DISCOVERY OF POLYGALACTURONASE PRODUCING BACILLUS TEQUILENSIS STRAIN ARMATI USING 16S rRNA GENE SEQUENCING
Objective: To identify a novel strain of polygalacturonase producing Bacillus tequilensis from poultry farm.
Methods: Poultry feces sample was serially diluted, and the pure isolate was subjected to morphological tests, biochemical tests, genomic DNA
isolation, polymerase chain reaction amplification, amplicon purification and 16S rRNA gene sequencing. The strain was screened for polygalacturonase
production using plate assay method. The nucleotide sequences obtained from the isolate were subjected to BLAST tool for the pairwise alignment.
RNA secondary structure was predicted through RNAStructure Web Server. Multiple antibiotic resistance (MAR) index was determined against six
antibiotics through disc diffusion method. Phylogenetic trees were inferred using the neighbor-joining algorithm in Molecular Evolution Genetic
Analysis software version 4.0.
Results: B. tequilensis strain ARMATI showed the production of polygalacturonase. MAR index of this novel strain was found to be zero. RNA
secondary structure with a minimum free energy of âˆ’281.40 kcal/mol was obtained. Phylogenetic tree of Bacillus 16S rRNA genes separate each
Bacillus according to their taxonomic positions and were supported statistically.
Conclusion: The present investigation clearly indicates the isolation, molecular characterization and sequence analysis of B. tequilensis strain ARMATI
from poultry farm, and industrial application of this novel strain for the production of the enzyme.
Keywords: Bacillus tequilensis, Multiple antibiotic resistance index, Polygalacturonase, Phylogenetic tree, RNA secondary structure, RNAStructure
Gummadi SN, Panda T. Purification and biochemical properties of microbial pectinases- A review. Process Biochem 2003; 38: 987â€“996.
Aguilar G, Trejo T, Garcia J, Huitron G. Influence of pH on endo and exo-pectinase production by Aspergillus species CH-Y-1043. Canad J Microbiol 1991; 37: 912â€“917.
Sakai T. Degradation of pectins in Microbial Degradation of Natural Products. G. Winkelmann, 1992, Ed: 57-81
Rehman HU, Shah AUQ, Aman A. Polygalacturonase: Production of pectin depolymerising enzyme from Bacillus licheniformis KIBGE IB-21. Carbohydrate Poly 2012; 90: 387â€“ 391.
Dosanjh N, Hoondal GS. Production of constitutive thermostable hyperactive exo-pectinase from Bacillus GK-8. Biotechnol Lett 1996; 18: 1435â€“1438.
Baracat MC, Vanetti MC, Araujo EF, Silva DO. Growth conditions of a pectinolytic Aspergillus fumigates for degumming of natural fibres. Biotechnol Lett 1991; 13(10): 693-696
Bruhlmann F, Leupin M, Erismann KH, Fiechter A. Enzymatic degumming of ramie bast fibres. J Biotechnol 2000; 76(1):43-50
Hoondal GS, Tiwari RP, Tiwari R, Dahiya N, Beg QK. Microbial alkaline pectinases and their industrial application: A review. Appl Microbiol Biotechnol 2002; 59: 409â€“418.
Maldonado MC, CÃ¡ceres S, Galli E, Navarro AR. Regulation of the production of polygalacturonase by Aspergillus niger. Folia Microbiologica (Praha) 2002; 47: 409â€“412.
Saleh M, Seedorf J, Hartung J. Total count of bacteria in the air of three different laying hen housing systems. Dtsch Tierarztl Wochenschr 2003; 110(9): 94-97.
Rahman MT, Saha S, Islam MA. Prevalence of Salmonella in Poultry feed of Mymensingh, Bangladesh. The Bangladesh Vet 1999; 16: 45-46.
