Microsoft Word - د.عامر خزعل -42 Alazawy Diyala Agricultural Sciences Journal, 5( 2 ) 25 – 37 ,2013 25   SEROLOGICAL STUDY BY USING THE ELISA TECHNIQUE TO IDENTIFICATION OF AVIAN INFECTIOUS BRONCHITIS DISEASE AMONG SOME FIELDS OF BROILER CHICKENS IN DIYALA PROVINCE. Amer Alazawy *Department of Pathology and Avian disease. - College of Veterinary Medicine - University of Diyala Amer_alazawy@yahoo.com ABSTRACT Infectious bronchitis (IB) is a common highly contagious viral disease of poultry; it causes major economic losses to the poultry industry. This study was conducted to investigate the prevalence of avian infectious bronchitis (IB) by evaluated the maternal derived antibodies (MDA) at 3 days of age and compared with the antibodies against the disease in 25 days of age, also to study the clinical signs and gross lesions of the suspected infected birds in some broiler chickens farms of Diyala governorate during the period from September 2012 to April 2013. Twenty broiler chicken flocks were reported in this study, blood samples were collected from each flocks two times to investigate the antibody titers by using Enzyme- Linked Immunosorbent Assay (ELISA). Significant variation (P<0.05) was observed in the means of antibody titers against IBV between the maternal derived antibodies at 3 days old and 25 days old, also 14 farms showed clinical signs of IB including mainly, severe respiratory signs with high morbidity and mortality, postmortem examination of affected flocks showed lesions mainly in trachea and kidney. The result of this study showed that IB was the most prevalence and widespread disease in broiler chickens farm particularly at 3 weeks of age. Therefore, because of the lack of a specific vaccine against endemic strains of IBV in Iraq, IB infection has remained a problem in the Diyala poultry farms. Key words: Infectious bronchitis, ELISA, maternal derived antibodies, Diyala. INTRODUCTION Avian infectious bronchitis virus (IBV) is a highly infectious pathogen of domestic fowl that replicates primarily in the respiratory tract but also in epithelial cells of ـــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــ Received for publication Sept. 10 , 2013 . Accepted for publication Nov. 10 , 2013 . Alazawy Diyala Agricultural Sciences Journal, 5( 2 ) 25 – 37 ,2013 26   the gut, kidney, and oviduct causes a highly contagious respiratory and sometimes urogenital disease of chickens that is characterized by respiratory signs, nephritis, or reduced egg production and quality in layer chickens (Cavanagh and Naqi, 1997). Avian infectious bronchitis (IB) was first described in the United States of America (USA) in the 1930s as an acute respiratory disease mainly of young chickens (Li et al., 2009). The virus is a member of the genus Coronavirus, family Coronaviridae, order Nidovirales. The genome of IBV is a single-stranded, linear positive sense RNA (Cavanagh, 2005; Cavanagh, 2001). IBVs have been identified from all over the world and more may found in the future, that lead to high morbidity in all ages of chickens and high mortality in chicks less than 3 weeks old(Case et al., 1983). Initially, it was believed all the isolates belong to a single prototype termed Massachusetts (Mass) serotype mostly isolated from commercial poultry (Cavanagh and Naqi, 1997). Subsequently, other serotypes were isolated and it is clear now that a considerable number of different serotypes with antigenic and pathogenic differences exist in different parts of the world (Gough et al., 1992). Since IBV was first described in 1930s, there are more than 30 serotypes of IBV and dozens of virulent strains, with a variation being developed from time to time cause economic loss in chickens. The little or no cross-protection occurs between different serotypes of