Hemosiderin-Laden Macrophages in Canine Mammary Carcinomas.

Macrophages are among the main actors in cancer immunoediting, with several functions, including recycling iron and packaging it in hemosiderin. Even though TAMs are widely studied in breast cancer and canine mammary tumors, hemosiderin-laden macrophages (HLMs) have not received as much attention. Considering the growing interest in iron metabolism in cancer, this study aims to evaluate the presence of HLMs in canine mammary tumors. Fifty cases of canine mammary carcinomas presenting aggregates of pigmented macrophages were chosen. Prussian blue and Meguro staining were performed to assess the presence of iron. Immunohistochemistry was carried out to try to identify macrophagic phenotypes and hypothesize their role. Evaluation of the H&E sections showed that pigmented macrophages were variously localized in peritumoral and stromal areas. These pigmented cells were variably stained with Prussian blue and reacted strongly with DAB in the Meguro staining method, thus confirming the presence of iron within them. In their immunohistochemistry, the HLMs were negative for the MAC387 but positive for CD 204 and VEGF. Considering their positivity for CD 204, HLMs could be M2 macrophages that supply iron to both the neoplastic cells and the tumor inflammatory microenvironment, promoting angiogenesis and protecting cancer cells from hypoxia.

Positive Impacts of Aphanizomenon Flos Aquae Extract on Obesity-Related Dysmetabolism in Mice with Diet-Induced Obesity.

The present study evaluated the ability of KlamExtra®, an Aphanizomenon flos aquae (AFA) extract, to counteract metabolic dysfunctions due to a high fat diet (HFD) or to accelerate their reversion induced by switching an HFD to a normocaloric diet in mice with diet-induced obesity. A group of HFD mice was fed with an HFD supplemented with AFA (HFD-AFA) and another one was fed with regular chow (standard diet-STD) alone or supplemented with AFA (STD-AFA). AFA was able to significantly reduce body weight, hypertriglyceridemia, liver fat accumulation and adipocyte size in HFD mice. AFA also reduced hyperglycaemia, insulinaemia, HOMA-IR and ameliorated the glucose tolerance and the insulin response of obese mice. Furthermore, in obese mice AFA normalised the gene and the protein expression of factors involved in lipid metabolism (FAS, PPAR-γ, SREBP-1c and FAT-P mRNA), inflammation (TNF-α and IL-6 mRNA, NFkB and IL-10 proteins) and oxidative stress (ROS levels and SOD activity). Interestingly, AFA accelerated the STD-induced reversion of glucose dysmetabolism, hepatic and VAT inflammation and oxidative stress. In conclusion, AFA supplementation prevents HFD-induced dysmetabolism and accelerates the STD-dependent recovery of glucose dysmetabolism by positively modulating oxidative stress, inflammation and the expression of the genes linked to lipid metabolism.

RNAseq Analysis of Livers from Pigs Treated with Testosterone and Nandrolone Esters: Selection and Field Validation of Transcriptional Biomarkers.

The use of anabolic-androgenic steroids (AASs) as growth promoters in farm animals is banned in the European Union, representing both an illicit practice and a risk for consumer health. However, these compounds are still illegally administered, often in the form of synthetic esters. This work aimed to characterize significant coding RNA perturbations related to the illicit administration of testosterone and nandrolone esters in fattening pigs. A total of 27 clinically healthy 90-day-old pigs were randomly assigned to test and control groups. Nine animals were treated with testosterone esters (Sustanon®) and other nine with nandrolone esters (Myodine®). At the end of the trial, liver samples were collected and analyzed using RNAseq, allowing the identification of 491 differentially expressed genes (DEGs). The transcriptional signature was further characterized by a smaller sub-cluster of 143 DEGs, from which a selection of 16 genes was made. The qPCR analysis confirmed that the identified cluster could still give good discrimination between untreated gilt and barrows compared to the relative testosterone-treated counterparts. A conclusive field survey on 67 liver samples collected from pigs of different breeds and weight categories confirmed, in agreement with testosterone residue profiles, the specificity of selected transcriptional biomarkers, showing their potential applications for screening purposes when AAS treatment is suspected, allowing to focus further investigations of competent authorities and confirmatory analysis where needed.

