Leptospirosis is a worldwide-spread zoonosis causing disease and death in dogs and in humans. A Leptospiral infection has been recorded in several wild carnivore species in Europe, but tissue pathological changes were not commonly described. The Grey wolf (Canis lupus) has been expanding its distribution range in north-eastern Italy during the last decade. A young wolf, representing the first individual handled in the region, was found road-killed and then submitted to necropsy. Pathological changes included erosive lesions of gingival mucosa, mild liver enlargement, and multifocal degenerative-necrotic areas along with hyperemic reactive lesions; multifocal interstitial nephritis and multifocal lung hemorrhages were observed. A Polymerase Chain Reaction (PCR) able to detect pathogenic species of Leptospira performed on a kidney sample was positive. Serological reactions for serogroup Gryppotyphosa (1:6400), Pomona (1:800), and Icterohaemorrhagiae (1:200) were evidenced by MAT. Genotyping by Multilocus Sequence Typing (MLST) performed on detected Leptospira characterized it as belonging to Sequence Type (ST) 117, which refers to L. kirschneri, serogroup Pomona, serovar Mozdok. Regardless of the role of Leptospira infection as an eventual predisposing factor to the road killing of this wolf, to the best of the authors’ knowledge, this is the first report of Leptospira-induced pathology in a wolf in Europe. Surveys on Leptospira infection in free-ranging wildlife species should be pursued in order to achieve further epidemiological knowledge on the circulation of the Leptospira strain.

Red mark syndrome (RMS) is a widespread skin disorder of rainbow trout in freshwater aquaculture, believed to be caused by a Midichloria-like organism (MLO). Here, we aimed to study the pathologic mechanisms at the origin of RMS by analyzing field samples from a recent outbreak through gene expression, MLO PCR, quantitative PCR, and a histopathological scoring system proposed for RMS lesions. Statistical analyses included a One-Way Analysis of Variance (ANOVA) with a Dunnett’s multiple comparisons test to assess differences among gene expression groups and a nonparametric Spearman correlation between various categories of skin lesions and PCR results. In short, the results confirmed the presence of a high quantity of 16S gene copy numbers of Midichloria-like organisms in diseased skin tissues. However, the number of Midichloria-like organisms detected was not correlated to the degree of severity of skin disease. Midichloria-like organism DNA was found in the spleen and head kidney. The spleen showed pathologic changes mainly of hyperplastic type, reflecting its direct involvement during infection. The most severe skin lesions were characterized by a high level of inflammatory cytokines sustaining and modulating the severe inflammatory process. IL-1 β, IL-6, IL-10, MHC-II, and TCR were upregulated in severe skin lesions, while IL-10 was highly expressed in moderate to severe ones. In the moderate form, the response was driven to produce immunoglobulins, which appeared crucial in controlling the skin disease’s severity. Altogether our results illustrated a complex immune interaction between the host and Midichloria-like organism.

Trachemys scripta is a turtle species native to Central America. Since the 1950s, pond sliders have been imported worldwide as companion animals, but have often ended up in foreign ecosystems with great ecological consequences. Moreover, both autochthonous and invasive species of turtles can be carriers of pathogens, including Chlamydiaceae. In the present study, pulmonary tissues collected from four Trachemys scripta were tested with a 23S-targeting real-time PCR (rPCR) specific for the Chlamydiaceae family. The turtles were hosted in a rescue center for wild exotic animals located in northeastern Italy, and were found dead after the hibernation period. Two out of four individuals resulted positive in rPCR for the presence of Chlamydiaceae. Further characterization of this positivity was performed by phylogenetic analysis of the 16S rRNA and outer membrane protein A genes. The phylogenetic tree showed that these chlamydial strains are identical to a novel Chlamydia reported in 2017 in Polish freshwater turtles, and closely related to Chlamydia pneumoniae and to other chlamydial strains found in reptiles. This first finding evidences the presence of this Chlamydia strain in Italian turtles, but further studies will be necessary to confirm the presence and the strain pathogenicity and to evaluate its prevalence in the local turtles’ population.

