Valentina Iovane

Serum samples were collected from wild boars (Sus scrofa) harvested during the 2005-2006 hunting season in Campania, southern Italy. Samples were tested for antibodies to Leptospira interrogan, Brucella spp., Salmonella spp., Aujeszky disease virus (ADV), porcine reproductive and respiratory stress syndrome virus (PRRSV), porcine parvovirus (PPV), classical swine fever virus (CSFV), and swine vesicular disease virus (SVDV). Of the 342 serum samples tested, 15 (4.4%) were seropositive to Brucella spp., nine (2.6%) were seropositive to L. interrogans, 66 (19.3%) were seropositive for Salmonella spp., 105 (30.7%) were seropositive for ADV, 27 (7.9%) were seropositive for PPV, and 129 (37.7%) were seropositive for PRRSV. All sera tested seronegative for SVDV and CSFV antibodies. These results, recorded for the first time in Campania, support the hypothesis that wild boar are reservoirs of certain infectious agents, but some infections in wild boars originate from their domestic counterparts.

Porcine coronaviruses and reproductive and respiratory syndrome (PRRS) are responsible for severe outbreaks that cause huge economic losses worldwide. In Italy, three coronaviruses have been reported historically: porcine epidemic diarrhea virus (PEDV), transmissible gastroenteritis virus (TGEV) and porcine respiratory coronavirus (PRCV). Although repeated outbreaks have been described, especially in northern Italy, where intensive pig farming is common, there is a worrying lack of information on the spread of these pathogens in Europe. In this work, we determined the seroprevalence of three porcine coronaviruses and PRRSV in the Campania region, southern Italy. A total of 443 samples were tested for the presence of antibodies against porcine coronaviruses and PRRSV using four different commercial ELISAs. Our results indicated that PEDV is the most prevalent among porcine coronaviruses, followed by TGEV, and finally PRCV. PRRSV appeared to be the most prevalent virus (16.7%). For coronaviruses, seroprevalence was higher in pigs raised in intensive farming systems. In terms of distribution, TGEV is more widespread in the province of Avellino, while PEDV and PRRSV are more prevalent in the province of Naples, emphasizing the epidemic nature of both infections. Interestingly, TGEV-positive animals are more common among growers, while seropositivity for PEDV and PRRSV was higher in adults. Our research provides new insights into the spread of swine coronaviruses and PRRSV in southern Italy, as well as a warning about the need for viral surveillance.

Abstract Imatinib (IM) is considered the gold standard for chronic myeloid leukemia (CML) treatment, although resistance is emerging as a significant problem. The proinflammatory cytokines interleukin‐6 (IL‐6) and interleukin‐8 (IL‐8) play an important role in cell proliferation, survival, and resistance to glucocorticoid‐mediated cell death. Several transcription factors such as NF‐KB and AP‐1 are activated in response to physiopathological increases and modulation of intracellular calcium levels. Our previous study demonstrated that lymphocytes from CML patients showed dysregulated calcium homeostasis and oxidative stress. Alteration in ionized calcium concentration in the cytosol has been implicated in the initiation of secretion, contraction, and cell proliferation. In this study, we hypothesized that IL‐6, IL‐8, NF‐kB, AP‐1, and intracellular calcium may be used as selective and prognostic factors to address the follow‐up in CML patients treated with imatinib. Our results demonstrated a significant down‐regulation in IL‐6 and IL‐8 release as well as NF‐kB and AP‐1 activation in lymphomonocytes from Imatinib‐treated patients, compared to samples from untreated patients. In parallel, IM treatment, in vivo and in vitro, were able to modulate the intracellular calcium concentration of peripheral blood mononuclear cells of CML patients by acting at the level of InsP 3 receptor in the endoplasmic reticulum and at the level of the purinergic receptors on plasma membrane. The results of this study show that measurements of NF‐kB, AP‐1, IL‐6, IL‐8, and intracellular calcium in CML patients treated with Imatinib may give important information to the hematologist on diagnostic criteria and are highly predictive in patients with newly diagnosed CML. J. Cell. Physiol. 227: 2798–2803, 2012. © 2011 Wiley Periodicals, Inc.

Hepatitis E virus (HEV) is a member of the genus Hepevirus within the family Hepeviridae. Hepatitis E is recognized as a zoonosis, and swine and wild boars (Sus scrofa) are known reservoirs of HEV infection. The aim of this study was to investigate the presence of HEV in wild boars and hunters exposed to infection in central Italy (Latium region). During the hunting season, blood samples were collected from 228 wild boars and 20 hunters. The seroprevalence of HEV infection was determined using a commercial enzyme-linked immunosorbent assay, previously validated for use in man, pigs and wild boars. The estimated HEV seroprevalence in wild boars and in hunters was 40.7% (93/228; 95% confidence interval [CI] 34.4-47.1%) and 25% (5/20; 95% CI 6.1-43.9%), respectively. Liver samples were collected from the boars and HEV RNA was detected by nested reverse transcriptase polymerase chain reaction. Fifty-five of 164 tested wild boar liver samples (33.5%; 95% CI 26.2-40.7%) and three of 20 (15.0%; 95% CI 1.3-28.7%) tested human serum samples were positive for HEV RNA. Phylogenetic analysis of the nucleotide sequences obtained from PCR products indicated that the HEV strains present in wild boars and the human population all belonged to genotype 3, supporting the zoonotic role of wild boars in the spread of HEV infection.

Following the COVID-19 epidemic outbreak in Ariano Irpino, Campania region (Italy), we tested lactating cows for the presence of SARS-CoV-2 on a cattle farm at which, prior to the investigation, 13 of the 20 farmworkers showed COVID-19-like symptoms, and one of them died. Twenty-four lactating cows were sampled to detect SARS-CoV-2. All nasal and rectal swabs and milk samples were negative for SARS-CoV-2 RNA. Of the 24 collected serum samples, 11 showed antibodies against SARS-CoV-2 nucleocapsid protein, 14 showed antibodies against SARS-CoV-2 spike protein, and 13 developed neutralising antibodies for SARS-COV-2; all samples were negative for Bovine Coronavirus (BCoV), another betacoronavirus. To our knowledge, this is the first report of natural serological evidence of SARS-CoV-2 infection in lactating cows. We hypothesise that this may be a case of reverse zoonosis. However, the role of cattle in SARS-CoV-2 infection and transmission seems to be negligible.

Sarcopenia, the age related loss of muscle mass and strength, is a multifactorial condition that occurs in a variety of species and represents a major healthcare concern for older adults in human medicine. In veterinary medicine, skeletal muscle atrophy is often observed in dogs as they reach old age, but the process is not well understood. Autophagy is a mechanism for degradation and recycling of cellular constituents and is potentially involved in sarcopenia. The aim of the present study was to evaluate the expression of three markers of autophagy, Beclin 1, LC3 and p62, in muscle wasting of geriatric dogs, to establish whether the levels of autophagy change with increasing age. Muscle biopsies from 25 geriatric dogs were examined and compared with those from five healthy young dogs. Samples from older dogs, assessed by routine histology, histoenzymatic staining and immunohistochemistry, showed evidence of muscle atrophy, sarcoplasmic vacuolisation and mitochondrial alterations. Furthermore, in 80% of the muscle samples from the older dogs, marked intracytoplasmic staining for Beclin 1 and LC3 was observed. Significantly greater expression of LC3 II and Beclin 1, but lower expression of p62, was found by Western blotting, comparing muscle samples from old vs. young dogs. The results of the study suggest that enhanced autophagy might be one of the factors underlying muscle atrophy in dogs as they age.

Abstract In this work, we investigated the effects of red orange and lemon extract (RLE) on ochratoxin A (OTA)‐induced nephrotoxicity. In particular, we analyzed the change in renal function and oxidative stress in Sprague–Dawley rats treated with OTA (0.5 mg/kg body weight, b.w.) and with RLE (90 mg/kg b.w.) by oral administration. After OTA treatment, we found alterations of biochemical and oxidative stress parameters in the kidney, related to a severe decrease of glomerular filtration rate. The RLE treatment normalized the activity of antioxidant enzymes and prevented the glomerular hyperfiltration. Histopathological examinations revealed glomerular damages and kidney cortex fibrosis in OTA‐rats, while we observed less severe fibrosis in OTA plus RLE group. Then, we demonstrated that oxidative stress could be the cause of OTA renal injury and that RLE reduces this effect.

