Search

Bovine tuberculosis and paratuberculosis are endemic in many areas worldwide. This work aims to study cytokines production and gene expression profiles of bovine macrophages infected with Mycobacterium bovis and Mycobacterium paratuberculosis subsp. avium (MAP) strains to identify potential diagnostic biomarkers. Bovine bone marrow stem cells were differentiated into macrophages and subsequently infected in vitro with different spoligotypes of M. bovis and MAP field strains (as single infections and coinfections), using different multiplicity of infection. Supernatant and cell pellets were collected 24 h, 48 h, and one week post-infection. Preliminarily, gene expression on cell pellets of IL-1β, IL-2, INFγ, IL-6, IL-10, IL-12, and TNFα was assessed by qRT-PCR one week p.i. Subsequently, IL-1β and IL-6 were measured by ELISA and qRT-PCR to investigated their production retrospectively 24 h and 48 h p.i. A variability in macrophages response related to the concentration of mycobacteria, the coinfection with MAP, and M. bovis spoligotypes was identified. An early and constant IL-6 increase was observed in the M. bovis infection. A lower increase in IL-1β was also detected at the highest concentration of the two M. bovis spoligotypes one week post-infection. IL-6 and IL-1 β production was reduced and differently expressed in the MAP infection. IL-6 appeared to be the earliest cytokines produced by bovine macrophages infected with M. bovis .

Introduction: Paratuberculosis (PTB) is a worldwide chronic, contagious enteric disease caused by Mycobacterium avium subsp. paratuberculosis (MAP) mainly affecting ruminant species. PTB is a WOAH-listed disease with direct and indirect economic losses in the livestock sector, negative impact on animal welfare and significant public health concerns. In spite of this, MAP prevalence in small ruminants is still unknown and the prevalence appears to be underestimated in many countries. The aim of this study is providing a first large-scale serological survey on MAP infection in small ruminants in Sicily, a region of Southern Italy with the 11.3 and 8.9% Italian national heritage of sheep and goats, respectively., Methods: For this purpose, we analyzed a total of 48,643 animals reared in 439 flocks throughout Sicily. MAP seroprevalence was estimated both at herd-level and animal-level within breeds reared in all the nine sampled provinces., Results: Our results revealed a high overall apparent prevalence at herd-level of 71.8% in sheep and 60.8% in goat farms with an animal-level prevalence of 4.5 and 5.1% in sheep and goats, respectively. Significant statistical differences were found between the provinces and within the breeds both in sheep and goats., Discussion: Our study provides the first large-scale serological survey on PTB infection in small ruminants in Sicily and showed a high prevalence of disease depending to the species, breed and province. This study represents the first step to better understand the MAP epidemiology in a typical Mediterranean breeding context, suggesting the need of in-depth study on the herds risk factors, including the eventual presence of candidate genes for resistance/susceptibility to PTB in native breeds., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2024 Di Marco Lo Presti, Ippolito, Migliore, Tolone, Mignacca, Marino, Amato, Calogero, Vitale, Vicari, Ciarello and Fiasconaro.)

Canine distemper is a contagious and severe systemic viral disease that affects domestic and wild carnivores worldwide. In this study, two adult female ferrets ( Mustela putorius furo ) were evaluated for cutaneous lesions. Scab, fur, and swab samples from the external auditory canal, cutaneous lesions, and scrapings were analyzed. Canine distemper virus (CDV)-positive samples underwent RT-PCR/RFLP with the restriction enzyme PsiI, and the hemagglutinin gene sequence was obtained. According to the restriction enzyme and sequence analyses, the viral strains were typed as CDV field strains that are included within the Europe lineage and distinct from those including vaccinal CDV strains. The sequence analysis showed the highest nucleotide identity rates in older Europe lineage CDV strains collected from dogs and a fox in Europe. This study is the first to report on CDV infection in ferrets in southern Italy and contributes to the current knowledge about natural CDV infection in this species. In conclusion, vaccination remains crucial for preventing the disease and counteracting cross-species infection. Molecular biology techniques can enable the monitoring of susceptible wild animals by ensuring the active surveillance of CDV spread.

