Time Series Resolution of the Fish Necrobiome Reveals a Decomposer Succession Involving Toxigenic Bacterial Pathogens

Briallen Lobb,Michael D J Lynch,Jiujun Cheng,Michael J Mansfield,Trevor C Charles,Josh D Neufeld,Andrew C Doxey,Rhiannon Hodgson,Paul M Craig,Simon Lax

doi:10.1128/msystems.00145-20

Abstract

Despite progress understanding microbial communities involved in terrestrial vertebrate decomposition, little is known about the microbial decomposition of aquatic vertebrates from a functional and environmental context. Here, we analyzed temporal changes in the "necrobiome" of rainbow darters, which are common North American fish that are sensitive indicators of water quality. By combining 16S rRNA gene and shotgun metagenomic sequence data from four time points, we studied the progression of decomposers from both taxonomic and functional perspectives. The 16S rRNA gene profiles revealed strong community succession, with early decomposition stages associated with Aeromonas and Clostridium taxa and later stages dominated by members of the Rikenellaceae (i.e., Alistipes/Acetobacteroides genera). These results were reproducible and independent of environmental perturbation, given that exposure to wastewater treatment plant effluent did not substantially influence the necrobiome composition of fish or the associated water sample microbiota. Metagenomic analysis revealed significant changes throughout decomposition in degradation pathways for amino acids, carbohydrates/glycans, and other compounds, in addition to putrefaction pathways for production of putrescine, cadaverine, and indole. Binning of contigs confirmed a predominance of Aeromonas genome assemblies, including those from novel strains related to the pathogen Aeromonas veronii These bins of Aeromonas genes also encoded known hemolysin toxins (e.g., aerolysin) that were particularly abundant early in the process, potentially contributing to host cell lysis during decomposition. Overall, our results demonstrate that wild-caught fish have a reproducible decomposer succession and that the fish necrobiome serves as a potential source of putative pathogens and toxigenic bacteria.IMPORTANCE The microbial decomposition of animal tissues is an important ecological process that impacts nutrient cycling in natural environments. We studied the microbial decomposition of a common North American fish (rainbow darters) over four time points, combining 16S rRNA gene and shotgun metagenomic sequence data to obtain both taxonomic and functional perspectives. Our data revealed a strong community succession that was reproduced across different fish and environments. Decomposition time point was the main driver of community composition and functional potential; fish environmental origin (upstream or downstream of a wastewater treatment plant) had a secondary effect. We also identified strains related to the putative pathogen Aeromonas veronii as dominant members of the decomposition community. These bacteria peaked early in decomposition and coincided with the metagenomic abundance of hemolytic toxin genes. Our work reveals a strong decomposer succession in wild-caught fish, providing functional and taxonomic insights into the vertebrate necrobiome.

Highlights

Despite progress understanding microbial communities involved in terrestrial vertebrate decomposition, little is known about the microbial decomposition of aquatic vertebrates from a functional and environmental context
Individual fish were subjected to decomposition with river water and sediment at room temperature for 1, 4, 8, and 10 days in sterile containers that acted as microcosms of a natural decomposition environment
Sample 16S rRNA gene profiles for fish decomposition microbiomes (“necrobiomes”) for these four time points and two water/sediment sources revealed reproducible microbial communities among independent replicates and between environments. This microbial succession was apparent at the order level of taxonomy (Fig. 2) and at the level of amplicon sequence variants (ASVs) (Fig. 3), variation in ASV composition was evident among fish samples and environments (Fig. 3)

Summary

Introduction

Despite progress understanding microbial communities involved in terrestrial vertebrate decomposition, little is known about the microbial decomposition of aquatic vertebrates from a functional and environmental context. The 16S rRNA gene profiles revealed strong community succession, with early decomposition stages associated with Aeromonas and Clostridium taxa and later stages dominated by members of the Rikenellaceae (i.e., Alistipes/Acetobacteroides genera) These results were reproducible and independent of environmental perturbation, given that exposure to wastewater treatment plant effluent did not substantially influence the necrobiome composition of fish or the associated water sample microbiota. A freshwater study of salmon carcasses reported that members of the Proteobacteria, Firmicutes, and Bacteroidetes dominated carcass decomposition [15] Despite these previous studies characterizing the taxonomic shifts associated with fish decomposition, much more work is needed to explore the microbial communities and their functions within fish necrobiomes and their associated aquatic ecosystems. Important research questions include the following. (i) What is the composition of aquatic vertebrate necrobiomes and how does it change over time? (ii) How do changes in environmental parameters affect necrobiome communities? (iii) What metabolic activities/functions are present in necrobiome communities and how do they change over time?

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