Peripheral Nerve Regeneration at 1 Year: Biodegradable Polybutylene Succinate Artificial Scaffold vs. Conventional Epineurial Sutures.

The utilization of a planar poly(1,4-butylene succinate) (PBS) scaffold has been demonstrated as an effective approach for preserving nerve continuity and facilitating nerve regeneration. In this study, we assessed the characteristics of a microfibrous tubular scaffold specifically designed and fabricated through electrospinning, utilizing PBS as a biocompatible and biodegradable material. These scaffolds were evaluated as nerve guide conduits in a rat model of sciatic nerve neurotmesis, demonstrating both their biodegradability and efficacy in enhancing the reconstruction process over a long-term period (1-year follow-up). Histological assay and electrophysiological evaluation were performed to compare the long-term outcomes following sutureless repair with the microfibrillar wrap to outcomes obtained using traditional suture repair.

Epidemiological and genetic analysis of Cetacean Morbillivirus circulating on the Italian coast between 2018 and 2021.

Cetacean morbillivirus (CeMV) has caused several outbreaks, unusual mortality events, and interepidemic single-lethal disease episodes in the Mediterranean Sea. Since 2012, a new strain with a northeast (NE) Atlantic origin has been circulating among Mediterranean cetaceans, causing numerous deaths. The objective of this study was to determine the prevalence of CeMV in cetaceans stranded in Italy between 2018 and 2021 and characterize the strain of CeMV circulating. Out of the 354 stranded cetaceans along the Italian coastlines, 113 were CeMV-positive. This prevalence (31.9%) is one of the highest reported without an associated outbreak. All marine sectors along the Italian coastlines, except for the northern Adriatic coast, reported a positive molecular diagnosis of CeMV. In one-third of the CeMV-positive cetaceans submitted to a histological evaluation, a chronic form of the infection (detectable viral antigen, the absence of associated lesions, and concomitant coinfections) was suspected. Tissues from 24 animals were used to characterize the strain, obtaining 57 sequences from phosphoprotein, nucleocapsid, and fusion protein genes, which were submitted to GenBank. Our sequences showed the highest identity with NE-Atlantic strain sequences, and in the phylogenetic study, they clustered together with them. Regarding age and species, most of these individuals were adults (17/24, 70.83%) and striped dolphins (19/24, 79.16%). This study improves our understanding on the NE-Atlantic CeMV strain in the Italian waters, supporting the hypothesis of an endemic circulation of the virus in this area; however, additional studies are necessary to deeply comprehend the epidemiology of this strain in the Mediterranean Sea.

Physiologic Cyclical Load on Inguinal Hernia Scaffold ProFlor Turns Biological Response into Tissue Regeneration.

Surgical repair of groin protrusions is one of the most frequently performed procedures. Currently, open or laparoscopic repair of inguinal hernias with flat meshes deployed over the hernial defect is considered the gold standard. However, fixation of the implant, poor quality biologic response to meshes and defective management of the defect represent sources of continuous debates. To overcome these issues, a different treatment concept has recently been proposed. It is based on a 3D scaffold named ProFlor, a flower shaped multilamellar device compressible on all planes. This 3D device is introduced into the hernial opening and, thanks to its inherent centrifugal expansion, permanently obliterates the defect in fixation-free fashion. While being made of the same polypropylene material as conventional hernia implants, the 3D design of ProFlor confers a proprietary dynamic responsivity, which unlike the foreign body reaction of flat/static meshes, promotes a true regenerative response. A long series of scientific evidence confirms that, moving in compliance with the physiologic cyclical load of the groin, ProFlor attracts tissue growth factors inducing the development of newly formed muscular, vascular and nervous structures, thus re-establishing the inguinal barrier formerly wasted by hernia disease. The development up to complete maturation of these highly specialized tissue elements was followed thanks to biopsies excised from ProFlor from the short-term up to years post implantation. Immunohistochemistry made it possible to document the concurrence of specific growth factors in the regenerative phenomena. The results achieved with ProFlor likely demonstrate that modifying the two-dimensional design of hernia meshes into a 3D outline and arranging the device to respond to kinetic stresses turns a conventional regressive foreign body response into advanced probiotic tissue regeneration.

