Proc. of the 3rd Animal Wealth Research Conf. in the Middle East & North Africa IMMUNOMODULATOR EFFECT OF LEVAMISOLE ON BROILERS VACCINATED WITH AVIAN INFLUENZA VACCINE (H5N1) Sabry, E.O.1; Arafa, A.S.2; Ali, M.M.A.3 and Lamya A.F. Attia4 1, 3, 4 Animal Health Research Institute, Benha branch Avian Diseases Dept.1; Chemistry Dept.3 & Virology Dept.4 National Laboratory for Veterinary Quality Control of Poultry Production 2 ABSTRACT T his study was undertaken to investigate the possible implications of levamisole in combination with avian influenza vaccine (H5N1), in broilers, in order to potentiate the immune response developed against that vaccine. One hundred, one-day old apparently healthy Hubbard chicks were used. On 15-day old, the chicks were divided into four equal groups, 25 chicks each. The first group was neither vaccinated nor treated; it was served as a control group. The other groups were vaccinated with inactivated reassortant avian influenza virus vaccine (H5N1 subtype, Re-1 strain) in a dose of 0.5 ml /chick subcutaneously on 15-day old. The second group was vaccinated only (vaccinated non-treated). The third group was vaccinated and treated with levamisole (10 mg\kg body weight) orally, on 16-day old (the second day post vaccination) then daily for five successive days. The fourth group was vaccinated and treated with levamisole (3 mg\kg bogy weight) subcutaneously, on 16-day old (the second day post vaccination) then daily for five successive days. On 42-day old, the levamisole10 and levamisole3 treated groups showed a significant increase of HI titer and bursa of Fabricius weight to body weight ratio when compared with vaccinated only (vaccinated non-treated) and control groups. There were no significant changes among the all treated groups in average spleen weight to body weight ratio. The levamisole10 and levamisole3 treated groups showed improvement in bursal index when compared with vaccinated only group. All treated groups showed spleen hypertrophy. Also, the levamisole10 and levamisole3 treated groups showed a significant increase of serum total proteins, serum albumin, serum globulin and total leucocytic count, on the 42-day old, when compared with vaccinated only (vaccinated non-treated) and control groups. It can be concluded that, levamisole stimulated both humeral and cellular immune response in avian influenza vaccinated chickens. It can be clearly seen from our results that, augmentation of immune response to avian influenza vaccine (H5N1) in chickens could be achieved by administration of levamisole either in drinking water or by subcutaneous injection. Key words: levamisole, Immunomodulator effect, avian influenza (H5N1), broilers vaccinated chickens. Foreign Agricultural Relations (FAR), Egypt, 29th Nov. – 1st Dec., (2010) pp. 384 - 395 IMMUNOMODULATOR EFFECT OF LEVAMISOLE ON BROILERS VACCINATED …… INTRODUCTION Avian influenza (AI) is a viral disease of poultry, human and avian species (Dongzi et. al 2007). AI caused by avian influenza virus (AIV). Influenza viruses belong to the family Orthomyxoviridae and are divided into three types A, B and C on the basis of differences in its nucleoprotein. Influenza A and B are able to cause seasonal epidemics, while all influenza pandemics of the last century have been caused by influenza type A (Nicholson et al., 2003). AIV belongs to the A type viruses which are responsible for major disease problem in birds as well as in humans (Normile, 2005). AI is affecting the respiratory, digestive and/or nervous system of many species of birds. AI causes high morbidity & mortality and reducing the productivity of the affected birds (Swayne et. al, 2000). No treatments