Comparative analysis of metabolic models of microbial communities reconstructed from automated tools and consensus approaches.

Genome-scale metabolic models (GEMs) of microbial communities offer valuable insights into the functional capabilities of their members and facilitate the exploration of microbial interactions. These models are generated using different automated reconstruction tools, each relying on different biochemical databases that may affect the conclusions drawn from the in silico analysis. One way to address this problem is to employ a consensus reconstruction method that combines the outcomes of different reconstruction tools. Here, we conducted a comparative analysis of community models reconstructed from three automated tools, i.e. CarveMe, gapseq, and KBase, alongside a consensus approach, utilizing metagenomics data from two marine bacterial communities. Our analysis revealed that these reconstruction approaches, while based on the same genomes, resulted in GEMs with varying numbers of genes and reactions as well as metabolic functionalities, attributed to the different databases employed. Further, our results indicated that the set of exchanged metabolites was more influenced by the reconstruction approach rather than the specific bacterial community investigated. This observation suggests a potential bias in predicting metabolite interactions using community GEMs. We also showed that consensus models encompassed a larger number of reactions and metabolites while concurrently reducing the presence of dead-end metabolites. Therefore, the usage of consensus models allows making full and unbiased use from aggregating genes from the different reconstructions in assessing the functional potential of microbial communities.

Microbiome heterogeneity in tissues of the coral, Fimbriaphyllia (Euphyllia) ancora.

Coral microbiomes differ in the mucus, soft tissue and skeleton of a coral colony, but whether variations exist in different tissues of a single polyp is unknown. In the stony coral, Fimbriaphyllia ancora, we identified 8,994 amplicon sequencing variants (ASVs) in functionally differentiated polyp tissues, i.e., tentacles, body wall, mouth and pharynx, mesenterial filaments, and gonads (testes and ovaries), with a large proportion of ASVs specific to individual tissues. However, shared ASVs comprised the majority of microbiomes from all tissues in terms of relative abundance. No tissue-specific ASVs were found, except in testes, for which there were only two samples. At the generic level, Endozoicomonas was significantly less abundant in the body wall, where calicoblastic cells reside. On the other hand, several bacterial taxa presented significantly higher abundances in the mouth. Interestingly, although without statistical confirmation, gonadal tissues showed lower ASV richness and relatively high abundances of Endozoicomonas (in ovaries) and Pseudomonas (in testes). These findings provide evidence for microbiome heterogeneity between tissues within coral polyps, suggesting a promising field for future studies of functional interactions between corals and their bacterial symbionts.

Chlamydiae as symbionts of photosynthetic dinoflagellates.

Chlamydiae are ubiquitous intracellular bacteria and infect a wide diversity of eukaryotes, including mammals. However, chlamydiae have never been reported to infect photosynthetic organisms. Here, we describe a novel chlamydial genus and species, Candidatus Algichlamydia australiensis, capable of infecting the photosynthetic dinoflagellate Cladocopium sp. (originally isolated from a scleractinian coral). Algichlamydia australiensis was confirmed to be intracellular by fluorescence in situ hybridization and confocal laser scanning microscopy and temporally stable at the population level by monitoring its relative abundance across four weeks of host growth. Using a combination of short- and long-read sequencing, we recovered a high-quality (completeness 91.73% and contamination 0.27%) metagenome-assembled genome of A. australiensis. Phylogenetic analyses show that this chlamydial taxon represents a new genus and species within the Simkaniaceae family. Algichlamydia australiensis possesses all the hallmark genes for chlamydiae-host interactions, including a complete type III secretion system. In addition, a type IV secretion system is encoded on a plasmid and has previously been observed for only three other chlamydial species. Twenty orthologous groups of genes are unique to A. australiensis, one of which is structurally similar to a protein known from Cyanobacteria and Archaeplastida involved in thylakoid biogenesis and maintenance, hinting at potential chlamydiae interactions with the chloroplasts of Cladocopium cells. Our study shows that chlamydiae infect dinoflagellate symbionts of cnidarians, the first photosynthetic organism reported to harbor chlamydiae, thereby expanding the breadth of chlamydial hosts and providing a new contribution to the discussion around the role of chlamydiae in the establishment of the primary plastid.

A targeted approach to enrich host-associated bacteria for metagenomic sequencing.

Multicellular eukaryotic organisms are hosts to communities of bacteria that reside on or inside their tissues. Often the eukaryotic members of the system contribute to high proportions of metagenomic sequencing reads, making it challenging to achieve sufficient sequencing depth to evaluate bacterial ecology. Stony corals are one such complex community; however, separation of bacterial from eukaryotic (primarily coral and algal symbiont) cells has so far not been successful. Using a combination of hybridization chain reaction fluorescence in situ hybridization and fluorescence activated cell sorting (HCR-FISH + FACS), we sorted two populations of bacteria from five genotypes of the coral Acropora loripes, targeting (i) Endozoicomonas spp, and (ii) all other bacteria. NovaSeq sequencing resulted in 67-91 M reads per sample, 55%-90% of which were identified as bacterial. Most reads were taxonomically assigned to the key coral-associated family, Endozoicomonadaceae, with Vibrionaceae also abundant. Endozoicomonadaceae were 5x more abundant in the 'Endozoicomonas' population, highlighting the success of the dual-labelling approach. This method effectively enriched coral samples for bacteria with <1% contamination from host and algal symbionts. The application of this method will allow researchers to decipher the functional potential of coral-associated bacteria. This method can also be adapted to accommodate other host-associated communities.