Silk-Based Matrices and c-Kit-Positive Cardiac Progenitor Cells for a Cellularized Silk Fibroin Scaffold: Study of an in vivo Model.

The production of a cellularized silk fibroin scaffold is very difficult because it is actually impossible to differentiate cells into a well-organized cardiac tissue. Without vascularization, not only do cell masses fail to grow, but they may also exhibit an area of necrosis, indicating a lack of oxygen and nutrients. In the present study, we used the so-called tyrosine protein kinase kit (c-Kit)-positive cardiac progenitor cells (CPCs) to generate cardiac cellularized silk fibroin scaffolds, multipotent cells isolated from the adult heart to date that can show some degree of differentiation toward the cardiac phenotype. To test their ability to differentiate into the cardiac phenotype in vivo as well, CPC and collagen organoid-like masses were implanted into nude mice and their behavior observed. Since the 3-dimensional structure of cardiac tissue can be preserved by scaffolds, we prepared in parallel different silk fibroin scaffolds with 3 different geometries and tested their behavior in 3 different models of immunosuppressed animals. Unfortunately, CPC cellularized silk fibroin scaffolds cannot be used in vivo. CPCs implanted alone or in collagen type I gel were destroyed by CD3+ lymphocyte aggregates, whereas the porous and partially oriented scaffolds elicited a consistent foreign body response characterized by giant cells. Only the electrospun meshes were resistant to the foreign body reaction. In conclusion, c-Kit-positive CPCs, although expressing a good level of cardiac differentiation markers in vitro with or without fibroin meshes, are not suitable for an in vivo model of cardiac organoids because they are degraded by a T-cell-mediated immune response. Even scaffolds which may preserve the survival of these cells in vivo also induced a host response. However, among the tested scaffolds, the electrospun meshes (F-scaffold) induced a lower response compared to all the other tested structures.

African Swine Fever and Its Control Measures in Wild Boar: A "De Iure Condito" Analysis in the European Union.

Recently, the African swine fever (ASF) epizootic has been reported in domestic pigs and wild boars in several European Union Member States (EU MS) and epidemiological evidence has accumulated which indicates that wild boar play a key role in maintaining and spreading the disease. Thanks to the experience gained when managing ASF outbreaks in Sardinia (Italy) and Eastern Europe, Directive 2002/60 CE was issued. This directive represented an important step forward in controlling the disease, particularly the risk of spreading the virus to wild animals. Since 2021, according to Regulation (EU) 2016/429, which is also called "Animal Health Law-AHL", when the MS competent authority suspects or confirms ASF (a cat. A listed disease) in wild animals, it is mandatory to conduct surveillance in the wild boar population and implement the necessary control measures. Within AHL, Regulations (EU) 2020/687 and 2023/594 established special ASF control measures in kept and wild porcine animals, and their products and by-products, focusing on and underlying old and new responsibilities that vets (both public and private ones) have to accomplish under the new regulations. The new change in the legal framework deals with specific measures to be applied in the wild and represents a great challenge for MS veterinary services. Some of these measures have been well established in the last two decades, particularly those related to application in the farming system, while other measures are still new to veterinary health management and require a holistic approach in terms of intensity, considering all geographical, ecological, productive, cultural and social features of the involved EU MS. In this contribution, the authors intend to focus on specific measures which have been issued in order to limit or stop the spread of ASF in a wild, "boundless" ecosystem. These measures expand the field of competence of the official veterinary service to wild areas in addition to farm activity.

Preliminary Study on the Host Response to Bivalent and Monovalent Autogenous Vaccines against Mycoplasma agalactiae in Dairy Sheep.