The intestines are recognized as the main source of chronic inflammation in chronic kidney disease (CKD) and, among other cells, macrophages are involved in modulating this process as well as in the impaired immune response which also occurs in CKD patients. In this study, we evaluated the effect of Indoxyl Sulfate (IS), a protein bound uremic toxin poorly eliminated by hemodialysis, on inflammatory, oxidative stress and pro-apoptotic parameters, at the intestinal level in mice, on intestinal epithelial cells (IEC-6) and on primary murine peritoneal macrophages. C57BL/6J mice were treated with IS (800 mg/kg i.p.) for 3 or 6 h and histopathological analysis showed that IS induced intestinal inflammation and increased cyclooxygenase-2 (COX-2), nitrotyrosine and Bax expression in intestinal tissue. In IEC-6 cells, IS (125–1000 µM) increased tumor necrosis factor-α levels, COX-2 and inducible nitric oxide synthase expression and nitrotyrosine formation. Moreover, IS increased pro-oxidant, pro-inflammatory and pro-apoptotic parameters in peritoneal macrophages from IS-treated mice. Also, the serum concentration of IS and pro-inflammatory levels of cytokines resulted increased in IS-treated mice. Our results indicate that IS significantly contributes to affect intestinal homeostasis, immune response, and to induce a systemic pro-inflammatory state thus highlighting its potential role as therapeutic target in CKD patients.

Cattle and water buffalo are the main livestock species that are raised in the Campania region, southern Italy, and they contribute significantly to the regional rural economy. Currently there are limited data on the prevalence of relevant impact infections, such as bovine coronavirus (BCov), an RNA virus that causes acute enteric and respiratory disease. Although these diseases are described primarily in cattle, there have been reports of spillovers to other ruminants, including water buffalo. Here, we determined the seroprevalence of BCoV in cattle and water buffalo in the Campania region of southern Italy. An overall seroprevalence of 30.8% was determined after testing 720 sampled animals with a commercial enzyme-linked immunosorbent assay. A risk factor analysis revealed that the seropositivity rates in cattle (49.2%) were higher than in water buffalo (5.3%). In addition, higher seroprevalence rates were observed in older and purchased animals. In cattle, housing type and location were not associated with higher seroprevalence. The presence of BCoV antibodies in water buffalo was associated with the practice of co-inhabiting with cattle, demonstrating that this practice is incorrect and promotes the transmission of pathogens between different species. Our study found a considerable seroprevalence, which is consistent with previous research from other countries. Our results provide information on the widespread distribution of this pathogen as well as the risk factors that are involved in its transmission. This information could be useful in the control and surveillance of this infection.

The administration of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) to a variety of cultured cells may alter their ability to proliferate and die. In a previous study we demonstrated that TCDD induced proliferation in Madin-Darby Bovine Kidney (MDBK) cells where no signs of apoptosis were observed, but herein, analysis of MDBK cell morphology, in a large number of exposed cells, revealed some alterations, as expanded cytoplasm, an increase of intercellular spaces and many pyknotic nuclei. Hence, the aim of the current study was to elucidate the influences of dioxin on cell proliferation and cell death. We found that dioxin increased proliferation, as well as, activated cell death with autophagy, as we detected by increased amount of LC3-II, an autophagosome marker. Furthermore, formation of acidic vesicular organelles was observed by fluorescence microscopy following staining with the lysosomotropic agent acridine orange. These results were accompanied by down-regulation of telomerase activity, bTERT and c-Myc. Key tumor-suppressor protein p53 and expression of cell cycle inhibitor p21Waf1/Cip1 were activated after TCDD exposure. These changes occurred with activation of ATM phosphorylation in the presence of a decrease in Mdm2 protein levels. Taken together, these results support the idea that TCDD in MDBK cells, may exert its action, in part, by enhancing cell proliferation, but also by modulating the incidence of induced cell death with autophagy.

TGF-beta1 has been shown to induce autophagy in certain cells but whether and how this action is exerted in muscle and whether this activity relates to TGF-beta1 control of muscle cell differentiation remains unknown. Here, we show that expression of the autophagy-promoting protein phosphoprotein enriched in diabetes/phosphoprotein enriched in astrocytes (PED/PEA-15) progressively declines during L6 and C2C12 skeletal muscle cell differentiation. PED/PEA-15 underwent rapid induction upon TGF-beta1 exposure of L6 and C2C12 myoblasts, accompanied by impaired differentiation into mature myotubes. TGF-beta1 also induced autophagy in the L6 and C2C12 cells through a PP2A/FoxO1-mediated mechanism. Both the TGF-beta1 effect on differentiation and that on autophagy were blocked by specific PED/PEA-15 ShRNAs. Myoblasts stably overexpressing PED/PEA-15 did not differentiate and showed markedly enhanced autophagy. In these same cells, the autophagy inhibitor 3-methyladenine rescued TGF-beta1 effect on both autophagy and myogenesis, indicating that PED/PEA-15 mediates TGF-beta1 effects in muscle. Muscles from transgenic mice overexpressing PED/PEA-15 featured a significant number of atrophic fibers, accompanied by increased light chain 3 (LC3)II to LC3I ratio and reduced PP2A/FoxO1 phosphorylation. Interestingly, these mice showed significantly impaired locomotor activity compared with their non-transgenic littermates. TGF-beta1 causes transcriptional upregulation of the autophagy-promoting gene PED/PEA-15, which in turn is capable to induce atrophic responses in skeletal muscle in vivo.

Cellular iron metabolism is essentially controlled by the binding of cytosolic iron regulatory proteins (IRP1 or IRP2) to iron-responsive elements (IREs) located on mRNAs coding for proteins involved in iron acquisition, utilization and storage. The 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) is one of the most potent toxins of current interest that occurs as poisonous chemical in the environment. TCDD exposure has been reported to induce a broad spectrum of toxic and biological responses, including significant changes in gene expression for heme and iron metabolism associated with liver injury. Here, we have investigated the molecular effects of TCDD on the iron metabolism providing the first evidence that administration of the toxin TCDD to mammalian cells affects the maintenance of iron homeostasis. We found that exposure of Madin-Darby Bovine Kidney cell to TCDD caused a divergent modulation of IRP1 and IRP2 RNA-binding capacity. Interestingly, we observed a concomitant IRP1 down-regulation and IRP2 up-regulation thus determining a marked enhancement of transferrin receptor 1 (TfR-1) expression and a biphasic response in ferritin content. The changed ferritin content coupled to TfR-1 induction after TCDD exposure impairs the cellular iron homeostasis, ultimately leading to significant changes in the labile iron pool (LIP) extent. Since important iron requirement changes occur during the regulation of cell growth, it is not surprising that the dioxin-dependent iron metabolism dysregulation herein described may be linked to cell-fate decision, supporting the hypothesis of a central connection among exposure to dioxins and the regulation of critical cellular processes. This article is part of a Special Issue entitled: 11th European Symposium on Calcium.

A previous study demonstrated that infection of a canine fibrosarcoma cell line (A-72 cells) by canine coronavirus (CCoV) resulted in apoptosis (Ruggieri et al., 2007). In this study, we investigated the cell death processes during infection and the underlying mechanisms. We found that CCoV-II triggers apoptosis in A-72 cells by activating initiator (caspase-8 and -9) and executioner (caspase-3 and -6) caspases. The proteolytic cleavage of poly(ADP-ribose) polymerases (PARPs) confirmed the activation of executioner caspases. Furthermore, CCoV-II infection resulted in truncated bid (tbid) translocation from the cytosolic to the mitochondrial fraction, the cytochrome c release from mitochondria, and alterations in the pro- and anti-apoptotic proteins of bcl-2 family. Our data indicated that, in this experimental model, both intrinsic and extrinsic pathways are involved. In addition, we demonstrated that the inhibition of apoptosis by caspase inhibitors did not affect CCoV replication, suggesting that apoptosis does not play a role in facilitating viral release.

Sarcopenia, the age-related loss of muscle mass and strength, is a multifactorial condition that represents a major healthcare concern for the elderly population. Although its morphologic features have been extensively studied in humans, animal models, and domestic and wild animals, only a few reports about spontaneous sarcopenia exist in other long-lived animals. In this work, muscle samples from 60 healthy Podolica-breed old cows (aged 15-23 years) were examined and compared with muscle samples from 10 young cows (3-6 years old). Frozen sections were studied through standard histologic and histoenzymatic procedures, as well as by immunohistochemistry, immunofluorescence, and Western blot analysis. The most prominent age-related myopathic features seen in the studied material included angular fiber atrophy (90% of cases), mitochondrial alterations (ragged red fibers, 70%; COX-negative fibers, 60%), presence of vacuolated fibers (75%), lymphocytic (predominantly CD8+) inflammation (40%), and type II selective fiber atrophy (40%). Immunohistochemistry revealed increased expression of major histocompatibility complex I in 36 cases (60%) and sarcoplasmic accumulations of β-amyloid precursor protein-positive material in 18 cases (30%). In aged cows, muscle atrophy was associated with accumulation of myostatin. Western blot analysis indicated increased amount of both proteins-myostatin and β-amyloid precursor protein-in muscles of aged animals compared with controls. These findings confirm the presence of age-related morphologic changes in cows similar to human sarcopenia and underline the possible role of amyloid deposition and subsequent inflammation in muscle senescence.