The interest for in ovo feeding has grown in the last decades mainly concerning probiotics, live microorganisms that can actively interact with the embryo. The aim of this study was to evaluate the effects of a multi-strain probiotic diluted in Marek’s disease vaccine (MDV) on zootechnical performances, intestinal morphology and Eimeria spp. infection. One hundred and twenty eggs of Ross 308 broiler chickens were incubated until 18 d, when 105 fertilised and vital eggs were randomly allocated into three groups. A control group (C) was inoculated with MDV; two treated groups (P1 and P2) were inoculated with MDV and different concentrations of probiotics: 1 × 10⁵ CFU/100 μL in P1 and 1 × 10⁶ CFU/100 μL in P2. After hatching, chickens were separated into three replicates (10/replicate). Zootechnical parameters were determined. At the end of the cycle (35 d), chickens were slaughtered, and the intestine was collected for morphological analysis from nine chickens per group (three/replicate). Eimeria spp. oocyst shedding was determined weekly, and parasitic lesions were analysed on the histological sample. In ovo treatment with probiotic did not influence hatching rate but significantly improved body weight and positively influenced intestinal morphometric data compared to C. Oocyst shedding in faeces resulted in an increase in C, with significant differences at sampling performed at 14, 21 and 28 d of age. These results suggest that the tested probiotic compound is safe for in ovo supplementation and effectively improves zootechnical performances and coccidian resistance.

Multiplex immunostaining (mIHC) allows the simultaneous detection of multiple antigenic targets within the same tissue section, providing a deeper understanding of spatial variation in cellular distribution. The aim of the present study is to apply this technique to examine the spatial variation of lymphocyte populations in swine lymph nodes during PCV2-SD and PRRSV lymphadenopathy compared with reactive lymphoid hyperplasia. A triple immunohistochemical stain with CD3, CD20 and IBA1 antibodies for the concurrent detection of T lymphocytes, B lymphocytes and macrophages, respectively, was performed. Multiplex immunohistochemistry (mIHC) revealed that, compared to reactive hyperplasia, the most significant changes in lymph node cell populations occurred in the follicles for both PCV2 and PRRSV infections. Additionally, in PCV2 cases, notable alterations were also observed in the interfollicular areas. In PCV2-affected lymph nodes, follicles not only significantly decreased in number but also showed a marked significant reduction in CD20⁺ and CD3⁺ cells. The interfollicular region in these cases also exhibited a significant reduction in CD3⁺ cells. In contrast, PRRSV-associated lymphadenopathy showed significantly increased CD20⁺ cells in the follicles, with a similar trend noted in the interfollicular region. mIHC provides more informative results on a single tissue section, thus preserving the topographical information of the tissue and allowing a comprehensive study of cellular composition, cellular functionality and cell–cell interactions, proving to be a valuable tool for studying and understanding disease dynamics.

Reighardia sternae is a pentastomid parasitising the respiratory system of larids and skuas, the pathological significance of which is still poorly understood. In this study, pentastomes found in the interclavicular air sac of a deceased and necropsied yellow-legged gull (Larus michahellis) were identified by parasitological and molecular methods and associated gross and histopathological lesions were evaluated. After morphological evaluation and molecular characterisation, the species R. sternae was identified. This is the first report of R. sternae infecting L. michahellis in Italy. Gross pathology showed generalised aerosacculitis, parabronchial edema, and enlargement of the heart, liver, and spleen. The systemic lesions found at histopathological examination were indicative of chronic cardiocirculatory failure, which led to the death of the gull. Moreover, inflammation was present throughout the lungs and air sacs in the form of some granulomas and diffuse small nodular aggregates rich in lymphocytes. The remnants of a parasite were observed within a granulomatous lesion in the lung, suggesting a primary host response to the parasite. These latter findings may suggest that the migration of R. sternae was the cause of the pathological granulomatous pictures in the examined yellow-legged gull.