Istituto Zooprofilattico Sperimentale del Mezzogiorno

Background: Brucella canis is an emerging zoonotic pathogen responsible for canine reproductive disorders and public health concerns. This study assessed the seroprevalence of B. canis in dogs from Campania, Southern Italy (2022-2025). Methods: Serum samples (n = 400) were retrospectively screened using an indirect immunofluorescence assay (IFAT), performed according to the manufacturer's instructions. Screening was conducted at a 1:40 cut-off, followed by serial dilutions to determine endpoint titres. Statistical analysis included chi-square tests for univariable screening, followed by nominal logistic regression models to evaluate the association between IFAT positivity and predictive factors (year, province, and sex of dogs). Additionally, a general linear model (GLM) was applied to the seropositive subset (n = 69) to analyse the magnitude of the antibody response, expressed as geometric mean titres (GMTs). Results: The overall seroprevalence was 17.3% (95% CI: 13.6-21.0%). Dog's sex, year of sampling, and province were not significant independent predictors of infection (p > 0.05), but GLM analysis showed that sampling year (p = 0.0024) and province (p = 0.0490) significantly influenced antibody intensity. A significant temporal increase in antibody intensity was observed towards 2025 (p = 0.037), suggesting an intensification of infection pressure. Conclusions: Our results confirm that Brucella canis is an endemic pathogen in the shelter dog population of southern Italy. The high seroprevalence and significant increase in antibody magnitude (GMT) over the study period indicate rising infection pressure, highlighting the urgent need for mandatory screening and a coordinated One Health surveillance strategy to manage zoonotic risk effectively.

Milk is a primary source of vital nutrients and bioactive components fundamental to the growth and development of both newborn animals and humans. Produced by economically significant livestock species (including cattle, buffaloes, goats, sheep and camels), milk is a complex matrix rich in caseins, vitamins, fats, and proteins. Beyond its classical nutritional profile, milk serves as a pivotal vehicle for milk-derived extracellular vesicles (mEVs). These specialized food-derived EVs (fEVs) exert pleiotropic effects that resonate with the One Health paradigm, linking animal well-being and human nutrition to broader ecosystem stability. mEVs offer unique advantages, such as high biocompatibility and gastrointestinal stability, also rendering them potential therapeutic tools as drug delivery systems. However, challenges remain regarding the standardization of mEVs and the variability of their molecular cargo. This review provides a comprehensive comparative analysis of mEVs across a diverse taxonomic range, including bovines, water buffaloes, yaks, camels, goats, pigs, horses, donkeys, and humans, highlighting their distinct functional signatures. Indeed, a critical issue in mEV research is the isolation process: recommendations to minimize contamination from milk fat globules and casein micelles (which can cover EV signals) are given. Finally, current detection methods and instrumentation, with a specific focus on advancing flow cytometry (FC) approaches are discussed. Key insights include the use of conventional FC (with fluorescence triggering, the necessity of rigorous controls and calibration, and the utility of bead-based assays to overcome resolution limits) and imaging flow cytometry (IFC). In both technical approaches, the application of different EV generic fluorescent markers and the strategic selection of tetraspanins (i.e., CD9, CD63, CD81), is mandatory: we emphasize that selecting the correct antibody clones and accounting for inter-species cross-reactivity are essential steps for ensuring the accuracy and reproducibility of mEV research across mammalian species.

