Screening and characterization of potent poly glutamic acid producing Bacillus sp. isolated from Kinema, water and soil samples

Microbially produced gamma poly glutamic acid (γ–PGA) is a commercially important biopolymer with many applications in foods and various other substances and are abundantly used in different parts of the world. With an aim to study the potent γ–PGA producing Bacillus species, a total of 47 different samples (Kinema, soil, and water) were randomly collected from different locations across the country, and Bacillus sp. were selectively isolated, screened, and characterized by performing physiological, biochemical, morphological, and 16S rRNA gene sequencing. The microbial production of γ–PGA was assayed with the selected isolates on the PGA medium and the metabolite obtained was recovered by ethanol precipitation method and further characterized by thin-layer chromatography (TLC). Thermotolerance (25–60 °C), pH tolerance (4–9), and NaCl tolerance (1–9%) tests were performed to optimize the bacterial growth and γ–PGA production and its viscosity were measured by Ostwald's viscometer. Out of 145 randomly selected colonies, 63 isolates were Gram-positive, rods, and endospore producers and were presumptively confirmed as genus Bacillus. Higher growth of γ–PGA producers were reported in 22 isolates and was found at optimum conditions such as temperature (30–37 °C), pH (6.5–7), incubation time (3 days), and NaCl concentration (3%) and γ–PGA thus produced was further verified by TLC with the retention factor (RF) value 0.27. The potent isolates were closely similar to Bacillus subtilis subsp. stercoris, Bacillus cereus, Bacillus paranthracis, and Bacillus licheniformis etc. Based on the findings of the study, B. licheniformis is the most potent γ–PGA producing Bacillus sp. which can further be used for the commercial production of γ–PGA. To the best of our knowledge, there is yet no published research from Nepal showing the production of the γ–PGA although microbially produced γ–PGA are the major constituents in some popular foods in particular communities of the country.