In Italy, dairy sheep farming represents a vital agro-industry sector, but it is still challenged by contagious agalactia (CA), which is endemic there, and vaccination is the most economical and sustainable tool for control. This study aimed to evaluate the combined Mycoplasma agalactiae (Ma)-Staphylococcus aureus (Sa) vaccine (Ma-Sa) against the Ma monovalent vaccine in ewes. Twelve primiparous Ma-free ewes were randomly grouped into three equal groups: first, the control group injected with placebo, second, the group vaccinated with the Ma monovalent vaccine, and third, the group vaccinated with Ma-Sa combined vaccine, with two S/C doses at 45-day intervals. The animals were examined for serological, hematological, and somatic cell count (SCC) changes for 17 successive weeks. A significant increase in anti-Ma antibody mean titers, leukocytes, and platelets was observed in the vaccinated animals, with the highest values in those who received the combined vaccine. Neutrophils were high only in the animals who received the combined vaccine. SCC was lower in the vaccinated animals during the first six weeks. This study concludes that the combined Ma-Sa vaccines enhance immune response and potentiate its efficacy against Ma. This improvement might be attributed to the sensitization/activation effect of S. aureus on platelets, which are recoded to act as a key regulator for the coordination of all components of the innate immune system. Even though this study included a small number of animals, its findings about the potentialities of this inactivated vaccine in the control of CA are strongly encouraging. Further confirmation might be needed through additional replicates and a challenge study is needed before proceeding with widespread use.

Study on the Canine Adenovirus Type 1 (CAdV-1) Infection in Domestic Dogs in Southern Italy.

Canine adenovirus type 1 (CAdV-1) is the causative agent of a systemic and potentially fatal viral disease of domestic and wild canids. In Italy, CAdV-1 infection has also been occasionally described in dogs, but information on the epidemiology and its genomic features is still limited. A study was conducted on 291 dogs suspected of infectious gastrointestinal disease. Samples collected from dogs in southern Italy between 2017 and 2020 were analyzed. Virological and histopathological assays were carried out. The presence of CAdVs and other canine viral enteropathogens was investigated, and sequence and phylogenetic analyses were performed. CAdV-1 was detected in six (2.1%) dead stray dogs alone or in mixed infections with other viruses. Gross lesions and histopathological findings referred to CAdV infection were observed, also involving the central nervous system tissues. All inoculated samples were successfully isolated. Sequence analysis evidenced divergences with the circulating strains previously described in Italy and a closer relation with older CAdV-1 strains collected from other countries, suggesting a genetic heterogeneity of CAdV-1 in Italy. The evidence of the circulation of CAdV-1 and its genomic features allows us to have more in-depth knowledge of the epidemiology and evolution of the CAdV-1 genomic variants.

Does TLS Exist in Canine Mammary Gland Tumours? Preliminary Results in Simple Carcinomas.

Neoplastic progression is influenced by the expression of tumour antigens that activate an anti-tumour immune response. Human medical studies show that this body defence is carried out in secondary lymphoid organs (SLOs) but also directly in the tumour through organized cellular aggregates that are called tertiary lymphoid structures (TLSs). However, their occurrence has different meanings in different tumour types. For example, the presence of TLSs in breast cancer is associated with the most aggressive subtypes. This paper aimed to study TLSs in canine mammary simple carcinomas. A morphological assessment of the inflammatory infiltrate was performed on H&E sections of fifty cases. Immunohistochemistry was then carried out to typify the inflammatory cells in the tumour microenvironment. Results showed that, sometimes, inflammatory infiltrates were organized in follicles close to high-grade carcinomas, simulating a lymphoid organization, as in breast cancer. Therefore, we can assume that even in canine mammary tumours, TLSs exist and they are entities to consider due to their presence in the most aggressive histotypes or tumours with a high degree of malignancy.

Dynamic Responsive Inguinal Scaffold Activates Myogenic Growth Factors Finalizing the Regeneration of the Herniated Groin.

