Agricultural Research Management Information System

Home
Research Summary
Search
About Us
- ARMIS
- Brochure
Contact Us
Report
User Request
Data Input
Help
- Operation Manual
  - PDF
  - Video
- Program Area & Commodity
We have reached 37600 number of research entries at this moment.

- Logout

Research Detail

Home
Research
Detail

Development of Slide Agglutination Test for the Rapid Diagnosis of Mycoplasma Infection in the Chicken

M. Arefin
Department of Pathology, Faculty of Veterinary Science, Bangladesh Agricultural University, Mymensingh-2202, Bangladesh

J. A. Begum,
Department of Pathology, Faculty of Veterinary Science, Bangladesh Agricultural University, Mymensingh-2202, Bangladesh,

R. Parvin
Department of Pathology, Faculty of Veterinary Science, Bangladesh Agricultural University, Mymensingh-2202, Bangladesh,

M. M. Rahman
Department of Pathology, Faculty of Veterinary Science, Bangladesh Agricultural University, Mymensingh-2202, Bangladesh,

M. A. H. N. A. Khan
Department of Pathology, Faculty of Veterinary Science, Bangladesh Agricultural University, Mymensingh-2202, Bangladesh,

E. H. Chowdhury
Department of Pathology, Faculty of Veterinary Science, Bangladesh Agricultural University, Mymensingh-2202, Bangladesh,

Abstract/ Executive Summary:

Mycoplasma gallisepticum is considered to be the most important pathogenic Mycoplasma sp. and occurs world-wide. For effective control of mycoplasmosis, early diagnosis is the cornerstone, but the cultural method is laborious, expensive and timeconsuming, whereas agglutination test is easy to perform in field conditions and cheaper. Mycoplasma gallisepticum was isolated and two types of stained antigen produced. The antigen produced distinct tiny clumps with positive antisera of Mycoplasma spp. The slide agglutination system was sensitive, reliable and cheap. This method may be used to screen Mycoplasma infections in poultry farms.

Keywords: Slide agglutination test, Rapid diagnosis, Mycoplasma infection, Chicken

Location: Bangladesh Agricultural University, Mymensingh

Start Date:

End Date:

Program Area: Animal Health and Management

Commodity: Chicken

Objectives:

To prepare Mycoplasma-coloured antigen, develop slide micro-agglutination and agglutination tests for the diagnosis of avian mycoplasmosis.

Methodology:

Production of seeds Mycoplasma was maintained in Mycoplasma agar in the laboratory and the isolates were plated on Mycoplasma agar, and a fried-egg-like single colony was selected for each isolate .This was transferred to Mycoplasma broth and incubated at 37⁰C for 48 hours and maintained by transferring from one agar plate to another for several months. Kanamycin was added to the medium to prevent the growth of Gram-negative bacteria, and horse serum was used for the growth of Mycolasma organism. This culture was used for the production of stained antigens. Preparation of mycoplasma-stained colour antigen The local isolate of Mycoplasma gallisepticum was transferred to a conical flask containing 50 ml of Mycoplasma broth base, which was incubated for 11 days at 37⁰C, at which time each flask revealed dense growth. Fifty ml broth culture was divided into two conical flasks each containing 25 ml. Tetrazolium blue and neotetrazolium chloride were added aseptically, 0.5% to each culture. The flasks were agitated and incubated for 24 hours, then 0.5% phenol was added to each flask, which were incubated for a further 2 hours for inactivation. The stained broth suspension was filtered through sterile gauze and poured into eppendorf tubes. These were centrifuged at 14,000 rpm for 15 minutes. The supernatant fluid was decanted and the packed cells were suspended in 0.5% phenol saline. The suspension was vortexed vigorously with a few sterile glass beads and transferred to another eppendorf tube. A dense suspension of deeply stained cells resulted. The solution was kept at 4°C as neotetrazolium- and tetrazolium-stained antigen for future use. The total protein concentration of stained antigen was measured by the Folin Phenol method as described by Lowry et al. (1951) and the protein concentration was adjusted to 6.4 ng/μl with the help of BSA standard curve. The antigen was packaged for field use. Collection of field samples A total of 110 serum samples were collected from local markets and poultry farms into eppendorf tubes. The slide micro-agglutination test was performed with the new Mycoplasma antigen. For development of slide micro-agglutination test, 20 μl of stained antigen and 20 μl chicken sera were placed on a sterile glass slide by a micropipette and mixed thoroughly by stirring. The results were read within 1 minute. Serum and water were used as negative control. For proper mixing, the antigen was shaken before use. The stain that used for preparation of antigen was sensitive to light so it kept away from the light.

Source of Information: The Bangladesh Veterinarian (2011), 28(2) : 80 – 84

Article Link (Online Journal):

Funding Source:

Total Budget (Tk.) :

Achievement/Findings:

A tetrazolium-stained Mycoplasma antigen from a local isolate was successfully developed. [With the elapse of time, the stained antigen may react with the nonspecific antibody present in the serum. Therefore, results within one minute were indicated suspecting cases of mycoplasmosis. After testing specificity with other closely related organism and sensitivity with some commercial antigen, a test kit may be prepared

Knowledge Management: Journal