for AI exist yet. The availability and use of effective vaccines can be a valuable tool in controlling outbreaks of avian influenza (AI). Numerous vaccines against AI have been developed and shown to be efficacious for the prevention of the disease (Alexander and Brown, 2000). Levamisole is an optic isomer of the phenylimidothiazole salts of tetramisole. It has been shown to have a high level of anthelminthic activity against many parasitic nematodes (Janssen, 1976). It has also been found to possess immunostimulating effects (Renoux and Renoux, 1971). Levamisole has been shown to stimulate cell mediated immunity through the enhanced maturation of cells (van Eygen et al., 1976) or through the enhanced function of macrophages (Kelly, 1978). Levamisole's mechanism of action for its immunostimulating effects is not well understood. It is believed that, it restores cell-mediated immune function in peripheral T lymphocytes and stimulates phagocytosis by monocytes. Under selective condition levamisole can enhance immune responses to viral antigens (Babiuk and Misra, 1981). Levamisole has immunostimulating effect in immunosuppressed animals and reduced the frequency of some diseases (Flesh et al., 1982). Levamisole vaccinated chickens showed higher serum immunoglobulin concentration (Vyas et al., 1987). Also, the chickens injected subcutaneously with levamisole at a dose of 7.5 mg/kg body weight on the second day post vaccination with Hitchner B1 vaccine showed a significant increase of serum total protein and serum protein fractions. (Bastami et al., 1991). This study was undertaken to investigate the possible implications of levamisole in combination with avian influenza vaccine (H5N1), in broilers, in order to potentiate the immune response developed against that vaccine. MATERIAL AND METHODS 1- Animals: One hundred, one-day old apparently healthy Hubbard chicks (male and female) were used. The chicks were purchased from Dakahlia Poultry Company. The chicks were divided into four equal groups (25 chicks each) on 15-day old. 385 Sabry, E.O et al 2- Vaccine: Inactivated reassortant avian influenza virus vaccine (H5N1 subtype, Re-1 strain). The live virus concentration per dose ≥ 108 EID50 / 0.2 ml before inactivation. The vaccine was produced by Yebio Bioengineering Co., China. It was given in a dose of 0.5 ml /chick subcutaneously (s/c) on 15-day old. 3- Drug: Levamisole: it was used in two forms: a- Water soluble powder: (Levamisole "avico"®, Arab Veterinary Industrial Co. (AVICO)). It was given in a dose of 10 mg\kg body weight (Armanious et al., 1991), orally, on 16-day old (the second day post vaccination) with 24 hours interval between doses for five successive days. b- Injectable solution: (Avisole®, Arab Veterinary Industrial Co. (AVICO)) It was given in a dose of 3 mg\kg bogy weight (Chenchev et. al., 1981) subcutaneously (s/c) on 16day old (the second day post vaccination) with 24 hours interval between doses for five successive days. 4- Ration and Water: Birds were fed on commercial standard ration from Cairo Poultry Company. The ration assumed to be balanced. It contained crude protein not less than 21%, crude fat not less than 2.7%, crude fibers not more than 2.7% and metabolizing energy not less than 2950 Kcal/kg ration. The ration and fresh water were offered to the birds ad libitum. 