Fabio Marino

Rapid evaporative ionization mass spectrometry (REIMS) coupled with a monopolar handpiece used for surgical resection and combined with chemometrics has been previously explored by our research group (Mangraviti et al. in Int J Mol Sci 23(18):10562, 2022) to identify several mammary gland pathologies. Here, the increased sample size allowed the construction of three statistical models to distinguish between benign and malignant canine mammary tumours (CMTs), facilitating a more in-depth investigation of changes in cellular metabolic phenotype during neoplastic transformation and biological behaviour. The results demonstrate that REIMS is effective in identifying neoplastic tissues with an accuracy of 97%, with differences in MS spectra characterized by the relative abundance of phospholipids compared to triglycerides more commonly identified in normal mammary glands. The increased rate of phospholipid synthesis represents an informative feature for tumour recognition, with phosphatidylcholine and phosphatidylethanolamine, the two major phospholipid species identified here together with sphingolipids, playing a crucial role in carcinogenesis. REIMS technology allowed the classification of different histotypes of benign CMTs with an accuracy score of 95%, distinguishing them from normal glands based on the increase in sphingolipids, glycolipids, phospholipids, and arachidonic acid, demonstrating the close association between cancer and inflammation. Finally, dysregulation of fatty acid metabolism with increased signalling for saturated, mono- and polyunsaturated fatty acids characterized the metabolic phenotype of neoplastic cells and their malignant transformation, supporting the increased formation of new organelles for cell division. Further investigations on a more significant number of tumour histotypes will allow for the creation of a more extensive database and lay the basis for how understanding metabolic alterations in the tumour microenvironment can improve surgical precision.

In mariculture facilities, bacterial infections pose significant production challenges, with potentially catastrophic impacts on fish species. Bacterial co-infections are a widespread phenomenon in the natural marine environment, although their impact on aquatic organisms remains poorly investigated. This study aimed to detail the pathological findings associated with a natural bacterial co-infection caused by three different pathogens, namely Photobacterium damselae subsp. piscicida, Tenacibaculum maritimum and Vibrio sp., as the cause of mass mortality in European sea bass. The fish had been reared in open-net cages in Sicily and later transferred for experimental research purposes to a user establishment after immunization with an inactivated vaccine. Macroscopic, cytological and histopathological examinations were performed on 109 animals, and bacterial species were identified by the 16S rRNA gene. Overall, ulcerative skin lesions, necrotizing myositis and tail rot with occasional tail loss were associated with tenacibaculosis and vibriosis, while P. damselae subsp. piscicida mainly caused granulomatous inflammation in the spleen and head kidney. Finally, an injection site reaction due to the oil-adjuvanted vaccine administered intraperitoneally was observed in the abdominal fat. Understanding the impact of bacterial pathogens is essential to manage the health and welfare of farmed fish, and the importance of a good health monitoring program cannot be overstated to avoid outbreaks and the possible emergence of new pathogens due to the intensification of the production systems, antibiotic resistance and climate changes. The study would also highlight the importance of the quarantine period when animals supplied for research come from aquaculture farms and how the main goal in the near future should be to better define the procedures to provide completely pathogen-free animals.

Bluntnose sixgill shark, Hexanchus griseus (Bonnaterre, 1788), is a little-known elasmobranch in the Mediterranean Sea. Given the lack of information about H. griseus, the aim of our study was to describe the helminth fauna of this species. In March 2023, one H. griseus juvenile female specimen was found off the coast of Messina (Italy) and referred by the Italian Coast Guard to our laboratory for necropsy and parasitological evaluation. After necropsy, the specimen’s gills, stomach and spiral valve were investigated for parasite presence. All collected parasites were stored in 70% ethanol for routine parasitological analysis. No lesions due to parasites were found in the gills or skin. Three species of helminths were found in one studied female specimen of Hexanchus griseus, namely, two cestode species (Phyllobothrium sinuosiceps and Nybelinia sp., larvae) and one trematode (Otodistomum veliporum). Among them, five Trypanorhyncha plerocercoid larvae were found attached to the stomach mucosa, and six adult cestodes and one digenean trematode were collected from the spiral valve. No other parasite taxa were found in the celomic organs. This study reports new information regarding the parasitic fauna of H. griseus from the central Mediterranean Sea.