Mastitis is an udder inflammation and infection causing several economic losses in dairy cows. The milk metabolomic changes associated with clinical or subclinical mastitis have been investigated. However, little is known on milk metabolome associated with intramammary infection. The aim of this study was to investigate the quarter-milk metabolomic profile affected by intramammary infection, subclinical mastitis, and clinical mastitis in dairy cows. A total of 80 quarter-milk samples of multiparous Holstein-Friesian dairy cows were used in this cross-sectional design study. Samples were equally divided into 4 groups: healthy (H; no clinical signs of mastitis, SCC <200,000 cells/mL, negative at bacteriological analysis); intramammary infection (IMI group; no clinical signs of mastitis, SCC <200,000 cells/mL, positive at bacteriological analysis); subclinical mastitis (SCM; no clinical signs of mastitis, SCC ≥200,000 cells/mL, positive at bacteriological analysis); and clinical mastitis (CM; clinical signs of mastitis, SCC ≥200,000 cells/mL, positive at bacteriological analysis). Statistical analysis was conducted by fitting a linear mixed model with the group as the fixed effect, quarter nested within animal as random effect, and DIM as covariate. Analysis identified 45 metabolites, and among them 34 were significantly different among groups. Among these, 18 metabolites (2-aminoadipate, Ala, creatine-phosphate, dimethylamine, N-acetyl-Gly, O-phosphocholine, glucose, lactose, maltose, cis-aconitate, carnitine, fumarate, lactate, phenylacetate, 2-ketobutyrate, acetoacetate, citicoline, and orotate) progressively changed from the H to the CM stage, and other 12 metabolites (Leu, taurine, Val, arabinose, galactose, ribose, acetate, formate, pyruvate, 5-dodecenoic acid, 3-hydroxybutyrate, and ascorbate) differed only in the CM group. These metabolites were related to blood-milk barrier damage, inflammation, oxidative stress, cell proliferation, energy and lipid metabolisms, the citrate (TCA) cycle, systemic energy status, and microorganism metabolism. These results suggest that metabolomic alterations in milk begin to occur when the mammary gland is infected with somatic cells within normal limits and progressively worsen.

IntroductionBovine tuberculosis (TB) in water buffalo (Bubalus bubalis) is primarily diagnosed using intra vitam tests, as the intradermal tuberculin test (IDT) and the interferon-gamma release assay (IGRA), both of which detect cell-mediated immunity (CMI). However, a subset of infected animals fails to mount a detectable CMI response, posing a significant risk of undetected transmission. Serological tests assessing the humoral immune response could provide a valuable complementary tool for identifying infected animals that escape detection through traditional CMI-based assays.MethodsThis study analyzed 895 serum samples from water buffaloes, including 393 from TB-free herds and 502 from TB-infected herds. Animals from TB-free herds were tested using IDT and an ELISA assay, whereas those from infected herds were also tested using IGRA. We developed an ELISA assay targeting MPB70, MPB83, ESAT6, CFP10, PPDB, and P22 antigens to investigate the role of the humoral response in TB diagnosis.Results and discussionThe ELISA showed a specificity of 98.2%. However, sensitivity differed based on the antigen used: among the most reactive proteins, sensitivity was 67.5% for MPB70, 69.8% for P22, and 74.4% for PPDB. Moreover, approximately 70% of samples with discordant IDT and IGRA results, as well as those with positive IDT but inconclusive IGRA results, tested positive by serology, highlighting the potential of antibody-based detection to improve TB diagnosis in buffaloes. Our findings suggest that integrating serological testing with standard diagnostic methods could enhance the detection of infected animals, ultimately contributing to better TB control in buffalo populations.

The Gulf of Pozzuoli, a marginal sub-basin of the Tyrrhenian Sea, has a longstanding tradition of bivalve mollusk farming. Despite its historical importance, the area faces significant anthropogenic pressures, particularly from the decommissioned industrial site at Bagnoli. Seasonal monitoring data indicate peaks in mussel contamination by polycyclic aromatic hydrocarbons (PAHs) in late autumn and early winter. This study analyzed the oceanographic and meteorological dynamics driving PAH contamination in 2016, when elevated PAH concentrations were recorded at one farming site. A combination of in situ measurements, satellite observations, numerical model outputs, and Lagrangian simulations was used to examine sea surface circulation, wind patterns, wave dynamics, and particle transport pathways. Contamination events were consistently preceded by approximately one week of northwestward surface currents, likely facilitating pollutant transport from Bagnoli to the farming area. Southerly waves exceeding 1.2 meters in height and periods longer than 8 seconds were recorded prior to contamination peaks, suggesting wave-induced resuspension of PAHs from sediments. These hydrodynamic conditions coincided with prevailing southwesterly winds. The findings support the hypothesis that specific meteorological and oceanographic conditions enhance PAH transport from Bagnoli to the mussel farms, posing risks to aquaculture and public health. Based on previous and current data, the study provides robust conclusions on PAH pollution in the Gulf of Pozzuoli and offers a rationale for improving monitoring and management strategies to mitigate contamination risks in vulnerable coastal areas.