IBV is well known, therefore, continuous determination of the epidemic serotype and production of new generations of vaccines are crucial for controlling IB in each country(Casais et al., 2003). Vertical transmissions within the embryo have never been reported. But virus may be present on the shell surface of hatching eggs via shedding from the oviduct or alimentary tract (Saif et al., 2008). The main method of protecting poultry from infectious bronchitis (IB) is the administration of both live attenuated and killed vaccines. Because of the lack of a specific vaccine against endemic strains of IBV. IBV infection has remained a problem in the Iraqi poultry industry causing mortality and adverse effects on quantity and quality of egg production as well as renal failure lead to high range of mortality in broiler chickens due to infections with strains serologically different from those used for vaccine serotypes (Lim et al., 2011). The purpose of this study was conducted to investigate the prevalence of avian infectious bronchitis (IB) by the evaluating the (MDA) at 3 days age and compared with the antibodies against the disease in 25 days age also to study the clinical signs and gross lesions on the suspected infected birds in some broiler chickens flocks of Diyala governorate. MATERIALS AND METHODS 1- Broiler flock To study the sero-prevalence of IB, blood sample were collected from twenty broiler farms of different selected flocks in different areas of Diyala province during the period from September 2012 to May 2013. Each flock involved Alazawy Diyala Agricultural Sciences Journal, 5( 2 ) 25 – 37 ,2013 27   between 5000 to 6000 chicks of Ross 308 breed which were received from different hatcheries, Diyala/Iraq. The chicks were reared for 35 days on floor under almost healthy environmental circumstances. Pelleted feed from a commercial source was provided ad libitum. ). All the flocks had been vaccinated with IB vaccines (mostly H-120) in 3 days age and also vaccinated against Newcastle and Gumboro disease in 12 and 14 days respectively. Commercial vials of live attenuated vaccines (Lyophilized) were used for vaccination program as follow: 1- Nobilis IB H120 is a live vaccine against Infectious Bronchitis (Massachusetts H120, 3 Log10 EID50, Intervet – Holland). 2- Nobilis ND Clone 30 is a live vaccine against Newcastle disease in chickens (NDV Clone30 strain/Lentogenic. 3- Nobilis® Gumboro E228, (A freeze-dried culture of a non-pathogenic strain of Infectious Bursal Disease virus. 2-Serological Examination: A total of 720 Blood samples were collected from 18 broiler of each flock for the separation of sera to measurement of (MDA) and also sera from chick suspected to be infected with IBV by indirect ELISA on 3rd, days of chick’s age (prior to vaccination) and day 25 (Synbiotic Corporation). Five ml blood was collected aseptically from the wing vein of birds by disposable syringes (Terumo, Japan). Collected blood centrifugation by using (Hettich Centrifuge, Germany) for 10 minutes at 3000 rpm. After collecting the serum, we prepared from detection the (MDA) and also to estimate the suspected infected chickens with infectious bronchitis virus by using indirect ELISA according to the manufacturer’s instruction using IBV pre-coated plates and pre-diluted, ready to use reagents and buffer commercial kit( Bio Teck, USA). 