The production demand of edible snails in the Mediterranean area is very high and the attention to snail borne diseases is increasing. Following mass mortality events, we have analyzed 240 samples of Cornu aspersum collected from farms across Italy. Anatomopathological examination showed the presence of alterations of the gastro-intestinal apparatus and of the digestive gland, while histopathological examination revealed the presence of Rickettsia-like organisms (RLOs) in 70% (168/240) of cases and Giemsa positive amoebae in the remaining 30% (72/240) of cases. RLOs were localized mainly at the level of the DG, where regressive changes or nodular inflammation was observed. TEM examination of RLOs samples revealed the presence of many rod-shaped electron dense microorganisms. Amoebal infection occurred in the kidney, intestine, lung, the DG and were associated to regressive events or infiltrative/nodular and encapsulation like inflammation. To date it is still unclear if the pathogens detected could represent a risk for humans and animals, therefore further studies are needed to better elucidate this point.

Parasitism of cephalopods is common, including infection with Aggregata spp., Ichthyobodo spp., dicyemids, cestodes of the orders Tetraphyllidea and Trypanorhynchidea, and various crustaceans. Cestodiasis in octopuses is reported, although a full histologic description of lesions has not been previously described. Cestodiasis was identified in 10 octopuses of 4 different species, which included 4 common octopuses (Octopus vulgaris), 3 Caribbean reef octopuses (Octopus briareus), 2 two-spot octopuses (Octopus bimaculoides), and 1 giant Pacific octopus (Enteroctopus dofleini). Larval cestodes were present in the cecum (n = 5), intestines (n = 4), digestive gland (n = 3), chitinous alimentary tract (n = 2), renal appendage (n = 1), and salivary duct (n = 1). In 5 cases, larval cestodes invaded tissue and were associated with hemocytic inflammation and tracts of necrotic tissue in the intestines (n = 3), digestive gland (n = 3), and/or renal appendage (n = 1). When present in the chitinous alimentary tract (esophagus, stomach) or cecum, larval cestodes were in the central lumen and not associated with lesions. One adult cestode was identified in the mantle cavity and was not associated with lesions. Other common concurrent parasitic infections included enteric Aggregata spp. infection, branchial Rickettsia-like organism infection, enteric nematodiasis, and an arthropod-associated branchitis.

A black eye syndrome (BES) was discovered in both captive and wild populations of Alaskan snow crabs Chionoecetes opilio and Tanner crabs C. bairdi. Field prevalences ranged from 0.37% (n = 594/161295) to 19.6% (n = 62/316) in snow crabs from the eastern Bering Sea and from 0.09% (n = 15/16638) to 0.7% (n = 133/18473) in Tanner crabs from the same trawl samples, with a slightly greater percentage (1.4%, n = 57/3945) in Tanner crabs from the Aleutian and Kodiak islands fisheries in the Gulf of Alaska. BES is not associated with crab mortality and has 2 distinct manifestations: abnormal black foci of internal eye pigment with no discernible histological lesions, which, in many cases, is followed by corneal shell disease with ulceration and distal eyestalk erosion. It is assumed for this study that these are early and late stages of BES that are somehow related. Our results suggest that early stages of abnormal pigmentation are noninfectious, possibly related to changing ocean conditions affecting crab endocrinology and neuropeptide control of secondary eye pigment. Potential light-induced photoreceptor damage of harvested crabs with dark-adapted eyes is another anthropogenic factor possibly contributing to the early changes in eye pigmentation. Normal eyestalk microanatomy specific for Chionoecetes spp. is provided as necessary baseline information for future studies. Early in the study, an unreported rickettsia-like organism (RLO) was discovered infecting dissected black eyestalks submitted for examination from 5 of 6 dead snow crabs, suggesting association with BES. Subsequent samples indicated the RLO was systemic, infected both black and normal-appearing eyestalks, and was unrelated to BES. However, the multiorgan infection and histopathology indicated the RLO could be a primary pathogen of snow crabs.

An infection caused by a rickettsia-like organism (RLO) was detected in the blue king crab Paralithodes platypus from the eastern Sea of Okhotsk. The external signs of the disease are lethargy and an empty gastrointestinal tract. Dissection of infected individuals revealed that their hepatopancreas was light yellow in color. The causative agent of infection is Gram-negative rod-shaped bacterium targeted exclusively at hepatopancreas tissues. In the cytoplasm of infected cells, the bacteria are enclosed in parasite vacuoles or located immediately in cytosol. An ultrastructural analysis showed two main morphological types corresponding to the life cycle stages in the RLO: the vegetative stage of intermediate bodies, characterized by growth and division processes, and the infection stage of elementary bodies, which are spore-like non-dividing short rods surrounded by a multilayered membrane and having an osmiophilic inclusion body. At the terminal stage of infection, as a result of lysis of the infected cells, the RLO enters the lumen of the hepatopancreatic tubules which contributes to the spread of infection. According to genetic analysis based on 16S rRNA gene sequences, the RLO from P. platypus is most closely related to the Candidatus Hepatobacter penaei, NCBI #JX981946 (94.7% similarity) and NCBI #KY363553 (94.1% similarity). The high level of genetic differences (more than 5%) of the studied pathogen, along with the structural features, allows characterizing the RLO isolated from P. platypus as a new species of the genus Candidatus Hepatobacter paralithodi nov. sp., NCBI #MK928971.

