The Halophilic Microbial Community of Bitter Lake National Wildlife Refuge

Background and study goals: Established in 1937, Bitter Lakes Wildlife Refuge in New Mexico is an important oasis in a seemingly barren desert filled with essential wetland and desert species. The Bitter Lakes serve as an essential water source for the local wildlife. The lakes themselves are hypersaline, resulting in a unique biodiverse ecosystem. The salt lakes are home to microorganisms adapted to saline environments. Studying these microorganisms can help gain important insights into the biogeochemical processes of hypersaline lakes, which are important to understanding the ecology of these lake systems, as well as the evolutionary adaptations associated with saline tolerance. Thus far, no studies have been performed on the halophilic microbial communities found at the Bitter Lakes. This thesis investigates the microbial community of this hypersaline environment by utilizing a combination of culture-dependent and culture-independent methods in microbiology. Microbial community structure was analyzed using 16S rDNA gene sequence analysis from environmental DNA (eDNA) to reveal how the community structure is influenced by seasonal variations and physicochemical/ environmental parameters such as pH, temperature, dissolved oxygen, and salinity levels. A total of 32 unique strains were isolated, characterized, and identified using microscopy, molecular methods, and phylogenetic analysis. In addition, the complete genomes of six of these selected isolates were sequenced and analyzed.

Results: Culture-independent 16S rDNA gene sequence analysis shows that Bacteria dominates across all sampling events and sites, primarily belonging to Proteobacteria, Bacteroidota, and Actinobacteria. These findings are inconsistent with previous studies in other hypersaline environments, which have mostly observed a dominance of archaea. For those studies that did find bacteria to be dominant over archaea, Proteobacteria was the dominant bacterial phylum, which is consistent with this study’s findings. Redundancy analysis (RDA) reveals that the microbiome composition at Bitter Lake National Wildlife Refuge is largely dependent upon salinity, temperature, and seasonal differences across samplings. Salinity differences between sites showed different levels of biodiversity, and temperature across samplings (seasonal differences) showed differences between biodiversity between samplings. Molecular and phylogenetic analysis of the isolated strains reveals identity with the halophile groups of Salimicrobium, Marinococcus, Salinicoccus, Alkalibacillus, Pontibacillus, and Halobacillus. Isolates were found to display orange to off-white coloration on media, consistent with findings in previous studies of halophilic bacteria. However, most isolates were coccus-shaped, whereas most bacteria found in other studies were bacillus-shaped. Whole-genome sequencing analysis identified six unique species of halophiles.