3- Symptoms and Gross lesion appearance Postmortem examination was carried out on the infected chicks and also on the dead carcass. Chickens brought to the poultry diagnosis laboratory, college of veterinary medicine, university of Diyala, by farmers. Five or more chicks per each flock were examined on day 10 and weekly thereafter for 2 weeks, those chickens were in varying ages. All cases from flocks started to show clinical signs such as depression, respiratory distresss (rales, sneezing, and gasping), mild serous nasal discharge and an occasional but infrequent gasping reflex accompanied by extension of the head and neck. Some dyspnea with abdominal breathing was observed, more frequently in groups with mixed infections of IBV and avian influenza, sometimes accompanied by lacramation, facial swelling, and death rate of 40%. Alazawy Diyala Agricultural Sciences Journal, 5( 2 ) 25 – 37 ,2013 28   Whereas chickens exhibit renal cases observed mild sneezing and tracheal rales which develop about 24 hours, after this time severe symptoms are evident. The chick stands in a hunched position, eating very little and drinking more than usual, whereas the vent feather are often slightly soiled with watery white droppings with a total mortality of 10% to 1 5 % . These chickens were necropsied to check different infected organ. RESULTS AND DISCUSSION 1-Maternal derived antibodies titers: The measurement of maternal-derived antibodies at 3days of age (as indicated by ELISA test) showed in table 1. Table 1: Mean titer of maternal derived antibodies (MDA) against IBV at 3 days age with the mean titer of 8331.10. No of flock Age (days) Means of maternal-derived antibody titer against IBV Coefficient of Variation %CV 1 3days 8185 19.23 2 3days 9310 20.19 3 3days 10193 23.23 4 3days 9864 19.45 5 3days 8720 17.35 6 3days 7649 61.54 7 3days 8749 21.82 8 3days 8300 20.90 9 3days 8178 19.44 10 3days 6574 14.35 11 3 days 9827 20 12 3 days 8906 34 13 3 days 8258 32 14 3 days 5214 30 15 3 days 9864 33 16 3 days 8213 23 17 3 days 6308 22 18 3 days 10527 33 19 3 days 6605 21 20 3 days 8392 30.02 According to the (Table 1) the uniformity of the ELISA titers is an excellent method of determining the (MDA) in a normally distributed flock. If the CV% Alazawy Diyala Agricultural Sciences Journal, 5( 2 ) 25 – 37 ,2013 29   is less than 30% the uniformity of the flock was excellent that means the chicks have good immunity from the hens. 2-The measurement of antibodies against IBVvirus on 25 days of ages as suspected infected flocks (indicated by ELISA test) showed in Table 3. Table 2: Mean titer of the flocks after 25 days was 372.2500. No. Age (days) Means antibodies against IB virus on 25 days of ages CV% 1 25 304 157.01 2 25 279 124.65 3 25 435 140.24 4 25 387 103.12 5 25 228 69.25 6 25 795 69.14 7 25 480 86.55 8 25 459 95.07 9 25 832 99.02 10 25 278 111.05 11 25 1177 114.56 12 25 191 254.85 13 25 149 291.10 14 25 130 133.09 15 25 279 124.65 16 25 173 198.13 17 25 306 198.97 18 25 161 203.89 19 25 228 340.08 20 25 174 193.23 According to the (Table 2) the uniformity of the ELISA titers is an excellent method of determining the suspected infected flocks. If the %CV is greater than 80%, it means that the flock either have a bimodal population or incomplete or non-uniformity of vaccine administration, and the important interpretation her were the flock have early exposure to IB virus. The measurement of antibodies titers on 25 days age as suspected infected flocks with IBV showed a significant variation (P<0.05) among the MDA at 3 days ages as showed in table 3. Alazawy Diyala Agricultural Sciences Journal, 5( 2 ) 25 – 37 ,2013 30   Table 3: Means of antibody titer against IBV virus on 3 days as (MDA) and also on 25 days as the flocks to be suspected infected with IBV. Within a row, the titers of antibodies that do not have common small letter superscripts vary from each other (P <0.05). Groups Means ± SE of antibody titer 95% Confidence Interval Lower Bound Upper Bound Treatment T1 (MDI at 3 days ages) 8281 ± 233.622a 7808.657 8754.543 Treatment T2 (suspected infected flocks at 25 days ages). 