajuli NA, Farjam AS, Ramiah SK, Amir AA, et al. Environmental temperature and stocking density effects on acute phase proteins, heat shock protein 70, circulating corticosterone and performance in broiler chickens. Int J Biometeorol 2015; 59(11): 1577–83.

Beloor J, Kang HK, Kim YJ, Subramani VK, Jang IS, Sohn SH, et al.The Effect of Stocking Density on Stress Related Genes and Telomeric Length in Broiler Chickens. Asian-Aust J Anim Sci 2010; 23(4): 437–43.

Basu N, Todgham AE, Ackerman PA, Bibeau MR, Nakano K, Schulte PM, et al.Heat shock protein genes and their functional signifi-cance in fish. Gene 2002; 295(2): 173–83.

Gornati R, Papis E, Rimoldi S, Terova G, Saroglia M, Bernardini G.Rearing density influ-ences the expression of stress-related genes in sea bass (Dicentrarchus labrax, L.). Gene 2004; 341: 111–8.

Aksakal E, Ekinci D, Erdoğan O, Bey-demir Ş, Alım Z, Ceyhun SB.Increasing stock-ing density causes inhibition of metabolic–antioxidant enzymes and elevates mRNA levels of heat shock protein 70 in rainbow trout. Livest Sci 2011; 141(1): 69–75.

Craig EA, Gross CA.Is hsp70 the cellular thermometer? Trends in Biochem Sci 1991; 16: 135–40.

Parsell DA, Lindquist S.The function of heat-shock proteins in stress tolerance: degrada-tion and reactivation of damaged proteins. An-nu Rev Genet 1993; 27: 437–96.

Sahin K, Sahin N, Onderci M, Gursu MF, Issi M.Vitamin C and E can alleviate negative effects of heat stress in Japanese quails. Food , Agric Envir 2003; 1(2): 244-–9.

Mahmoud KZ, Edens FW, Eisen EJ, Ha-venstein GB.Ascorbic acid decreases heat shock protein 70 and plasma corticosterone response in broilers (Gallus gallus domesticus) subjected to cyclic heat stress. Comp Biochem Physiol B Biochem Mol Biol 2004; 137(1): 35–42.

Jang IS, Ko YH, Moon YS, Sohn SH.Effects of Vitamin C or E on the Pro-inflammatory Cytokines, Heat Shock Protein 70 and Antioxidant Status in Broiler Chicks under Summer Conditions. Asian-Australas J Anim Sci 2014; 27(5): 749–56.

Sahin N, Tuzcu M, Orhan C, Onderci M, Eroksuz Y, Sahin K.The effects of vitamin C and E supplementation on heat shock protein 70 response of ovary and brain in heat-stressed quail. Br poult sci 2009; 50(2): 259–65.

Khassaf M, McArdle A, Esanu C, Va-silaki A, McArdle F, Griffiths RD, et al.Effect of vitamin C supplements on antioxidant de-fence and stress proteins in human lymphocytes and skeletal muscle. J Physiol 2003; 549(Pt 2): 645–52.

Fangue NA, Hofmeister M, Schulte PM.Intraspecific variation in thermal tolerance and heat shock protein gene expression in common killifish, Fundulus heteroclitus. J Exp Biol 2006; 209(Pt 15): 2859–72.

Copat C, Brundo MV, Arena G, Grasso A, Oliveri Conti G, Ledda C, et al.Seasonal var-iation of bioaccumulation in Engraulis encra-sicolus (Linneaus, 1758) and related biomarkers of exposure. Ecotoxicol Environ Saf 2012; 86: 31–7.

Yu J, Bao E, Yan J, Lei L.Expression and localization of Hsps in the heart and blood vessel of heat-stressed broilers. Cell stress chaperones 2008; 13(3): 327–35.

Kim M, Ahn IY, Kim H, Cheon J, Park H.Molecular characterization and induction of heat shock protein 90 in the Antarctic bivalve Laternula elliptica. Cell stress chaperones 2009; 14(4): 363–70.

Gu XH, Hao Y, Wang XL.Overexpression of heat shock protein 70 and its relationship to intestine under acute heat stress in broilers: 2. Intestinal oxidative stress. Poult Sci 2012; 91(4): 790–9.




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Veterinary Medicine and The One Health Concept