Preliminary assessment of persistent organic pollutants (POPs) in tissues of Risso's dolphin (Grampus griseus) specimens stranded along the Italian coasts.

Ecotoxicological and pathological research on Grampus griseus (Cuvier, 1812) (Risso's dolphins) is scarce both globally and in the Mediterranean Sea. This species has been classified as "Vulnerable" by the International Union for Conservation of Nature (IUCN) in the Mediterranean Sea. To evaluate the presence of "persistent organic pollutants" (POPs), especially organochlorine compounds (OCs), in the animals, chemical analyses were performed on tissues and organs of Risso's dolphin stranded along the Italian coasts between 1998 and 2021. Toxic contaminants such as hexachlorobenzene (HCB), polychlorinated biphenyls (PCBs), dichlorodiphenyltrichloroethane and its metabolites (DDTs) were examined in the blubber, liver, muscle, and brain of 20 animals, and data was correlated with sex, age, and stranding locations.

Parasitological and pathological findings in fin whales Balaenoptera physalus stranded along Italian coastlines.

Mediterranean fin whales Balaenoptera physalus face many threats to their conservation, including both anthropogenic and natural issues. There are few records of the parasitic fauna of this species in this geographical area. To partially fill in this gap of knowledge, we investigated the presence and potential impact of parasitic diseases in Mediterranean fin whales. Seven animals stranded along Italian coastlines between 2006 and 2015 were submitted for necropsy and parasitological examination. The protozoan parasite Toxoplasma gondii was detected in 1 fin whale and, for the first time in mysticetes, it was successfully genotyped as a type II strain with 15 microsatellite markers. One crustacean (Pennella spp.) and 4 helminth taxa (Crassicauda boopis, Ogmogaster antarcticus, Tetrabothrius ruudi and Bolbosoma sp.) were detected and morphologically identified. Different degrees of ectoparasitism by adult P. balaenoptera were recorded. Immature stages of Pennella sp. were also detected in 2 animals and are described here for the first time in cetaceans. Infestation by C. boopis was confirmed or suspected in 5 cases. Parasitic thrombi, involving renal veins and caudal vena cava, and fibrosis of renal parenchyma were associated with C. boopis and likely resulted in some degree of renal dysfunction. Larval nematodes were found within foci of mesenteric endarteritis. Further research to evaluate the prevalence of this potentially fatal endoparasitosis in Mediterranean fin whales is warranted.

Evaluation of PCDD/Fs, dioxin-like PCBs and PBDEs in sperm whales from the Mediterranean Sea.

Numerous studies to date have reported concentrations of Persistent Organic Pollutants (POPs) in different marine mammal species worldwide. Yet data on sperm whales are scarce from rich and unique biodiverse areas such as the Mediterranean Sea. This work aimed to assess levels of dioxin-like polychlorinated biphenyls (dl-PCBs), polybrominated diphenyl ethers (PBDEs), and polychlorodibenzo-p-dioxins and furans (PCDD/Fs) in blubber of sperm whales stranded along the Italian coast between 2008 and 2016. POP mean concentrations (dl-PCBs: 6410 ng/g l.w.; PBDEs: 612 ng/g l.w.; PCDD/Fs: 57.8 pg/g l.w.) were mostly in line with what has been previously reported on the same species in the Mediterranean environment and tended to be higher than those reported from other geographical regions. The relative abundance followed the order dl-PCBs > PBDEs ≫ PCDD/Fs. Interestingly, the non-ortho dl-PCB pattern (126 > 169 > 77) was similar to that described in other studies worldwide and different from what is described in its main prey. This could be linked to particular metabolic activities in sperm whales against these highly toxic contaminants. Total TEQs ranged from 275 to 987 pg/g l.w. and showed the pattern Σnon-ortho-dl-PCBs > Σortho-dl-PCBs > PCDDs > PCDFs, with PCBs' contribution about 96%. These findings highlight the high abundance of PCBs still found in the Mediterranean environment despite having been banned for decades. All sperm whales analyzed in this study surpassed the threshold of 210 pg WHO-TEQ/g l.w. proposed as starting point of immunosuppression in harbour seals; a level of contamination that may have contributed to an impairment of their immune system.

Neuronal and astrocytic involvement in striped dolphins (Stenella coeruleoalba) with morbilliviral encephalitis.

