Bacterioplankton Communities in Dissolved Organic Carbon-Rich Amazonian Black Water

Abstract

The Amazon River basin sustains dramatic hydrochemical gradients defined by three water types: white, clear, and black waters. In black water, important loads of allochthonous humic dissolved organic matter (DOM) result from the bacterioplankton degradation of plant lignin. However, the bacterial taxa involved in this process remain unknown, since Amazonian bacterioplankton has been poorly studied. Its characterization could lead to a better understanding of the carbon cycle in one of the Earth's most productive hydrological systems. Our study characterized the taxonomic structure and functions of Amazonian bacterioplankton to better understand the interplay between this community and humic DOM. We conducted a field sampling campaign comprising 15 sites distributed across the three main Amazonian water types (representing a gradient of humic DOM), and a 16S rRNA metabarcoding analysis based on bacterioplankton DNA and RNA extracts. Bacterioplankton functions were inferred using 16S rRNA data in combination with a tailored functional database from 90 Amazonian basin shotgun metagenomes from the literature. We discovered that the relative abundances of fluorescent DOM fractions (humic-, fulvic-, and protein-like) were major drivers of bacterioplankton structure. We identified 36 genera for which the relative abundance was significantly correlated with humic DOM. The strongest correlations were found in the Polynucleobacter, Methylobacterium, and Acinetobacter genera, three low abundant but omnipresent taxa that possessed several genes involved in the main steps of the β-aryl ether enzymatic degradation pathway of diaryl humic DOM residues. Overall, this study identified key taxa with DOM degradation genomic potential, the involvement of which in allochthonous Amazonian carbon transformation and sequestration merits further investigation. IMPORTANCE The Amazon basin discharge carries an important load of terrestrially derived dissolved organic matter (DOM) to the ocean. The bacterioplankton from this basin potentially plays important roles in transforming this allochthonous carbon, which has consequences on marine primary productivity and global carbon sequestration. However, the structure and function of Amazonian bacterioplanktonic communities remain poorly studied, and their interactions with DOM are unresolved. In this study, we (i) sampled bacterioplankton in all the main Amazon tributaries, (ii) combined information from the taxonomic structure and functional repertory of Amazonian bacterioplankton communities to understand their dynamics, (iii) identified the main physicochemical parameters shaping bacterioplanktonic communities among a set of >30 measured environmental parameters, and (iv) characterized how bacterioplankton structure varies according to the relative abundance of humic compounds, a by-product from the bacterial degradation process of allochthonous DOM.

Keywords: Acinetobacter; Methylobacterium; Polynucleobacter; bacterioplankton; carbon cycle; dissolved organic carbon; dissolved organic matter; humic acids; microbiome.

FIG 1

Main steps of biological degradation of lignin. Humic dissolved organic matter (DOM) (in red) is produced after the initial oxidation of lignin-derived compounds (58, 68). CDOM, chromophoric dissolved organic matter.

FIG 2

Distance-based redundancy analyses (RDAs) of global and transcriptionally active bacterioplankton communities, and their functional profiles significantly cluster according to water type. RDA ordination plots of sampling sites according to the global bacterioplankton taxonomic structure (a), transcriptionally active bacterioplankton taxonomic structure (b), and inferred functional repertory (c). The inferred functional repertory is based on the taxonomy of the 16S rRNA amplicon data. The samples are colored according to their water type. The P and F values are from the analysis of variance (ANOVA) tests done on the RDA solution. Environmental parameters associated with dissolved organic carbon (DOC) concentration or fluorescent dissolved organic matter (FDOM) components are in red. ***, P < 0.001. Selection of environmental parameters was done according to their variance inflation factors and by using stepwise selection via ordistep.

FIG 3

Random forest (RF) machine-learning analyses identified the 40 amplicon sequence variants (ASVs) showing the most important differentiation between water types. The results are shown in heat maps for the taxonomical structure of the global bacterioplankton community (a) and the transcriptionally active bacterioplankton community data set (b). The heat map columns represent samples, and rows are different ASVs. The taxonomic annotation of each ASV was done at the highest resolution possible for each ASV. The colored tag at the left of each row corresponds to the ASV taxonomic assignation. Black boxes identify to which water type the taxa were associated (i.e., in which water type they were most abundant). The relative abundance (color intensity) of each ASV in each sample is scaled according to the abundance of the same ASV in all other samples.

FIG 4

Black water sites showed unique DOC and FDOM profiles, characterized by a higher relative abundance of aromatic, high molecular weight humic carbon. (a) Principal component analysis showing how sampling sites cluster according to their FDOM optical characteristics. The three bigger dots refer to group centroids. The environmental variables are as follows: “DOC” refers to concentration (mg/liter) of DOC; “protein” refers to percentage of protein-like FDOM; “humic” refers to percentage of humic-like FDOM; “fulvic” refers to percentage of fulvic-like FDOM; and SAC340, SUVA254, and abs254.365 refer to absorbance ratios detailed in Table 2. (b to d) Parallel factor analysis (PARAFAC) model components show the presence of humic-like (b), protein-like (c), and fulvic-like (d) FDOM fractions in the sites sampled.

FIG 5

The concentration of humic FDOM correlated with the structure of global and transcriptionally active bacterioplankton communities. (a, b) Ordisurf plots (i.e., RDA plots fitted with isolines describing humic FDOM relative abundance variations) based on the structure of global (a) and transcriptionally active bacterioplankton (b). The samples are colored according to their water type. (c, d) Spearman correlation-based networks of coabundance between humic FDOM and bacterial ASVs from the global bacterioplankton (c) or transcriptionally active bacterioplankton (d) communities. For ease of viewing, only the taxa that correlated directly with humic FDOM or that were direct neighbors of such taxa were kept for network construction. The nodes are colored according to their taxonomic assignation (at the best level possible). Labels in the legend for panels c and d refer to the best level of taxonomic assignation reached for these ASVs. The black lines in panels c and d represent direct correlations between humic FDOM and specific ASVs, while gray lines represent indirect correlations. All correlations in panels c and d are positive correlations. In panel d, lines from the three nodes directly correlated with humic FDOM are colored according to their node taxonomic assignation to highlight the indirect effect of humic FDOM on the transcriptionally active bacterioplankton community. These lines sre not colored in panel c for ease of viewing. FDOM, fluorescent dissolved organic matter; ASV, amplicon sequence variant.

FIG 6

Potential presence of different pathways of the main steps of humic acids’ degradation in the genera that abundance correlated with humic FDOM. Spearman correlation between the relative abundances of humic FDOM and bacterial genera are plotted on a heat map from least to most correlated (left to right, respectively). Polynucleobacter is the genus showing the highest correlation to humic FDOM concentration, followed by Methylobacterium and Acinetobacter. The complete list of pathways that were screened for in the functional repertory can be found in Table S3; however, only the pathways that were found in at least one genus are represented in the heat map for ease of viewing. FDOM, fluorescent dissolved organic matter.

FIG 7

Map of the 15 sampling sites distributed throughout the Brazilian Amazon basin. The colors of the dots represent the water type (black, white, or clear).