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Determination of Serum Antibody Titres and Immune Status of Layer Flocks Against Newcastle Disease Virus at Chittagong District of Bangladesh

Kashem MA
Department of Microbiology, Chittagong Veterinary and Animal Sciences University (CVASU), Khulshi-4202, Chittagong

Parvej M
Department of Microbiology, Chittagong Veterinary and Animal Sciences University (CVASU), Khulshi-4202, Chittagong

Hashem MA
Veterinary Surgeon, Chittagong City Corporation, Chittagong

Moula MM
Veterinary Surgeon, Community based Animal Health Care Services, Mirshari, Chittagong, Bangladesh.

Kibria ASMG
Department of Anatomy and Histology, Chittagong Veterinary and Animal Sciences University (CVASU), Khulshi-4202, Chittagong

Abstract/ Executive Summary:

A study was conducted to assess the level of serum antibody titres and immune status of layer birds against Newcastle Disease virus by Haemagglutination Inhibition (HI) test in different areas of Chittagong district during November to December, 2010. Sixteen layer flocks were selected based on different ages of birds. A total of 235 serum samples were collected and tested at Microbiology laboratory of CVASU. HI test was performed using commercial Newcastle Disease vaccine (Avinew®) as a source of 4HAU virus antigen. The antibody titre (GMT) levels in 18-26 weeks age group were found to be 70.198, followed by 47.551, 34.776, 17.281 and 18.855 in 27-40, 41-57, 58-73 and >73 weeks age groups, respectively. Moreover, 100% specific immunity against ND was found in 18-26, 27-40 and 41-57 weeks age groups of birds, whereas 93.33 and 94.73% specific immunity was found in 58-73 and >73 weeks age groups, respectively. On an average, 97.87% layer birds showed specific immunity and 2.13% showed nonspecific immunity against NDV. We considered HI titre of 1:8 or above as specific immunity and less than 1:8 as non specific immunity. Highest HI titre was found at the age of 18-26 weeks and lowest titre was at 58-73 weeks of age. The lower level of HI titre seemed to be directly related to some important factors relating to vaccination which have been highlighted in this paper.

Keywords: Antibody titers; Immune status; HI test; Newcastle disease virus; Layer birds.

Location: Chittagong district of Bangladesh

Start Date: 00-11-2010

End Date: 00-12-2010

Program Area: Pest Management

Non-commodity: Performance

Objectives:

The present study was undertaken with the following objectives: to determine the serum antibody titres and identify the specific immune response of layer birds against Newcastle Disease virus at some selected areas of Chittagong district of Bangladesh.

Methodology:

Study areas, sample size and duration: The study was conducted in four selected areas of Chittagong District namely, Chandgaon, Mohra, Kalorghat, and Sitakunda. All laboratory activities were performed at the Department of Microbiology, Faculty of Veterinary Medicine, Chittagong Veterinary and Animal Sciences University, Chittagong, Bangladesh, from November to December, 2010. A total of 235 samples were collected from layer birds with the age ranging 18- 26, 27-40, 41-57, 58-73 and >73 weeks. For collection of blood samples the birds were randomly selected from flocks. Collection and preservation of samples: From each bird 2 ml of blood was collected aseptically from wing vein, placed at room temperatures for 30 minutes and then kept in chilling temperatures for separation of serum from clot. The separated sera were collected in Eppendorf tubes and stored at -20°C until used for further analysis. Preparation of 1% chicken RBC suspension: For making 1% chicken RBC suspension, 5ml of blood was taken aseptically from 4 weeks old specific pathogen free chickens with a disposable syringe containing ethylene diamine tetra acetic acid as an anticoagulant. After transferring the blood into centrifuge tube, equal volume of PBS was added and centrifuged at 1000 rpm for 10 minutes. The supernatant was poured off. Again 30 volumes of PBS added with blood cells and centrifugation steps repeated for twice. After washing thrice with PBS, 1% chicken RBC was prepared by adding 1 ml RBC to 99 ml of PBS. Haemagglutination test and determination of 4HA unit virus for HI test: A 50µl of PBS was dispended into each well of one row of the plastic v-bottomed 96 well plate. Then 50l of virus suspension (Avinew® vaccine, Advance Animal Science Co. Ltd.) was placed into the first well and made two-fold dilutions of the suspension across the row by transferring 50µ of fluid from one well to the next. Then 50µ fluid was discarded from the last well so that the volume in each well remained the same. A control row was also made by the same procedure only by using PBS instead of virus antigen. Then 50µ of PBS was added to each well including control. Finally, 50µ of 1% Chicken RBCs suspension was added to each well, tilted gently and allowed to stand at room temperature for 40 minutes by covering to stop dehydration. The result was read and recorded that 8 th well was the last to show haemagglutination (thin film) which indicated 1 HA unit. Determination of 4HA unit virus antigen from this HA results was done as described by Ilaria and Alexander. Haemagglutination Inhibition (HI) test: HI test was performed according to the procedure of OIE. In brief, serial two fold dilution of field sera (50µl) was made with phosphate buffer saline in 96- wells microtitre plate up to 10th well. Equal volume of 4HA units of ND virus was added into each well up to 11th well. For facilitating the antigen antibody reaction, the plates were kept at room temperature for more than 30 minutes. Then 50µl of 1% (v/v) chicken RBCs suspension was added up to 12th well. The 11th well contained antigen and RBCs as a positive control whereas 12th well contained only RBCs as a negative control. The RBCs were allowed to settle down at room temperature for 40 minutes by slightly agitating the plate. After that the test result was assessed by tilting the plates and appearance of a sharp button because of settling of intact RBCs was recorded as positive. Maximum dilution of each sample was considered as the end point of HI test from which serum antibody titre was determined by reciprocal measurement of the observed result.

Source of Information: International Journal of Natural Sciences (2011), 1(2):35-38

Article Link (Online Journal):

Funding Source:

Total Budget (Tk.) :

Achievement/Findings:

From this study we conclude that, the antibody titer against Newcastle Disease virus in commercial layer flocks of Chittagong District was apparently protective due to the maintaining of proper vaccination schedule and also due to the previous exposure of Newcastle Disease virus.

Knowledge Management: Journal