Dolphin morbillivirus (DMV), a highly pathogenic agent, may cause peculiar, "brain-only" forms of infection (BOFDI), in which viral antigen and/or genome is found exclusively in the brain from striped dolphins (Stenella coeruleoalba). These BOFDIs show morphopathological similarities with subacute sclerosing panencephalitis and old dog encephalitis (ODE) in measles virus-infected patients and in canine distemper virus-infected dogs, respectively. The brain tissue from 3 BOFDI-affected striped dolphins was investigated by means of double labelling-indirect immunofluorescence (DL-IIF) and ultrastructurally, in order to characterize the DMV-targeted neuronal and non-neuronal cell populations, along with the associated submicroscopic findings. Viral colonization of calbindin-immunoreactive (IR) and nitric oxide synthase-IR neurons was detected in the cerebral parenchyma from the 3 DMV-infected dolphins under study, associated with nuclear (chromatin) and cytoplasmic (mitochondrial) ultrastructural changes. Furthermore, a limited viral targeting of brain astrocytes was found in these animals, all of which exhibited a prominent astrogliosis/astrocytosis. To the best of our knowledge, those herein reported should be the first submicroscopic pathology and neuropathogenetic data about BOFDI in striped dolphins. In this respect, the marked astrogliosis/astrocytosis and the low viral colonization of brain astrocytes in the 3 DMV-infected dolphins under investigation are of interest from the comparative pathology and viral neuropathogenesis standpoints, when compared with ODE-affected dogs, in whose brain a non-cytolytic, astrocyte-to-astrocyte infectious spread has been recently documented. Further studies aimed at characterizing the complex DMV-host interactions in BOFDI-affected striped dolphins are needed.

Candidate gene molecular markers as tools for analyzing genetic susceptibility to morbillivirus infection in stranded Cetaceans.

Morbilliviruses, such as Cetacean morbillivirus (CeMV) or Phocine distemper virus (PDV), represent a growing threat for marine mammals on both hemispheres. Because free-ranging animal populations strongly rely on natural resistance mechanisms, innate immunity-related genes and virus cell entry receptor genes may represent key factors involved in susceptibility to CeMV in Cetaceans. Using the next generation sequencing technology, we have sequenced 11 candidate genes in two model species, Stenella coeruleoalba and Phocoena phocoena. Suitable single nucleotide polymorphism markers of potential functional importance, located in genes coding for basigin (BSG, CD147), the signaling lymphocyte activating molecule (SLAMF1), the poliovirus-related receptor-4 (NECTIN4, PVRL4), toll-like receptors 3, 7, 8 (TLR3, TLR7, TLR8), natural resistance-associated macrophage protein (SLC11A1) and natural cytotoxicity triggering receptor 1 (NCR1), were identified in each model species, along with MHC-DQB haplotypes unique for each species. This set of molecular markers represents a potentially useful tool for studying host genetic variation and susceptibility to morbillivirus infection in Cetaceans as well as for studying functionally important genetic diversity of selected Cetacean populations.

Western blot expression of 5-lipoxygenase in the brain from striped dolphins (stenella coeruleoalba) and bottlenose dolphins (tursiops truncatus) with or without encephalitis/meningo-encephalitis of infectious nature.

Dolphin Morbillivirus (DMV), Toxoplasma gondii and Brucella ceti are pathogens of major concern for wild cetaceans. Although a more or less severe encephalitis/meningo-encephalitis may occur in striped dolphins (Stenella coeruleoalba) and bottlenose dolphins (Tursiops truncatus) infected by the aforementioned agents, almost no information is available on the neuropathogenesis of brain lesions, including the neuronal and non-neuronal cells targeted during infection, along with the mechanisms underlying neurodegeneration. We analyzed 5-lipoxygenase (5-LOX) expression in the brain of 11 striped dolphins and 5 bottlenose dolphins, affected or not by encephalitic lesions of various degrees associated with DMV, T. gondii and B. ceti. All the 8 striped dolphins with encephalitis showed a more consistent 5-LOX expression than that observed in the 3 striped dolphins showing no morphologic evidence of brain lesions, with the most prominent band intensity being detected in a B. ceti-infected animal. Similar results were not obtained in T. gondii-infected vs T. gondii-uninfected bottlenose dolphins. Overall, the higher 5-LOX expression found in the brain of the 8 striped dolphins with infectious neuroinflammation is of interest, given that 5-LOX is a putative marker for neurodegeneration in human patients and in experimental animal models. Therefore, further investigation on this challenging issue is also needed in stranded cetaceans affected by central neuropathies.