BACKGROUND: Postoperative chronic pain caused by fixation and/or fibrotic incorporation of hernia meshes are the main concerns in inguinal herniorrhaphy. As inguinal hernia is a degenerative disease, logically the treatment should aim at stopping degeneration and activating regeneration. Unfortunately, in conventional prosthetic herniorrhaphy no relationship exists between pathogenesis and treatment. To overcome these incongruences, a 3D dynamic responsive multilamellar scaffold has been developed for fixation-free inguinal hernia repair. Made of polypropylene like conventional flat meshes, the dynamic behavior of the scaffold allows for the regeneration of all typical inguinal components: connective tissue, vessels, nerves, and myocytes. This investigation aims to demonstrate that, moving in tune with the groin, the 3D scaffold attracts myogenic growth factors activating the development of mature myocytes and, thus, re-establishing the herniated inguinal barrier. METHODS: Biopsy samples excised from the 3D scaffold at different postoperative stages were stained with H&E and Azan-Mallory; immunohistochemistry for NGF and NGFR p75 was performed to verify the degree of involvement of muscular growth factors in the neomyogenesis. RESULTS: Histological evidence of progressive muscle development and immunohistochemical proof of NFG and NFGRp75 contribution in neomyogenesis within the 3D scaffold was documented and statistically validated. CONCLUSION: The investigation appears to confirm that a 3D polypropylene scaffold designed to confer dynamic responsivity, unlike the fibrotic scar plate of static meshes, attracts myogenic growth factors turning the biological response into tissue regeneration. Newly developed muscles allow the scaffold to restore the integrity of the inguinal barrier.

Potential SARS-CoV-2 Susceptibility of Cetaceans Stranded along the Italian Coastline.

Due to marine mammals' demonstrated susceptibility to SARS-CoV-2, based upon the homology level of their angiotensin-converting enzyme 2 (ACE2) viral receptor with the human one, alongside the global SARS-CoV-2 occurrence and fecal contamination of the river and marine ecosystems, SARS-CoV-2 infection may be plausibly expected to occur also in cetaceans, with special emphasis on inshore species like bottlenose dolphins (Tursiops truncatus). Moreover, based on immune and inflammatory responses to SARS-CoV-2 infection in humans, macrophages could also play an important role in antiviral defense mechanisms. In order to provide a more in-depth insight into SARS-CoV-2 susceptibility in marine mammals, we evaluated the presence of SARS-CoV-2 and the expression of ACE2 and the pan-macrophage marker CD68. Aliquots of tissue samples, belonging to cetaceans stranded along the Italian coastline during 2020-2021, were collected for SARS-CoV-2 analysis by real-time PCR (RT-PCRT) (N = 43) and Immunohistochemistry (IHC) (N = 59); thirty-two aliquots of pulmonary tissue sample (N = 17 Tursiops truncatus, N = 15 Stenella coeruleoalba) available at the Mediterranean Marine Mammal Tissue Bank (MMMTB) of the University of Padua (Legnaro, Padua, Italy) were analyzed to investigate ACE2 expression by IHC. In addition, ACE2 and CD68 were also investigated by Double-Labeling Immunofluorescence (IF) Confocal Laser Microscopy. No SARS-CoV-2 positivity was found in samples analyzed for the survey while ACE2 protein was detected in the lower respiratory tract albeit heterogeneously for age, gender/sex, and species, suggesting that ACE2 expression can vary between different lung regions and among individuals. Finally, double IF analysis showed elevated colocalization of ACE2 and CD68 in macrophages only when an evident inflammatory reaction was present, such as in human SARS-CoV-2 infection.

In vivo efficacy of verteporfin loaded gold nanorods for combined photothermal/photodynamic colon cancer therapy.

The high incidence of cancer recurrences and the frequent occurrence of multidrug resistance often stem from a poorly selective and inefficient antineoplastic therapy, responsible for the onset of undesired side effects as well. A combination of minimal-invasive approaches could thus be a useful strategy to surmount these shortcomings, achieving a safe and solid cancer therapy. Herein, a multi-therapeutic nanotool was designed by merging the photothermal properties of gold nanorods (AuNRs) with the photodynamic activity of the photosensitizer verteporfin. AuNRs were coated with the natural materials lipoic acid and gellan gum (AuNRs_LA,GG) and subsequently loaded with verteporfin (AuNRs_LA,GG/Vert) producing stable colloidal dispersions. AuNRs_LA,GG/Vert were characterized in terms of stability, size and morphology. The hyperthermia exhibited after NIR excitation (810 nm) was also evaluated to highlight the effect on increasing the drug released profile in intra-tumoral mimicking media, as well as cytotoxicity on human colon cancer cell line (HCT116). In vivo studies on HCT116 murine xenograft models were carried out to prove the ability of AuNRs_LA,GG to arrest the tumor growth via NIR laser-triggered hyperthermia. Furthermore, the complete xenograft depletion was demonstrated upon AuNRs_LA,GG/Vert administration by combined photothermal (PTT) and photodynamic (PDT) effects.