Essential oils (EOs) are natural compounds used in various fields such as traditional medicine, gastronomy, and agriculture, often used against plant and animal parasites. In the present study, the toxic and pathological effects of basil (Ocimum basilicum) essential oil on the development of zebrafish embryos was investigated. The manufacturer datasheet reported linalool and eugenol as major compounds. The effects of the essential oil were evaluated through a zebrafish embryo toxicity test (ZFET) following the OECD n°236 guidelines. Embryos were exposed to different essential oil concentrations (50, 100, and 200 μL/L). Results showed mortality effects in the experimental groups in a dose-dependent manner. Moreover, zebrafish exposed to the higher concentration showed several pathological alterations; particularly, delayed hatching, pericardial edema, and a significant reduction of heart rate after 96 h post-fertilization (hpf) compared to the control group. Results reported here suggest an anaesthetic effect of the O. basilicum at low concentrations, due to the possible synergist effect of the main components such as linalool and eugenol which act on the GABAergic system evoking bradycardia.

The present work proposes the use of a fast analytical platform for the mass spectrometric (MS) profiling of canine mammary tissues in their native form for the building of a predictive statistical model. The latter could be used as a novel diagnostic tool for the real-time identification of different cellular alterations in order to improve tissue resection during veterinary surgery, as previously validated in human oncology. Specifically, Rapid Evaporative Ionization Mass Spectrometry (REIMS) coupled with surgical electrocautery (intelligent knife—iKnife) was used to collect MS data from histologically processed mammary samples, classified into healthy, hyperplastic/dysplastic, mastitis and tumors. Differences in the lipid composition enabled tissue discrimination with an accuracy greater than 90%. The recognition capability of REIMS was tested on unknown mammary samples, and all of them were correctly identified with a correctness score of 98–100%. Triglyceride identification was increased in healthy mammary tissues, while the abundance of phospholipids was observed in altered tissues, reflecting morpho-functional changes in cell membranes, and oxidized species were also tentatively identified as discriminant features. The obtained lipidomic profiles represented unique fingerprints of the samples, suggesting that the iKnife technique is capable of differentiating mammary tissues following chemical changes in cellular metabolism.

Corrigendum: In the Wake of the Ongoing Mass Mortality Events: Co-occurrence of Mycobacterium, Haplosporidium and Other Pathogens in Pinna nobilis Collected in Italy and Spain (Mediterranean Sea)

Following the Mass Mortality Events (MMEs) of the pen shell P. nobilis in Campania region and Sicily, a survey of moribund P. nobilis specimens was also conducted in other Italian regions (Campania, Tuscany, Sardinia, and Apulia) and Spain (Catalunya). Histopathological and molecular examination of 27 specimens of P. nobils revealed different types of pathogens associated with tissue lesions, morbidity and mortality. Presence of Mycobacterium, Vibrio species, Haplosporidium pinnae and Perkinsus sp. were detected, differently distributed into the areas. The Mycobacterium sp., previously reported in Campania and Sicily samples, was observed in all the analyzed areas and individuals, associated to systemic inflammatory lesions. In Spain, H. pinnae was observed in 36% of cases, always associated to the Mycobacterium sp. Molecular study using hsp65 genes and Internal Transcriber Spacer ITS support that a new species of Mycobacteria is infecting P. nobilis, close to M. triplex and belonging to the group of M. simiae complex with M. sherrisi. Presence of Perkinsus spp. resembling P. mediterraneus was observed in 2 out of 13 Italian individuals whose presence should be addressed as potential risk for shellfish aquaculture of the area. Vibrio spp. were also detected in some case. The preliminary results of this study suggest that Mycobacterium sp., Vibrio spp., H. pinnae and Perkinsus sp. cooperate to disease pathogenesis, being Mycobacterium and Haplosporidium most of the time involved. Vigilant inspection of those areas where MME is now starting, along with continuous systematic surveys, are crucial to define the spatiotemporal progress of mortality and the role of every single pathogen in the disease outcome.

This study reports the presence of cartilaginous cysts in zebrafish gills, purchased at the end of 2012 before the application of the Italian Legislative Decree 26/2014 and used for experimental research. It highlights the importance of using specific pathogen-free organisms from reliable production establishments to avoid obtaining false results in experiments. Of 20 samples analyzed, 4 showed cysts in gills, with abnormal tissue modification and cartilaginous metaplasia induced by prolonged parasitic insult. The results obtained from the periodic health monitoring program showed the presence of metacercariae of digenean trematodes, specifically identified as Centrocestus formosanus.