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Rickettsiales‐like organisms (RLOs) are intracellular bacteria that have been reported to be symbionts of over 60 species of bivalves and marine gastropod molluscs of aquaculture importance around the world. Morphologically, some of these prokaryotes are similar to bacteria from the order Rickettsiales, and in most cases, they form intracytoplasmic inclusions similar to those observed in bacteria of the family Anaplasmataceae. Genetically, only Candidatus Xenohaliotis californiensis (CXc), a parasite of several abalone species, has been classified as a member of the order Rickettsiales. The other two characterized RLOs are distantly related; one belongs to the same class (α‐proteobacteria), and the other belongs to the γ‐proteobacteria. Hyperparasitism of RLOs by bacteriophages has been reported in several species of mollusc and appears to have a negative effect on RLOs. The intracellular lifestyle of RLOs has been associated with cell alterations including hypertrophy and compression of the cytoplasmic organelles, and in most cases, no host reaction is observed by histology. In some cases, a high infection intensity of RLOs has been related to dysfunction of the affected organs that can result in disease, and in other cases, high burdens of RLOs have been associated with mortality, but most of these cases have yet to be confirmed. To date, only CXc has been confirmed as the aetiological agent of a chronic and potentially lethal disease of several abalone species known as withering syndrome. This disease is listed by the World Organization of Animal Health. With the ever‐increasing interest in preserving fisheries and expanding the aquaculture production of mollusc species, more efforts should be focused on the characterization and study of RLOs to elucidate the type of symbiosis they maintain with their host and determine when they are acting as true pathogens that should be subjected to control in aquaculture. This review provides an overview of the literature on RLOs that infect bivalves and gastropods of aquaculture importance, and we discuss the most important aspects of these unique prokaryotes. [ABSTRACT FROM AUTHOR]

Bacteriophages are recognized as major mortality agents of microbes, among them intracellular marine rickettsiales-like bacteria. Recently, a phage hyperparasite of Candidatus Xenohaliotis californiensis (CXc) has been described. This bacterium is considered the causal agent of Withering Syndrome (WS) which is a chronic and potentially lethal disease of abalone species from California, USA and the peninsula of Baja California, Mexico. This hyperparasite which infects CXc could be used as a biocontrol agent for WS. Therefore, it is necessary to obtain genomic information to characterize this phage. In this study, the first complete genome sequence of a novel phage, Xenohaliotis phage (pCXc) was determined. The complete genome of pCXc from red abalone (Haliotis rufescens) is 35,728 bp, while the complete genome of pCXc from yellow abalone (Haliotis corrugata) is 35,736 bp. Both phage genomes consist of double-stranded DNA with a G + C content of 38.9%. In both genomes 33 open reading frames (ORFs) were predicted. Only 10 ORFs encode proteins that have identifiable functional homologues. These 10 ORFs were classified by function, including structural, DNA replication, DNA packaging, nucleotide transport and metabolism, life cycle regulation, recombination and repair, and additional functions. A PCR method for the specific detection of pCXc was developed. This information will help to understand a new group of phages that infect intracellular marine rickettsiales-like bacteria in mollusks. [ABSTRACT FROM AUTHOR]

Candidatus Xenohaliotis californiensis ( C Xc) is a Rickettsiales-like prokaryote that is considered the causal agent of Withering Syndrome (WS), a chronic disease of abalone, from the west coast of North America and it is listed by the International Organization for Animal Health (OIE) as a reportable agent due to its pathogenicity. This bacterium in red abalone Haliotis rufescens , black abalone Haliotis cracherodii , and yellow abalone Haliotis corrugata from California, US and Baja California, Mexico has been found to be infected by a bacteriophage. To date, there is no information on the epizootiology of C Xc and its bacteriophage in natural populations of abalone; furthermore, it is unknown if the bacteriophage was also present in C Xc infecting blue abalone Haliotis fulgens . The objective of this study was to determine the distribution, prevalence and intensity of C Xc, as well as to determine the distribution and prevalence of the bacteriophage and to study interactions between host sex and hyperparasitism in blue abalone and yellow abalone. Tissue samples were obtained from seven localities where the commercial capture of wild abalone is carried out. Samplings were conducted throughout the 2012–2013 capture seasons and a total of 182 blue abalone and 170 yellow abalone were obtained. The prevalence and intensity of C Xc and the prevalence of the bacteriophage were determined by histology. The identity of C Xc was confirmed by PCR, product sequence analysis and in situ hybridization while the identity of the bacteriophage was corroborated by TEM. The prevalence of C Xc infected and uninfected by the bacteriophage was 80% in blue abalone and 62% in yellow abalone. Low infection intensities were found in 86% of blue abalone and 82% of yellow abalone. Infection intensity was significantly higher in undifferentiated yellow abalone. The bacteriophage in C Xc showed a prevalence of 22% and 31% in blue abalone and yellow abalone respectively. These results show that C Xc and its bacteriophage are widely distributed in the peninsula of Baja California and that they are well established in natural populations of blue abalone and yellow abalone. Additionally, this data constitutes the first record of a bacteriophage in blue abalone. [ABSTRACT FROM AUTHOR]

Candidatus Xenohaliotis californiensis (C Xc) is a Rickettsiales-like organism (RLO) that is considered the causal agent of withering syndrome (WS), a chronic and potentially fatal disease for several abalone species. C Xc can be infected by a bacteriophage (p C Xc), and both phage-infected and non-phage-infected C Xc can coexist with a presumable different species of RLO tentatively named stippled Rickettsiales-like organism (ST-RLO). Currently, the taxonomic affiliation of the ST-RLO is unknown, and no studies have been conducted to determine its presence in infected abalones in the Peninsula of Baja California, Mexico. To determine whether the ST-RLO is a histological variant of C Xc or a new species of RLO, we performed histology and species-specific in situ hybridization (ISH) for C Xc and p C Xc in red abalone, Haliotis rufescens , infected with C Xc and hyperparasitized by p C Xc, which also showed the presence of the ST-RLO. Additionally, a histological analysis of the presence of all of these RLOs from 2008 to 2019 from an aquaculture facility was performed. The results showed that all ST-RLOs were negative by ISH for C Xc and p C Xc, suggesting that the ST-RLO is a different species of bacterium and that it is not susceptible to infection by p C Xc. Histologically similar alterations to those produced by C Xc in the post esophagus were observed. The three RLOs regularly coexist in all the infected abalones, and there is a relatively constant proportion of each kind of RLO over time, suggesting that equilibrium is reached during coexistence; the largest percentage corresponded to C Xc non phage infected (81%), followed by C Xc infected by p C Xc (16%) and the ST-RLO (3%). In all cases, the ST-RLO infection remained at a subclinical level making it difficult to determine if it acts as pathogen or as a commensal. Therefore, the genetic characterization of the ST-RLO and pathogenicity studies are needed to further elucidate the role of this new RLO in the development of of WS in abalone species of North America. • Confirmation of the presence of an unidentified rickettsia in abalone. • This unidentified rickettsia produces similar damages to those attributed to Candidatus Xenohaliotis californiensis. • This rickettsia coexists with Candidatus Xenohaliotis californiensis non infected and infected by the bacteriophage pCXc. • The role of this rickettsia in the development of abalone Withering Syndrome is unknown. • This is the first study to perform ISH for pCXc and to confirm that the ST-RLO is not susceptible to infection by pCXc. [ABSTRACT FROM AUTHOR]