Faced with the national emergency linked to the spread of COVID-19 in Italy, digital technologies have made it possible to carry out the ordinary activities of the various educational agencies through the main tool of Distance Learning (DaD). The intensive use of information and communication technologies (ICT) and the guarantee of accessibility represent an enabling prerogative for current education systems, enriching training in a variety of ways and opportunities which must be accessible to all. ICTs take on a propulsive function for change in schools because they make it possible to affect the school setting, to transform the learning environment by redesigning space, reorganizing time, modifying communication and socialization processes, encouraging, in students, the development of key competencies in digital literacy and media education. Considering the context of a school transformed and renewed by the teaching and training potential of ICT, it becomes central to reconstruct the requests and needs developed by the practitioners of educational policies to cope with the reorganization of teaching methods and times at the time of DaD. Starting from these premises, the paper focuses attention on the social impact (Stern, 2016) of DaD to evaluate: the extent and intensity of the methodological-didactic innovation required of teachers for the organization and conducting remote lessons; the increase-in students-of transversal and digital literacy skills (team working, problem solving, etc.) potentially associated with the use of ICTs; the involvement and collaboration of families in the process of assessing and verifying learning. The reflection is part of a broader research project by the University of Sapienza University of Rome entitled "The social impact assessment of DaD after COVID-19"; a 3 year evaluation research addressed to a typological sample of upper secondary schools in Rome classified on the basis of the Infrastructure and Equipment indicator of the Rav and the social effect of the school (school effect) on the academic performance of students in the tests Invalsi. The evaluation aimed to identify-from the DaD experience-indications useful in re-designing the school's intervention strategy in the phases following the pandemic; for this reason it adopted an analysis perspective that valorized the positive and most successful aspects in the testimonies of the teachers and students involved in the first phase of the research (conducted in May-June 2021). Within the framework of the Positive Thinking Evaluation, the empirical evidence-collected through the administration/conduction of semi-structured interviews, focus groups, online ethnographic observation of the lessons in DaD-will allow us to reflect on some dimensions of success and of particular social innovation for the teachers' teaching practices and the students' learning processes in DaD. In the Positive Thinking Evaluation, success is a positive effect (not just a "good practice"), even an unexpected one, of an activity that has produced a positive change in the context of program implementation., Competing Interests: The author declares that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest. The reviewer EC declared a past collaboration with the author to the handling editor., (Copyright © 2023 Lo Presti.)

Introduction: The persistence of animal tuberculosis (TB) in livestock is a major concern in Sicily, Italy. The objective of this study was to elucidate the transmission dynamics of M. bovis infection in a highly circumscribed, and at the same time geographically diverse, high-risk area of the island through an in-depth geo-epidemiological investigation of TB in cattle and black pigs raised in small-scale extensive farms across the district of Caronia., Methods: We used genotype analysis coupled with geographic information system (GIS) technology and phylogenetic inference to characterize the spatial distribution of TB and M. bovis genotypes in livestock and the genetic relationships between M. bovis isolates. A total of 589 M. bovis isolates collected from slaughtered cattle ( n  = 527) and Sicilian black pigs ( n  = 62) over a 5-year period (2014-2018) were included in the study., Results: TB was widespread throughout the district and was most frequent in the north-central area of the district, especially along one of the district's streams. We identified a total of 62 M. bovis genotypes. Identical genetic profiles were isolated from both neighboring and non-neighburing herds. The 10 most frequent genotypes, accounting for 82% of M. bovis isolates, showed geographic specificities in that they tended to cluster in specific spatial niches. The landscape structure of these niches-i.e. steep slopes, rocky ridges, meadows and streams-is likely to have had a significant influence on the distribution of TB among livestock in Caronia. Higher concentrations of TB were observed along streams and in open meadows, while rocky ridges and slopes appeared to have hampered the spread of TB., Discussion: The geographical distribution of TB cases among livestock in Caronia is consistent with several epidemiological scenarios (e.g., high density of infected herds along the streams or in hilly plateau where livestock share pastures). Landscape structure is likely to play an important role in the transmission and persistence of M. bovis infection across the district. Additional potential risk factors, such as livestock trading and extensive breeding methods, are also discussed. Our results will contribute to the improvement of surveillance, control and eradication activities of TB in Sicily by the implementation of ad hoc TB control measures, especially in farms located along streams, sharing common pastures or with mixed animal species., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Marianelli, Verrubbi, Pruiti Ciarello, Ippolito, Pacciarini and Di Marco Lo Presti.)

Mycobacterium avium ssp. paratuberculosis (MAP), causes Johne's disease (JD), or paratuberculosis, a chronic enteritis of ruminants, which in goats is characterized by ileal lesions. The work described here is a case-control association study using the Illumina Caprine SNP50 BeadChip to unravel the genes involved in susceptibility of goats to JD. Goats in herds with a high occurrence of Johne's disease were classified as healthy or infected based on the level of serum antibodies against MAP, and 331 animals were selected for the association study. Goats belonged to the Jonica (157) and Siriana breeds (174). Whole-genome association analysis identified one region suggestive of significance associated with an antibody response to MAP on chromosome 7 (p-value = 1.23 × 10 -5 ). These results provide evidence for genetic loci involved in the antibody response to MAP in goats., (© 2022 Stichting International Foundation for Animal Genetics.)