Swine brucellosis is a zoonotic disease that affects both domestic pigs (Sus scrofa) and wild pigs such as wild boar (Sus scrofa). We evaluated the presence of antibodies against Brucella spp. in wild boar hunted in the Campania region, Italy, during the hunting season of 2016–17. Serum samples were collected from 434 wild boars and tested for antibodies against Brucella spp. For detection of antibodies to swine brucellosis, the rose Bengal test (RBT) and a blocking enzyme-linked immunosorbent assay (ELISA) were performed. Of 434 serum samples, 22 (5.1%) tested by RBT and 58 (13.5%) tested using the blocking ELISA were positive for brucella. The seroprevalences of Brucella spp. did not differ between the sexes or age classes, while the prevalence of infected animals was positively correlated with the Avellino province. Our data showed that wild boar in the Campania region are exposed to brucella infection, and that the prevalence of the disease has increased in recent years.

Aujeszky's disease (AD, pseudorabies) is a viral disease of suids caused by Suid Herpesvirus 1 (SHV-1) also referred as Aujeszky's disease virus (ADV) or Pseudorabies virus (ADV). Domestic pig and Wild boar (Sus scrofa) are the natural host, but many species can be infected with ADV. The aim of our study was to evaluate seroprevalence of AD in wild boar hunted in the Campania Region, during the 2016-2017 hunting season. A total of 503 serum samples from wild boars hunted in the provinces of Campania Region (Southern Italy) were collected and were tested for antibody against ADV using an AD, blocking ELISA assay. A Seroprevalence of 23.85% (120/503, 95% Confidence Interval (CI): 20.15-27.55) was found. Gender was not significantly associated with of ADV seropositivity (p > 0.05), while the presence of ADV antibodies was statistically associated with age (>36-month, p < 0.0001) and location (Avellino, p = 0.0161). Our prevalence values are like those obtained in 2010 in our laboratory (30.7%), demonstrating a constant circulation of ADV in the area.

Following the coronavirus disease 2019 (COVID-19) epidemic peak in Ariano Irpino, Campania region (Italy), we tested cattle for the presence of Severe Acute Respiratory Syndrome Coronavirus-2 (SARS-CoV-2) on a cattle farm at which, prior to the investigation, 13 of the 20 farmworkers showed COVID-19-like symptoms, and one of them died. Twenty-four cows were sampled to detect SARS-CoV-2. All nasal and rectal swabs and milk samples were negative for SARS-CoV-2 RNA. Of the 24 collected serum samples, 11 were positive for SARS-CoV-2 nucleocapsid protein, 14 were positive for SARS-CoV-2 spike protein, and 13 were positive for SARS-COV-2-neutralising antibodies; all samples were negative for Bovine Coronavirus (BCoV), another betacoronavirus. To our knowledge, this is the first report of natural serological evidence of SARS-CoV-2 infection in cattle. We hypothesise that this may be a case of reverse zoonosis. However, the role of cattle in SARS-CoV-2 infection and transmission seems to be negligible.

Abstract Idiopathic inflammatory myopathies (IIMs) are inflammatory disorders of unknown origin. On the basis of clinical, histopathological, and immunological features, they can be differentiated into three major and distinct subsets: dermatomyositis; polymyositis; and inclusion‐body myositis. Although a few animal models for IIM are currently available, they lack several characteristic aspects of IIMs. The aim of our study was to examine skeletal muscle involvement in an experimental animal model of visceral leishmaniasis, a disseminated infection caused by the protozoan parasite Leishmania infantum , and to compare features of associated inflammation with those of human IIM. Syrian hamsters infected intraperitoneally with amastigotes of L. infantum were killed at 3 or 4 months post‐infection, and the skeletal muscles were studied. Focal inflammation was predominantly observed in the endomysium and, to a lesser extent, in perivascular areas. Degenerating muscle fibers were also found, as well as myonecrosis. Immunofluorescence with confocal laser scanning microscopy was used to characterize the phenotype of inflammatory infiltrates and the distribution of MHC class I and II in muscle biopsies. The infiltrating inflammatory cells consisted mainly of T cells, and CD8 + T cells were found in non‐necrotic muscle fibers that expressed MHC class I on the sarcolemma. In addition to T cells, several macrophages were present. The model we are proposing closely resembles polymyositis and may be useful in studying certain aspects of this disease such as the role of T cells in muscle inflammation and myocytotoxicity, while also providing novel therapeutic targets. Muscle Nerve, 2009

There is sufficient evidence that both bovine herpesvirus (BoHV-1) and bubaline herpesvirus (BuHV-1) can overcome the species barrier represented by their respective hosts, cattle and buffalo. Although several studies have focused on the impact of BoHV-1 on buffalo, little is known about the impact of BuHV-1 on cattle. In this work, we evaluated the seroprevalence of BuHV-1 in the cattle population in an area where intensive buffalo farming is highly developed (Campania region, Italy). BuHV-1 seroprevalence of cattle sampled in this study was estimated to be 21.4% using a specific commercial ELISA for the detection of antibodies against glycoprotein E of the virus. Risk factor assessment by univariate analysis revealed a correlation between housing type and higher prevalence. Similarly, cattle housed with buffalo and adult animals had a higher likelihood of being seropositive. BoHV-1 vaccination did not prove to be a protective factor against BuHV-1 exposure. The role of age, grazing, and co-living with buffalo in influencing BuHV-1 exposure was also confirmed by multivariate analysis. All BuHV-1 positive animals were also tested with cross-serum neutralization aimed at evaluating the specific antibody titers against BoHV-1 and BuHV-1. We, therefore, assessed the potential cross-reaction between BoHV-1 and BuHV-1, the co-infection rate, and the agreement of the assays used. This study described the presence of BuHV-1 in the cattle population of the Campania region (Italy) and indicated the requirement to take BuHV-1 into consideration for any measures and control and/or eradication plans to be applied against BoHV-1.

Given the emergence of the coronavirus disease 2019 (COVID-19), zoonoses have raised in the spotlight of the scientific community. Animals have a pivotal role not only for this infection, but also for many other recent emerging and re-emerging viral diseases, where they may represent both intermediate hosts and/or vectors for zoonoses diffusion. Today, roughly two-thirds of human infections are derived from animal origins; therefore, the search for new broad-spectrum antiviral molecules is mandatory to prevent, control and eradicate future epidemic outbreaks. Host defense peptides, derived from skin secretions of amphibians, appear as the right alternative to common antimicrobial drugs. They are cationic peptides with an amphipathic nature widely described as antibacterial agents, but less is reported about their antiviral potential. In the present study, we evaluated the activity of five amphibian peptides, namely RV-23, AR-23, Hylin-a1, Deserticolin-1 and Hylaseptin-P1, against a wide panel of enveloped animal viruses. A strong virucidal effect was observed for RV-23, AR-23 and Hylin-a1 against bovine and caprine herpesviruses, canine distemper virus, bovine viral diarrhea virus, and Schmallenberg virus. Our results identified these three peptides as potential antiviral-led candidates with a putative therapeutic effect against several animal viruses.

The role of the Toll like receptor 4 (TLR4) is of recognising intracellular and extracellular pathogens and of activating the immune response. This process can be compromised by single nucleotide polymorphisms (SNPs) which might affect the activity of several TLRs. The aim of this study is of ascertaining whether SNPs in the TLR4 of Bubalus bubalis infected by Brucella abortus, compromise the protein functionality. For this purpose, a computational analysis was performed. Next, computational predictions were confirmed by performing genotyping analysis. Finally, NMR-based metabolomics analysis was performed to identify potential biomarkers for brucellosis. The results indicate two SNPs (c. 672 A > C and c. 902 G > C) as risk factor for brucellosis in Bubalus bubalis, and three metabolites (lactate, 3-hydroxybutyrate and acetate) as biological markers for predicting the risk of developing the disease. These metabolites, together with TLR4 structural modifications in the MD2 interaction domain, are a clear signature of the immune system alteration during diverse Gram-negative bacterial infections. This suggests the possibility to extend this study to other pathogens, including Mycobacterium tuberculosis. In conclusion, this study combines multidisciplinary approaches to evaluate the biological and structural effects of SNPs on protein function.

Tuberculosis (TB) is one of the deadliest infectious disorders in the world. To effectively TB manage, an essential step is to gain insight into the lineage of Mycobacterium tuberculosis (MTB) and the distribution of drug resistance. Although the Campania region is declared a cluster area for the infection, to contribute to the effort to understand TB evolution and transmission, still poorly known, we have generated a dataset of 159 genomes of MTB strains, from Campania region collected during 2018-2021, obtained from the analysis of whole genome sequence. The results show that the most frequent MTB lineage is the 4 according for 129 strains (81.11%). Regarding drug resistance, 139 strains (87.4%) were classified as multi susceptible, while the remaining 20 (12.58%) showed drug resistance. Among the drug-resistance strains, 8 were isoniazid-resistant MTB, 4 multidrug-resistant MTB, while only one was classified as pre-extensively drug-resistant MTB. This dataset expands the existing available knowledge on drug resistance and evolution of MTB, contributing to further TB-related genomics studies to improve the management of this disease.

