Habitat specificity and co-occurrence network of bacterial communities in the Xincuntun Cave, Guilin, Guangxi

Abstract

Although karst systems comprise approximately 5%-20% of the Earth's ice-free land surface, only 10% of them have so far been explored. Karst caves are important components of karst systems and represent one of the most unique and poorly understood ecosystems on Earth. They are extreme environments in terms of nutrition deficiency, permanent darkness, and isolation from other ecosystems, and serve as a natural laboratory to study the deep biosphere. Caves are also ideal places to discover microbial dark matter such as microbial biodiversity and novel metabolic pathways and metabolites. Microorganisms in caves have been demonstrated to be involved in the dissolution and formation of minerals, biogeochemical cycles, and degradation of organic matter and thus have a fundamental role in sustaining the cave ecosystems. However, knowledge about microbial distribution, diversity, potential functions, and interactions in subsurface caves has remained limited to date particularly in different niches, which limits the utilization of microbial resources and hampers our understanding of biogeochemical processes in the deep biosphere. To address these issues, bacterial communities from seven different niches including cave wall, weathered crust, loose sediments, biofilms from the wet surface of stalagmite, biofilms from the dry surface of stalagmite, microbial mat on rock surface, and overlying soils in the Xincuntun Cave, Southwest China, were investigated via high-throughput sequencing and bioinformatics. Alpha diversities of bacterial communities significantly differed from one niche to the other. Specifically, bacterial diversity was the highest in the overlying soil, followed by those in weathering crust, sediments, wet surface of stalagmite, cave wall, dry surface of stalagmite, and microbial mat. Bacterial communities were dominated by Actinobacteria and Proteobacteria and showed high specificity for their niches as indicated by principal coordinates analysis (PCoA) and bacterial indicator groups identified by the linear discriminant analysis effect size (LEfSe) method. Nitrospira and Polycyclovoran were indicator genera on the dry surface of stalagmite. Candidatus Udaeobacter, Vicinamibacter, Bacillus, and Longimicrobium were indicator genera in overlying soils, microbial mat, sediments, and weathering crust, respectively. Redundancy analysis was conducted to investigate the relationship between environmental factors and bacterial communities. Results showed that pH, TOC, and Mg/Si (chemical weathering index) were of significance to construct bacterial communities. The pH had significantly positive association with Gammaproteobacteria, Rokubacteria, and Nitrospirae, whereas it was negatively related with Actinobacteria. TOC had a significantly positive correlation with Alphaproteobacteria, Gammaproteobacteria, and Planctomycetes and negative correlation with Acidobacteria. Bacterial interactions among different groups were analyzed via co-occurrence networks. The results indicated a high-modularity network with Gammaproteobacteria, Candidatus Udaeobacter, and Thermoleophilia, being the top-three keystone members, which may contribute to the stability of microbial network. The high proportion of positive links between nodes inferred fundamental cooperation among different microbial groups surviving under the extreme conditions in karst caves. Our results revealed habitat specificity of bacterial communities, microbial interactions, and environmental driving forces for bacterial communities in the Xincuntun Cave, enhancing our understanding of the deep terrestrial biosphere.