5- Experimental design: One hundred, one-day old apparently healthy Hubbard chicks were used. The chicks were divided into four equal groups on 15-day old, 25 chicks each. The first group was neither vaccinated nor treated; it was served as a control group. The other groups were vaccinated with inactivated reassortant avian influenza virus vaccine (H5N1 subtype, Re-1 strain) in a dose of 0.5 ml /chick (according to the produced company, Yebio) subcutaneously (s/c) on 15-day old. The second group was vaccinated only. The third group was vaccinated and treated with levamisole in a dose of 10 mg\kg body weight, orally, on 16-day old (the second day post vaccination) with 24 hours interval between doses for five successive days. The fourth group was vaccinated and treated with levamisole in a dose of 3 mg\kg body weight subcutaneously (s/c) on 16-day old (the second day post vaccination) with 24 hours interval between doses for five successive days. The design was shown in table (1). 6- Serum Haemagglutination inhibition (HI) antibody assay: The blood samples were collected on the 15th. and 42th. day of age. Blood samples were collected from the jugular vein of five birds of each group. Blood samples were allowed to separate the serum and stored at -20 until use. Haemagglutination inhibition technique was used for detection of antibody level against avian influenza virus as described (Swayne et al 1998) The HI test was a standard beta test using 4 heamagglutinating units of antigen in 96-well plates, where the test serum had been diluted two-fold dilution. HI end point titers were determined as the reciprocal of the highest serum dilution that produced complete inhibition of the haemagglutinating activity. 386 IMMUNOMODULATOR EFFECT OF LEVAMISOLE ON BROILERS VACCINATED …… 7- Determination of immunity indices: Immunity indices included bursal and splenic index. Immunity indices were determined on the 15th. and 42th. day of age. On the 15th day of age, five birds were randomly selected and slaughtered. Bursa of Fabricius and spleen collected, from each slaughtered bird, and separately weighed. On the 42th. day of age, Five birds from each experimental group were randomly selected and slaughtered. Each slaughtered bird was individually weighed. Bursa of Fabricius and spleen collected, from each slaughtered bird, and separately weighed. Immunity indices: According to Lucio and Hitchner (1979): i- Burasl index: Bursal index equal to or less than 0.7 was considered bursal atrophy. ii- Spleen index: The spleen index was lower than the lowest spleen to body weight ratio of the control group was considered spleen atrophy. The spleen index was higher than the highest spleen to body weight ratio of the control group was considered as spleen hypertrophy. 8- Haematological and Biochemical examinations: Blood samples were collected from the jugular vein of five birds of each group. The samples were collected on the 42th. day of age (four weeks post vaccination). Each sample was divided into two parts: the first part was collected in tube containing EDTA (1mg / ml fresh blood) for haematological study which included total leucocytic counts (WBCs) (Schalm et al., 1975). The second part was allowed to separate the serum and kept at -20 ºC till biochemical analysis to determine total protein (Weichselbaum, 1946), albumin (Doumas, 1971), globulins (difference between total protein and albumin) (Coles, 1974) and albumin / globulins ratio (A/G ratio). 