In recent decades, the global rise of viral emerging infectious diseases has posed a substantial threat to both human and animal health worldwide. The rapid spread and accumulation of mutations into viruses, and the limited availability of antiviral drugs and vaccines, stress the urgent need for alternative therapeutic strategies. Antimicrobial peptides (AMPs) derived from natural sources present a promising avenue due to their specificity and effectiveness against a broad spectrum of pathogens. The present study focuses on investigating the antiviral potential of oreochromicin-1 (oreoch-1), a fish-derived AMP obtained from Nile tilapia, against a wide panel of animal viruses including canine distemper virus (CDV), Schmallenberg virus (SBV), caprine herpesvirus 1 (CpHV-1), and bovine herpesvirus 1 (BoHV-1). Oreoch-1 exhibited a strong antiviral effect, demonstrating an inhibition of infection at concentrations in the micromolar range. The mechanism of action involves the interference with viral entry into host cells and a direct interaction between oreoch-1 and the viral envelope. In addition, we observed that the peptide could also interact with the cell during the CDV infection. These findings not only highlight the efficacy of oreoch-1 in inhibiting viral infection but also emphasize the potential of fish-derived peptides, specifically oreoch-1, as effective antiviral agents against viral infections affecting animals, whose potential to spill into humans is high. This research contributes valuable insights to the ongoing quest for novel antiviral drugs with the potential to mitigate the impact of infectious diseases on a global scale.

Bovine herpesvirus 1 (BoHV-1) can provoke conjunctivitis, abortions and shipping fever. BoHV-1 infection can also cause immunosuppression and increased susceptibility to secondary bacterial infections, leading to pneumonia and occasionally to death. Herein, we investigated the influence of MG-132, a proteasome inhibitor, on BoHV-1 infection in bovine kidney (MDBK) cells. Infection of MDBK cells with BoHV-1 induces apoptotic cell death that enhances virus release. Whereas, MG-132 inhibited virus-induced apoptosis and stimulated autophagy. Protein expression of viral infected cell protein 0 (bICP0), which is constitutively expressed during infection and is able to stimulate Nuclear factor kappa B (NF-κB), was completely inhibited by MG-132. These results were accompanied by a significant delay in the NF-κB activation. Interestingly, the efficient virus release provoked by BoHV-1-induced apoptosis was significantly reduced by MG-132. Overall, this study suggests that MG-132, through the activation of autophagy, may limit BoHV-1 replication during productive infection, by providing an antiviral defense mechanism.

The mechanism responsible for the anti-seizure effect of ketogenic diets is poorly understood. Because the substantia nigra pars reticulata (SNr) is a “gate” center for seizures, the aim of the present experiment was to evaluate if a ketogenic diet modifies the neuronal response of this nucleus when a seizure-inducing drug is administered in rats. Two groups of rats were given a standard diet (group 1) or a ketogenic diet (group 2) for four weeks, then the threshold for seizure induction and the firing rate of putative GABAergic neurons within the SNr were evaluated with progressive infusion of pentylenetetrazole under general anesthesia. The results demonstrated that the ketogenic diet abolished the correlation between the firing rate response of SNr-neurons and the seizure-threshold. This result suggests that the anti-seizure effect of ketogenic diets can be due to a decrease in reactivity of GABAergic SNr-neurons.

Ulcerative dermatitis (UD) is an idiopathic, spontaneous and progressive disease typically affecting C57BL/6 aged mice with an unknown aetiopathogenesis. For this study, we evaluated 25 cases of UD in C57BL/6NCrl-Tg(HMGA1P6)1Pg mice. Formalin-fixed, paraffin-embedded skin samples were submitted to morphological investigations. Immunohistochemical analysis was performed to characterize and quantify inflammatory cells using CD3, CD45/B220, CD4, CD8 and IL-17 antibodies. Mast cell-bound IgE was investigated by immunofluorescence, whereas serum and cryopreserved skin samples were collected for molecular analysis. Student's t-test (two-tailed) was performed to assess significant differences between the two groups. Affected skin showed extensive areas of ulceration and diffuse, severe and mixed inflammatory infiltrates. No relevant changes were observed in control mice. Immunohistochemical analysis showed a predominant CD3 + CD4 + leukocyte population with fewer CD45/B220 and IL-17 immunolabelled cells and mast cell-bound IgE. Increases in TNFα, IL-1β and Il-6 mRNA expression were observed in the skin of affected animals compared to controls. Serum TNFα and IL-6 did not vary between affected and control mice. Inflammatory infiltrates and cytokine expression were consistent with both Th2/IgE and Th17 differentiation, a typical pattern of a type I hypersensitivity reaction. Overall, our data suggest an allergic-based aetiopathogenesis of UD in C57BL/6NCrl-Tg(HMGA1P6)1Pg mice.

Myocarditis associated with infectious diseases may occur in dogs, including those caused by the protozoa Neospora caninum, Trypanosoma cruzi, Babesia canis, and Hepatozoon canis. However, although cardiac disease due to Leishmania infection has also been documented, the immunopathological features of myocarditis have not been reported so far. The aim of this study was to examine the types of cellular infiltrates and expression of MHC classes I and II in myocardial samples obtained at necropsy from 15 dogs with an established intravitam diagnosis of visceral leishmaniasis. Pathological features of myocardium were characterized by hyaline degeneration of cardiomyocytes, necrosis, and infiltration of mononuclear inflammatory cells consisting of lymphocytes and macrophages, sometimes with perivascular pattern; fibrosis was also present in various degrees. Immunophenotyping of inflammatory cells was performed by immunohistochemistry on cryostat sections obtained from the heart of the infected dogs. The predominant leukocyte population was CD8+ with a fewer number of CD4+ cells. Many cardiomyocytes expressed MHC classes I and II on the sarcolemma. Leishmania amastigote forms were not detected within macrophages or any other cell of the examined samples. Our study provided evidence that myocarditis in canine visceral leishmaniasis might be related to immunological alterations associated with Leishmania infection.

Infectious diseases are a common cause of death in young dogs. Several factors are thought to predispose young dogs to microbiological infections. Identifying the cause of death is often a challenge, and broad diagnostic analysis is often needed. Here, we aimed to determine the infectious causes of death in young dogs aged up to 1 year, examining how it relates to age (under and over 6 months), lifestyle (owned versus ownerless), breed (purebred and crossbreed), and gender. A retrospective study was conducted in a 3-year period (2015-2017) on 138 dead dogs that had undergone necropsy and microbiological diagnostics. Enteritis and pneumonia were the most commonly observed lesions. Polymicrobism was more prevalent (62.3%) than single-agent infections and associated with a higher rate of generalised lesions. Ownerless dogs showed over a three-fold higher predisposition to viral coinfections than owned dogs. Above all, canine parvovirus was the most prevalent agent (77.5%), followed by canine coronavirus (31.1%) and canine adenovirus (23.9%); ownerless pups had a higher predisposition to these viruses. Escherichia coli (23.9%), Clostridium perfringens type A (18.1%), and Enterococcus spp. (8.7%) were the most commonly identified bacteria, which mostly involved in coinfections. A lower prevalence of CDV and Clostridium perfringens type A was observed in puppies under 6 months of age. In conclusion, this study is the first comprehensive survey on a wide panel of microbiological agents related to necropsy lesions. It lays the groundwork for future studies attempting to understand the circulation of infectious agents in a determined area.

Bovine herpesvirus 1 (BoHV-1) is an important cattle pathogen, that may cause rhinotracheitis, abortions and shipping fever. Virus establishes latency in sensory neurons, but periodically could reactivate. Recent studies identified mouse neuroblastoma (Neuro-2A) cells as a novel cell culture model to study factors that regulate BoHV-1 productive infection in neuronal cells. Herein, following BoHV-1 infection in Neuro-2A, a reduced cell viability occurred. Membrane damage and death morphological alterations, features of apoptosis and necrosis, were distinguished in infected cells. In addition, biochemical signs of apoptosis (caspase 3 activation and PARP cleavage) were observed. These results were accompanied by incomplete autophagy due to enhanced amounts of autophagic markers (LC3-II, ATG5 and Beclin 1), in the presence of increased levels of p62. Interestingly, protein expression of viral infected cell protein 0 (bICP0) was detected in Neuro-2A cells, although BoHV-1 inefficiently replicates in these cells, because just low levels of viral yield were found. Taken together, our results suggest that BoHV-1 may exert its potential neurotoxicity through a combined mechanism of necrosis and apoptosis. Moreover, incomplete autophagy occurred during BoHV-1 replication in Neuro-2A cells, which were favourable for viral persistence.

