Linfeng Gong

Five-Year Monitoring of Bacterial Communities in Dripping Water from the Heshang Cave in Central China: Implication for Paleoclimate Reconstruction and Ecological Functions

ABSTRACT Microbial lipids in stalagmites are increasingly used to reconstruct the paleoclimate change, necessitating the investigations on microbial communities in dripping waters. A pilot molecular survey was conducted on bacterial communities of dripping waters at two sites (1D and 3D) in Heshang Cave of Hubei Province in central China for a period of 2008 to 2013. The samples were subjected to genome DNA extraction, 16S rRNA gene amplification, clone library construction and phylogenetic analysis to explore the seasonal variation of bacterial communities and their association with environmental factors including regional air temperature, precipitation, cave temperature, pH, conductivity and dripping rate of the water samples. Seasonal variations were clearly observed in components and diversities of bacterial communities at both sites. Bacterial community was dominated by Gammaproteobacteria in autumn and winter, whereas Betaproteobacteria became dominant in samples collected in summer and spring. Among the environmental factors investigated, regional air temperature was found to have a strong impact on bacterial communities indicated by cluster and redundancy analysis. Moreover the bacterial biodiversity was observed to increase with the temperature rising. Bacteria identified in dripping water were either oligotrophs or able to acquire nutrients from minerals under oligotrophic conditions. They may also be able to induce calcite precipitation in cave systems. Our data shed light on the potential of microbes used as a tool for the reconstruction of paleo-temperature as well as on the ecological functions of bacterial communities in oligotrophic caves.

Phylogenetic diversity of culturable fungi in the Heshang Cave, central China

Caves are nutrient-limited and dark subterranean ecosystems. To date, attention has been focused on geological research of caves in China, whilst indigenous microbial diversity has been insufficiently characterized. Here, we report the fungal diversity in the pristine, oligotrophic, karst Heshang Cave, central China, using a culture-dependent method coupled with the analysis of the fungal rRNA-ITS gene sequences. A total of 194 isolates were obtained with six different media from 14 sampling sites of sediments, weathered rocks, and bat guanos. Phylogenetic analysis clustered the 194 sequenced isolates into 33 genera within 15 orders of three phyla, Ascomycota, Basidiomycota, and Zygomycota, indicating a high degree of fungal diversity in the Heshang Cave. Notably, 16 out of the 36 fungal genera were also frequently observed in solution caves around the world and 23 genera were previously found in carbonate cave, indicating potential similarities among fungal communities in cave ecosystems. However, 10 genera in this study were not reported previously in any solution caves, thus expanding our knowledge about fungal diversity in cave ecosystems. Moreover, culturable fungal diversity varied from one habitat to another within the cave, being the highest in sediments, followed by weathered rocks and bat guanos as indicated by α-diversity indexes. At the genus level, Penicillium accounted for 40, 54, and 52% in three habitats of sediments, weathered rocks, and bat guanos, respectively. Trichoderma, Paecilomyces, and Aspergillus accounted for 9, 22, and 37% in the above habitats, correspondingly. Despite of the dominance of Penicillium in all samples, β-diversity index indicated significant differences between each two fungal communities in the three habitats in view of both the composition and abundance. Our study is the first report on fungal communities in a natural pristine solution cave system in central China and sheds light on fungal diversity and functions in cave ecosystems.

pH Shaping the Composition of sqhC-Containing Bacterial Communities

Sediment and tailings samples were collected from sites with a contrasting physicochemical gradient to investigate microbial squalene-hopene cyclase (sqhC) composition and distribution in terrestrial environments. Acidobacteria (66%), Alphaproteobacteria (96%) and Gammaproteobacteria (55%) were found to dominate sqhC communities, respectively, at the acidic Dajiuhu Peatland, the alkaline Heshang Cave and strongly acidic tailings of Tongling copper mine in China. Statistical analysis confirmed that pH was the important factor impacting the geographical distribution of sqhC at phylum level. sqhC gene abundance is comparable at the three sites. However, the total amount of hopanoids in per gram total organic matter content (TOC) is 1.75 times higher in the acidic peatland than that in the alkaline Cave and it is below the detecting limit in tailings of Tongling copper mine, inferring the potential impact of pH in regulating the hopanoid production.

Distribution of Bathyarchaeota Communities Across Different Terrestrial Settings and Their Potential Ecological Functions

High abundance and widespread distribution of the archaeal phylum Bathyarchaeota in marine environment have been recognized recently, but knowledge about Bathyarchaeota in terrestrial settings and their correlation with environmental parameters is fairly limited. Here we reported the abundance of Bathyarchaeota members across different ecosystems and their correlation with environmental factors by constructing 16S rRNA clone libraries of peat from the Dajiuhu Peatland, coupling with bioinformatics analysis of 16S rRNA data available to date in NCBI database. In total, 1456 Bathyarchaeota sequences from 28 sites were subjected to UniFrac analysis based on phylogenetic distance and multivariate regression tree analysis of taxonomy. Both phylogenetic and taxon-based approaches showed that salinity, total organic carbon and temperature significantly influenced the distribution of Bathyarchaeota across different terrestrial habitats. By applying the ecological concept of ‘indicator species’, we identify 9 indicator groups among the 6 habitats with the most in the estuary sediments. Network analysis showed that members of Bathyarchaeota formed the “backbone” of archaeal community and often co-occurred with Methanomicrobia. These results suggest that Bathyarchaeota may play an important ecological role within archaeal communities via a potential symbiotic association with Methanomicrobia. Our results shed light on understanding of the biogeography, potential functions of Bathyarchaeota and environment conditions that influence Bathyarchaea distribution in terrestrial settings.

