Diversity and protease production potential of haloalkaliphilic bacteria isolated from the Kesongo Mud Volcano of Indonesia

Aims: Extremophiles are microorganisms capable of surviving in extreme environments and producing stable enzymes called extremozymes, such as proteases.One example of extremophilic bacteria is haloalkaliphilic bacteria, which can thrive under high salinity and alkaline pH conditions.The Kesongo Mud Volcano in Blora, Central Java, Indonesia, offers potential as a source of haloalkaliphilic bacteria with protease-producing capability.The aim of this study was to isolate, characterize morphologically, screen proteolytic haloalkaliphilic bacteria, determine the optimal temperature and pH for protease activity of the best isolate, and identify the best isolate using molecular methods.Methodology and results: Samples were collected from two points in the Kesongo Mud Volcano, and bacterial isolation was performed using modified Luria Bertani media.Proteolytic activity was screened using Skim Milk Agar media, and the best isolate was selected based on its proteolytic index.Further tests were conducted to optimize protease production in terms of temperature and pH.Molecular identification involved 16S rRNA gene amplification and phylogenetic analysis.Out of 35 isolates, 17 showed proteolytic activity, with BK4 exhibiting the highest activity (proteolytic index 1.66).Optimization showed BK4 produced maximal protease at pH 11 and 37 °C.Molecular analysis revealed 99.07%similarity of BK4 with Halomonas sp.SSL14.Conclusion, significance and impact of study: These findings highlight the potential of haloalkaliphilic bacteria from extreme environments as a source of industrially relevant enzymes, emphasizing the value of microbial bioprospecting in geothermal ecosystems.