372.250 ± 233.622b -100.693 845.193 2- Gross lesions determination Mortality in infected flocks reached between 20-40% during 7-9 days, the noticeable lesions of infected chickens showed lesions that identified primarily in the upper respiratory tract which characterized by severe congestion with serous, catarrhal, or caseous exudates in the trachea, nasal passages, and sinuses (Fig 1). Air sacs may appear mild to severe cloudy or contain yellow caseous exudates that appeared when the infection has been complicated with secondary bacterial infection ( Escherichia Coli or mycoplasma) that lead to, airsacculitis, pericarditis and perihepatitis (Fig 2). Alazawy Diyala Agricultural Sciences Journal, 5( 2 ) 25 – 37 ,2013 31   Similar lesions observed with the presence e of mycoplasma and E coli that led to more sever clinical signs resulting of pericarditis, perihepatitis and airsacculitis with depressed growth (Fig, 3). In severe infected chickens with IBV, the trachea showed tubular cast formations in the tracheal bifurcation extended into the primary and secondary bronchi were observed in all dead birds in 25 day- old (Fig, 4 ). Alazawy Diyala Agricultural Sciences Journal, 5( 2 ) 25 – 37 ,2013 32   Following infection with nephropathogenic strains, the birds are in good condition demonstrating the rapid course o f the disease. The kidneys were pale and were uniformly enlarged from two to four times normal size, with mottled dark and light areas. Focal areas of uric acid precipitation appear in the kidney and may also accumulate in the ureters (Fig, 5). Infectious bronchitis virus (IBV) is prevalent in all countries with an intensive poultry industry, with the incidence of infection approaching 100%. Vaccination is only partially successful because new serotypes continue to develop due to the continual emergence of antigenic variants (Hopicins, 1978). In recent years a considerable amount of laboratory investigation has been directed at the development of ELISAs for the seroepidemiological detection of exposure to IBV and other viral diseases (Cavanagh, 2007). Alazawy Diyala Agricultural Sciences Journal, 5( 2 ) 25 – 37 ,2013 33   Maternal immunity against IB in this study as revealed by ELISA showed high titer (mean titer 8331.10) which appear on table 1. The result of this finding was in agreement with previous published study (Sharma et al., 2000) who mentioned that the development of the reproductive tract of the hens, some of the stimulated lymphocytes localized in the lamina propria of the oviduct and in the stroma of the ovary. Antibody produced locally in these organs usually represented a significant increase of the transferred antibody to the eggs and constantly transfer from the hens to the chick with high titer. Also this finding was in agreement with previous studies (Luis et al., 1982) which showed that most breeder flocks are periodically vaccinated for both ND and IB, parental antibodies are often transmitted to the progeny at significant levels lead to high titer of maternal antibodies. Previous study showed that vaccinated flocks against IBV were often applied to reduce economic losses due to an infection with field strains of IBV. Also vaccination procedure, such as using spray vaccination of broiler flocks at 1 day old of age play important role to reduce the viral diseases and induces good clinical protection upon challenge with a virulent IBV strain of the same serotype (Cavanagh and Naqi, 2003; Davelaar et al., 1982). Previous study showed that vaccines against IBV, such as H-120 are often applied to reduce economic losses due to an infection with field strains of IBV (Parsons et al.,1992), whereas in current study, it appeared that, despite the regular vaccination of infectious bronchitis vaccine mostly used H-120 strain, the mean titer were decreased and cannot protected the flocks, and also this finding was in line with(Matthijs