Lack of Overt Genome Reduction in the Bryostatin-Producing Bryozoan Symbiont “Candidatus Endobugula sertula”

ABSTRACTThe uncultured bacterial symbiont “CandidatusEndobugula sertula” is known to produce cytotoxic compounds called bryostatins, which protect the larvae of its host,Bugula neritina. The symbiont has never been successfully cultured, and it was thought that its genome might be significantly reduced. Here, we took a shotgun metagenomics and metatranscriptomics approach to assemble and characterize the genome of “Ca. Endobugula sertula.” We found that it had specific metabolic deficiencies in the biosynthesis of certain amino acids but few other signs of genome degradation, such as small size, abundant pseudogenes, and low coding density. We also identified homologs to genes associated with insect pathogenesis in other gammaproteobacteria, and these genes may be involved in host-symbiont interactions and vertical transmission. Metatranscriptomics revealed that these genes were highly expressed in a reproductive host, along withbrygenes for the biosynthesis of bryostatins. We identified two new putativebrygenes fragmented from the mainbryoperon, accounting for previously missing enzymatic functions in the pathway. We also determined that a gene previously assigned to the pathway,bryS, is not expressed in reproductive tissue, suggesting that it is not involved in the production of bryostatins. Our findings suggest that “Ca. Endobugula sertula” may be able to live outside the host if its metabolic deficiencies are alleviated by medium components, which is consistent with recent findings that it may be possible for “Ca. Endobugula sertula” to be transmitted horizontally.IMPORTANCEThe bryostatins are potent protein kinase C activators that have been evaluated in clinical trials for a number of indications, including cancer and Alzheimer's disease. There is, therefore, considerable interest in securing a renewable supply of these compounds, which is currently only possible through aquaculture ofBugula neritinaand total chemical synthesis. However, these approaches are labor-intensive and low-yielding and thus preclude the use of bryostatins as a viable therapeutic agent. Our genome assembly and transcriptome analysis for “Ca. Endobugula sertula” shed light on the metabolism of this symbiont, potentially aiding isolation and culturing efforts. Our identification of additionalbrygenes may also facilitate efforts to express the complete pathway heterologously.

Biologically active substances from water invertebrates: a review

Some species of invertebrates especially bryozoans (Bryozoa syn. Ectoprocta) and marine sponges (Porifera) are very important sources of pharmacologically exploitable compounds. These substances are probably produced to protect themselves from fish predators and may be an advantage in competition. The real sources of compounds with these antipredatory effects are probably not marine invertebrates themselves, but microscopic symbionts or food which they feed on. Bryostatins from bryozoan species Bugula neritina are produced by a bacterial symbiont called Candidatus Endobugula sertula. They have significant anti-cancer effects, but also other therapeutic benefits. Compounds with the structure of bryostatins were also discovered in some other invertebrates. Sponges are a source of many compounds, e.g., ara-A (vidarabine), manzamine, lasonolides, spongistatins, peloruside and others with antimicrobial, anti-cancer, immunosuppressive and similar activities. Other important sources of compounds with medical effects are tunicates (Tunicata syn. Urochordata) and some snails (Mollusca). One drug was developed from tunicates – Yondelis against refractory soft-tissue sarcomas. Certain other drugs originate from snails: e.g., prialt, which acts against chronic pain in spinal cord injury.  

Isolation of Two Polyketide Synthase Gene Fragments from the Uncultured Microbial Symbiont of the Marine Bryozoan Bugula neritina

ABSTRACT “Candidatus Endobugula sertula,” the uncultured microbial symbiont of the bryozoan Bugula neritina, produces ecologically and biomedically important polyketide metabolites called bryostatins. We isolated two gene fragments from B. neritina larvae that have high levels of similarity to polyketide synthase genes. These gene fragments are clearly associated with the symbiont and not with the host.

“Candidatus Endobugula glebosa,” a Specific Bacterial Symbiont of the Marine Bryozoan Bugula simplex

ABSTRACT The bryozoans Bugula neritina and Bugula simplex harbor bacteria in the pallial sinuses of their larvae as seen by electron microscopy. In B. neritina, the bacterial symbiont has been characterized as a gamma-proteobacterium, “Candidatus Endobugula sertula.” “Candidatus E. sertula” has been implicated as the source of the bryostatins, polyketides that provide chemical defense to the host and are also being tested for use in human cancer treatments. In this study, the bacterial symbiont in B. simplex larvae was identified by 16S rRNA-targeted PCR and sequencing as a gamma-proteobacterium closely related to and forming a monophyletic group with “Candidatus E. sertula.” In a fluorescence in situ hybridization, a 16S ribosomal DNA probe specific to the B. simplex symbiont hybridized to long rod-shaped bacteria in the pallial sinus of a B. simplex larva. The taxonomic status “Candidatus Endobugula glebosa” is proposed for the B. simplex larval symbiont. Degenerate polyketide synthase (PKS) primers amplified a gene fragment from B. simplex that closely matched a PKS gene fragment from the bryostatin PKS cluster. PCR surveys show that the symbiont and this PKS gene fragment are consistently and uniquely associated with B. simplex. Bryostatin activity assays and chemical analyses of B. simplex extracts reveal the presence of compounds similar to bryostatins. Taken together, these findings demonstrate a symbiosis in B. simplex that is similar and evolutionarily related to that in B. neritina.