LAUSR

Simple Summary The physicochemical conditions of thermal springs are one of the most significant barriers for detecting microbial life. According to the findings of various studies, high-throughput DNA sequencing technology can be utilized to perform more precise and detailed microbiome assessments. The main goal of this paper was to determine the microbiome in a thermal spring by metabarcoding environmental DNA obtained from four different sources and revealing how temperature and chemical composition affect the microbiome. This research also aimed to gather information that will aid in determining the best gene region and bioinformatic pipeline. The findings revealed a link between four different thermal springs’ physicochemical parameters and microbial composition and we found various manipulable steps in this study. This research is also first comprehensive thermal spring metabarcoding study conducted in Turkey. Abstract One of the most significant challenges for detecting microbial life in thermal springs by conventional techniques such as culturing is these places’ physicochemical (temperature, heavy metal content, pH, etc.) conditions. Data from several studies suggest that high-throughput DNA sequencing technologies can be used to perform more accurate and detailed microbiome analyses. The primary aim of this paper was to determine the microbiome in the thermal source by metabarcoding environmental DNA isolated from four different sources and reveal the reflection of differences caused by temperature and chemical content on the microbiome. DNA was extracted from water filtered with enclosed filters and using the Illumina high-throughput sequencing platform, V3 and V4 regions of the 16S rRNA gene were sequenced. The results showed a correlation between physicochemical conditions and microorganism composition of four different thermal springs. Springs with extremely high temperature (89–90 °C) were dominated by hyperthermophiles such as Hydrogenobacter and Thermus, while a spring with a high temperature (52 °C) was dominated by thermophiles such as Thermoanaerobaculum and Desulfurispora, and a spring with a low temperature (26 °C) and high salinity was dominated by halophiles and sulfur-oxidizers such as Hydrogenovibrio and Sulfirimonas. With this research, we observed many manipulable steps according to the work of interest. This study sought to obtain data that will help decide the right gene region and choose the optimal bioinformatic pipeline.