A rickettsia-like organism, designated NZ-RLO2, was isolated from Chinook salmon (Oncorhynchus tshawytscha) farmed in the South Island, New Zealand. In vivo growth showed NZ-RLO2 was able to grow in CHSE-214, EPC, BHK-21, C6/36 and Sf21 cell lines, while Piscirickettsia salmonis LF-89T grew in all but BHK-21 and Sf21. NZ-RLO2 grew optimally in EPC at 15°C, CHSE-214 and EPC at 18°C. The growth of LF-89 T was optimal at 15°C, 18°C and 22°C in CHSE-24, but appeared less efficient in EPC cells at all temperatures. Pan-genome comparison of predicted proteomes shows that available Chilean strains of P. salmonis grouped into two clusters (p-value = 94%). NZ-RLO2 was genetically different from previously described NZ-RLO1, and both strains grouped separately from the Chilean strains in one of the two clusters (p-value = 88%), but were closely related to each other. TaqMan and Sybr Green real-time PCR targeting RNA polymerase (rpoB) and DNA primase (dnaG), respectively, were developed to detect NZ-RLO2. This study indicates that the New Zealand strains showed a closer genetic relationship to one of the Chilean P. salmonis clusters; however, more Piscirickettsia genomes from wider geographical regions and diverse hosts are needed to better understand the classification within this genus.

A histopathological survey revealed parasites and pathological conditions affecting lagoon cockles Cerastoderma glaucum along the Galician coast; serious pathological threats were not detected because the potentially pathogenic conditions (infections with a Marteilia-like parasite and bucephalid sporocysts, disseminated neoplasia and a condition involving large foci of heavy haemocytic reaction) were rare, while more prevalent parasites had negligible or limited pathogeny. Considering that C. edule and C. glaucum are sympatric in some Galician rias, it is remarkable that C. glaucum was not seriously affected by Marteilia cochillia while C. edule suffered an intense outbreak of this parasite associated with massive mortality. Comparison of the digestive gland between cockle species showed co-occurrence of digestive tubules in different phases, with abundant disintegrated tubules, in the case of C. glaucum, while C. edule showed synchronicity and absence of fully disintegrated tubules; these differences could influence their susceptibility to M. cochillia because the main location of this parasite in common cockles is the epithelia of the digestive gland. Moreover, the observation of histological sections through the digestive gland easily allows differentiating the 2 cockle species.

Atlantic salmon Salmo salar L. farmed in south-east Tasmania, Australia, are susceptible to infection by the Tasmanian Rickettsia-like organism (TRLO), a Gram-negative bacterium. Here, we report the first isolation of TRLO from south-east Tasmania in pure culture and show that the bacterium is culturable on both specialised enriched agar and in cell culture using the CHSE-214 cell line. In vitro cultured TRLO was used to reproducibly elicit disease in Atlantic salmon parr held in fresh water. In inoculated fish, TRLO was observed intracytoplasmically in peripheral blood leucocytes, suggesting that these cells are responsible for haematogenous dispersal of the bacterium within the host. Fish with experimentally induced disease presented with gross and histopathological changes similar to TRLO-infected fish at commercial marine farms. TRLO was also isolated in culture from farmed Atlantic salmon in the Tamar River and Macquarie Harbour production areas in Tasmania, both of which have no history of TRLO-associated disease. These TRLO isolates appear to be serologically distinct from each other as well as from isolates obtained from south-east Tasmania, linking each serotype to a specific geographical location within Tasmania. Despite the lack of clinical evidence of TRLO-linked disease in fish grown in the Tamar River and Macquarie Harbour, experimental infection trials demonstrably showed the pathogenic potential of these TRLO serovars. Together, these data provide evidence that TRLO is a fastidious, facultative intracellular bacterium and confirm TRLO as a pathogen of Atlantic salmon, causing a disease designated Tasmanian salmonid rickettsiosis.

Cyclophilins (CyPs) are a family of proteins that bind the immunosuppressive agent cyclosporin A (CsA) with high-affinity and belong to one of the three superfamilies of peptidyl-prolyl cis-trans isomerases (PPIase). In this report, three cyclophilin genes (Ca-CyPs), including Ca-CyPA, Ca-CyPB and Ca-PPIL3, were identified from oyster, Crassostrea ariakensis Gould in which Ca-CyPA encodes a protein with 165 amino acid sequences, Ca-CyPB encodes a protein with 217 amino acid sequences and Ca-PPIL3 encodes a protein with 162 amino acid sequences. All of the three Ca-CyPs genes contain a typical CyP-PPIase domain with its signature sequences and Ca-CyPB contains an N-signal peptide sequences. Tissue distribution study revealed that Ca-CyPs were ubiquitously expressed in all examined tissues and the highest levels were observed in hemocytes. RLO incubation upregulated the mRNA expression levels of Ca-CyPs, indicating that three Ca-CyPs might be involved in oyster immune response against RLO infection.