Background: Immunotherapy in high-risk neuroblastoma (HR-NBL) does not live up to its full potential due to inadequate (adaptive) immune engagement caused by the extensive immunomodulatory capacity of HR-NBL. We aimed to tackle one of the most notable immunomodulatory processes in neuroblastoma (NBL), absence of major histocompatibility complex class I (MHC-I) surface expression, a process greatly limiting cytotoxic T cell engagement. We and others have previously shown that MHC-I expression can be induced by cytokine-driven immune modulation. Here, we aimed to identify tolerable pharmacological repurposing strategies to upregulate MHC-I expression and therewith enhance T cell immunogenicity in NBL., Methods: Drug repurposing libraries were screened to identify compounds enhancing MHC-I surface expression in NBL cells using high-throughput flow cytometry analyses optimized for adherent cells. The effect of positive hits was confirmed in a panel of NBL cell lines and patient-derived organoids. Compound-treated NBL cell lines and organoids were cocultured with preferentially expressed antigen of melanoma (PRAME)-reactive tumor-specific T cells and healthy-donor natural killer (NK) cells to determine the in vitro effect on T cell and NK cell cytotoxicity. Additional immunomodulatory effects of histone deacetylase inhibitors (HDACi) were identified by transcriptome and translatome analysis of treated organoids., Results: Drug library screening revealed MHC-I upregulation by inhibitor of apoptosis inhibitor (IAPi)- and HDACi drug classes. The effect of IAPi was limited due to repression of nuclear factor kappa B (NFκB) pathway activity in NBL, while the MHC-I-modulating effect of HDACi was widely translatable to a panel of NBL cell lines and patient-derived organoids. Pretreatment of NBL cells with the HDACi entinostat enhanced the cytotoxic capacity of tumor-specific T cells against NBL in vitro, which coincided with increased expression of additional players regulating T cell cytotoxicity (eg, TAP1/2 and immunoproteasome subunits). Moreover, MICA and MICB, important in NK cell cytotoxicity, were also increased by entinostat exposure. Intriguingly, this increase in immunogenicity was accompanied by a shift toward a more mesenchymal NBL cell lineage., Conclusions: This study indicates the potential of combining (immuno)therapy with HDACi to enhance both T cell-driven and NKcell-driven immune responses in patients with HR-NBL., Competing Interests: Competing interests: None declared., (© Author(s) (or their employer(s)) 2022. Re-use permitted under CC BY-NC. No commercial re-use. See rights and permissions. Published by BMJ.)

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.

Immunotherapies targeting checkpoint inhibition and cell therapies are considered breakthroughs for cancer therapy. However, only a part of patients benefit from these treatments and resistance has been observed. Combining both approaches can potentially further enhance their efficacy. With the advent of gene editing techniques, such as clustered regularly interspaced short palindromic repeats-CRISPR associated protein 9 (CRISPR-Cas9), the elimination of checkpoint molecules became available as an option in good manufacturing practice conditions to improve persistence and efficacy. However, no data of CRISPR-Cas9 application have been reported in cord blood (CB)-derived immune cells, potentially usable for allogeneic cell therapy purposes. In this article, we describe the optimization of a protocol to deplete checkpoint molecules at the genomic level using CRISPR-Cas9 technology from CB-dendritic cells (DCs) and CB-CD8 + T cells. The protocol is based on the electroporation of a ribonucleoprotein complex, easily translatable to clinical settings. In both cell types, the knock-out (KO) was successful and did not affect cell viability. CB-DCs showed a decrease in expression of the targeted protein ranging from 50% to 95%, while CB-CD8 + T cells showed a reduction in the range of 25-45%. The procedure did not affect the stimulatory function of the CB-DCs or the response of CB-CD8 + T cells (proliferation or TNF-α production). In conclusion, we optimized a protocol to eliminate checkpoint molecules from CB-derived DCs and CD8 + T cells, with the aim to further implement allogeneic cell therapies for cancer.

