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Title: Biocontrol of Brucella Abortus Using Brucellaphages
Authors: , Arfat
Keywords: Biological & Medical Sciences
Issue Date: 2021
Publisher: University of Veterinary and Animal Sciences, Lahore.
Abstract: ISOLATION AND PHYSICOCHEMICAL CHARACTERIZATION OF BRUCELLAPHAGES Bovine brucellosis, caused by Brucella abortus, is an economically significant bacterial disease causing enormous economic losses in developing countries. Due to emerging antibacterial resistance in the current use of antibiotics and insufficient immunity by WHO-recommended vaccine strategies, it is recommended to cull the positive animals to control the disease. In such circumstances, the use of host-specific bacteriophages could be an alternate option to control the disease. In the present study, brucellaphages were isolated from slurry samples (n=50) of livestock farms. Seven samples were found positive in the spot method, while two samples gave the positive plaques of pinpoint size (0.5 mm) with round and clear appearance in a plaque assay. Isolated brucellaphages (BaP1 and BaP2) did not produce plaques against Staphylococcus aureus, Streptococcus, Bacillus, Escherichia coli, Salmonella, and Pasteurella multocida. Physicochemical characterization revealed that lytic activity of phages was present up to 60oC which started to decrease at 70oC and maximum stability was between 7 to 9 pH. Exposure of sunlight, normal fluorescent, and UV light inactivated these phages within 3 hours, 24 hours, and 15 minutes, respectively. Phages become inactivated in 15 minutes when treated with Sodium Dodecyl Sulphate, chloroform, Lysozyme, and Proteinase K, however, no effect of normal saline, Trypsin, EDTA and RNAse was observed on brucellaphages. In conclusion, the results have laid the foundation to standardize practical applications of brucellaphages after detailed in-vitro and in-vivo experimental evaluations. Keywords: Brucellosis, Brucella abortus, brucellaphage, plaque assay, physicochemical characterization ISOLATION, PROPAGATION AND BIOCONTROL ACTIVITY OF INDIGENOUS BACTERIOPHAGES AGAINST BRUCELLA ABORTUS Brucellosis is an important zoonotic disease with public health significance. Its control is a serious challenge, despite the availability of vaccines; also the ever-increasing antimicrobial resistance puts pressure on disease treatment. Alternate therapy to control brucellosis using biological substances such as brucellaphages can be an effective method. This study was conducted to isolate bacteriophages against Brucella abortus from sewage/slurry samples (n=50), collected from cattle farms. Isolation and propagation of brucellaphages were made through spot and plaque assay using the double-agar overlay technique. Two samples (n=2) were found positive on the plates as circular, clear, and pinpoint plaques (0.3 to 0.5 mm) against Brucella abortus. Isolated brucellaphages (ϕP1 and ϕP2) were unable to lyse heterogeneous bacterial species i.e. Staphylococcus aureus, Streptococcus sp., Salmonella sp., Escherichia coli, Bacillus subtilis, and Pasteurella multocida. Titre of brucellaphages was determined to be as 4.6×106 PFU/ mL. Thermal and pH stability of phages showed them to be stable up to 60oC and between 7 to 9 pH. Nucleic acid and protein characterization displayed these phages with double stranded DNA of approximately 40 kb size and two prominent protein bands of 45 kDa and 70 kDa. Biocontrol activity of brucellaphages showed average Brucella abortus count reduction to 4.5×103 from 5.0×108 CFU/ g of soil sample within 48 hr when treated with maximum phage titer of 5.0×1011 PFU/ mL. Hence, this study revealed that brucellaphages are present in our environment and can potentially be considered for their cost-effective practical applications in the inactivation of Brucella abortus after comprehensive experimental evaluations. Key words: Brucellosis, Brucellaphages, Brucella abortus, Overlay technique, Characterization, Biocontrol activity
Gov't Doc #: 24342
Appears in Collections:PhD Thesis of All Public / Private Sector Universities / DAIs.

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