The Italian Network of Laboratories for Veterinary Oncology (NILOV) 2.0: Improving Knowledge on Canine Tumours.

Advances in tumour research are crucial, and comparative oncology can improve the knowledge in several ways. Dogs are not only models of specific naturally occurring tumours but can also be sentinels of environmental exposures to carcinogens, as they share the same environment with their owners. The purpose of this work was to describe the data collected by The Italian Network of Laboratories for Veterinary Oncology in the first 9 years of activity (2013-2021) and to evaluate their potential epidemiological significance. Frequencies of tumour topographies and main morphologies in dogs were described, analysed and compared, calculating age-adjusted proportional morbidity ratios and considering several risk factors (breed, sex, period and region of residence). These observations allowed us to highlight differences not only in morphology and topography of some tumours but also to formulate hypotheses on the potential role of some risk factors, e.g., neutering/spaying or geographical location. In our opinion, the results of this case series confirm the importance of initiating and consolidating animal cancer registration initiatives that would facilitate the possibility of conducting multicentric collaborative studies to deepen the knowledge of the epidemiology of tumours in dogs from a comparative perspective.

Alternative Molecular Tools for the Fight against Infectious Diseases of Small Ruminants: Native Sicilian Sheep Breeds and Maedi-Visna Genetic Susceptibility.

Maedi-visna (MV) is a disease caused by small ruminant lentiviruses. It is included in the list of notifiable terrestrial animal diseases due to economic losses and animal welfare harm in the sheep sector. To date, control programs remain the onliest approach to avoiding infection. The allelic variant p.Glu35Lys (E35K) of the TMEM154 gene has been strongly associated with host vulnerability to MV illness. The present study aimed to investigate the association of TMEM154 E35K allele frequencies with MV susceptibility in native Sicilian sheep breeds. More than 400 animals from 14 local sheep were serologically tested and genotyped for the TMEM154 E35K polymorphism. The local breeds displayed different values of MV seroprevalence, with the lowest antibody prevalence in Barbaresca and Pinzirita breeds. TMEM154 protective allele (K35) was less frequent than the risk allele (E35) in Valle del Belìce breed, whereas the other three breeds showed a more balanced alleles distribution. A positive association between seroprevalence and genotype was found in the entire sample set. The risk of infection resulted in more than 3-fold times as high in sheep with EK and EE genotype compared to the KK genotype. Our data could be helpful in establishing selection breeding programs aimed at reducing MV infection in Sicilian sheep farming and encouraging the breeding of native breeds.

Mycobacterium bovis Tuberculosis in Two Goat Farms in Multi-Host Ecosystems in Sicily (Italy): Epidemiological, Diagnostic, and Regulatory Considerations.

Mycobacterium bovis (M. bovis) is the causative agent of animal tuberculosis (bTB), infecting and causing disease in several animal species. In areas where there are complex interactions between reservoir hosts and susceptible species, the control of this pathogen is a challenge. The authors report two outbreaks of goat tuberculosis caused by M. bovis in multi-host ecosystems within two protected natural areas of Sicily, where TB is historically endemic. The first outbreak (Farm A) was identified after the incidental detection at the slaughterhouse of TB-like lesions in goat viscera ready to be disposed. Single intradermal cervical tuberculin test (SICT) was performed in Farm A on 205 goats, resulting positive in 10 (4.9%). After slaughtering, six out of ten animals showed TB-like lesions, from which M. bovis spoligotype SB0841 was isolated. The typing did not reveal any epidemiological connection with the neighboring cattle, suggesting that free-ranging type of management exposed the affected goat livestock or wildlife infected with other strains. The second outbreak (Farm B) was detected in a mixed farm (bovine, caprine, and ovine), where relapsing outbreaks of TB in cattle were registered in the previous years after performing the SICT in cohabiting goats. SICT resulted positive in 6/153 (3.9%), and two animals showed bTB-like lesions. No mycobacteria were cultured, and the final diagnosis of TB was achieved by histopathology and immunohistochemistry. The reported outbreaks highlight the importance of assessing the epidemiological, diagnostic, and regulatory critical issue, which is fundamental to optimizing the strategies of eradicating TB in the endemic multi-host ecosystem described.