et al., 2008) who observed that the majority (67 per cent) of these isolates which causes IBV belonged to serotypes other than the Massachusetts type. The current study were differ from those (Hofsted and Yoder, 1992)) who mentioned that IBV H-120 vaccine is one of the commonly used live IB vaccines in commercial poultry, and spray vaccination of broilers at 1 day or 15 days of age generally induces good clinical protection upon challenge with a virulent IBV strain of the same serotype. The present study revealed that the, antibody levels obtained from vaccinated flocks at 25 days of age demonstrated presence of significant differences of the mean titer at 25 days age at level (P< 0.05) from MDA at 3 days age, this differences could be due to the flocks vaccinated with H-120 which had the lowest antibody response resulted from the failure of H-120 to induce active immune response in the infected flocks or the flocks showed a very early exposure to disease agent causes by IBV. In contrast to the above findings, the results of current study differ from that of (Luis, 1982) who found that vaccinated the chickens with H-120 led to highest antibody response suggesting a typical response of antibody that make good flocks protection against IBV. Alazawy Diyala Agricultural Sciences Journal, 5( 2 ) 25 – 37 ,2013 34   This study also in agreement with the earlier works of (Lue et al., 1993; Arathy et al., 2001) who showed that the reason for outbreaks in vaccinated chickens may be due to the tendency of the virus to undergo continuous genomic drift and shift which led to emerge of new serotypes differences from vaccine strain especially in areas of intensive poultry farming. The present study showed that, the clinical signs and gross lesions such as respiratory distress (rales, sneezing, gasping and dyspnea) were most sever in bird infected with IBV, also post mortem lesions showed trachael congestion with serous, catarrhal exudates and cast formation in tracheal bifurcation abruptly reduced airflow, resulting in asphyxiation that was responsible for mortality in dead birds. The findings in the current study are consistent with those of previous works (Jahromi et al.,2008; Seifi et al., 2012) who consistently found that the cast formation in tracheal bifurcation were most severe in the AIV and IBV infected chickens which lead to asphyxiation and subsequently responsible for mortality in dead birds. Whereas previous work of (Tavakkoli et al., 2001; Karimi et al., 2010) observed that infectious bronchitis live vaccine, H-120 strain enhancing the virulence of low pathogenic H9N2 avian influenza virus in broiler chicken flocks which lead to cast formation in tracheal bifurcation. The current study revealed that the incidence and severity of airsacculitis, pericarditis and perihepatitis may be higher observed in the suspected mycoplasma infection which mixed with IBV in infected flocks and are in agreement with other study of (Chu and Uppal, 1975) who observed that IB infection may precipitate latent mycoplasma infection which showed increased spread and severity of infection. Whereas previous work of (Kleven et al., 1978) observed that the presence of virus, even the vaccine strain, enhanced the number of mycoplasma isolations and precipitated the incidence of airsacculitis. The result of present study showed that in some cases, variant strains of IBV seem to be the only agents involved in infectious primary kidney lesions in broiler chickens with mortality of up 25%, which led to focal area of uric acid precipitation appear in the kidneys and may also accumulated in the ureters. The present finding seem to be consistent with other published studies (Darbyshire et al., 1978) who observed that the initial and principal site of viral replication