The chemical and microbiological characteristics of filets of Spaurus aurata L. specimens fed with diets containing a Hermetia illucens meal (HIM) at the 25, 35, and 50%, as a partial replacement for fish meal (FM) were evaluated. The diets, formulated to satisfy the nutritional needs of fish, were isoenergetic (22 MJ/kg gross energy), isonitrogenous (43 g/100 g, a.f.), and isolipidic (19 g/100 g, a.f.). Seventy-two specimens were randomly killed after 186 days of growing trials. Then, the filets were analyzed for chemical profile, fatty acids, amino acids, minerals, and microbial flora. Data were subjected to statistical analysis. No significant differences were observed in chemical composition. The sum of polyunsaturated fatty acids (PUFAs) showed a similar content in the filets; eicosapentaenoic acid was similar in the filets of HIM0, HIM35%, and HIM50%, whereas docosahexaenoic acid was higher in filets of the HIM0 group. n3/n6 PUFA ratio and the sum of EPA + DHA showed a high value ( p < 0.001) in filets of the group fed with FM. No significant difference was observed in thrombogenic index and hypocholesterolaemic/hypercholesterolaemic ratio in the groups; the atherogenic index showed a higher value ( p = 0.001) in the HIM50% group. Indispensable amino acids showed some significant ( p < 0.0001) differences in the groups; arginine and phenylalanine content was higher in the filets of fish fed with FM; isoleucine and valine content was higher in the filets of HIM50%; leucine, lysine and methionine content was lower in the filets of HIM35%; histidine content was lower in the filets of HIM25%; tryptophan content was lower in filets of the HIM50% group. EAA/NEAA ratio showed highest value in the filets of the group that received FM. The presence of HIM in the three diets kept chromium, manganese, iron, copper, zinc, and nickel levels lower than those recommended by various authorities. Ca/P ratio showed a higher level ( p < 0.0001) in the group fed with FM than those fed with diets containing HIM. The insect meal in the diets did not influence the microbiological profile of fish. Use of HIM as an unconventional feed ingredient in Sparus aurata diet looks promising, although the quality of filets may be affected., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Oteri, Chiofalo, Maricchiolo, Toscano, Nalbone, Lo Presti and Di Rosa.)

Bovine Tuberculosis (bTB) is a chronic disease caused by Mycobacterium bovis, affecting cattle and other mammalian species, such as pigs. In the present work, we developed a novel multi-antigen assay (The TB-Luminex multiplex test) to diagnose bTB in pig sera. Moreover, we investigated the seroreactivity to the different antigens employed (MPB83, MPB70, CFP10 and ESAT6) and the possible correlation with bTB lesions distribution in the positive pigs. The serum samples were collected from 59 bTB positive pigs and 186 pigs reared in an officially Tuberculosis free area. Sera were processed according to an optimized protocol for the detection of antibodies by a multiantigen assay using Luminex technology. The positive group showed visible lesions with localized (54.2%) or generalized (45.8%) distribution. Culture confirmed the infection in 62.7% of the cases, and histopathology and intra-vitam assays were used as additional confirmatory tests. Within the set of antigens tested, the immunodominant was MPB83 (positive in 94.9% of the affected pigs), followed by CFP10, MPB70 and ESAT6 (positivity shown in 81.3%, 67.8% and 25.4% of the positive pigs tested, respectively). The best antigens combination was MPB83/CFP10, with a 96.6% sensitivity and 96.8% specificity. Overall, the test showed high sensitivity (98.3% and 86.4%) and specificity (96.2% and 97.8%), if sera were considered positive according to the positivity to a single antigen or at least two antigens, respectively. The TB-Luminex multiplex test results did not give significantly different outcomes according to lesions distribution. Given the present study results, the TB-Luminex multiplex test is a reliable test capable of detecting bTB in most infected pigs with good Se and Sp, regardless of the stage of the disease. In conclusion, multi-antigen tests can be used as individual tests and screening tools for domestic and wild suids within bTB eradication programs., (Copyright © 2022 Elsevier B.V. All rights reserved.)

Bovine leptospirosis is an infectious zoonotic disease causing reproductive problems and economic losses in livestock. This work reports, for the first time in Sicily (South Italy), an outbreak of Leptospira interrogans serogroup Pomona that occurred in cattle farms within the Nebrodi Park and was mainly characterized by full-term abortion. Blood and urine samples were collected at different time points from animals of six different farms (Farms A-F) sharing the same grazing area. Research of antibodies against pathogenic Leptospira species in serum samples was carried out via Micro Agglutination Test (MAT). Urine samples were subjected to pathogen isolation and molecular analyses via TaqMan Real Time-PCR. Genotyping of Leptospira species was obtained by Multi-locus sequence typing. MAT detected antibodies against Leptospira interrogans serogroup Pomona in serum samples of all the farms. Pathogenic Leptospira spp. DNA and culture isolation was obtained from urine samples. Genotyping confirmed the excretion of L. interrogans serogroup Pomona. This study describes clinical manifestations, diagnostic implications and epidemiological characteristics of an outbreak in cattle due to L. interrogans Pomona in a protected multi-host area, where domestic and wild animals share the same habitat, suggesting a role of wild species in transmission and persistence of Pomona serogroup among cattle.

