Historical contingencies and phage induction diversify bacterioplankton communities at the microscale

Abstract

In many natural environments, microorganisms decompose microscale resource patches made of complex organic matter. The growth and collapse of populations on these resource patches unfold within spatial ranges of a few hundred micrometers or less, making such microscale ecosystems hotspots of heterotrophic metabolism. Despite the potential importance of patch-level dynamics for the large-scale functioning of heterotrophic microbial communities, we have not yet been able to delineate the ecological processes that control natural populations at the microscale. Here, we address this challenge by characterizing the natural marine communities that assembled on over 1,000 individual microscale particles of chitin, the most abundant marine polysaccharide. Using low-template shotgun metagenomics and imaging, we find significant variation in microscale community composition despite the similarity in initial species pools across replicates. Chitin-degrading taxa that were rare in seawater established large populations on a subset of particles, resulting in a wide range of predicted chitinolytic abilities and biomass at the level of individual particles. We show, through a mathematical model, that this variability can be attributed to stochastic colonization and historical contingencies affecting the tempo of growth on particles. We find evidence that one biological process leading to such noisy growth across particles is differential predation by temperate bacteriophages of chitin-degrading strains, the keystone members of the community. Thus, initial stochasticity in assembly states on individual particles, amplified through ecological interactions, may have significant consequences for the diversity and functionality of systems of microscale patches.

Keywords: community assembly; historical contingencies; marine particles; microscale; prophages.

Fig. 1.

Modeling POM degradation with a laboratory system of enriching of marine microbes on chitin particles. (A) Microscale marine particles are discrete, spatially separated nutrient-rich habitats dynamically populated and degraded by complex communities of heterotrophic bacteria. The interparticle distance range is estimated from data reported in Simon et al. (16). (B) Schematic depicting experimental design and analysis. Microscale chitin particles were individually incubated in seawater, and the DNA content of particle-attached communities was quantified and submitted for shotgun metagenomic sequencing. Communities were characterized using MAGs, which were classified into three predicted ecological roles for this ecosystem: chitin degraders, chitooligosaccharide exploiters, and metabolic byproduct scavengers.

Fig. 2.

High compositional variability across replicate endpoint particles is driven by conditionally rare degrader taxa. Smaller black dots indicate the relative abundance of each MAG per endpoint particle (n = 149), with larger white dots indicating the log₁₀[mean relative abundance] across the particles on which the MAG was found. Light red highlights indicate which MAGs display the “jackpot” phenomenon. MAGs are sorted from left to right by their prevalence across particles (i.e., the percent of particles on which they are detected; top bar graph), with the number of chitinases encoded in each MAG indicated in the bar graph beneath. The annotations at the bottom of the plot show each MAG’s taxonomic order (E, Enterobacterales; R, Rhodobacterales; P, Pseudomonadales; F, Flavobacteriales; C, Cytophagales; O, other). See SI Appendix, Fig. S1 for additional details.

Fig. 3.

Endpoint particles diverge in community-level functional potential and biomass. (A) Ternary plot of the relative abundances of organisms occupying the three ecological roles (degrader, exploiter, scavenger) on each endpoint particle (n = 149), calculated by summing the relative abundances of MAGs classified into each role. Red dots represent jackpot particles, and black ones represent nonjackpot particles. Jackpot particles harbored significantly higher degrader populations than nonjackpot particles (79.8% vs. 47.4% on average; Mann–Whitney U test: P < 2.2 × 10⁻¹⁶). (B) Estimates of absolute bacterial cell counts on endpoint particles through qPCR of the 16S rRNA gene in DNA extracted from particle-attached communities. Jackpot particles (red dots) harbored significantly higher numbers of cells (Mann–Whitney U test: P = 2.3 × 10⁻⁷) than nonjackpot particles (black dots). (C) Representative images of endpoint particles that were harvested after 167 h of incubation in seawater and stained with the DNA-intercalating dye SYTO 9. (Scale bar, 50 µm.) Particle-attached communities spanned a range of growth states, from sparsely to densely populated.

Fig. 4.

Bacteriophages become increasingly activated during community development and contribute to variability in bacterial abundances on endpoint particles. (A) Schematic of approach to detect productive phage infections from metagenomic data. (Left) During lysogenic infections, prophages replicate with their bacterial hosts (VMR ≈ 1; Top); lysogenic phage contigs have read coverage values similar to those of most bacterial contigs of their host MAG (Bottom). (Right) During productive infections, prophages replicate much more than their hosts (VMR ≫ 1; Top); productive phage contigs have read coverage values much higher than those of most bacterial contigs of their host MAG (Bottom). (B) Representative examples of phages with lysogenic coverage patterns on all endpoint particles (top three rows), and of phages with productive coverage patterns on a subset of particles (bottom three rows). For each phage contig, VMR is shown across endpoint particles on which each MAG is present; gray dots, particles on which the phage contig is not a coverage outlier; red dots, particles on which the phage is a high coverage outlier; dashed line, VMR = 1. (C) Total VMRs for productive phages over time. The first time point shows productive VMRs of initial seawater samples; subsequent time points show productive VMRs for chitin particle-attached communities incubated in seawater; smaller red dots, values for individual samples; larger white dots, mean VMR for each time point. (D) Metabolomic profiles of the seawater surrounding chitin particles as a function of incubation duration. Values are depicted in terms of fold-change at each time point relative to the first time point (dashed line, no change); red line (and shading), mean (±1 SD) weighted ion intensity (Materials and Methods); blue line, number of unique metabolites. (E) Absolute bacterial cell counts on endpoint particles (n = 142), estimated through qPCR, vs. each particle’s total VMR for productive phages. Cell counts were negatively correlated with productive VMRs (Spearman’s ρ = −0.56, P = 3.3 × 10⁻¹³; red line and shading, log–log linear regression and 95% CI; R² = 0.23, P = 1.3 × 10⁻⁹). Marginal histograms are distributions of productive VMRs (red) and bacterial cell counts (light gray). (Inset) Bar plot of values of Spearman’s ρ between cell counts and productive VMRs of bacterial populations by ecological role (blue, degraders; green, exploiters; yellow, scavengers; see SI Appendix, Fig. S14B for details).

Fig. 5.

Conceptual model of key processes contributing to the diversification of communities on microscale particles. Schematics of community development over time and the resultant features measured on particles are shown for different scenarios highlighting how historical contingencies and phage induction, particularly affecting keystone degraders, can contribute to variability in microscale community assembly. Bacterial populations are indicated by their ecological role (blue, degraders; green, exploiters; yellow, scavengers), and phages are depicted in red. (A) Jackpot particles are those on which degraders arrive early and proliferate, leading to communities with high relative and absolute degrader abundances and low species diversity. (B) Low-biomass particles are those on which degraders are not able to proliferate, either because they become established on a particle relatively late due to stochastic arrival (Top) or because phage induction impedes the establishment of a robust population (Bottom).