primarily in the trachea and from this site the virus spreads to other internal organ such as the kidneys with highest titer led to enlarged the kidneys and may be pale or marbled and deposits of urates may be present in the ureters of some chickens. In conclusion this study obviously demonstrates that the widespread nature of the IBV present in poultry flocks in Diyala province. Future work should included the isolation and serotyping of IBV in any area where this disease is not adequately controlled, so that a suitable vaccination program by using the Alazawy Diyala Agricultural Sciences Journal, 5( 2 ) 25 – 37 ,2013 35   common filed serotypes as vaccines, can be adopted to protect against IBV caused disease. REFERENCES Cavanagh, D., S and Naqi. 1997. Infectious bronchitis, p. 511–526. In B. W. Calnek, H. J. Barnes, C. W. Beard, W. M. Reid, and H. W. Yoda (ed.), Diseases of poultry, 10th ed. Iowa State University Press, Ames, Iowa. Li, L., C. Xue., F.Chen., J. Qin., Q. Xie and Y. Bi. (2009). Isolation and genetic analysis revealed no predominant new strains of avian infectious bronchitis virus circulating in South China during 2004–2008. Vet Microbiol. 2010; 143:145–54. Cavanagh, D. 2003. Severe acute respiratory syndrome vaccine development: experiences of vaccination against avian infectious bronchitis coronavirus. Avian Pathol., 32, 567–582. Cavanagh, D. 2001. A nomenclature for avian coronavirus isolates and the question of species status. Avian Pathol. 30:109–115. Case, J.T., A.A.Ardans., D.C.Bolton and B.J.Reynolds.1983. Optimization of parameters for detecting antibodies against infectious bronchitis virus using an enzyme-linked immunosorbent assay: temporal response to vaccination and challenge with live virus. Avian Diseases 27:196-210. Gough, R.E., M.C. Randall., D. J. Dagless., W. J. Cox, and D. Pearson. 1992. A “new” strain of infectious bronchitis virus infecting domestic fowl in Great Britain. Vet. Rec., 130: 493–494. Casais, R., B. Dove., D. Cavanagh, and P. Britton.2003. Recombinant avian infectious bronchitis virus expressing a heterologous spike gene demon- strates that the spike protein is a determinant of cell tropism. J. Virol., 77: 9084-9089. Saif, Y.M., A. M. Fadly., J. R.Glisson., L.R. McDougald., L.K. Nolan and D.E.Swayne. 2008. Diseases of poultry, 12th edn. Iowa State University Press, Ames, Iowa, pp 117–130. Lim, T.H., H. J.Le., D. H. Lee., Y. N. Lee., J. K.Park and H. N. Youn. 2011. An emerging recombinant cluster of nephropathogenic strains of avian infectious bronchitis virus in Korea. Infect Genet Evol. 11:678– 85. ELISA – SYNBIOTICS Corporation. (1997-2003). www. Symbiotic.com. Hopkins, S.R .1978. Typing field isolates of infectious bronchitis by the plaque- reduction test. Avian Dis. 22:71-81. Cavanagh, D.2007. Coronavirus avian infectious bronchitis virus. EDP Sciences., Vet. Res., 38: 281-297. Sharma, J.M., I. J. Kim., S. Rautenschlein and H. Y.Yeh.2000. Infectious bursal disease virus of chicken: pathogenesis and immunosuppression. Develop. Comp. Immunol., 24: 223-235. Alazawy Diyala Agricultural Sciences Journal, 5( 2 ) 25 – 37 ,2013 36   Luis, F., A. P.Andrade and O.J. Fletcher.1982. Vaccination of Day-Old Broilers against Infectious Bronchitis: Effect of Vaccine Strain and Route of Administration.Avian Dis., 27,178-187. Cavanagh, D and S. A. Naqi. 2003. Infectious bronchitis. In: Saif YM, Barnes HJ, Glisson JR, Fadly AM, McDougald LR, Swayne DE (eds). Diseases of Poultry, 11th Ed. Pp: 101-119. Ames, Iowa States University Press, USA. Davelaar, F.G., B. Kouwenhoven and A. G.Burger.1984. Occurrence and significance of infectious bronchitis variant strains in egg- and broiler production in the Netherlands. Vet. Q., 6: 114–120. Parsons, D., M.M.Ellis.,D. Cavanagh and j. k.Cook.1992. Characterisation of an infectious bronchitis virus isolated from vaccinated broiler breeder flocks. Vet. Rec. 31;131(18):408-11. Matthijs, M. G. R., A. Bouma., F. C. Velkers., J. H. H. van Eck and J. A. Stegeman .2008. Transmissibility of Infectious Bronchitis Virus H120 Vaccine Strain among Broilers under Experimental Conditions. Avian: 3, 461-466. Hofstad, M.S. and H. W. Yoder. 