Objective: This systematic review and meta-analysis aimed to summarise the available evidence on the pre- and intra-operative risk factors for anastomotic leakage (AL) after bowel resection and anastomosis for ovarian cancer (OC)., Study Design: We searched online databases from Pubmed, Scopus, ScienceDirect, and Cochrane Library from inception to October 2020. Pre- and intra-operative risk factors for AL were considered as the primary outcomes. Research heterogeneity and bias were evaluated by I 2 and by the Newcastle Ottawa scale, respectively. The study was registered with PROSPERO, CRD42018095225., Results: The overall AL rate after OC surgery (median ± SD) was 5.3 ± 12% (277 AL on 5178 anastomoses). Thirteen non-randomised studies were included in the meta-analysis enrolling a total of 3274 patients. Pre albumin level ≤ 3 gr/dl, multiple bowel resections and primary cytoreductive surgery were associated with a significantly high risk of AL with a pooled OR of 5.29 (95% CI: 1.51-18.59), OR = 4.4 (95% CI: 1.19-16.66) and OR = 1.71 (95% CI: 1.05-2.77), respectively. Optimal cytoreduction, ASA score, ascites, and protective stoma were not associated with an increased risk of AL., Conclusion: Based on the best available evidence, preoperative albumin level <3 gr/dl, multiple bowel resections and primary cytoreductive surgery were associated with an increased risk for AL after bowel surgery for OC., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2021. Published by Elsevier B.V.)

Background: Aujeszky's disease is caused by Suid Herpes Virus-1 and species belonging to the genus Sus scrofa are the main reservoir hosts. This virus, however, is capable of infecting and causing severe disease, with an almost constant fatal outcome in other species, both domestic and wild (carnivores, monogastric herbivores and ruminants). Moreover, the possibility of transmission to humans has been demonstrated. This study reports and describes the clinical, diagnostic, pathological and phylogenetic aspects of two cases of Aujeszky's disease in two hunting dogs following the ingestion of infected wild boar raw meat. These cases are contextualized in the province of Messina (Sicily), where a high prevalence of Aujeszky's disease has been recorded (average of 12,20% in the period 2010-2019) in farmed pig, and with evidence of spread to other species. A severe outbreak in cattle has recently been reported in these areas. Nevertheless, cases of Aujeszky's disease in dogs are rarely reported and this study represents the first well-documented report in this species in Sicily., Case Presentation: After a wild boar hunt, two dogs showed neurological symptoms and intense itching unresponsive to therapy. Diagnosis of Aujeszky's disease was made based on clinical suspicion, anamnestic information and confirmed by the isolation of the virus from the brain of both dogs. In addition, molecular typing, sequencing and phylogenetic analysis of the Real-Time PCR products were performed. The sequences studied were placed in the Italian Clade 1 along with the sequences obtained from wild boars and hunting dogs from Italy and France., Conclusions: The finding of this disease in non-natural hosts in Sicilian multi-host epidemiological contexts suggests that the risk of inter-species transmission is concrete and that attention should be paid to developing disease control programs in these territories. The data obtained from genome sequencing of the two SuHV-1 isolates contribute to the enrichment of the GenBank with unknown sequences and the phylogenetic analysis implementation., (© 2022. The Author(s).)

Aujeszky's disease is caused by Suid alphaherpesvirus 1 , and its main reservoir host is the pig. However, other species are also susceptible. Infection with this virus causes a severe neurological clinical picture named Aujeszky's disease, usually accompanied by itching and death a few days after the onset of symptoms. This study reports a multi-species outbreak of Aujeszky's disease that occurred in Sicily, which led to the death of 2 goats, 15 sheep, 2 dogs, 2 cats and 2 foxes. The diagnosis was made by culture, indirect immunofluorescence on brain samples and confirmed by biological test on rabbits. This study reports the first cases of Aujeszky's disease in Italy in cats, goat and sheep. The finding of Aujeszky's disease in several species in Sicily suggests a potential epizootic risk. In such areas where a multi-host system is recognised, an analysis of the risk factors should be carried out in order to develop targeted strategies for the control and eradication of the disease. The critical issues that hinder the control of Aujeszky's disease in the studied territory and perspectives for eradication in the light of EU regulation 429/2016 are also discussed.

