Nanopore Sequencing of Amoebophrya Species Reveals Novel Collection of Bacteria Putatively Associated With Karlodinium veneficum.

Abstract

The dinoflagellate parasite Amoebophrya sp. ex Karlodinium veneficum plays a major role in controlling populations of the toxic bloom-forming dinoflagellate K. veneficum and is one of the few cultured representatives of Marine Alveolate Group II. The obligate parasitic nature of this Amoebophrya spp. precludes isolation in culture, and therefore, genomic characterization of this parasite relies on metagenomic sequencing. Whole-genome sequencing of an Amoebophrya sp. ex K. veneficum-infected culture using Nanopore long reads revealed a diverse community of novel bacteria as well as several species previously reported to be associated with algae. In sum, 39 metagenome-assembled genomes were assembled, and less than half of these required binning of multiple contigs. Seven were abundant but of unknown genera, 13 were identifiable at the generic level by BLAST (8 of which were apparently complete single-contig genomes), and the remaining 19 comprised less abundant (individually accounting for <2% of the total bacterial reads in the culture) and often rarer and/or novel species. Attempts to culture strains identified through sequencing revealed that only two of these bacterial isolates were readily amenable to cultivation, stressing the importance of a dual culture- and sequencing-based approach for robust community analysis. Functional annotations of metagenome-assembled genomes are presented here to support the characterization of a microbial community associated with K. veneficum and/or Amoebophrya sp. ex K. veneficum cultured from the Chesapeake Bay and give preliminary insights into the nature of the associations these bacteria have with this parasite-host complex.