Limited Metabolomic Overlap between Commensal Bacteria and Marine Sponge Holobionts Revealed by Large Scale Culturing and Mass Spectrometry-Based Metabolomics: An Undergraduate Laboratory Pedagogical Effort at Georgia Tech

Abstract

Sponges are the richest source of bioactive organic small molecules, referred to as natural products, in the marine environment. It is well established that laboratory culturing-resistant symbiotic bacteria residing within the eukaryotic sponge host matrix often synthesize the natural products that are detected in the sponge tissue extracts. However, the contributions of the culturing-amenable commensal bacteria that are also associated with the sponge host to the overall metabolome of the sponge holobiont are not well defined. In this study, we cultured a large library of bacteria from three marine sponges commonly found in the Florida Keys. Metabolomes of isolated bacterial strains and that of the sponge holobiont were compared using mass spectrometry to reveal minimal metabolomic overlap between commensal bacteria and the sponge hosts. We also find that the phylogenetic overlap between cultured commensal bacteria and that of the sponge microbiome is minimal. Despite these observations, the commensal bacteria were found to be a rich resource for novel natural product discovery. Mass spectrometry-based metabolomics provided structural insights into these cryptic natural products. Pedagogic innovation in the form of laboratory curricula development is described which provided undergraduate students with hands-on instruction in microbiology and natural product discovery using metabolomic data mining strategies.

Pseudobulbiferamides: Plasmid-Encoded Ureidopeptide Natural Products with Biosynthetic Gene Clusters Shared Among Marine Bacteria of Different Genera

“…The isolation of Microbulbifer sp. MKSA007 used in this study from a marine sponge has been previously described . Liquid fermentation and extraction were the same as previously described .…”

“…The Pseudovibrio and Microbulbifer genera coinhabit the commensal microbiomes of marine sponges. , We, among others, have previously reported the discovery of bulbiferamides 1 – 4 from marine Microbulbifer spp. bacteria isolated from sponges Aplysina fulva and Aiolochroia crassa that were collected in the Florida Keys (Figure A). , The mass spectrometric fragmentation spectra (MS/MS spectra) for 1 – 4 demonstrated 165 and 26 Da neutral losses characteristic of the Phe residue connected via the ureidopeptide bond.…”

“…In extracts of the bacterium Microbulbifer sp. MKSA007 that was isolated from the sponge Smenospongia aurea also collected in the Florida Keys, we detected metabolites demonstrating similar features in their fragmentation spectra (Figure B). S.…”

“…The Pseudovibrio and Microbulbifer genera coinhabit the commensal microbiomes of marine sponges. 11,12 We, among others, have previously reported the discovery of bulbiferamides 1−4 from marine Microbulbifer spp. bacteria isolated from sponges Aplysina fulva and Aiolochroia crassa that were collected in the Florida Keys (Figure 1A).…”

“…In extracts of the bacterium Microbulbifer sp. MKSA007 that was isolated from the sponge Smenospongia aurea also collected in the Florida Keys, 11 we detected metabolites demonstrating similar features in their fragmentation spectra (Figure 1B). S. aurea, A. fulva, and A. crassa are high microbial abundance sponges that are ubiquitous on shallow reefs in the Florida Keys.…”