Multidrug-resistant Acinetobacter baumannii is an important pathogen causing healthcare-associated infections. The onset of Pan Drug Resistant (PDR) strains of A. baumannii, which are also resistant to colistin due to its excessive consumption, makes these strains difficult to eradicate. The appearance of new aggressive phenotypes among sentinel microorganisms in the control of healthcare-associated infections (HAI) requires the molecular approach to define a database of local nosocomial strains. Whole Genome Sequencing (WGS) is a useful tool to monitor the emergence of new aggressive bacterial phenotypes among sentinel microorganisms for the control of HAI. We describe a case report of colistin-resistant A. baumannii strain isolated from a blood-stream infection and evaluate the presence of antibiotic resistance genes (ARGs) by whole-genome sequencing (WGS). A colistin-resistant, PDR A. baumannii strain was isolated from blood cultures, bile fluid and wound in a patient with bacteremia in "Santa Maria delle Grazie Hospital" , Pozzuoli Italy. The strain was characterized by both biochemical and Whole Genome Sequencing (WGS) analysis. We identified a colistin-resistant strain of A. baumannii from a human bloodstream infection. WGS results were integrated with diagnostic procedures for a better characterization of the co-occurrence of several distinct antibiotic resistance mechanisms.

The presence of mycotoxins in cheese is a significant concern due to their potential health risks. Mycotoxins can contaminate cheese through two main routes: indirectly via contaminated animal feed, and/or directly, because of mold growth on dairy products. It has been reported that cheese may contain metabolites of aflatoxin B1 such as aflatoxin M1 (AFM1), aflatoxicol (AFL), and, its precursor, sterigmatocystin (STC). This study presents a reliable method for the simultaneous determination of AFM1, AFL, and STC in cheeses made from ovine, goat, or buffalo milk. The method was developed using single liquid extraction, clean-up by an immunoaffinity column (IAC), and liquid chromatography-tandem mass spectrometry (LC-MS/MS) determination. The method was subjected to initial validation according to EU regulations, which outline the required performance parameters and criteria of analytical methods for official food control. The limits of quantification (LOQs) of the method for AFM1, AFL, and STC are 2.0 ng/kg, 5.0 ng/kg, and 1.0 ng/kg, respectively. The method was applied in a study for the assessment of mycotoxin transfer from milk to cheeses and also their growth.

There are four genotypes of ovine papillomaviruses (OaPVs): OaPV1, OaPV2, and OaPV4, which are ovine delta papillomaviruses responsible for epithelial and mesenchymal cell infections, and OaPV3, an epitheliotropic Dyokappapapillomavirus associated with cutaneous tumors in sheep, including squamous cell carcinoma. Vaginal swabs of healthy mares were evaluated for the presence of PVs to investigate whether the vaginal virobiota of asymptomatic mares harbored OaPVs. High-performance digital droplet polymerase chain reaction (ddPCR) was used to quantitatively detect OaPV types 1–4 DNA in 94 vaginal swabs collected at the National Reference Center for Veterinary and Comparative Oncology (CEROVEC), Genoa, Italy. All samples were comparatively evaluated for OaPV DNA loading using real-time quantitative PCR. ddPCR detected OaPV DNA in 25 vaginal swab samples (26.6%), whereas qPCR revealed 13 vaginal swabs (11.7%). Differences between the two molecular protocols were determined to be statistically significant using McNemar’s test (p < 0.0005). The detected OaPV types were OaPV1 and OaPV3. Both methods failed to detect OaPV2 or OaPV4 DNA, which could be attributed to the limited number of samples examined. OaPV1 is the most prevalent OaPV in equine vaginal virobiota . This study is the first to provide evidence of the presence of OaPV DNA in vaginal swabs of healthy mares. This comparative detection approach underscores the superior sensitivity of ddPCR over qPCR.