227 Pakistan Veterinary Journal ISSN: 0253-8318 (PRINT), 2074-7764 (ONLINE) Accessible at: www.pvj.com.pk Optimization of Quality Control Factors for Indigenous Mycoplasma Gallisepticum Bacterin Preparation and Their Impact on Immunoprophylaxis in Broilers Asim Raza 1 , Arfan Ahmad 1 , Masood Rabbani 1 , Altaf Mahmood 2 , Muhammad Younus 3 , Zakir Ali 2 and Abdul Ahad 4 ,* 1 Department of Microbiology, University of Veterinary and Animal Sciences, Lahore, Pakistan; 2 Livestock and Dairy Development Department, Govt. of Punjab, Pakistan; 3 Department of Pathology, University of Veterinary and Animal Sciences, Lahore, Pakistan; 4 Department of Microbiology and Veterinary Public Health, Chittagong Veterinary and Animal Sciences University Chittagong, Khulshi–4225, Bangladesh *Corresponding author:ahadvet1969@yahoo.co.uk ARTICLE HISTORY (14-535) ABSTRACT Received: Revised: Accepted: Published online: October 20, 2014 August 18, 2015 September 01, 2015 December 31, 2015 To inactivate locally isolated Mycoplasma gallisepticum (MG), phenol, formaldehyde and binary ethylenimine were optimized for their concentrations and exposure time. Their effect on immunoprophylaxis induction was then assessed by using different levels of immunogen/PCV and different adjuvants. 0.10M, 0.125% and 0.6% concentrations of binary ethylenimine, formaldehyde and phenol inactivated MG colonies within 24, 8 and 4 hours respectively. Formaldehyde inactivated vaccine with 1.5% Immunogen/PCV and montanide oil as adjuvant induced significantly (P<0.05) higher anti MG antibody titer than other locally prepared vaccines. Conclusion is that montanide oil adjuvated indigenous formalized bacterin can induce desired immune response. ©2015 PVJ. All rights reserved Key words: Adjuvants Bacterins Broilers Inactivants Mycoplasma gallisepticum (MG) To Cite This Article: Raza A, Ahmad A, Rabbani M, Mahmood A, Younus M, Ali Z and Ahad A, 2016. Optimization of quality control factors for indigenous Mycoplasma gallisepticum bacterin preparation and their impact on immunoprophylaxis in broilers. Pak Vet J, 36(2): 227-229. INTRODUCTION Mycoplasma gallisepticum (MG) is cause of chronic respiratory disease. Young birds are more susceptible than adults. The portals of entry of the pathogen are inhalation and conjunctiva whereas those of exit are respiratory tract, eggs and semen. How the organism overcomes bird’s natural defense mechanism is still not clearly understood (Bradbury, 2001; Ahmad et al., 2008). The pathogen is transmitted horizontally through contaminated dust, droplets and feathers and vertically through contaminated egg yolk. Economic losses occur due to high morbidity, reduced growth rate, poor feed conversion ratio, decreased egg production and poor hatchability (Sarkar et al., 2005; Noel et al., 2012). Live and inactivated imported vaccines are mostly used in northern areas of Pakistan but the disease is not uncommon even in vaccinated flocks which may be due to subtle antigenic variation in immunogen of the vaccine. This situation necessitated the development of indigenous vaccine through present experiment in order to achieve desired immunoprophylaxis. MATERIALS AND METHODS Local MG isolate was obtained from University Diagnostic Lab. Different concentrations of inactivants including phenol (0.6, 0.30 & 0.125%); formaldehyde (0.50, 0.25 & 0.125%) and binary ethylenimine (0.05, 0.10 & 0.15M) were tested to determine the optimum concentration and exposure time for inactivation of MG culture. These mentioned concentrations were added in separate tubes containing viable MG isolates following the procedures of OIE (2015). Tubes were then incubated at room temperature. After exposure of 04, 08, 12, 24 and 48 hours, the viability of MG isolate was tested by culturing the MG containing broth on PPLO agar leading to incubation and observation under stereoscope microscope. Frey’s agar plates were observed 10 days post streaking. The concentration of each inactivant with exposure time was recorded. Three optimum concentrations (one for each inactivant) with least exposure time were selected and nine types of vaccines were prepared using 1% PCV with montanide oil as adjuvant, i.e., F-Vac: Formaldehyde inactivated; P-Vac: Phenol inactivated; B-Vac: Binary ethylenimine inactivated; Using formaldehyde and montanide oil (4:1 ratio) with different PCV concentrations adjusted according to procedure of Ghany (2011); PCV1-Vac: 0.5% PCV; PCV2- Vac: 1% PCV; PCV3-Vac: 1.5% PCV; Using formaldehyde as inactivant with 1% PCV but different adjuvants M-Vac: Montanide oil as adjuvant; A-Vac: Aluminium hydroxide gel; W-Vac: Water SHORT COMMUNICATION