9- Statistical analysis: The data was analyzed using one-way ANOVA test according to Snedecor and Cochran (1969) by computerized statistical program (SPSS version 13.0). Data are expressed as the mean ± SE. Duncan's multiple range test was used to determine the difference among various groups according to Duncan (1955). Difference between means was considered significant at P < 0.05. RESULTS 1- Effect of levamisole on humoral immune response (HI titer): Effect of levamisole on humeral immune response (HI titer) of broilers vaccinated with avian influenza vaccine (H5N1) were detected on 15-day old and 42-day old (four weeks post vaccination) and the data were shown in table (2). On 15-day old, the geometric mean of HI titer of detected chicks was (0.31± 0.02). On 42-day old, the levamisole 10 (7.50 ± 0.09) and levamisole 3 (7.62 ± 0.11) treated groups showed a significant increase at (P < 0.05) of HI titer when compared with vaccinated only (6.60 ± 0.08) and control groups (0.00 ± 0.00).. The levamisole 10 (7.50 ± 0.09) treated group not showed any significant changes at (P < 0.05) of HI titer when compared with levamisole 3 (7.62 ± 0.11) treated group.. The vaccinated only group (6.60 ± 0.08) showed a significant increase at (P < 0.05) when compared with control group (0.00 ± 0.00).. 387 Sabry, E.O et al 2- Effect of levamisole on immunity indices: 1- Effect on the average bursa of Fabricius weight to body weight ratio: Effect of levamisole on the average bursa of Fabricius weight to body weight ratio (gm/kg) in broilers vaccinated with avian influenza vaccine (H5N1) were detected on 15-day old and 42-day old (four weeks post vaccination) and the data were shown in table (3). On 42-day old, the levamisole 10 (2.19 ± 0.03) and levamisole 3 (2.16 ± 0.02) treated groups showed a significant increase at (P < 0.05) of the average bursa of Fabricius weight to body weight ratio when compared with vaccinated (1.70 ± 0.01) only and control (1.68 ± 0.04) groups. 2- Effect on the average spleen weight to body weight ratio: Effect of levamisole on average spleen weight to body weight ratio (gm/kg) in broilers vaccinated with avian influenza vaccine (H5N1) were detected on 15-day old and 42day old (four weeks post vaccination) and the data were shown in table (3). On 42-day old, there were no significant changes among the all treated groups in average spleen weight to body weight ratio. 3- Effect of levamisole on bursal index: Effect of levamisole on bursal index in broilers vaccinated with avian influenza vaccine (H5N1) was detected on 42-day old (four weeks post vaccination) and the data were shown in table (4). No bursal atrophy was observed in all treated groups. The levamisole 10 and levamisole 3 treated groups showed improvement in the bursal index when compared with vaccinated only group. 4- Effect of levamisole on splenic index: Effect of levamisole on spleen in broilers vaccinated with avian influenza vaccine (H5N1) was detected on 42-day old (four weeks post vaccination) and the data were shown in table (4). All treated groups showed splenic hypertrophy. 3- Effect on Serum proteins & total leucocytic count: Effect of levamisole on serum proteins and total leucocytic count of broilers vaccinated with avian influenza vaccine (H5N1) were evaluated on 42-day old (four weeks post vaccination) and the data were represented in table (5).. The levamisole 10 and levamisole 3 treated groups showed a significant increase at (P < 0.05) of serum total proteins, serum albumin and serum globulin on 42-day old when compared with vaccinated only and control groups.. All experimental groups (four groups) not showed any significant changes of serum albumin/globulin ratio on the 42-day old. The levamisole 10 and levamisole 3 treated groups showed a significant increase at (P < 0.05) of total leucocytic count on the 42-day old when compared with vaccinated only and control groups.. DISCUSSION This study was undertaken to investigate the possible implications of levamisole in combination with avian influenza vaccine, in broilers, in order to potentiate the immune response developed against that vaccine. 