Caprine Herpesvirus type 1 (CpHV-1) is a species-specific herpes virus able to induce apoptosis in several biological systems. In the present study we aimed to investigate the ability of CpHV-1 to reduce cells viability, to replicate and to cause cell death also in human cancer cell lines. We tested the CpHV-1 effects on HEL-299, Vero, MDA-MB-468, HeLa, U2OS, PC3, A549 and K562 neoplastic cell lines and on MDBK cells. Firstly, we evaluated the effect of CpHV-1 infection on cell viability by MTT assay and our data showed that CpHV-1 can induce a marked cytopathic effect (CPE) in most of cell lines tested, except for HEL-299, Vero and K562 cells. The reduction of cell viability was associated with a significant increase of viral production. We next investigated if CpHV-1 was able to induce cell death and so through western blotting analysis we evaluated cleaved caspase 3, LC3II and p62 protein levels after infection. Caspase 3 activation was detected in MDBK cells and, even if at different times p.i., also in MDA-MB-468, U2OS, and PC3 cell lines, while LC3II increase and concomitant p62 protein reduction were observed only in U2OS, and A549 cells, no significant alteration of these proteins was observed in the other cell lines tested. Finally, to confirm virus ability to trigger apoptosis we performed an Annexin-V apoptosis test after 24 h p.i. Although we need to further explore mechanisms underlying CpHV-1 treatment, this study could serve as the basis for the development of new treatment options aiming to fight several cancer types.

Background Google Glass is a head-mounted device designed in the shape of a pair of eyeglasses equipped with a 5.0-megapixel integrated camera and capable of taking pictures with simple voice commands. Objective The objective of our study was to determine whether Google Glass is fit for veterinary forensic pathology purposes. Methods A total of 44 forensic necropsies of 2 different species (22 dogs and 22 cats) were performed by 2 pathologists; each pathologist conducted 11 necropsies of each species and, for each photographic acquisition, the images were taken with a Google Glass device and a Nikon D3200 digital single-lens reflex (DSLR) camera. The pictures were collected, divided into 3 groups (based on the external appearance of the animal, organs, and anatomical details), and evaluated by 5 forensic pathologists using a 5-point score system. The parameters assessed were overall color settings, region of interest, sharpness, and brightness. To evaluate the difference in mean duration between necropsies conduced with Google Glass and DSLR camera and to assess the battery consumption of the devices, an additional number of 16 necropsies were performed by the 2 pathologists. In these cases, Google Glass was used for photographic reports in 8 cases (4 dogs and 4 cats) and a Nikon D3200 reflex camera in the other 8 cases. Statistical evaluations were performed to assess the differences in ratings between the quality of the images taken with both devices. Results The images taken with Google Glass received significantly lower ratings than those acquired with reflex camera for all 4 assessed parameters (P&lt;.001). In particular, for the pictures of Groups A and B taken with Google Glass, the sum of frequency of ratings 5 (very good) and 4 (good) was between 50% and 77% for all 4 assessed parameters. The lowest ratings were observed for the pictures of Group C, with a sum of frequency of ratings 5 and 4 of 21.1% (342/1602) for region of interest, 26% (421/1602) for sharpness, 35.5% (575/1602) for overall color settings, and 61.4% (995/1602) for brightness. Furthermore, we found a significant reduction in the mean execution time for necropsy conduced with the Google Glass with respect to the reflex group (P&lt;.001). However, Google Glass drained the battery very quickly. Conclusions These findings suggest that Google Glass is usable in veterinary forensic pathology. In particular, the image quality of Groups A and B seemed adequate for forensic photographic documentation purposes, although the quality was lower than that with the reflex camera. However, in this step of development, the high frequency of poor ratings observed for the pictures of Group C suggest that the device is not suitable for taking pictures of small anatomical details or close-ups of the injuries.

Abstract Cyclosporin A (CsA) is the prototype of immunosuppressant drugs that have revolutionized the management of all transplantation and autoimmune diseases. Side effects of CsA mainly affecting the kidney but also observed in liver and heart, limit the therapeutic use of this drug after organ transplantation. The renal toxicity of CsA is attributed to reduced renal blood flow which leads to hypoxia‐reoxygenation injury accompanied by excessive generation of oxygen‐derived free radicals. In several therapeutic protocols, CsA is used in association with corticosteroids to obtain better therapeutic results. Recently, our studies showed that hydrocortisone (HY) has a protective effect on CsA‐induced cardiotoxicity. In fact our previous results demonstrated that in rat cardiomyocytes, CsA toxicity is due to a calcium overload, which in turn induce lipid peroxidation and determines oxidative stress‐induced cell injury. Treatment with HY effectively inhibits CsA‐induced toxicity, decreasing lipid peroxidation as well as calcium intracellular concentration. In this study we evaluated in vivo the effects of CsA, used alone or in association with HY, on some parameters of renal dysfunction (blood urea nitrogen; BUN, creatinine, and cholesterol), malondialdheyde (MDA) levels, antioxidant enzyme catalase (CAT), superoxide dismutase (SOD), glutathione peroxidase (GPx), and apoptosis. CsA administration for 24 days resulted in a marked renal oxidative stress, which significantly deranged the renal functions. Treatment with CsA in association with HY significantly improved the renal dysfunction and renal oxidative status. This study clearly suggests the role of oxidative stress in the pathogenesis of CsA‐induced nephrotoxicity. J. Cell. Biochem. 113: 997–1004, 2012. © 2011 Wiley Periodicals, Inc.

Water buffalo (Bubalus bubalis) are susceptible to bovine herpesvirus type 1 (BoHV-1) and a species-specific herpesvirus, bubaline herpesvirus type 1 (BuHV-1). In this study, an attenuated marker BoHV-1 based vaccine against BuHV-1 challenge was evaluated to determine whether it induces protection from viral replication. One group of water buffalo calves was immunized with an attenuated BoHV-1 marker vaccine. A second group was not vaccinated and used as the control. During the post-vaccination period, we monitored the humoral immune response. The efficacy of the vaccine was tested after intranasal challenge of the calves with a BuHV-1 strain. The experiment showed that after vaccination, BuHV-1 replication was significantly reduced by approximately three titer points compared to the controls. The control animals showed high levels of viral shedding and mild signs associated with BuHV-1 infection. Therefore, our study provides evidence for the existence of cross-protection between BoHV-1 and BuHV-1 in buffalo calves.

Background: A retrospective sero-survey for evidence of West Nile virus (WNV) infection in European wild boar (Sus scorfa) was conducted in the Latium region, Italy, on stored serum samples of the period November 2011 to January 2012. Methods: Sera were collected from 168 European wild boars and screened for antibodies to WNV and other Flaviviruses by competitive enzyme linked immunosorbent assay (cELISA). All sera positive for Flavivirus antibodies by cELISA were further examined by virus neutralization test (VNT). To test the presence of Flavivirus RNA in samples, an RT-PCR was performed using a pan-Flavivirus primers pair. Results: Thirteen wild boars (7.73%) were seropositive for Flaviviruses. The hemolysis of serum samples limited the interpretation of the VNT for 7 samples, confirming the presence of specific antibody against WNV in a single European wild boar serum sample. The presence of ELISA positive/VNT negative samples suggests the occurrence of non-neutralizing antibodies against WNV or other antigen-related Flaviviruses. No samples resulted positive for Flavivirus by RT-PCR assay. Conclusion: Although a moderately high percentage of animals with specific antibody for WNV has been detected in wild boar in other surveillance studies in Europe, this has not been reported previously in Italy. Together, these data indicate that European wild boar are exposed to WNV and/or other related-Flavivirus in central Italy and confirm the usefulness of wild ungulates, as suitable Flavivirus sentinels.

Canine coronavirus (CCoV), an alphacoronavirus, may cause self-limiting enteric disease in dogs, especially in puppies. The noteworthy plasticity of coronaviruses (CoVs) occurs through mutation and recombination processes, which sometimes generate new dangerous variants. The ongoing SARS-CoV-2 pandemic and the isolation of a novel canine-feline recombinant alphacoronavirus from humans emphasizes the cross-species transmission ability of CoVs. In this context, exploring antiviral compounds is essential to find new tools for fighting against CoVs infections. Fungi produce secondary metabolites, which are often developed as antibiotics, fungicides, hormones, and plant growth regulators. Previous examinations of benzo-γ-pyrone 3-O-methylfunicone (OMF), obtained from Talaromyces pinophilus, showed that it reduces the infectivity of hepatitis C virus and bovine herpesvirus 1. Based on this evidence, this study evaluated the antiviral ability of OMF against CCoV infection in a canine fibrosarcoma (A72) cell line. During CCoV infection, a non-toxic dose of OMF markedly increased features of cell viability. Moreover, OMF induced a significant reduction in virus yield in the presence of an intense downregulation of the viral nucleocapsid protein (NP). These findings occurred in the presence of a marked reduction in the aryl hydrocarbon receptor (AhR) expression. Taken together, preliminary findings suggest that OMF inhibiting AhR shows promising activity against CCoV infection.