The determination of alkaline phosphatase (ALP) in cheeses has become an official method for controlling cheeses with a protected designation of origin (PDO), all of which use raw milk. PDO cheeses, characterized by high craftsmanship, usually have an uneven quality. However, for these cheeses, it is necessary to establish ALP values so that they can be defined as a raw milk product. In this study, a dataset with Pecorino Siciliano PDO samples was analyzed to determine ALP both at the core and under the rind. The results showed that there was no significant difference between the different zones in Pecorino cheese. A second dataset of 100 pecorino cheese samples determined that ALP was only at the core of the cheese. Moreover, there was a statistically significant difference between the ALP values of cheeses produced with raw milk and those produced with pasteurized milk. Furthermore, according to the temperatures, a wide variability of ALP values was observed in the Pecorino Siciliano PDO samples from the core of the cheeses. This was a result of several under scotta whey cooking methodologies adopted by cheesemakers, which do not permit a clear range. Therefore, further investigation is desirable.

Competing Interests: Competing interests: None declared.

With the aim to explore the use of A. hypochondriacus seeds for animal feeding, the agronomic traits, nutrients, and bioactive compounds of four accessions with different origin (India, Nebraska, Iowa, and Pennsylvania) grown in a Mediterranean environment were studied. Proximate composition was determined using the official methods of analyses, fatty acid profile by gas chromatography, total phenolic content (TPC) and the scavenging activity (DPPH • and ABTS •+ ) by colorimetric method. A one-way ANOVA model was performed to determine the differences between accessions. The four A. hypochondriacus accessions showed interesting seed yield results. No significant differences were observed for crude protein and crude fiber; the oil content showed the significant highest values in the seeds from Nebraska and Pennsylvania, but their nutritional characteristics were significantly different. The accession from Nebraska showed the highest oleic and linoleic acid levels, the highest values of polyunsaturated fatty acids, the best atherogenic and thrombogenic indices and hypocholesterolemic/hypercholesterolaemic ratio, and the highest TPC content. The accession from Pennsylvania showed the highest antioxidant activity and lowest peroxidation index. On the whole, A. hypochondriacus seeds can be used as pseudo-cereal to balance the animal diet and the accession should be chosen according to the different metabolic pathways of unsaturated fatty acids in ruminant and monogastric animals.

Adoptive T cell therapy utilizing tumor-specific autologous T cells has shown promising results for cancer treatment. However, the limited numbers of autologous tumor-associated antigen (TAA)-specific T cells and the functional aberrancies, due to disease progression or treatment, remain factors that may significantly limit the success of the therapy. The use of allogeneic T cells, such as umbilical cord blood (CB) derived, overcomes these issues but requires gene modification to induce a robust and specific anti-tumor effect. CB T cells are readily available in CB banks and show low toxicity, high proliferation rates, and increased anti-leukemic effect upon transfer. However, the combination of anti-tumor gene modification and preservation of advantageous immunological traits of CB T cells represent major challenges for the harmonized production of T cell therapy products. In this manuscript, we optimized a protocol for expansion and lentiviral vector (LV) transduction of CB CD8 + T cells, achieving a transduction efficiency up to 83%. Timing of LV treatment, selection of culture media, and the use of different promoters were optimized in the transduction protocol. LentiBOOST was confirmed as a non-toxic transduction enhancer of CB CD8 + T cells, with minor effects on the proliferation capacity and cell viability of the T cells. Positively, the use of LentiBOOST does not affect the functionality of the cells, in the context of tumor cell recognition. Finally, CB CD8 + T cells were more amenable to LV transduction than peripheral blood (PB) CD8+ T cells and maintained a more naive phenotype. In conclusion, we show an efficient method to genetically modify CB CD8 + T cells using LV, which is especially useful for off-the-shelf adoptive cell therapy products for cancer treatment., Competing Interests: N.P.v.T. is an employee of AVROBIO, Inc., Cambridge, MA, USA. J.K. is cofounder and shareholder of Gadeta; is an inventor on multiple patents dealing with γδTCR topics as well as isolation of engineered immune cells; and received research support from Miltenyi Biotec, Novartis, and Gadeta. The other authors declare no competing interests., (© 2021 The Authors.)