Claus D, Berkeley RCW. Genus Bacillus Cohn 1872, 174AL. In Bergeyâ€™s Manual of Systematic Bacteriology.1986. vol. 2, pp. 1105â€“1139. Edited by P. H. A. Sneath, N. S. Mair, M. E. Sharpe & J. G. Holt. Baltimore: Williams & Wilkins
Gatson JW, Bruce F, Benz BF, Chandrasekaran C, Satomi M, Venkateswaran K et al. Bacillus tequilensis sp. nov., isolated from a 2000-year-old Mexican shaft-tomb, is closely related to Bacillus subtilis. Int J Syst Evolution Microbiol 2006; 56: 1475â€“1484
Shah KP, Chandok HK, Rathore P, Sharma MV, Yadav M, Nayarisseri SA. Screening, isolation and identification of Polygalacturonase producing Bacillus tequilensis strain EMBS083 using 16S rRNA gene sequencing. Europ J Biolog Sci 2013; 5(1): 09-13
Tamura K, Dudley J, Nei M, Kumar S. MEGA4: Molecular Evolutionary Genetics Analysis (MEGA) software version 4.0. Mol Biol Evol 2007; 24: 1596â€“1599.
Krumperman PH. Multiple antibiotic resistance indexing E.coli to identifying riok sources of faecal contamination of foods. Appl Environ Microbiol 1983; (46): 165-170
Xu D, Cote JC. Phylogenetic relationships between Bacillus species and related genera inferred from comparison of 3â€² end 16S rDNA and 5â€² end 16S-23S ITS nucleotide sequences. Int J Syst Bacteriol 2003; 53: 695â€“704.
Emanuilova EL, Toda K. Î±-amylase production in batch and continuous cultures by Bacillus caldolyticus. Appl Microbiol Biotechnol 1984; 19: 301-305.
Miranda CA, Martins OB, Clementino MM. Species-level identification of Bacillus strains isolates from marine sediments by conventional biochemical, 16S rRNA gene sequencing and inter-tRNA gene sequence lengths analysis. Anton Van Leeuwenhoek 2008; 93: 297â€“304.
Khusro A, Preetamraj JP, Panicker SG. A Comparative analysis of antibacterial activity of Citrus limonium juice extracts, antibiotics and commercially available citric acid against new strains of bacteria for the prevention of eye infections. Int J Adv Res 2013; 1(9): 104-111.
Khusro A, Aarti C, Preetamraj JP, Panicker SG. In vitro Studies on antibacterial activity of aqueous extracts of spices and vegetables against Bacillus licheniformis strain 018 and Bacillus tequilensis strain ARMATI. Int J Curr Microbiol App Sci 2013; 2(9): 79-88
Bonala KC, Mangamoori LN. Production and optimization of lipase from Bacillus tequilensis NRRL B-41771. Int J Biotechnol Appl 2012; 4(1): 134-136
Lin X. Kelemen DW, Miller ES, Shih JCH. Nucleotide sequence and expression of kerA, the gene encoding a keratinolytic protease of Bacillus licheniformis PWD-1. Appl Environ Microbiol 1995; 61: 1469-1474.
Jahan Z, Khan SN, Hoq MM. Screening of keratinolytic bacteria from poultry wastes. Bangladesh J Sci Ind Res 2010; 45(3): 261-266
Mehta RS, Jholapara RJ, Sawant CS. Isolation of a novel feather-degrading bacterium and optimization of its cultural conditions for enzyme production. Int J Pharm Pharm Sci 2014; 6 (1): 194-201
Fox GE, Wisotzkey JD, Jurtshunk JP. How close is close: 16S rRNA sequence identity may not be sufficient to guarantee species identity. Int J Syst Bacteriol 1992; 42: 166â€“170.
Mollet C, Drancourt M, Raoult D. rpoB sequence analysis as a novel basis for bacterial identification. Mol Microbiol 1997; 26: 1005â€“1011.
Blackwood KS, Turenne CY,Harmsen D, Kabani AM. Reassessment of sequence based targets for identification of Bacillus species. J Clin Microbiol 2004; 42: 1626â€“1630.
Shangkuan YH, Yang JF, Lin HC, Shaio MF. Comparison of PCR-RFLP, ribotyping and ERIC-PCR for typing Bacillus anthracis and Bacillus cereus strains. J Appl Microbiol 2000; 89: 452â€“462.
Goto K, Omura T, Hara Y, Sadaie Y. Application of the partial 16S rDNA sequence as an index for rapid identification of species in the genus Bacillus. J Gen Appl Microbiol. 2000; 46: 1â€“8.
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