388 IMMUNOMODULATOR EFFECT OF LEVAMISOLE ON BROILERS VACCINATED …… Our results revealed that, the administration of levamisole to avian influenza vaccinated birds either orally (10 mg/kg body weights) or by subcutaneous (s/c) injection (3 mg/kg body weight) on 16-day old (the second day post vaccination) daily for five successive days resulted in potentiation of chicken immune response to avian influenza disease vaccination. This was evidenced by a significant increase in HI-geometeric means in levamisole treated groups (levamisole10 and levamisole3) versus non-treated groups (vaccinated only and control groups). This result was in accordance with that obtained by Kulkarni et al. (1973) and Vyas et al. (1987) who concluded that, levamisole treated groups showed a significant increase in HI-titers than non-treated ones. Also, Goldberg (1979) stated that, levamisole could be used to improve both humeral and cell mediated immune responses for inactivated viral vaccines. On the other hand, the obtained result disagreed with that previously obtained by Chenchev et al. (1981) and Angelov et al. (1984) who reported that, levamisole had no effect on HI-titers. Also, Sanda et al. (2008) studied the effect of levamisole as an immunomodulator in Cockerels vaccinated with Newcastle disease vaccine. They inferred that, levamisole is not an efficient immunomodulator for enhancement of response to Newcastle disease vaccine by chicken. The obtained results showed that, there were significant increases in bursa of Fabricius weight to body weight ratio and spleen weight to body weight ratio in levamisole treated groups (levamisole10 and levamisole3) versus non-treated groups (vaccinated only and control groups). Also levamisole treated birds showed improvement in both bursal and splenic index. The results of estimation of serum total proteins, albumin and globulin revealed higher significant values in levamisole treated groups (levamisole10 and levamisole3) versus non-treated groups (vaccinated only and control groups).. These findings can be supported by those of Soppi et al. (1979) who reported increased immunoglobulin M and G due to levamisole treatment. Vyas et al. (1987) found also higher serum immunoglobulin concentration in levamisole vaccinated chickens. Bastami et al. (1991) inferred that, chickens injected subcutaneously with levamisole at a dose of 7.5 mg/kg body weight on the second day post vaccination with Hitchner B1 vaccine showed a significant increase of serum total protein and serum protein fractions. The levamisole treated groups (levamisole10 and levamisole3) showed a significant increase of total leucocytic count. This result coincided with those obtained by Soppi et al. (1979) and Maheswaran et al. (1980) who concluded that, levamisole enhance cellular immune response in chickens and turkeys. Abd El-Salam and Nourelhuda (1988) concluded that, the routine use of levamisole in poultry management is highly beneficial in terms of its non specific immunomodulation. Bastami et al. (1991) reported that, levamisole treated chickens showed a significant increase of total leucocytic count as well as heterophils, lymphocytes and monocytes. It can be concluded that, levamisole stimulated both humeral and cellular immune response in avian influenza vaccinated chickens. It can be clearly seen from our results that, augmentation of immune response to avian influenza vaccine (H5N1) in chickens could be achieved by administration of levamisole either in drinking water (10 mg/kg body weights) or by s/c injection (3 mg/kg body weight). 