The role of pigs in the maintenance of bovine tuberculosis caused by Mycobacterium bovis has been demonstrated in many settings; however, the current control programs usually do not state any intra-vitam diagnostic procedure in this species, as for the cattle. Carcass inspection has shown to be insufficient to detect infection in swine; thus, the assessment of intradermal tuberculin test and interferon-gamma release assay (IGRA) in this species is mandatory. The current study compares the performances of the single intradermal comparative cervical tuberculin (SICCT) test and IGRA. A total of 628 Nebrodi Black pigs raised in free-roaming farms were subjected to the two tests simultaneously. Besides, 124 animals underwent postmortem examination for the detection of tuberculous lesions and isolation of mycobacteria from target organs. The two tests showed a concordance of 94.42% with a Cohen's k coefficient of 0.786 and McNemar chi-square of 4.83 ( P = 0.03). Slightly lower levels of concordance (90.32%) between SICCT and IGRA were obtained in the group of 124 animals, with a Cohen's k = 0.797 and McNemar chi-squared value of 0.69 with a non-significant P = 0.41. Moreover, the results showed how IGRA tends to result positive in higher rates, mostly when non-tuberculous mycobacteria (NTM) were isolated, suggesting a possible impairment of specificity in the event of coinfections in the swine. In conclusion, the results obtained support the possibility of the strategic use of IGRA or SICCT in combination or alternatively one to the other, particularly IGRA which showed lower specificity but has evident advantages over SICCT., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2020 Ippolito, Fiasconaro, Pruiti Ciarello, Aliberti, Vitale, Amato, Pasquali and Di Marco Lo Presti.)

Recent developments in gene engineering technologies have drastically improved the therapeutic treatment options for cancer patients. The use of effective chimeric antigen receptor T (CAR-T) cells and recombinant T cell receptor engineered T (rTCR-T) cells has entered the clinic for treatment of hematological malignancies with promising results. However, further fine-tuning, to improve functionality and safety, is necessary to apply these strategies for the treatment of solid tumors. The immunosuppressive microenvironment, the surrounding stroma, and the tumor heterogeneity often results in poor T cell reactivity, functionality, and a diminished infiltration rates, hampering the efficacy of the treatment. The focus of this review is on recent advances in rTCR-T cell therapy, to improve both functionality and safety, for potential treatment of solid tumors and provides an overview of ongoing clinical trials. Besides selection of the appropriate tumor associated antigen, efficient delivery of an optimized recombinant TCR transgene into the T cells, in combination with gene editing techniques eliminating the endogenous TCR expression and disrupting specific inhibitory pathways could improve adoptively transferred T cells. Armoring the rTCR-T cells with specific cytokines and/or chemokines and their receptors, or targeting the tumor stroma, can increase the infiltration rate of the immune cells within the solid tumors. On the other hand, clinical "off-tumor/on-target" toxicities are still a major potential risk and can lead to severe adverse events. Incorporation of safety switches in rTCR-T cells can guarantee additional safety. Recent clinical trials provide encouraging data and emphasize the relevance of gene therapy and gene editing tools for potential treatment of solid tumors.

Essential oils (EOs) obtained from aromatic plants are rich in natural components with interesting antimicrobial effects. The aim of this study was to evaluate the chemical composition of EOs extracted from Origanum majorana (OM-EO), Mentha suaveolens (MS-EO), Rosmarinus officinalis (RO-EO), Salvia officinalis (SO-EO) and Mentha pulegium (MP-EO). Their antioxidant properties and antibacterial activity against Listeria monocytogenes and different serotypes of Salmonella enterica subsp. enterica were also studied. The EOs were extracted from plants by hydro-distillation and their chemical composition was determined by GC-MS. Terpinen-4-ol, 1,8-Cineole, Camphor, Limonene and Cinerone were the main chemical components found in OM-EO, RO-EO, SO-EO, MP-EO and MS-EO, respectively. To the best of our knowledge, Limonene and Cinerone were reported, for the first time, as the major components of MP-EO and MS-EO. Moreover, our results showed that MS-EO had the best antioxidant activity with an IC 50 of 0.78 ± 0.05 mg/mL, EC 50 of 1.53 ± 0.07 mg/mL, and RC 50 of 0.98 ± 0.04 mg/mL, and the higher antibacterial activity using microdilution broth method with MIC of 0.5% for Salmonella and 0.25% for L. monocytogenes, while OM-EO had the best antibacterial activity using disc diffusion method (inhibition diameters ranged between 15.3 ± 0.3 mm and 18.5 ± 0.3 mm for Salmonella and between 20.1 ± 0.2 mm and 25.4 ± 0.4 mm for L. monocytogenes). However, OM-EO and MS-EO present the higher percentage of sub-lethally injured cells against S. enterica (5.50 ± 0.11%) and L. monocytogenes (5.23 ± 0.07%), respectively. From this study, we can conclude that the investigated EOs are rich in components with interesting antibacterial activity and they could be applied in food preparations as natural preservatives to extend the shelf life of food products and to inhibit the growth of food-borne pathogens., (Copyright © 2020 Elsevier Ltd. All rights reserved.)