The measles virus (MeV) and canine distemper virus (CDV) belong to the genus Morbillivirus of the Paramyxoviridae family. They are enveloped viruses harboring a non-segmented negative-sense RNA. Morbilliviruses are extremely contagious and transmitted through infectious aerosol droplets. Both MeV and CDV may cause respiratory infections and fatal encephalitis, although a high incidence of brain infections is unique to CDV. Despite the availability of a safe and effective vaccine against these viruses, in recent years we are witnessing a strong resurgence of Morbillivirus infection. Measles still kills more than 100,000 people each year, and CDV causes widespread outbreaks, especially among wild animals, including non-human primates. No drugs are currently approved for MeV and CDV. Therefore, the identification of effective antiviral agents represents an unmet medical need. Here, we have investigated the potential antiviral properties of nitazoxanide (NTZ) against MeV and CDV. Antiviral activity was explored with live virus and cell-based assays. NTZ is a thiazolide that is approved by the FDA as an antiprotozoal agent for the treatment of Giardia intestinalis and Cryptosporidium parvum. Further, nitazoxanide and its metabolite tizoxanide have recently emerged as broad-spectrum antiviral agents. We found that NTZ blocks the MeV and CDV replication, acting at the post-entry level. Moreover, we showed that NTZ affects the function of the viral fusion protein (F), impairing viral spread. Our results indicate that NTZ should be further explored as a therapeutic option in measles and canine distemper virus treatment.

This report describes a case of chronic and deep pyodermitis in a 4-year-old male dog with a 3-month skin problems history that had been treated unsuccessfully with fluoroquinolone therapy, prescribed by a private medical veterinary practice, without an early diagnosis. Microbiological examination and antimicrobial susceptibility testing were performed in our laboratory (Faculty of Veterinary Medicine) and a diagnosis of Streptococcus constellatus-associated pyoderma in the dog was made. A new antimicrobial treatment, with tetracyclines, was designed after the definitive diagnosis and antimicrobial susceptibility testing performed by the Kirby-Bauer disc diffusion method. The dog remained free of clinical illness at completion of therapy. To our knowledge, this is the first case of a canine pyoderma caused by S. constellatus, a commensal organism which may also cause pyogenic infections. Furthermore, this study confirms that a fluoroquinolone represents a poor empirical choice for initial therapy of canine pyoderma.

This report describes the histopathology of two hundred and fifty-three mesenchymal tumors of the urinary bladder in cattle grazing on lands rich in bracken fern. Approximately 80% were hemangiomas and angiosarcomas. Hemangioma (capillary, cavernous, and large vessels) was the most frequent mesenchymal tumor and was more common than angiosarcoma. Although the appearance of endothelial cells can vary remarkably, epithelioid angiosarcomas, often containing multinucleated cells, were the most frequent malignant vascular tumors. Hemangiopericytoma and tumors of muscle and soft connective tissue origin, alone and/or in association with tumor-like lesions, were less frequently seen. Furthermore, forty-five cases of intravascular papillary endothelial hyperplasia (IPEH), a lesion not previously reported in the urinary bladder of cattle, were also described. Bovine papillomavirus type-2 DNA was amplified in tumor samples. Forty vascular tumors were investigated by dual-labeling immunofluorescence, and, for the first time, a coexpression of E5 and platelet-derived growth factor β receptor (PDGF β R) was shown to occur. The results show that the BPV-2 E5 oncoprotein binds to the activated form of the PDGF β receptor thus playing an important role in mesenchymal as well as epithelial carcinogenesis of the urinary bladder. Furthermore, these findings demonstrate that BPV-2 infects both epithelial and mesenchymal cells.

Infection of kidney cells (MDBK) with Bovine Herpesvirus 1 (BoHV-1) is affected by 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), which accelerates BoHV-1-induced apoptosis and increases virus replication. Herein, to elucidate the mechanism through TCDD modifies BoHV-1 infection, we analyzed the modulation of a members of Sirtuin proteins family in MDBK cells. We found that mitochondrial SIRT3 was upregulated during infection. This change was accompanied by cytoskeletal rearrangements and cell extensions. All these trends were drastically modified by TCDD. We hypothesize that, taken together, these results might further clarify the processes responsible for the action of TCDD on the BoHV-1 replication, resulting in enhanced virus production.

In human medicine, "sudden death" has been defined by the World Health Organization (WHO) as a non-violent, unexpected death occurring less than 24 h from the onset of symptoms. The aims of this study were: (1) to estimate the proportional mortality ratio for "sudden and unexpected death" (SUD) in young dogs; (2) to investigate the pathological and microbiological findings in SUD cases in young dogs. For these purposes, a retrospective study of a total of 145 cases of young dead dogs was performed. For each case, we collected information about the age, medical history and the gross and microbiological findings of the animals. The results of this study found 21 cases of SUD. The most frequently observed clinical symptoms in the cases of sudden death were acute respiratory symptoms, followed by acute gastroenteric symptoms, non-specific symptoms and neurological symptoms. The evaluation of necropsy reports allowed us to observe enteritis in 18 out of 21 cases and pneumonia in seven out of 21 cases. Viral infection with Canine parvovirus type 2 was the most common cause of SUD observed. These results could provide a valuable tool for the investigation of sudden death in young dogs.

We investigated the antibiotic resistance of a collection of 94 strains (55.6%) of haemolytic Escherichia coli (E. coli) isolated in 169 diarrhoeic faecal samples from buffalo (Bubalus Bubalis) calves. Bacterial colonies on McConkey and EHLY agar that showed the morphology of E. coli were biochemically tested and then, furtherly classified as haemolytic, using PCR-based assays for enterohemorrhagic E. coli hly (hlyEHEC) virulence gene. When the pathogenic isolates were tested for their susceptibility to 13 different antibiotics, each tested isolate was found to be highly resistant to more than three antibiotics. In fact, absolute resistance (100% of resistance) to penicillin G, lincomycin, neomycin, was detected. Amoxicillin/clavulonic acid and ampicillin were found to be moderately effective against the majority of isolates (46.8% of resistance). Thirty-two (34%) of the haemolytic E. coli strains were phenotypically resistant to tetracycline. None of the isolated strains of E. coli was resistant to colistin sulfate. We conclude that the high prevalence of antimicrobial resistance detected in our study is a source of concern, and cautious use of antibiotics in food producing animals is highly recommended.

A comprehensive pathological analysis of inbred strains is essential to define strain-specific spontaneous lesions and to understand whether a specific phenotype results from experimental intervention or reflects a naturally occurring disease. This study aimed to report and describe a novel condition affecting the skeletal muscles of an inbred C57BL/6NCrl mouse colony characterised by large sarcoplasmic vacuoles in the muscle fibres of male mice in the subsarcolemmal spaces and the intermyofibrillary network. There was no muscle weakness, loss of ambulation or cardiac/respiratory involvement. Post-mortem evaluation and histological analysis excluded the presence of pathological accumulations or lesions in other tissues and organs. Changes were seen in fibre size, with many hypotrophic and some slightly hypertrophic fibres. Histological, immunohistochemical and molecular analyses of the vacuolar content revealed dysregulation of the autophagy machinery while ruling out a morphologically similar condition marked by the accumulation of tubular aggregates.

Lentiviruses, including equine infectious anemia virus (EIAV), are considered viral quasispecies because of their intrinsic genetic, structural and phenotypic variability. Immunoenzymatic tests (ELISA) for EIAV reported in the literature were obtained mainly by using the capsid protein p26, which is derived almost exclusively from a single strain (Wyoming), and do not reflect the great potential epitopic variability of the EIAV quasispecies. In this investigation, the GenBank database was exploited in a systematic approach to design a set of representative protein antigens useful for EIAV serodiagnosis. The main bioinformatic tools used were clustering, molecular modelling, epitope predictions and aggregative/ solubility predictions. This approach led to the design of two antigenic proteins, i.e. a full sequence p26 capsid protein and a doublestrain polypeptide derived from the gp45 transmembrane protein fused to Maltose Binding Protein (MBP) that were expressed by recombinant DNA technology starting from synthetic genes, and analyzed by circular dichroism (CD) spectroscopy. Both proteins were used in an indirect ELISA test that can address some of the high variability of EIAV. The novel addition of the gp45 double-strain antigen contributed to enhance the diagnostic sensitivity and could be also useful for immunoblotting application.

