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Identification and characterization of thermophilic amylase producing bacterial isolates from the brick kiln soil

ABSTRACT: The present experiment was designed to isolate bacterial strains from the brick kiln soil and to check the activity and enzyme kinetics of amylase from these isolates. The bacterial colonies were isolated from soil samples through the serial dilution method. The bacterial isolates were identified through morphological, electron microscopic and molecular analysis. The 16S ribosomal RNA sequences of the isolates IR-1, IR-2, IR-3, IR-8, and IR-9 showed high similarities with Bacillus tequilensis, Bacillus paramycoides, Proteus alimentorum, Bacillus wiedmannii, and Pseudomonas aeruginosa, respectively. All of the bacterial isolates showed a positive catalase activity except IR-9. Furthermore, the isolates showed variable antagonistic effects against different bacterial pathogens. All of the strains produced indole acetic acid (IAA), and the concentrations increased in the presence of tryptophan application. The isolates showed the amylase enzyme activity and maximum activity of isolates was achieved in 4% starch concentration. The IR-9 isolate showed the highest amylase activity of 5.9 U/ml. The Vmax values of the extracellular amylase from different bacterial isolates ranged between 12.90 and 50.00 IU ml?1. The lowest Km value of 6.33 mg starch was recorded for IR-8 and the maximum Kcat value of 2.50 min?1 was observed for IR-3. The amylase activity of the isolates was significantly affected by a range of different incubation time, temperature, and pH values. Further tests are required before the potential utilization of these isolates for amylase production, and in the biopesticide and biofertilizer applications.

SUBMITTER: Ullah I

PROVIDER: S-EPMC7783820 | biostudies-literature | 2020 Nov

REPOSITORIES: biostudies-literature

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Project description:Polychlorinated biphenyls (PCBs) are persistent organic pollutants widely distributed in the environment and possess deleterious health effects. The main objective of the study was to obtain bacterial isolates from PCB-contaminated soil for enhanced biodegradation of PCB-77. Selective enrichment resulted in the isolation of 33 strains of PCB-contaminated soil nearby Bhilai steel plant, Chhattisgarh, India. Based on the prominent growth using biphenyl as the sole carbon source and the confirmation of its degradation by GC-MS/MS analysis, four isolates were selected for further study. The isolates identified by 16S rRNA gene sequencing were Pseudomonas aeruginosa MAPB-2, Pseudomonas plecoglossicida MAPB-6, Brucella anthropi MAPB-9, and Priestia megaterium MAPB-27. The isolate MAPB-9 showed a degradation of 66.15% biphenyl, while MAPB-2, MAPB-6, and MAPB-27 showed a degradation of 62.06, 57.02, and 56.55%, respectively in 48 h. Additionally, the degradation ability of these strains was enhanced with addition of co-metabolite glucose (0.2%) in the culture medium. Addition of glucose showed 100% degradation of biphenyl by MAPB-9, in 48 h, while MAPB-6, MAPB-2, and MAPB-27 showed 97.1, 67.5, and 53.3% degradation, respectively as analyzed by GC-MS/MS. Furthermore, in the presence of inducer, PCB-77 was found to be 59.89, 30.49, 27.19, and 4.43% degraded by MAPB-6, MAPB-9, MAPB-2, and MAPB-27, respectively in 7 d. The production of biosurfactants that aid in biodegradation process were observed in all the isolates. This was confirmed by ATR-FTIR analysis that showed the presence of major functional groups (CH2, CH3, CH, = CH2, C-O-C, C-O) of the biosurfactant. The biosurfactants were further identified by HPTLC and GC-MS/MS analysis. Present study is the first to report PCB-77 degradation potential of Pseudomonas aeruginosa, B. anthropi, Pseudomonas plecoglossicida, and Priestia megaterium. Similarly, this is the first report on Pseudomonas plecoglossicida and Priestia megaterium for PCB biodegradation. Our results suggest that the above isolates can be used for the biodegradation of biphenyl and PCB-77 in PCB-contaminated soil.

Project description:BACKGROUND:One of the most promising technologies to sustainably produce energy and to mitigate greenhouse gas emissions from combustion of fossil energy carriers is the anaerobic digestion and biomethanation of organic raw material and waste towards biogas by highly diverse microbial consortia. In this context, the microbial systems ecology of thermophilic industrial-scale biogas plants is poorly understood. RESULTS:The microbial community structure of an exemplary thermophilic biogas plant was analyzed by a comprehensive approach comprising the analysis of the microbial metagenome and metatranscriptome complemented by the cultivation of hydrolytic and acido-/acetogenic Bacteria as well as methanogenic Archaea. Analysis of metagenome-derived 16S rRNA gene sequences revealed that the bacterial genera Defluviitoga (5.5 %), Halocella (3.5 %), Clostridium sensu stricto (1.9 %), Clostridium cluster III (1.5 %), and Tepidimicrobium (0.7 %) were most abundant. Among the Archaea, Methanoculleus (2.8 %) and Methanothermobacter (0.8 %) were predominant. As revealed by a metatranscriptomic 16S rRNA analysis, Defluviitoga (9.2 %), Clostridium cluster III (4.8 %), and Tepidanaerobacter (1.1 %) as well as Methanoculleus (5.7 %) mainly contributed to these sequence tags indicating their metabolic activity, whereas Hallocella (1.8 %), Tepidimicrobium (0.5 %), and Methanothermobacter (<0.1 %) were transcriptionally less active. By applying 11 different cultivation strategies, 52 taxonomically different microbial isolates representing the classes Clostridia, Bacilli, Thermotogae, Methanomicrobia and Methanobacteria were obtained. Genome analyses of isolates support the finding that, besides Clostridium thermocellum and Clostridium stercorarium, Defluviitoga tunisiensis participated in the hydrolysis of hemicellulose producing ethanol, acetate, and H2/CO2. The latter three metabolites are substrates for hydrogentrophic and acetoclastic archaeal methanogenesis. CONCLUSIONS:Obtained results showed that high abundance of microorganisms as deduced from metagenome analysis does not necessarily indicate high transcriptional or metabolic activity, and vice versa. Additionally, it appeared that the microbiome of the investigated thermophilic biogas plant comprised a huge number of up to now unknown and insufficiently characterized species.

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Identification and characterization of thermophilic amylase producing bacterial isolates from the brick kiln soil

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