A comprehensive census of microbial diversity in hot springs of Tengchong, Yunnan Province China using 16S rRNA gene pyrosequencing

Abstract

The Rehai and Ruidian geothermal fields, located in Tengchong County, Yunnan Province, China, host a variety of geochemically distinct hot springs. In this study, we report a comprehensive, cultivation-independent census of microbial communities in 37 samples collected from these geothermal fields, encompassing sites ranging in temperature from 55.1 to 93.6°C, in pH from 2.5 to 9.4, and in mineralogy from silicates in Rehai to carbonates in Ruidian. Richness was low in all samples, with 21-123 species-level OTUs detected. The bacterial phylum Aquificae or archaeal phylum Crenarchaeota were dominant in Rehai samples, yet the dominant taxa within those phyla depended on temperature, pH, and geochemistry. Rehai springs with low pH (2.5-2.6), high temperature (85.1-89.1°C), and high sulfur contents favored the crenarchaeal order Sulfolobales, whereas those with low pH (2.6-4.8) and cooler temperature (55.1-64.5°C) favored the Aquificae genus Hydrogenobaculum. Rehai springs with neutral-alkaline pH (7.2-9.4) and high temperature (>80°C) with high concentrations of silica and salt ions (Na, K, and Cl) favored the Aquificae genus Hydrogenobacter and crenarchaeal orders Desulfurococcales and Thermoproteales. Desulfurococcales and Thermoproteales became predominant in springs with pH much higher than the optimum and even the maximum pH known for these orders. Ruidian water samples harbored a single Aquificae genus Hydrogenobacter, whereas microbial communities in Ruidian sediment samples were more diverse at the phylum level and distinctly different from those in Rehai and Ruidian water samples, with a higher abundance of uncultivated lineages, close relatives of the ammonia-oxidizing archaeon "Candidatus Nitrosocaldus yellowstonii", and candidate division O1aA90 and OP1. These differences between Ruidian sediments and Rehai samples were likely caused by temperature, pH, and sediment mineralogy. The results of this study significantly expand the current understanding of the microbiology in Tengchong hot springs and provide a basis for comparison with other geothermal systems around the world.

Figure 1. A map that shows the nine springs from both the Ruidian and Rehai locations.

The open symbols represent the geochemistry and the closed symbols represent the microbial communities. The circles represent the water and the squares represent the sediment geochemistry. The colors of each symbol represent the grouping of either microbial communities (from Figure 2) or water and sediment geochemistry (from Figure S1 and S3 for water and sediment/sinter, respectively) based on hierarchical clustering. The microbial community, sediment geochemistry, and water geochemistry were analyzed separately and color comparisons across different symbols cannot be made; however, a similar color within one symbol (for example solid circle) depicts microbial community or geochemical similarities across the sampling sites. XRD mineralogical analysis of the sediment geochemistry at Dagunguo (Dgg) failed. Huitaijing (Htj) did not have any sediment. There were no microbial data for the Zhenzhuquan (Zzq) sediments because of PCR failure. Abbreviations are Jimeiquan (Jmq), Zhenzhuquan (Zzq), Guminquan (Gmq), Huitaijing (Htj), Jinze (Jz), Diretiyanqu (Drty), Gongxiaoshe (Gxs), Shuirebaozhaqu (Srbz), and Dagunguo (Dgg).

Figure 2. Microbial community composition grouped by pH and temperature. A.

UPGMA cluster tree based on Bray-Curtis dissimilarity obtained based on the 97% cutoff level; B. microbial compositions at the phylum level. Filled circles at nodes represent jackknife values. “TFF”, “Sin”, “Sed”, “Str” and “W” in sample ID refer to the tangential flow filtration (TFF) sample (only from Dagunguo), sinter, sediment, streamer and water samples, respectively. The numbers in the site name are the pH and temperature of each site. The prefix “Bac” denotes bacterial phyla, and the prefix “Arch” denotes archaeal phyla. Only the microbial groups with abundance higher than 2% are displayed. The groups with abundances lower than 2% are included as “Others”.

Figure 3. Percentages of different microbial groups in sediments (left panels) and spring water (right panels) from Tengchong.

The size of the green circle on each plot serves as a scale and represents 100% of abundnace. A. Archaeal and Bacterial distributions: higher percentage of Archaea tended to occur in springs with higher temperature and lower pH. B. Aquificae genera distributions: distributions of Hydrogenobacter spp. and Hydrogenobaculum spp.; Hydrogenobacter-related sequences were abundant in neutral or alkaline springs, whereas Hydrogenobaculum-related sequences were abundant in acidic springs. C. Crenarcaheal order distributions: Desulfurococcales and Thermoproteales within class Thermopotei of phylum Crenarchaeota were abundant in neutral-alkaline springs, whereas Sulfolobales within class Thermopotei of phylum Crenarchaeota was abundant in acidic springs. These two groups did not co-exist in the same spring.