389 Sabry, E.O et al REFERENCES Abd El-Salam, E.B. and Nourelhuda, E.M. (1988): "Levamisole-induced resistance to Raillietina tetragona infection in young chicks". Acta Vet. Bron, 57: 147-152. Alexander, D.J. and Brown, I.H. (2000): "Recent zoonoses caused by influenza A viruses". Rev. Sci. Tech., 19(1): 197-225. Angelov, A.T.; Arnaudov, K.H. and Boyadzheva, I. (1984): "Effect of levamisole on antihaemagglutinin, serum sialic acid and lysozome in fowls exposed to aersol of Newcastle disease". Vet. Nouki, 21 (10): 63. Armanious, W.; Gergis, S.M.; Soliman, R.; Rizkallah, E.H. and Kaffagy, A.A.R. (1991): Effect of levamisole on the immune response of chickens to fowl cholera vaccines. J. Egypt. Vet. Med. Ass. 51, No. 1&2, 363-374. Babiuk, I.A. and Misra, V. (1981): "Levamisole and bovine immunity in vitro and in vivo effect on immuno responses to herpes viruses immununization". Can. J. Microbiol.,.27: 1312-1319. Bastami, M.A.; Aziza, M. Amer and El-Kady, M.F. (1991): "Stimulation of chicken immune system to Newcastle disease vaccine by levamisole treatment". Beni-Suef, Vet. Med. Res., 1: 161-169. Chenchev, I.; Koster, G.; Cholankova, R. and Bonovska, M. (1981): "Attempts to stimulate immunity to Newcatle disease by parenteral injection of levamisole". Vet. Nouki, 18 (10): 104. Coles, E.H. (1974): "Determination of globulin". Vet. Clin. Path. 2 nd. Ed., Saunders Company, Philadelphia and London, PP. 560-568. Dongzi, Lin, Jingfang LAN, and Zhizhen ZHANG (2007): Structure and function of the NSI protein of influenza A virus .Acta Biochim Biophys 39:155-162 Doumas, B. (1971): "Colourimetric method for albumin determination". Clin Chem. Acta, 31: 87-92. Duncan, D. R. (1955): "Multiple range and multiple F tests". Biometrics, 11: 31-42. Goldberg, M.E. (1979): Drug profile: Pharmacological and biochemical considerations. Apo Academy of pharmaceutical sciences, Pharmacology and toxicology section. Flesh, J.; Harel, W. and Nelken, D. (1982): "Immunopotentiating effect of levamisole in the prevention of bovine mastitis, fetal death and endometritis". Vet.Rec., 111: 56-57 Janssen, P.A.J., (1976):"The Levamisole story". Prod. Drug Res., 20: 347-383. Kelly,M,T. (1978):Modulation of microphage function by levamisole .J.Reticuloendothol Soc. 24,139. Kulkarni, V.B.; Mulbagal, A.N.; Khet, J.B. and Manda, A.V. (1973): "immunostimulating effect of tetramisole against Newcastle disease virus in chicks". Ind. Vet. J., 50 (3): 225. Lucio, B. and Hitchner, S.B. (1979):"Infectious bursal disease emulsified vaccine: effect upon neutralizing antibody levels in the dam." Avian Diseases, 23: 466-478. Maheswaran, S.K.; Dua, S.K. and Thies, E.S. (1980): "Levamisole induced augmentation of immune response to a live fowl cholera vaccine". poultry Sci. 59: 742-755. Nicholson, K.G; Wood, J.M.; Zambon, M. (2003): "Influenza". Lancet, 362:1733-1745. Normile, D. (2005): "North Korea collaborates to fight bird flu". Science, 308(5719): 175. 390 IMMUNOMODULATOR EFFECT OF LEVAMISOLE ON BROILERS VACCINATED …… Renoux, G. and M. Renoux (1971): "Effect immunostimulant d’un imidothiazole dans l’immunization des souris contre I’infection per Brucella abortus". Cr. Acad. Sci. (D) Paris, 272: 349-350. Sanda, M.E.; Anene, B.M. and Owoade, A. (2008): "Effect of levamisole as an immunomodulator in cockerels vaccinated with Newcatle disease vaccine". Int. J. Poult. Sci., 7(11): 1042-1044. Schalm, O W , Jain, N C and Carroll, E T (1975): "Veterinary haematology" 3rd Ed., Lea and Febiger, Philadelphia, USA. Snedecor, G. W. and Cochran, W. G. (1969): "Statistical methods". Sixth Ed., Iowa State University Press, Ames, Iowa,USA. Soppi,E.; Lassila,O.; Vilzanen, M.k.; Lentonem, O.P. and Eskola, J. (1979): "In vivo effect of Levamisole on cellular and humeral immunity in normal chickens". Clinical and experimental immunology, 38 (3): 609-614. Swayne, D.E.; D.A. Senne, and C.W. Beard (1998): "Influenza: Isolation and identification of avian pathogens, 4th. ed. D.E. Swayne, J.R. Glisson, M. W. Jackwood, J.E. Pearson, and W.M. Reed, eds. American Association of Avian Pathologists, Kennett Square, PA. pp. 150-155. Swayne, D.E.; Joan R. Beck, Micheal L. Perdue, and Charles W. Beard (2000): Efficacy of vaccines in chickens against highly pathogenic Hong Kong H5N1 Avian influenza, Avian diseases, 45:455-365. Van Eygen, M,.Znamensky, P.Y., Heck,E. and Raymaekers,I. (1976): Levamisole in prevention of recurrent upper respiratory tract infection in children .Lancet, I,380. Vyas, G.P.; Dholakia, P.M. and Kathria, L.G. (1987): "Studies on immunomodulation by levamisole along with vaccination in chicks against Ranikhet Disease". Ind. Vet. J., 64 (6): 456-462. Weichselbaum, P.E. (1946): "Colourimetric determination of total protein". Am. J. Clin. Path, 16: 40-46. 391 Sabry, E.O et al Table (1): Experimental design Experimental groups Group No. 1 2 3 4 No. of chicks/group 25 25 25 25 Vaccine (H5N1) + + + Levamisole 10 Levamisole 3 + - + Experimental groups' name Group's number 1 2 3 4 Group's name Control (non vaccinated non treated) Vaccinated only (vaccinated non treated) Levamisole 10 (vaccinated & treated with levamisole 10 mg/k.b.w1 orally for five successive days) Levamisole 3 (vaccinated & treated with levamisole 3 mg/k.b.w subcutaneously for five successive days) Treatments Dose Interval bet. Doses Duration of treatment Route of administration Time of administration Type Vaccine (H5N1) 0.5 ml /chick Non One injection S/C2 15-day old H5N1 Levamisole 10 Levamisole 3 10 mg/k.b.w 24 hours (daily) 5 successive days Orally 16-day old4 --- 3 mg/k.b.w 24 hours (daily) 5 successive days S/C 16-day old --- Sampling Parameter HI7 test Serum proteins Total leucocytic count Immunity indices Number of samples /group 5 5 5 5 1 Time of sampling 15-day old5 & 42-day old6 42-day old 42-day old 15-day old & 42-day old 2 k.b.w = kilogram body weight 16-day old = on the second day post vaccination 5 15-day old = just before the vaccination 6 42-day old = four weeks post vaccination 7 HI = Haemagglutination inhibition 4 392 S/C = Subcutaneously IMMUNOMODULATOR EFFECT OF LEVAMISOLE ON BROILERS VACCINATED …… Table (2): Effect of levamisole on humoral immune response (HI titer) of broilers vaccinated with avian influenza vaccine (H5N1). (n = 5, Mean ± SE) Geometeric mean titer (GMT) of HI Groups Control Duration Vaccinated only 0.31 15-day old 0.00 c ± 0.00 42-day old Levamisole 10 ± 0.02 7.50 a ± 0.09 6.60 b ± 0.08 Levamisole 3 7.62 a ± 0.11 Data were analyzed by one-way ANOVA Means with different alphabetical superscripts in the same row are significantly different at P ≤ 0.05 Table (3): Effect of levamisole on the average bursa of Fabricius & spleen weight to body weight ratio (gm/kg) in broilers vaccinated with avian influenza vaccine (H5N1). (n = 5, Mean ± SE) Average bursa of Fabricius & spleen weight to body weight ratio (g/kg) Groups Duration organ Control Vaccinated only Bursa 15-day old 42-day old 2.27 ± 0.05 0.77 ± 0.03 2.19 a ± 0.03 0.90 a ± 0.07 Spleen Bursa Spleen 1.68 b ± 0.04 0.91 a ± 0.02 Levamisole 10 1.70 b ± 0.01 0.92 a ± 0.04 Levamisole 3 2.16 a ± 0.02 0.95 a ± 0.06 Data were analyzed by one-way ANOVA Means with different alphabetical superscripts in the same row are significantly different at P ≤ 0.05 393 Sabry, E.O et al Table (4): Effect of levamisole on the bursal & splenic indices in broilers vaccinated with avian influenza vaccine (H5N1). (n = 5, Mean ± SE) Bursal and splenic indices Duration 42-day old Organ Bursa Spleen Groups Vaccinated only 1.03 1.01 Levamisole 10 1.33 0.99 Levamisole 3 1.31 1.04 The judgment of the