Importance of B4 Medium in Determining Organomineralization Potential of Bacterial Environmental Isolates

Abstract

B4 precipitation medium has been used as the preferred medium for studying mineral precipitation using bacterial strains in vitro since pioneer studies were performed by Boquet and coworkers in 1973. Using this medium, several authors have demonstrated that some environmental isolates were able to precipitate minerals, yet others did not. The main goal of the current study is to understand whether pH and buffer conditions would have a significant effect on mineral precipitation results for environmental isolates grown on B4. For this study, a total of 49 strains isolated from natural environments from Puerto Rico were grown on B4 plates, and their CaCO₃ precipitation potential was investigated. Our findings revealed a strong correlation between a lack of CaCO₃ precipitation and the acidification of the B4 plates by the colonies. The ability to precipitate CaCO₃ could be restored by buffering the B4 medium to a pH of 8.2. Buffering capacity of the medium was proposed to be involved in CaCO₃ precipitation: acid-base titrations conducted on the individual ingredients of B4 showed that yeast extract has a poor buffering capacity between pH 6.5–7.5. This pH range corresponds to the pH of B4 plates [6.87 (±0.05)] prior to the inoculation. This might explain why B4 is such a good precipitation medium: a small variation in the H⁺/OH⁻ balance during microbial growth and precipitation produces rapid changes in the pH of the medium. Finally, an amorphous matrix was distributed within 90% of the examined crystals generated on B4 medium by the environmental strains. Supplemental materials are available for this article. Go to the publisher's online edition of Geomicrobiology Journal to view the free supplemental file.

Keywords:

• bacterial mineralization
• B4 medium
• calcium carbonate precipitation

Acknowledgments

We would like to acknowledge Dr. Roger Ristau of UConn's IMS for his help with the SEM analyses. We also thank Dr. Brunella Perito and Daniele Nosi for the images with the confocal microscope. This work was supported by the Institutional Grant 2010 from the Pontifical Catholic University of Puerto Rico (PUCPR). Partial support for undergraduate students helping in the project was provided by PRL-SAMP (SNF) program at PUCPR. This is UConn's Center for Integrative Geosciences contribution number 18, Grants NSF EAR 1052974 and NSF MCB 0455620.