Inhibition of bacterial oxidation of ferrous iron by lead nitrate in sulfate-rich systems

Inhibition of bacterial oxidation of ferrous iron (Fe(II)) by Pb(NO(3))(2) was investigated with a mixed culture of Acidithiobacillus ferrooxidans. The culture was incubated at 30 °C in ferrous-sulfate medium amended with 0-24.2 mM Pb(II) added as Pb(NO(3))(2). Anglesite (PbSO(4)) precipitated immediately upon Pb addition and was the only solid phase detected in the abiotic controls. Both anglesite and jarosite (KFe(3)(SO(4))(2)(OH)(6)) were detected in inoculated cultures. Precipitation of anglesite maintained dissolved Pb concentrations at 16.9-17.6 μM regardless of the concentrations of Pb(NO(3))(2) added. Fe(II) oxidation was suppressed by 24.2 mM Pb(NO(3))(2) addition even when anglesite was removed before inoculation. Experiments with 0-48 mM KNO(3) demonstrated that bacterial Fe(II) oxidation decreased as nitrate concentration increased. Therefore, inhibition of Fe(II) oxidation at 24.2 mM Pb(NO(3))(2) addition resulted from nitrate toxicity instead of Pb addition. Geochemical modeling that considered the initial precipitation of anglesite to equilibrium followed by progressive oxidation of Fe(II) and the precipitation of jarosite and an amorphous iron hydroxide phase, without allowing plumbojarosite to precipitate were consistent with the experimental time-series data on Fe(II) oxidation under biotic conditions. Anglesite precipitation in mine tailings and other sulfate-rich systems maintains dissolved Pb concentrations below the toxicity threshold of A. ferrooxidans.

The Physiological Response of Synechococcus elongatus to Salinity: A Potential Biomarker for Ancient Salinity in Evaporative Environments

The purpose of this study was to characterize the physiological response of Synechococcus elongatus, a brackish-water cyanobacterium, to salt stress. S. elongatus was grown in artificial sea water medium with different salinities. The response was measured by analysis of extracellular polymeric substances (EPS) and membrane lipids. The EPS yields were positively correlated (r2 = 0.99) with the salinity. The ratio of unsaturated to saturated fatty acids (U/S) increased with salinity in the range of 2.1‰ to 31.5‰ and decreased at 52.5‰. A positive linear correlation (r2 = 0.92) was observed between the average chain length (ACL) of fatty acids and the salinity. These data indicate that S. elongatus adapted to salt stress by the secretion of EPS and by adjusting the membrane fluidity through the changes in ACL or desaturation of fatty acids. These variations in EPS and fatty acids may be used as geochemical biomarkers in sediments to unravel changes in the salinity of ancient evaporative environments.

Vertical variations and associated ecological function of bacterial communities from Sphagnum to underlying sediments in Dajiuhu Peatland

To investigate vertical changes of bacterial communities from living plants to the associated sediments and bacterial biogeochemical roles in peatland ecosystem, samples of different part of individual Sphagnum palustre and the different layers of the underlying sediments were collected from Dajiuhu Peatland in central China. All samples were subject to 16S rRNA gene clone libraries and quantitive PCR analysis. Even though bacteria vary in abundance at the same order of magnitude in all samples, they show great profile difference in composition from the top part of S. palustre to the low layer of the sediments. Cyanobacteria and alpha-Proteobacteria dominate at the top part whereas Acidobacteria at the middle part of S. palustre. Alpha-Proteobacteria and Acidobacteria are the dominant phyla at the bottom part of S. palustre and in the surface peat sediment. In contrast, bacterial communities in the subsurface sediments are dominated by Acidobacteria. These profile distributions of different bacterial communities are closely related to their ecological functions in the peatland ecosystem. Specifically, most Cyanobacteria were observed at the top green part of S. palustre, a horizon where the active photosynthesis of the moss occurs, which infers their endosymbiosis. In contrast, Acidobacteria, dominant in the subsurface sediments, are able to decompose the specific compounds on the cell wall of Sphagnum moss and thus might play an important role in the formation of the peatland, including the acidic condition. Methane oxidizing process might have been underestimated in Sphagnum peatland due to the identification of Methylocystaceae in all parts of the moss investigated here. The vertical difference in bacterial composition and bacterial ecological functions presented here sheds light on the understanding of biogeochemical processes, in particular the CH4 flux, in peat ecosystems.

Temporal and spatial variations of microbial carbon utilization in water bodies from the Dajiuhu Peatland, central China

To investigate the microbial utilization of organic carbon in peatland ecosystem, water samples were collected from the Dajiuhu Peatland and nearby lakes, central China across the year of 2014. The acridine orange (AO) staining and Biolog Eco microplates were used to numerate microbial counts and determine the carbon utilization of microbial communities. Meanwhile, physicochemical characteristics were measured for subsequent analysis of the correlation between microbial carbon utilization and environmental factors. Results indicated that total microbial counts were between 106–107 cells/L. Microbial diversities and carbon utilization rates showed a similar pattern, highest in September and lowest in November. Microbial communities in the peat pore waters preferred to utilize N-bearing carbon sources such as amines and amino acids compared with microbial communities in lakes. The network analysis of microbial utilization of 31 carbon substrates clearly distinguished microbial communities from peat pore waters and those from lakes. Redundancy analysis (RDA) showed the total organic nitrogen content (P=0.03, F=2.5) and daily average temperature (P=0.034, F=2.4) significantly controlled microbial carbon utilization throughout the sampling period. Our report is the first one to address the temporal and spatial variations of carbon utilization of microbial communities which are closely related to the decomposition of organic matter in the Dajiuhu Peatland in context of climate warming.