1996. Avian infectious bronchitis-virus distribution in tissues of chicks. Avian Dis., 10: 230-239. Lu, Y. S., H. W. Shieh., H. J. Tsai., D. E. Lin. and Y. L. Lee. 1993. The incidence and virus isolation of infectious bronchitis in chickens in Taiwan. J. Chinese Soc. Vet. Sci. 19:119-129. Arathy, S. A., S .Gopalakr., S.M. Kumthekar., D. Thomas and R.N. Sharma. 2011. Seroprevalence of Infectious Bronchitis Virus in Birds of Grenada. Inter. J. Poult. Sci.,10 (4): 266-268. Jahromi, M., K. Asasi, K., Nili, H., Dadras, H and H. Shooshtari. 2008. Coinfectionof avian influenza virus (H9N2 subtype) with infectious bronchitis live vaccine. Arch. Virol., 153: 651-655. Seifi, S., K. Asasi and A. Mohammadi. 2012. An experimental study on broiler chicken co-infected with the specimens containing avian influenza (H9 subtype) and infectious bronchitis (4/91 strain) viruses. Iranian J. Vet. Res, 39: 138-142. Tavakkoli, H., K. Asasi., A. Mohammadi. 2011. Effectiveness of two H9N2 low pathogenic avian influenza conventional inactivated oil emulsion vaccine on H9N2 viral replication and shedding in broiler chickens. Iranian J. Vet. Res., 12: 214-221. Karimi, M.,M. Ansari., K. Asasi and H.Nili .2010. Risk factors for detection of bronchial casts, most frequently seen in endemic H9N2 avian influenza infection, in poultry flocks. Prev. Vet. Med., 95: 275-280. Chu, H.P and P. K.Uppal .1975. Single and mixed infections of avian infectious bronchitis virus and Mycoplasma gallisepticum. Dev Biol Stand : 28:101- 114. Alazawy Diyala Agricultural Sciences Journal, 5( 2 ) 25 – 37 ,2013 37   Kleven, S.H., C. S.Eidson and O.J.Fletcher. 1978. Airsacculitis induced in broilers with a combination of Mycoplasma gaO.Jllinarum and respiratory viruses .Avian Diseases, 22, 707-716. Darbyshire J.H., J.K. Cook and R.W. Peters R.W.1978. Growth comparisons of avian infectious bronchitis vims strains in organ cultures of chicken tissues. Arch. Virol, 56, 317-325. باستخدام تقنية االليزا للتحري عن مرض التهاب القصبات الخمجي بين مجموعة ةدراسة سيرولوجي . من حقول الدواجن في محافظة ديالى عامر خزعل صالح العزاوي . جامعة ديالى - آلية الطب البيطري - لدواجنا وأمراض األمراضفرع * المستخلص من االمراض الشائعة الحدوث والشديدة العدوى والذي يسبب القصبات الهوائية الخمجي فايروس التهاب خسائر اقتصادية فادحة في مجال تربية الدواجن وهو شائع الحدوث في جميع البلدان التي تربى فيها سة لمعرفة مدى شيوع مرض االلتهاب القصبات الهوائية الخمجي في صصممت هذه الدرا. الدواجن اجريت . حقول تربية دجاج اللحم في محافظة ديالى من خالل دراسة سيرولوجية باستخدام تقنية االليزا تم حساب المناعة االموية . معة ديالىجا/آلية الطب البيطري –الدراسة في مختبر االحياء المجهرية لمعرفة التفاوت في هبوط مستوى االجسام ًايوم 25وآذلك تم حسابها بعمر أيام 3لالفراخ بعمر بهذا المرض والمصنع من سرآة العدة التشخيصية الخاصةالمناعيةاستنادا الى التعليمات المرفقة مع إلى ًايوم 25بعمر ) P< 0.05( االجسام المناعية سمبايوتك االمريكية حيث دل االنخفاض في مستوى دراسة تتعرض القطيع لالصابة المبكرة من خالل الهبوط في مستوى االجسام المناعية وآذلك تم بصعوبة التنفس والسعال والغرغرة وخروج مواد مخاطية من المنخرين السريرية والمتمثلة األعراض افقة للحاالت المصابة من خالل دراسة الصفات التشريحية لكل وآذلك دراسة التغيرات المرضية المر حالة والتي اظهرت وجود مواد سائلة ومتجبنة في القصبة الهوائية وآذلك تغيرات في االآياس الهوائية شيوع هذا المرض في حقول الدواجن في محافظة ديالى وخاصة في عمر اظهرت نتائج الدراسة. والكلى .ة اللقاحات المستخدمة ضد المرضاسابيع وعدم فاعلي 3 .ىديالالتهاب القضبات الخمجي، االليزا ، المناعة االموية ، : الكلمات المفتاحية