&lt;p&gt;The figure shows the immunostaining analysis of Ki67 in organs from Cdkn1b T197A/T197A mice treated with Vehicle or Bortezomib&lt;/p&gt;

&lt;p&gt;The file contains a detailed description of the generation of the Cdkn1b T197A/T197A mice&lt;/p&gt;

&lt;p&gt;The figure reports the Tunel staining of tissues from Cdkn1b WT/WT and Cdkn1b T197A/T197A mice&lt;/p&gt;

&lt;p&gt;The figure describes the effects of BAY 11-7082 on cell proliferation in the organs from Cdkn1b WT/WT and Cdkn1b T197A/T197A mice&lt;/p&gt;

&lt;p&gt;The figure illustrates the construct used for the generation of Cdkn1b T197A/T197A mice and the subsequent genotype validation.&lt;/p&gt;

&lt;div&gt;Abstract&lt;p&gt;The CDK inhibitor, p27&lt;sup&gt;kip1&lt;/sup&gt;, encoded by the &lt;i&gt;Cdkn1b&lt;/i&gt; gene can negatively modulate cell proliferation. The control of p27 activity during the cell cycle is regulated at multiple levels, including transcription, translation, and protein stability. The last residue of p27 (threonine 198 in human, threonine 197 in mouse) is involved in the control of protein stability. We have generated a murine knock-in model (&lt;i&gt;Cdkn1b&lt;/i&gt;&lt;sup&gt;T197A&lt;/sup&gt;) in which threonine 197 is replaced by alanine, which renders p27 protein highly unstable due to a high rate of proteasomal degradation. Expectedly, &lt;i&gt;Cdkn1b&lt;/i&gt;&lt;sup&gt;T197A/T197A&lt;/sup&gt; mice present with increased body size and weight, organomegaly, and multiple organ hyperplasia, similar to what is observed in &lt;i&gt;Cdkn1b&lt;/i&gt;&lt;sup&gt;KO/KO&lt;/sup&gt; mice. We investigated the effects exerted by the restoration of normal levels of p27 protein in the tissue of &lt;i&gt;Cdkn1b&lt;/i&gt;&lt;sup&gt;T197A/T197A&lt;/sup&gt; mice. We found that proteasome inhibition with bortezomib rescues the hyperplasia induced by the lack of p27 expression in &lt;i&gt;Cdkn1b&lt;/i&gt;&lt;sup&gt;T197A/T197A&lt;/sup&gt; but not in &lt;i&gt;Cdkn1b&lt;/i&gt;&lt;sup&gt;KO/KO&lt;/sup&gt; mice. However, BAY 11-7082, a proteasome inhibitor that stabilizes IκB but not p27, fails to rescue hyperplasia in &lt;i&gt;Cdkn1b&lt;/i&gt;&lt;sup&gt;T197A/T197A&lt;/sup&gt; mice. Bortezomib increases p27 half-life and reduces the proliferation in MEFs derived from &lt;i&gt;Cdkn1b&lt;/i&gt;&lt;sup&gt;T197A/T197A&lt;/sup&gt; but not from &lt;i&gt;Cdkn1b&lt;/i&gt;&lt;sup&gt;WT/WT&lt;/sup&gt; mice, whereas BAY 11-7082 had no effect on the protein levels of p27 and on the proliferation rate of &lt;i&gt;Cdkn1b&lt;/i&gt;&lt;sup&gt;T197A/T197A&lt;/sup&gt; MEFs.&lt;/p&gt;&lt;p&gt;The results presented here demonstrate that &lt;i&gt;Cdkn1b&lt;/i&gt;&lt;sup&gt;T197A/T197A&lt;/sup&gt; mice represent an attractive &lt;i&gt;in vivo&lt;/i&gt; model to investigate whether the targeting of p27 degradation machinery might prove beneficial in the treatment of a variety of human proliferative disorders caused by increased turnover of p27 protein.&lt;/p&gt;&lt;/div&gt;

&lt;p&gt;The figure reports the Tunel staining of tissues from Cdkn1b WT/WT and Cdkn1b T197A/T197A mice&lt;/p&gt;

&lt;div&gt;Abstract&lt;p&gt;The CDK inhibitor, p27&lt;sup&gt;kip1&lt;/sup&gt;, encoded by the &lt;i&gt;Cdkn1b&lt;/i&gt; gene can negatively modulate cell proliferation. The control of p27 activity during the cell cycle is regulated at multiple levels, including transcription, translation, and protein stability. The last residue of p27 (threonine 198 in human, threonine 197 in mouse) is involved in the control of protein stability. We have generated a murine knock-in model (&lt;i&gt;Cdkn1b&lt;/i&gt;&lt;sup&gt;T197A&lt;/sup&gt;) in which threonine 197 is replaced by alanine, which renders p27 protein highly unstable due to a high rate of proteasomal degradation. Expectedly, &lt;i&gt;Cdkn1b&lt;/i&gt;&lt;sup&gt;T197A/T197A&lt;/sup&gt; mice present with increased body size and weight, organomegaly, and multiple organ hyperplasia, similar to what is observed in &lt;i&gt;Cdkn1b&lt;/i&gt;&lt;sup&gt;KO/KO&lt;/sup&gt; mice. We investigated the effects exerted by the restoration of normal levels of p27 protein in the tissue of &lt;i&gt;Cdkn1b&lt;/i&gt;&lt;sup&gt;T197A/T197A&lt;/sup&gt; mice. We found that proteasome inhibition with bortezomib rescues the hyperplasia induced by the lack of p27 expression in &lt;i&gt;Cdkn1b&lt;/i&gt;&lt;sup&gt;T197A/T197A&lt;/sup&gt; but not in &lt;i&gt;Cdkn1b&lt;/i&gt;&lt;sup&gt;KO/KO&lt;/sup&gt; mice. However, BAY 11-7082, a proteasome inhibitor that stabilizes IκB but not p27, fails to rescue hyperplasia in &lt;i&gt;Cdkn1b&lt;/i&gt;&lt;sup&gt;T197A/T197A&lt;/sup&gt; mice. Bortezomib increases p27 half-life and reduces the proliferation in MEFs derived from &lt;i&gt;Cdkn1b&lt;/i&gt;&lt;sup&gt;T197A/T197A&lt;/sup&gt; but not from &lt;i&gt;Cdkn1b&lt;/i&gt;&lt;sup&gt;WT/WT&lt;/sup&gt; mice, whereas BAY 11-7082 had no effect on the protein levels of p27 and on the proliferation rate of &lt;i&gt;Cdkn1b&lt;/i&gt;&lt;sup&gt;T197A/T197A&lt;/sup&gt; MEFs.&lt;/p&gt;&lt;p&gt;The results presented here demonstrate that &lt;i&gt;Cdkn1b&lt;/i&gt;&lt;sup&gt;T197A/T197A&lt;/sup&gt; mice represent an attractive &lt;i&gt;in vivo&lt;/i&gt; model to investigate whether the targeting of p27 degradation machinery might prove beneficial in the treatment of a variety of human proliferative disorders caused by increased turnover of p27 protein.&lt;/p&gt;&lt;/div&gt;

&lt;p&gt;The figure describes the effects of BAY 11-7082 on cell proliferation in the organs from Cdkn1b WT/WT and Cdkn1b T197A/T197A mice&lt;/p&gt;

&lt;p&gt;The figure shows the immunostaining analysis of Ki67 in organs from Cdkn1b T197A/T197A mice treated with Vehicle or Bortezomib&lt;/p&gt;

&lt;p&gt;The figure illustrates the construct used for the generation of Cdkn1b T197A/T197A mice and the subsequent genotype validation.&lt;/p&gt;

&lt;p&gt;The file contains a detailed description of the generation of the Cdkn1b T197A/T197A mice&lt;/p&gt;

Download This Paper Open PDF in Browser Add Paper to My Library Share: Permalink Using these links will ensure access to this page indefinitely Copy URL Copy DOI

The CDK inhibitor, p27kip1, encoded by the Cdkn1b gene can negatively modulate cell proliferation. The control of p27 activity during the cell cycle is regulated at multiple levels, including transcription, translation, and protein stability. The last residue of p27 (threonine 198 in human, threonine 197 in mouse) is involved in the control of protein stability. We have generated a murine knock-in model (Cdkn1bT197A) in which threonine 197 is replaced by alanine, which renders p27 protein highly unstable due to a high rate of proteasomal degradation. Expectedly, Cdkn1bT197A/T197A mice present with increased body size and weight, organomegaly, and multiple organ hyperplasia, similar to what is observed in Cdkn1bKO/KO mice. We investigated the effects exerted by the restoration of normal levels of p27 protein in the tissue of Cdkn1bT197A/T197A mice. We found that proteasome inhibition with bortezomib rescues the hyperplasia induced by the lack of p27 expression in Cdkn1bT197A/T197A but not in Cdkn1bKO/KO mice. However, BAY 11-7082, a proteasome inhibitor that stabilizes IκB but not p27, fails to rescue hyperplasia in Cdkn1bT197A/T197A mice. Bortezomib increases p27 half-life and reduces the proliferation in MEFs derived from Cdkn1bT197A/T197A but not from Cdkn1bWT/WT mice, whereas BAY 11-7082 had no effect on the protein levels of p27 and on the proliferation rate of Cdkn1bT197A/T197A MEFs.The results presented here demonstrate that Cdkn1bT197A/T197A mice represent an attractive in vivo model to investigate whether the targeting of p27 degradation machinery might prove beneficial in the treatment of a variety of human proliferative disorders caused by increased turnover of p27 protein.

