Characterisation and distribution of the bacterial genus Endozoicomonas in a threatened surf clam

The toheroa Paphies ventricosa is a large Aotearoa New Zealand (ANZ) endemic surf clam of cultural importance to many Māori, the Indigenous people of ANZ. Extensive commercial and recreational harvesting in the 20th century dramatically reduced populations, leading to the collapse and closure of the fishery. Despite being protected for >40 yr, toheroa have inexplicably failed to recover. In 2017, intracellular microcolonies (IMCs) of bacteria were detected in 'sick' toheroa in northern ANZ. Numerous mass mortality events (MMEs) have recently been recorded in ANZ shellfish, with many events linked by the presence of IMCs resembling Rickettsia-like organisms (RLOs). While similar IMCs have been implicated in MMEs in surf clams elsewhere, the impact of these IMCs on the health or recovery of toheroa is unknown. A critical first step towards understanding the significance of a pathogen in a host population is pathogen identification and characterisation. To begin this process, we examined 16S rRNA gene sequences of the putative IMCs from 4 toheroa populations that showed 97% homology to Endozoicomonas spp. sequences held in GenBank. Phylogenetic analysis identified closely related Endozoicomonas strains from the North and South Island, ANZ, and in situ hybridization, using 16S rRNA gene probes, confirmed the presence of the sequenced IMC gene in the gill and digestive gland tissues of toheroa. Quantitative PCR revealed site-specific and seasonal abundance patterns of Endozoicomonas spp. in toheroa populations. Although implicated in disease outbreaks elsewhere, the role of Endozoicomonas spp. within the ANZ shellfish mortality landscape remains uncertain.