Optimization of Bio-Calcification Process for a Newly Isolated Urease Producing Bacterial Strain Advenelle sp. AV1

Abstract

Bio-calcification is a natural phenomenon catalyzed by the urease enzyme, which is produced by a wide range of bacterial strains. The bio-calcification process relies on several physicochemical factors like pH, Ca²⁺, inoculum size (serve as a nucleation site), temperature, etc. In this study, seven urease-producing bacteria (BS-1-BS-7) were isolated from the garden soil of the two locations and the soil collected from a cave located at Sahastradhara, Dehradun Uttarakhand, India. These isolates were tested for urease activity and CaCO₃ precipitation. Among the bacterial isolates, BS-6 produced the maximum ammonium ions and CaCO₃ precipitate (14.6 mg/50 ml) and therefore it was selected for the optimization study. The two important factors – inoculum size (4%, 12%, and 20%; OD value 1.2 at 600 nm) and CaCl₂ concentration (25, 50, and 100 mM) were optimized for bio-calcification process (CaCO₃ precipitation). The optimum inoculum size and CaCl₂ concentration for the CaCO₃ precipitation by the isolate BS-6 were found to be 12% and 50 mM, respectively. Two-way ANOVA (analysis of variance) indicates that both factors, inoculum size and CaCl₂ concentration significantly (p < 0.05) affect the CaCO₃ precipitation. The present study signifies the role of Ca²⁺ and inoculum size in the bio-calcification process. The selected bacterial strain BS-6 was further identified as a novel strain Advenelle sp. AV1 based on the 16S rRNA sequencing. This is the first report that shows Advenelle sp. AV1 as urease producing and bio-calcite forming bacteria.

Keywords:

• Advenelle sp. AV1
• bio-calcification
• CaCO₃ precipitation
• soil bacteria
• urease activity
• urease producing bacteria
• ureolysis

Authors’ contributions

SG & PT: Conducted experiments, collected data, and analyzed samples. VS: Experimental design, supervision, Data analysis, Writing – draft, review, and editing. AD: Conceptualized, Supervision, Writing – review and editing, Funding resources, Project administration, Funding acquisition.

Disclosure statement

No potential conflict of interest was reported by the author(s).

Additional information

Funding

This research work is supported by the Science and Engineering Research Board (SERB), India under the Grant agreement [EEQ/2017/000641].