First Molecular Survey to Detect Mycoplasma gallisepticum and Mycoplasma synoviae in Poultry Farms in a Strategic Production District of Sicily (South-Italy).

Mycoplasmas are recognized as avian pathogens, which may cause both respiratory disease and synovial infections in poultry, resulting in severe economic losses. Our study aims to determine the occurrence of Mycoplasma gallisepticum (MG) and Mycoplasma synoviae (MS) among commercial and rural laying hens located in Ragusa province (South Italy), using a duplex real time PCR. Four hundred tracheal swabs were collected from seven commercial (200 swabs) and 25 rural (200 swabs) farms without any clinical disease history. Out of 400 swabs collected, 50 (12.5%) and 93 (23.25%) were positive for MG and MS, respectively. In particular, 9 (18%) and 22 (23.65%) positive swabs for MG and MS, respectively, originated from commercial farms, compared to 41 (82%) and 71 (76.34%) obtained from rural farms. Data obtained show a lower prevalence of MG than MS in the studied farms. Moreover, both pathogens were spread in rural and commercial farms. PCR could be concluded as a rapid and sensitive method for the identification of MG and MS in areas where commercial farms that are declared Mycoplasma-free and rural flocks coexist. These data highlight the importance of surveillance also in rural poultry to monitoring the occurrence of mycoplasmas strains in strategic productive districts.

Neuropathological Characterization of Dolphin Morbillivirus Infection in Cetaceans Stranded in Italy.

Cetacean morbillivirus (CeMV) is responsible for epidemic and endemic fatalities in free-ranging cetaceans. Neuro-inflammation sustained by CeMV is a leading cause of death in stranded cetaceans. A novel dolphin morbillivirus (DMV) strain of Atlantic origin circulating in Italian waters since early 2016 has caused acute/subacute lesions associated with positive immunolabelling of the virus. To date, myelin damage has not been fully documented and investigated in cetaceans. This study describes neuropathological findings in the brain tissue of 31 cetaceans found stranded along the Italian coastline and positive for DMV infection on molecular testing. Cell changes in the areas of myelinopathy were revealed by double indirect immunofluorescence. The most frequent DMV-associated lesions were astro-microgliosis, neuronal necrosis, spongiosis, malacia, and non-suppurative meningoencephalitis. Myelin reduction and areas of demyelination were revealed by means of a specific myelin biomarker. Morbilliviral antigen immunolabelling was mainly observed in neurons and microglial cells, in association with a marked activation of microglia and astrocytes. These findings extend our knowledge of DMV-associated brain lesions and shed light on their pathogenesis.

Investigation of an Autochthonous Outbreak of Bovine Besnoitiosis in Northwestern Sicily.

Bovine besnoitiosis is a parasitic disease caused by the protozoan Besnoitia besnoiti, leading to infertility in bulls and abortions in cows. In Italy, it is considered an emerging disease, recently introduced by the importation of animals from Spain and France. In the last decade, many outbreaks have been reported and confirmed in native cattle, mostly in northern and central Italy. This study reports on an autochthonous outbreak of bovine besnoitiosis in two nearby farms located in northwestern Sicily. A 15-month-old Limousine bull born on the farm showed typical clinical signs of the chronic disease phase, such as edema of the scrotum with subsequent hyperkeratosis associated with eschars and thickening of the skin. A histopathological examination revealed the presence of Besnoitia tissue cysts containing bradyzoites in the eyes, tendons, testicles, dermis, and nictitating membrane. A serological investigation using a commercial ELISA kit revealed a high seroprevalence of the antibody anti-B. besnoiti (79.2% for the farms in this study). Clinical disease showed low prevalence (1.5%) despite the high seroprevalence of specific antibodies in the herd, confirming that bovine besnoitiosis is an emergent endemic pathogen in Sicily, but its clinical behavior still remains sporadic.