Cosmopolitan Distribution of Endozoicomonas-Like Organisms and Other Intracellular Microcolonies of Bacteria Causing Infection in Marine Mollusks
Irene Cano, David Ryder, Steve C. Webb, Brian J. Jones, Cara L. Brosnahan, Noèlia Carrasco-Querol, Barbara Bodinier, Dolors Furones, Tobia Pretto, Francesca Carella, Bruno Chollet, Isabelle Arzul, Deborah Cheslett, Evelyn Collins, Karin B. Lohrmann, Ana L Valdivia, Georgia M. Ward, Marı́a J. Carballal, António Villalba, Ionan Marigómez, Stein Mortensen, Kevin W. Christison, Wakeman C. Kevin, Eduardo Bustos, Lyndsay Christie, Matthew Green, Stephen W. Feist
Abstract
Intracellular microcolonies of bacteria, in some cases developing large extracellular cysts, have been historically reported infecting a wide diversity of economically important mollusc species worldwide, sometimes associated with severe lesions and mass mortality events. As an effort to characterise those organisms, traditionally named as Rickettsia or Chlamydia -like organisms (RLO/CLO), via international collaboration, 98 samples comprising 20 mollusc species were collected over 10 countries and examined using histology and phylogenetic analysis. A 16S rRNA gene amplicon library-based sequencing showed the presence of different species of Endozoicomonas-like organisms (ELO) in all the mollusc species analysed, infecting primarily gill and digestive glands. Co-infections of ELOs with other intracellular bacteria were also observed. Subsequent phylogenetic analysis of Operational Taxonomic Units (OTU) revealed a novel microbial diversity associated with molluscan RLO/CLOs infection distributed along different taxa, including Spirochaetes phyla, Rickettsiales order, Simkaniaceae family, Mycoplasma and Francisella genera, and sulfur-oxidizing endosymbionts. Sequences like Francisella halioticida/philomiragia and Candidatus Brownia rhizoecola were also obtained. The presence of ELO sequences in the RLO/CLO infection was confirmed by standard PCR, Sanger sequencing, and by in situ hybridisation in a selection of samples. The phylogenetic analysis conducted in this study will allow for further characterization of the microbial community associated with Rickettsia and Chlamydia-like infection in marine molluscs and their correlation with severity of the lesions in order to reveal their role as endosymbionts, commensals or true pathogens.