Int. J. BioRes. 1(3): 13--16 March, 2010 Maniruzzaman et al. 13 Full Length Research Paper ISOLATION AND IDENTIFICATION OF BACTERIAL FLORA FROM MILK OF APPARENTLY HEALTHY BUFFALO-COWS M Maniruzzaman, M F R Khan*, M M Amin, A K Paul and M Islam Department of Microbiology and Hygiene, Bangladesh Agricultural University, Mymensingh, Bangladesh *Correspondence: frkhanbau80@yahoo.com Accepted on 11 March 2010 ABSTRACT The study was conducted in 50 milk samples collected from apparently healthy lactating buffalo-cows during the period of July to October 2009 for isolation, identification of bacterial flora on basis of their morphology, staining, cultural and biochemical properties. Antibiotic sensitivity of the isolated organisms were also studied. Out of 50 milk samples examined for bacterial isolates. Total isolates were 40, out of which Lactobacillus spp. was 10 (25%), Bacillus spp. 10 (25%), Staphylococcus spp. 13 (32.5%), E. coli 5 (12.5%) and two 2 (5%) isolates were unclassified. Results of antibiotic sensitivity in this investigation showed that Lactobacillus spp. and Bacillus spp. were more or less sensitive to erythromycin, azithromycin, nalidixic acid but resistant to ampicillin, penicillin, trimethoprim, amoxicillin and metronidazole. Staphylococcus spp. was more or less sensitive to erythromycin, azithromycin, ampicillin, amoxycillin, nalidixic acid and penicillin but resistant to metronidazole and trimethoprim. E. coli was more or less sensitive to all types of antibiotics used for experiment. Judging from the parameter it was concluded that bacteria of fresh milk are not harmful but pathogenic bacteria may invade milk through various sources which is harmful. Key words: Bacterial flora, Buffalo cow, Antibiotic sensitivity, Milk INTRODUCTION Milk of buffaloes constituting an important source of market milk has some unusual qualities. It meets certain specific food requirements of human population in India and elsewhere. The fat content can exceptionally be as high as 15 percent and the overall average may be 7 %. Milk is also an excellent medium for the growth of a large variety of bacteria. Bacteria need considerable amounts of nutrient such as water, carbohydrate, fat and other substances for their growth and milk contains all of these nutrients (Harrigan et al. 1976). Microorganisms are always undesirable in milk and its products. These are capable of causing deterioration in flavor, physical appearance of milk and transmission of infectious diseases to the consumers. The various organisms get into milk through unhygienic, carelessness and unsanitary practices of the farmers, processors and distributors. Discoloration, sliminess, rapines, putrefaction, rancidity and many other defects are caused by various microorganisms growing in the milk and milk products. Bacterial contamination of raw and pasteurized milk is considered to be a great problem for dairy milk. The important genera of bacteria normally found in milk are, Microbacterium, Micrococcus, Streptococcus, Staphylococcus, Lactobacillus, Bacillus, Clostridium, Arthrobacter, Actinomyces, Coxiella, Pseudomonus etc. (Adesiyun, 1994). Most of these organisms are free living, widely distributed in soil, feeds, cows, buffaloes, goats, dairy utensils etc. Contamination usually occurs at the farm where milk is produced. So, the present research works was undertaken to isolate the bacteria from fresh buffalo milk, to study the cultural and biochemical properties of the isolated bacteria and to study the antibiotic sensitivity of the isolated bacteria. MATERIALS AND METHODS This study was conducted in the Department of Microbiology and Hygiene, Faculty of Veterinary Science, Bangladesh Agricultural University (BAU), Mymensingh during the period of July to October 2009. Sample collection Samples were collected directly from apparently healthy buffalo-cows in Senbari union of Trisal upazila under Mymensingh district. A total of 50 milk samples were collected from apparently healthy lactating buffalo-cows. Foremilk samples (up to 10 ml) were collected aseptically from individual quarters from all buffalo-cows for bacteriological culture, using routine techniques for aseptic infection. The samples were carried to the laboratory in an ice box contained ice and processed for the isolation and characterization of bacteria subsequently. The remaining samples were stored at 0˚C for further use. Isolation and identification of bacterial flora from milk of apparently healthy buffalo-cows For isolation and identification of bacterial flora, the procedure suggested by Cheesbrough, (1984) were followed through out the whole experiment. Isolation of bacterial flora Samples were incubated at 37°C for 24 hours. The incubated tubes were examined for turbidity that indicating bacterial growth. Samples were then inoculated onto NA plates and incubated at 37°C for 24 hours and subcultured onto NA plates to obtain pure culture. The subculture technique was followed up to obtaining pure culture.