Bacterioplankton community composition along a salinity gradient of sixteen high-mountain lakes located on the Tibetan Plateau, China

Abstract

The influence of altitude and salinity on bacterioplankton community composition (BCC) in 16 high-mountain lakes located at altitudes of 2,817 to 5,134 m on the Eastern Qinghai-Xizang (Tibetan) Plateau, China, spanning a salinity gradient from 0.02% (freshwater) to 22.3% (hypersaline), was investigated. Three different methods, fluorescent in situ hybridization, denaturing gradient gel electrophoresis (DGGE) with subsequent band sequencing, and reverse line blot hybridization (RLB) with probes targeting 17 freshwater bacterial groups, were used for analysis of BCC. Furthermore, the salt tolerances of 47 strains affiliated with groups detected in or isolated from the Tibetan habitats were investigated. Altitude was not found to influence BCC significantly within the investigated range. Several groups of typical freshwater bacteria, e.g., the ACK-M1 cluster and the Polynucleobacter group, were detected in habitats located above 4,400 m. Salinity was found to be the dominating environmental factor controlling BCC in the investigated lakes, resulting in only small overlaps in the BCCs of freshwater and hypersaline lakes. The relative abundances of different classes of Proteobacteria showed a sharp succession along the salinity gradient. Both DGGE and RLB demonstrated that a few freshwater bacterial groups, e.g., GKS98 and LD2, appeared over wide salinity ranges. Six freshwater isolates affiliated with the GKS98 cluster grew in ecophysiological experiments at maximum salinities of 0.3% to 0.7% (oligosaline), while this group was detected in habitats with salinities up to 6.7% (hypersaline). This observation indicated ecologically significant differences in ecophysiological adaptations among members of this narrow phylogenetic group and suggested ecological significance of microdiversity.

FIG. 1.

(A to C) Relative abundances of Alphaproteobacteria (A), Betaproteobacteria (B), and Gammaproteobacteria (C) in Tibetan lakes along a salinity gradient as determined by FISH. (D) Number of DGGE bands (gray squares) and number of RLB probes with positive hybridization signals (black diamonds). Trends in changes of numbers with salinity are indicated by first (gray line) and second (black line) order regression lines. (E) Maximum salinity at which the growth of cultivated representatives of nine bacterial taxa was observed in laboratory experiments (black bars) and maximum salinity at which the respective bacterial groups were detected in the environment by RLB (gray bars). The results shown for the nine different taxa represent results from 2 to 15 investigated strains (see Table S3 in the supplemental material). The black bars indicate the average maximum salinities at which growth of strains was observed, and the narrow extension indicates the highest salinity at which a member of the group grew. The open squares overlaying the respective bars indicate the salinity of the habitats from which the Tibetan strains were obtained. RLB probes were available only for the lower 10 bacterial groups. Sph. sp. Oulin, Sphingomonas sp. “Oulin”; Sphing. sp. Xin, Sphingomonas sp. “Xin”; Polynuc., Polynucleobacter.

FIG. 2.

CCA biplot based on FISH and environmental data of 15 investigated lakes (see Table S1 in the supplemental material). For Lake Qinghai and Namocuo, only data from the epilimnion were used. The analyzed bacterial groups were Alphaproteobacteria, Betaproteobacteria, Gammaproteobacteria, Actinobacteria, and Bacteroidetes (CFB bacteria). Salinity was the dominant factor significantly (P < 0.05) explaining the differences in composition of major bacterial groups. The x axis explained 37% of the observed variation.

FIG. 3.

CCA biplot based on DGGE data and environmental factors of 11 investigated lakes (see Table S1 in the supplemental material). The plot illustrates the differences in BCC in relation to the two environmental factors significantly (P < 0.05) influencing BCC. For Lake Qinghai and Namocuo, only data from the epilimnion were used. The eigenvalues of the x and y axes were 0.97 and 0.63, respectively. The two axes explained 45% of the observed variation in BCC. Salinity was strongly correlated with the x axis.

FIG. 4.

Phylogenetic affiliation of the 88 sequences obtained from DGGE bands. The origins of sequences from habitats with salinities of ≤2 g liter⁻¹ (freshwater and oligosaline) and >22 g liter⁻¹ (polysaline and hypersaline) were distinguished. The pie chart shows the contribution of the two habitat types to the total number of analyzed sequences.