Platelet-derived growth factors and receptors in Canine Lymphoma.

Platelet-derived growth factors (PDGFs) belong to a family of polypeptide growth factors that signal through cell surface tyrosine kinase receptors to stimulate growth, proliferation and differentiation. Platelet-derived growth factor receptors (PDGFRs) are also considered important targets for specific kinase inhibitors in the treatment of several human tumours. The aim of this study was to investigate the role of PDGF-A, PDGF-B, PDGFR-α and PDGFR-ß in canine lymphoma by determining gene and protein expression in lymph nodes of dogs with diffuse large B-cell lymphoma (DLBCL), peripheral T-cell lymphoma (PTCL), T-lymphoblastic lymphoma (T-LBL) and in healthy control dogs. One lymph node was also studied at the end of therapy in a subset of dogs in remission for DLBCL. In controls, PDGF-A, PDGFR-α and PDGFR-ß mRNA levels were significantly higher than in DLBCLs, PTCLs and T-LBLs. However, PDGFR-α and PDGFR-ß were minimally expressed by lymphocytes and plasma cells in normal lymph nodes as determined by immunohistochemistry, while neoplastic B and T cells showed the highest score (P <0.05). This discordant result may be compatible with the constitutive expression of these molecules by endothelial cells and fibroblasts in normal lymph nodes, thereby influencing gene expression results. Furthermore, these cells were not included in the immunohistochemical analysis. Similarly, dogs with DLBCL that were in remission at the end of therapy showed significantly higher gene expression of PDGFs and receptors than at the time of diagnosis and with an opposite trend to the protein assay. PDGF-B protein and mRNA were overexpressed in PTCLs and T-LBLs when compared with DLBCLs and controls (P <0.05). Additionally, there was a correlation between protein expression of PDGF-B and both PDGFRs in PTCLs and T-LBLs, suggesting an autocrine or paracrine loop in the aetiology of aggressive canine T-cell lymphomas. These data provide a rationale for the use of PDGFR antagonists in the therapy of aggressive T-cell lymphomas, but not in DLBCLs.

Myiasis by Megaselia scalaris (Diptera: Phoridae) in a python affected by pulmonitis.

Myiases are caused by the presence of maggots in vertebrate tissues and organs. Myiases have been studied widely in humans, farm animals, and pets, whereas reports of myiasis in reptiles are scarce. We describe a case of myiasis caused by the Megaselia scalaris (Loew) in an Indian python (Python molurus bivittatus, Kuhl) (Ophida: Boidae). The python, 15 yr old, born and reared in a terrarium in the mainland of Venice (Italy), was affected by diffuse, purulent pneumonia caused by Burkholderia cepacia. The severe infestation of maggots found in the lungs during an autopsy indicated at a myiasis.

Prion search and cellular prion protein expression in stranded dolphins.

The recent description of a prion disease (PD) case in a free-ranging bottlenose dolphin (Tursiops truncatus) prompted us to carry out an extensive search for the disease-associated isoform (PrPSc) of the cellular prion protein (PrPC) in the brain and in a range of lymphoid tissues from 23 striped dolphins (Stenella coeruleoalba), 5 bottlenose dolphins and 2 Risso s dolphins (Grampus griseus) found stranded between 2007 and 2012 along the Italian coastline. Three striped dolphins and one bottlenose dolphin showed microscopic lesions of encephalitis, with no evidence of spongiform brain lesions being detected in any of the 30 free-ranging cetaceans investigated herein. Nevertheless, we could still observe a prominent PrPC immunoreactivity in the brain as well as in lymphoid tissues from these dolphins. Although immunohistochemical and Western blot investigations yielded negative results for PrPSc deposition in all tissues from the dolphins under study, the reported occurrence of a spontaneous PD case in a wild dolphin is an intriguing issue and a matter of concern for both prion biology and intra/inter-species transmissibility, as well as for cetacean conservation medicine.