An outbreak of neurological disorders in a flock of 20 sheep coming from a rural farm in Civitella Roveto, Italy, occurred in winter 2015. All the animals showed tonic-clonic convulsions followed by muscle paralysis associated with dilated pupils, tremor, tachycardia, tachypnea and diarrhea. The presence of bundles of dry broom of Spartium junceum L. in the feed, eaten by the animals supported the hypothesis of plant intoxication. Two animals died after worsening of clinical signs. The anatomopathological findings and the laboratory results ruled out viral or bacterial infections or accidental exposure to other toxics. Phytochemical study showed the presence of large amount of cytisine, a nicotinic acetylcholine receptor agonist, in all parts of the plant eaten by the animals. Clinical and pathological findings, the complete remission of clinical signs after the exclusion of dry broom from the diet, together with the results of phytochemical analyses results corroborated the hypothesis of S. junceum L. intoxication.

AbPasrtartuabcetrculosis is a chronic infection of domestic and wild ruminants, caused by Mycobacterium avium subsp. paratuberculosis. The persistence of paratuberculosis infection for months up to years without exhibiting any clinical signs makes the diagnosis and control program a difficult proposition. Limited informations on prevalence of paratuberculosis in water buffaloes ( Bubalus bubalis ) are available. We carried out a study on 1350 buffaloes belonging to 56 herds in the Caserta province, of Campania region, Italy. The prevalence of infected buffalo dairy herds was estimated by a commercial ELISA kit of individual blood samples of animals over 24 months of age. On the basis of performance (sensitivity 43%, specificity 99.3%) of ELISA test on serum, the resulting true prevalence at animal level and at herd level was 4% (95% CI 3% to 5%) and 74,1% (95% CI 71.8% to 76%). Considering the paucity of epidemiological reports in the region our results could be a useful contribution towards the prevention of buffalo paratuberculosis in the area.

Twenty-eight urothelial tumours of the urinary bladder were studied in 4-16-year-old cattle grazing on lands rich in bracken fern. Four primary adenocarcinomas, twelve papillary carcinomas, ten invasive carcinomas, two urothelial carcinomas with endophytic growth were examined. They were classified using morphological parameters which have been recently suggested in the report on the new histological classification of urothelial tumours of the urinary bladder of cattle. Furthermore, some incidental benign reactive processes such as cystitis glandularis and intestinal metaplasia, coexisting with urothelial tumours, were also investigated. E5 oncoprotein of bovine papillomavirus type 2 (BPV-2) was detected by immunoprecipitation in twenty-two tumours (79%). The remaining six tumours (21%) were E5-negative. Neuroendocrine differentiation was evident in fifteen tumours, four adenocarcinomas and eleven urothelial tumours; twelve of them, that is 80% (four adenocarcinomas and eight urothelial cancers) were E5-positive. Primary adenocarcinomas were composed of a number of multiple glands embedded in a loose stroma. Glandular lumina were lined by tall columnar cells which showed a marked immunoreactivity to chromogranin A, synaptophysin and serotonin. In eight papillary carcinomas containing varying degrees of a glandular or glandular-like differentiation, immunoreactivity was focally scattered among glandular structures. Two of them were E5-negative. Finally, immunoreactivity was also present in three invasive urothelial carcinomas. They were composed of glandular-like structures, cords and nests of neoplastic cells. Glandular-like structures showed a more marked immunoreactivity. A weak immunoreactivity was also seen in the cells at the edges of the nests. One of them appeared to be E5-negative. This study widens the spectrum of histological neoplastic urothelial lesions of cattle and affords further evidence that microscopic patterns of bovine urinary bladder tumours share striking morphological similarities with the human counterparts.

the subject of the thesis research project, aims to characterize a murine model for spontaneous muscle pathology comparable to human Vacuolar myopathy with Excessive Autophagy (XMEA). Along with Danon disease, these myopathies are characterized by the accumulation of vacuoles within the myofiber. In 1988, Kalimo et al. have described five cases (all males) who had progressive proximal myopathy, which did not involve the heart muscle. The muscle biopsies of all patients showed numerous vacuoles, sarcoplasmic and subsarcolemmal, many of which were positive for the lysosomal enzyme, for the morphological appearance described, was proposed the term: X-Linked Vacuolar myopathy with Excessive Autophagy (XMEA). We used crosses of strain C57/BL6 mice of different ages and sex, we processed triceps muscles, quadriceps femoris and the cranial tibial muscle and made morphological, histochemical and immunohistochemical staining on this samples. From laboratory tests performed on muscle biopsies, we found the presence of numerous vacuoles within muscle fibers, only in males. All these characters are comparable to XMEA. The availability of a well characterized mouse model it may help define the etiopathogenesis of this disease.

Constituting an integral part of a heme's porphyrin ring, iron is essential for supplying cells and tissues with oxygen. Given tight links between oxygen delivery and iron availability, it is not surprising that iron deprivation and oxygen deprivation (hypoxia) have very similar consequences at the molecular level. Under hypoxia, the expression of major iron homeostasis genes including transferrin, transferrin receptor, ceruloplasmin, and heme oxygenase-1 is activated by hypoxia-inducible factors to provide increased iron availability for erythropoiesis in an attempt to enhance oxygen uptake and delivery to hypoxic cells. Iron-response proteins (IRP1 and IRP2) and “cap-n-collar” bZIP transcriptional factors (NE-F2 p45; Nrf1, 2, and 3; Bach1 and 2) also control gene and protein expression of the key iron homeostasis proteins. In this article, we give an overview of the mechanisms by which iron pathways are regulated by hypoxia at multiple levels. In addition, potential clinical benefits of manipulating iron pathways in the hypoxia-related conditions anemia and ischemia are discussed.

<sec> <title>BACKGROUND</title> Google Glass is head-mounted device designed in the shape of a pair of eyeglasses equipped with 5.0 Mpx integrated camera and capable to take pictures with simple voice commands </sec> <sec> <title>OBJECTIVE</title> The aim of the study was to determine if Google Glass is fit for Veterinary forensics pathology purposes. </sec> <sec> <title>METHODS</title> A total of 44 forensic necropsies of 2 different species (22 dogs and 22 cats) were performed by 2 pathologists; each pathologist conducted 11 necropsies of every species and, for each photographic acquisition, the images were taken with Google Glass device and reflex camera Nikon D3200. Furthermore, the pictures were collected, divided in 3 groups (external appearance of the animal, organs and anatomical details) and evaluated by 5 forensic pathologists with a 5-point score system. The parameters assessed were: overall color settings, region of interest, sharpness and brightness. Moreover, to evaluate the difference in mean duration between necropsies conduced with glass and DSLR camera and to assess the battery consumption of the devices, an additional number of 16 necropsies were performed by 2 pathologists. In these cases, Glass was used for photographic report in 8 cases (4 dogs and 4 cats) and a reflex camera Nikon D3200 for the other 8 cases. Statistical evaluations were performed to assess the differences in ratings between the quality of the pictures taken by both devices. </sec> <sec> <title>RESULTS</title> the images taken by Google Glass received significantly lower ratings than those acquired by reflex camera for all 4 parameter assessed (P&lt;.01). In particular, in the pictures of the Groups A and B taken by Glass, the sum of the ratings for 5 (very good) and 4 (good) was comprised between 50 and 77% for all 4 parameter assessed. The lowest ratings were observed in the pictures of group C with a sum of ratings for "5 and 4" of 21.1 % for region of interest, 26 % for sharpness, 35.5 % for overall color settings and 61.4% for brightness. Furthermore, we found a significant reduction in mean necropsy execution time conduced with the Google Glass with respect to the reflex group (P&lt;.01). But, on the other hand, the former drained the battery very quickly. </sec> <sec> <title>CONCLUSIONS</title> These findings suggested that Glass was usable in veterinary forensic pathology. In particular, the images quality of the groups A and B seemed adequate for forensic photographic documentation purpose, although the images quality was lower if compared to a reflex camera. However, in this step of development, the high frequency of poor ratings observed in the pictures of the group C suggested that the device was not suitable for taking pictures of small anatomical details or close-up of the injuries. </sec>