results occurred according to Lucio and Hitchner (1979) as follows: The bursa of fabricius considered to be atrophied when bursal index become equal to or less than 0.7 The splenic index was lower than the lowest spleen to body weight ratio of the control group (0.77) was considered spleen atrophy. The splenic index was higher than the highest spleen to body weight ratio of the control group (0.91) was considered as spleen hypertrophy. Table (5): Effect of levamisole on serum proteins & total leucocytic count of broilers vaccinated with avian influenza vaccine (H5N1). (n = 5, Mean ± SE) Serum proteins & total leucocytic count Parameter Total Protein (gm/dl) Serum albumin (gm/dl) Serum globulin (gm/dl) A/G ratio WBCs (cell x 103/ml) Groups Control 4.28 b ± 0.06 1.55 b ± 0.04 2.74 b ± 0.07 0.57 a 0.01 19.30 b ± 0.14 Vaccinated only 4.14 b ± 0.04 1.48 b ± 0.03 2.66 b ± 0.04 0.56 a 0.03 19.09 b ± 0.24 Levamisole 10 4.75 a ± 0.15 1.70 a ± 0.02 3.05 a ± 0.08 0.56 a ± 0.02 21.08 a ± 0.37 Levamisole 3 4.85 a ± 0.21 1.72 a ± 0.06 3.13 a ± 0.07 0.55 a ± 0.01 21.44 a ± 0.43 A/G ratio = Albumin/Globulin ratio Data were analyzed by one-way ANOVA Means with different alphabetical superscripts in the same row are significantly different at P ≤ 0.05. 394 ‫…… ‪IMMUNOMODULATOR EFFECT OF LEVAMISOLE ON BROILERS VACCINATED‬‬ ‫ﺗﺄﺛﯿﺮ اﻟﻠﯿﻔﺎﻣﯿﺰول اﻟﻤﺤﻔ ﺰ ﻟﻠﻤﻨﺎﻋ ﺔ ﻋﻠ ﻰ دﺟ ﺎج اﻟﻠﺤ ﻢ اﻟﻤﺤﺼ ﻦ ﺑﻠﻘ ﺎح أﻧﻔﻠ ﻮﻧﺰا اﻟﻄﯿ ﻮر‬ ‫)‪(H5N1‬‬ ‫ﺻﺒﺮي اﻟﺴﯿﺪ ﻋﻤﺮ ﺳﺎﻟﻢ‪ ،1‬ﻋﻠﻲ ﻣﺤﻤﺪ ﻣﺤـﻤﺪ أﺣﻤـﺪ‪، 3‬ﻋﺒﺪ اﻟﺴﺘﺎر ﻋﺮﻓﺔ ﻣﺤﻤﺪ‪ 2‬وﻟﻤﯿ ﺎء ﻋﻄﯿ ﮫ ﻣﺤﻤ ﺪ‬ ‫‪4‬‬ ‫ﻓﺘﺤﻲ‬ ‫‪1‬‬ ‫‪431‬‬ ‫ﻣﻌﮭﺪ ﺑﺤﻮث ﺻﺤﺔ اﻟﺤﯿﻮان ‪ -‬ﻓﺮع ﺑﻨﮭﺎ ﻗﺴﻢ أﻣﺮاض اﻟﺪواﺟﻦ ‪ ،‬ﻗﺴﻢ اﻟﻜﯿﻤﯿﺎء اﻟﺤﯿﻮﯾﺔ واﻟﺴ ﻤﻮم‬ ‫‪4‬‬ ‫وأﻣﺮاض اﻟﻨﻘﺺ اﻟﻐﺬاﺋﻰ‪ ،3‬ﻗﺴﻢ اﻟﻔﯿﺮوﻟﻮﺟﻲ‬ ‫‪2‬‬ ‫اﻟﻤﻌﻤﻞ اﻟﻘﻮﻣﻲ ﻟﻠﺮﻗﺎﺑﺔ اﻟﺒﯿﻄﺮﯾﺔ ﻋﻠﻰ اﻹﻧﺘﺎج اﻟﺪاﺟﻨﻲ‬ ‫اﻟﮭﺪف ﻣﻦ ھﺬه اﻟﺪارﺳﺔ ھﻮ ﻣﺤﺎوﻟﺔ ﻣﻌﺮﻓﺔ ﺗ ﺄﺛﯿﺮ اﺳ ﺘﺨﺪام دواء اﻟﻠﯿﻔ ﺎﻣﯿﺰول ﻟﺰﯾ ﺎدة اﻹﺳ ﺘﺠﺎﺑﺔ‬ ‫اﻟﻤﻨﺎﻋﯿ ﺔ ﻟﻠﻘ ﺎح أﻧﻔﻠ ﻮﻧﺰا اﻟﻄﯿ ﻮر )‪ (H5N1‬ﻓ ﻲ دﺟ ﺎج اﻟﻠﺤ ﻢ‪ .‬اﺟﺮﯾ ﺖ ھ ﺬه اﻟﺪراﺳ ﺔ ﻋﻠ ﻰ ﻋ ﺪد ‪100‬‬ ‫ﻛﺘﻜﻮت ﻋﻤﺮ ﯾﻮم ﻣﻦ ﻧﻮع اﻟﮭﺎﺑﺮد‪ .‬ﻗﺴﻤﺖ ھﺬه اﻟﻜﺘﺎﻛﯿﺖ إﻟﻰ أرﺑﻌﺔ ﻣﺠﻤﻮﻋﺎت ﻣﺘﺴﺎوﯾﺔ ﻛﻞ ﻣﻨﮭ ﺎ ﺑ ﮫ‬ ‫‪ 25‬ﻛﺘﻜ ﻮت‪ .‬اﺳ ﺘﺨﺪﻣﺖ اﻟﻤﺠﻤﻮﻋ ﺔ اﻷوﻟ ﻲ ﻛﻀ ﺎﺑﻄﺔ ﺳ ﺎﻟﺒﺔ ﻟﻠﺘﺠﺮﺑ ﺔ )ﻏﯿ ﺮ ﻣﺤﺼ ﻨﺔ وﻏﯿ ﺮ ﻣﻌﺎﻟﺠ ﺔ‬ ‫ﺑﺪواء اﻟﻠﯿﻔﺎﻣﯿﺰول(‪ .‬ﺑﺎﻗﻲ اﻟﻤﺠﻤﻮﻋ ﺎت ﺗ ﻢ ﺗﺤﺼ ﯿﻨﮭﺎ ﺑﻠﻘ ﺎح أﻧﻔﻠ ﻮﻧﺰا اﻟﻄﯿ ﻮر )‪ (H5N1‬ﺑﺠﺮﻋ ﺔ ‪0.5‬‬ ‫ﻣﻠ ﻲ‪/‬ﻛﺘﻜ ﻮت ﺗﺤ ﺖ اﻟﺠﻠ ﺪ ﻋﻨ ﺪ ﻋﻤ ﺮ ‪ 15‬ﯾ ﻮم‪ .‬اﻟﻤﺠﻤﻮﻋ ﺔ اﻟﺜﺎﻧﯿ ﺔ ﻣﺤﺼ ﻨﺔ ﺑﻠﻘ ﺎح أﻧﻔﻠ ﻮﻧﺰا اﻟﻄﯿ ﻮر‬ ‫)‪ (H5N1‬ﻓﻘ ﻂ‪ .‬اﻟﻤﺠﻤﻮﻋ ﺔ اﻟﺜﺎﻟﺜ ﺔ ﻣﺤﺼ ﻨﺔ ﺑﻠﻘ ﺎح أﻧﻔﻠ ﻮﻧﺰا اﻟﻄﯿ ﻮر )‪ (H5N1‬وﻣﻌﺎﻟﺠ ﺔ ﺑ ﺪواء‬ ‫اﻟﻠﯿﻔﺎﻣﯿﺰول ﺑﺠﺮﻋﺔ ‪ 10‬ﻣﻠﯿﺠﺮام‪/‬ﻛﯿﻠﻮﺟﺮام ﻣﻦ وزن ﺟﺴﻢ اﻟﻄ ﺎﺋﺮ ﻓ ﻲ ﻣﯿ ﺎه اﻟﺸ ﺮب ﻓ ﻲ اﻟﯿ ﻮم اﻟﺜ ﺎﻧﻲ‬ ‫ﻟﻠﺘﺤﺼﯿﻦ‪ ،‬وﻟﻤﺪة ﺧﻤﺴﺔ أﯾﺎم ﻣﺘﺘﺎﻟﯿ ﺔ‪ .‬اﻟﻤﺠﻤﻮﻋ ﺔ اﻟﺮاﺑﻌ ﺔ ﻣﺤﺼ ﻨﺔ ﺑﻠﻘ ﺎح أﻧﻔﻠ ﻮﻧﺰا اﻟﻄﯿ ﻮر )‪(H5N1‬‬ ‫وﻣﻌﺎﻟﺠﺔ ﺑﺪواء اﻟﻠﯿﻔﺎﻣﯿﺰول ﺑﺠﺮﻋﺔ ‪ 3‬ﻣﻠﯿﺠﺮام‪/‬ﻛﯿﻠﻮﺟﺮام ﻣﻦ وزن ﺟﺴﻢ اﻟﻄ ﺎﺋﺮ وذﻟ ﻚ ﺑ ﺎﻟﺤﻘﻦ ﺗﺤ ﺖ‬ ‫اﻟﺠﻠﺪ ﻓﻲ اﻟﯿﻮم اﻟﺜﺎﻧﻲ ﻟﻠﺘﺤﺼﯿﻦ‪ ،‬وﻟﻤﺪة ﺧﻤﺴﺔ أﯾﺎم ﻣﺘﺘﺎﻟﯿﺔ‪.‬‬ ‫ﻗﺪ أﻇﮭﺮت اﻟﻨﺘﺎﺋﺞ أن اﺳﺘﺨﺪام اﻟﻠﯿﻔ ﺎﻣﯿﺰول )ﻓ ﻲ ﻣﯿ ﺎه اﻟﺸ ﺮب ﺑﺠﺮﻋ ﺔ ‪ 10‬ﻣﻠﯿﺠ ﺮام‪/‬ﻛﯿﻠ ﻮﺟﺮام‬ ‫ﻣﻦ وزن ﺟﺴﻢ اﻟﻄﺎﺋﺮ أو ﺑﺎﻟﺤﻘﻦ ﺗﺤ ﺖ اﻟﺠﻠ ﺪ ﺑﺠﺮﻋ ﺔ ‪ 3‬ﻣﻠﯿﺠ ﺮام‪/‬ﻛﯿﻠ ﻮﺟﺮام ﻣ ﻦ وزن ﺟﺴ ﻢ اﻟﻄ ﺎﺋﺮ(‬ ‫ﯾﺰﯾ ﺪ اﻹﺳ ﺘﺠﺎﺑﺔ اﻟﻤﻨﺎﻋﯿ ﺔ ﻟﻠﻄﯿ ﻮر اﻟﻤﺤﺼ ﻨﺔ ﺑﻠﻘ ﺎح أﻧﻔﻠ ﻮﻧﺰا اﻟﻄﯿ ﻮر وﺗﺠﻠ ﻲ ھ ﺬا ﻣ ﻦ ﺧ ﻼل اﻟﺰﯾ ﺎدة‬ ‫اﻟﻤﻌﻨﻮﯾ ﺔ ﻓ ﻲ ﻣﺴ ﺘﻮﯾﺎت اﻷﺟﺴ ﺎم اﻟﻤﻨﺎﻋﯿ ﺔ وزﯾ ﺎدة ﻧﺴ ﺒﺔ اﻟﺒ ﺮوﺗﯿﻦ اﻟﻜﻠ ﻲ وﻧﺴ ﺒﺔ اﻷﻟﺒ ﻮﻣﯿﻦ وﻧﺴ ﺒﺔ‬ ‫اﻟﺠﻠﻮﺑﯿﻮﻟﯿﻦ ﻓﻲ ﻣﺼﻞ اﻟﻄﯿﻮر‪ .‬ﻛﺬﻟﻚ أﻇﮭﺮ اﺳﺘﺨﺪام اﻟﻠﯿﻔﺎﻣﯿﺰول زﯾﺎدة ﻣﻌﻨﻮﯾﺔ ﻓ ﻲ ﻧﺴ ﺒﺔ وزن ﻛ ﯿﺲ‬ ‫ﻓﺎﺑﺮﯾﺸ ﯿﺲ إﻟ ﻰ وزن ﺟﺴ ﻢ اﻟﻄ ﺎﺋﺮ‪ .‬ﻛ ﺬﻟﻚ أﻇﮭ ﺮت اﻟﻤﺠﻤﻮﻋ ﺎت اﻟﻤﻌﺎﻟﺠ ﺔ ﺑ ﺪواء اﻟﻠﯿﻔ ﺎﻣﯿﺰول زﯾ ﺎدة‬ ‫ﻣﻌﻨﻮﯾﺔ ﻓﻲ اﻟﻌﺪد اﻟﻜﻠﻲ ﻟﻜﺮات اﻟﺪم اﻟﺒﯿﻀﺎء‪.‬‬ ‫ﻣﻦ ھﺬه اﻟﻨﺘﺎﺋﺞ ﯾﺘﻀﺢ أن اﺳﺘﺨﺪام اﻟﻠﯿﻔﺎﻣﯿﺰول )ﻓﻲ ﻣﯿﺎه اﻟﺸﺮب ﺑﺠﺮﻋﺔ ‪ 10‬ﻣﻠﯿﺠﺮام‪/‬ﻛﯿﻠ ﻮﺟﺮام‬ ‫ﻣ ﻦ وزن ﺟﺴ ﻢ اﻟﻄ ﺎﺋﺮ أو ﺑ ﺎﻟﺤﻘﻦ ﺗﺤ ﺖ اﻟﺠﻠ ﺪ ﺑﺠﺮﻋ ﺔ ‪ 3‬ﻣﻠﯿﺠ ﺮام‪/‬ﻛﯿﻠ ﻮﺟﺮام ﻣ ﻦ وزن ﺟﺴ ﻢ اﻟﻄ ﺎﺋﺮ(‬ ‫ﯾﺰﯾﺪ اﻹﺳﺘﺠﺎﺑﺔ اﻟﻤﻨﺎﻋﯿﺔ ﻟﻠﻄﯿﻮر اﻟﻤﺤﺼﻨﺔ ﺑﻠﻘﺎح أﻧﻔﻠﻮﻧﺰا اﻟﻄﯿﻮر )‪.(H5N1‬‬ ‫‪395‬‬ ‫‪View publication stats‬‬