Aujeszky's disease in cattle is caused by Suid herpes virus 1. The natural infection has been reported worldwide in bovine species and it is related to direct and indirect contact with infected pigs, which represent the main reservoir of the virus. Here, it is reported the first documented outbreak of Aujeszky's disease in cattle in Sicily (Italy). Severe itching and nonspecific neurological symptoms were the main reported clinical signs. No characteristic gross and histological features were reported other than cutaneous lesions caused by excessive pruritus and hyperaemia, haemorrhages and inflammation in the central nervous system. Diagnosis was confirmed by real time PCR and immunohistochemistry on the nervous tissue. The route of infection remained unknown, but serological data observed in pigs living in close cohabitation with cattle revealed a circulation of a wild strain of the virus in the area. This study contributes to a better knowledge of this disease in a non-conventional host and suggests the need to increase the prophylaxis control plans in specific breeding contexts.

β-Thalassemia intermedia is a disorder characterized by ineffective erythropoiesis (IE), anemia, splenomegaly, and systemic iron overload. Novel approaches are being explored based on the modulation of pathways that reduce iron absorption (ie, using hepcidin activators like Tmprss6-antisense oligonucleotides [ASOs]) or increase erythropoiesis (by erythropoietin [EPO] administration or modulating the ability of transferrin receptor 2 [Tfr2] to control red blood cell [RBC] synthesis). Targeting Tmprss6 messenger RNA by Tmprss6-ASO was proven to be effective in improving IE and splenomegaly by inducing iron restriction. However, we postulated that combinatorial strategies might be superior to single therapies. Here, we combined Tmprss6-ASO with EPO administration or removal of a single Tfr2 allele in the bone marrow of animals affected by β-thalassemia intermedia (Hbbth3/+). EPO administration alone or removal of a single Tfr2 allele increased hemoglobin levels and RBCs. However, EPO or Tfr2 single-allele deletion alone, respectively, exacerbated or did not improve splenomegaly in β-thalassemic mice. To overcome this issue, we postulated that some level of iron restriction (by targeting Tmprss6) would improve splenomegaly while preserving the beneficial effects on RBC production mediated by EPO or Tfr2 deletion. While administration of Tmprss6-ASO alone improved the anemia, the combination of Tmprss6-ASO + EPO or Tmprss6-ASO + Tfr2 single-allele deletion produced significantly higher hemoglobin levels and reduced splenomegaly. In conclusion, our results clearly indicate that these combinatorial approaches are superior to single treatments in ameliorating IE and anemia in β-thalassemia and could provide guidance to translate some of these approaches into viable therapies., (© 2020 by The American Society of Hematology.)

Hematopoietic cell transplantation (HCT) is a last resort, potentially curative treatment option for pediatric patients with refractory acute myeloid leukemia (AML). Cord blood transplantation (CBT) results in less relapses and less graft-versus-host disease when compared to other sources. Nevertheless, still more than half of the children die from relapses. We therefore designed a strategy to prevent relapses by inducing anti-AML immunity after CBT, using a CB-derived dendritic cell (CBDC) vaccine generated from CD34+ CB cells from the same graft. We here describe the optimization and validation of good manufacturing practice (GMP)-grade production of the CBDC vaccine. We show the feasibility of expanding low amounts of CD34+ cells in a closed bag system to sufficient DCs per patient for at least three rounds of vaccinations. The CBDCs showed upregulated costimulatory molecules after maturation and showed enhanced CCR7-dependent migration toward CCL19 in a trans-well migrations assay. CBDCs expressed Wilms' tumor 1 (WT1) protein after electroporation with WT1 -mRNA, but were not as potent as CBDCs loaded with synthetic long peptides (peptivator). The WT1-peptivator loaded CBDCs were able to stimulate T-cells both in a mixed lymphocyte reaction as well as in an antigen-specific (autologous) setting. The autologous stimulated T-cells lysed not only the WT1+ cell line, but most importantly, also primary pediatric AML cells. Altogether, we provide a GMP-protocol of a highly mature CBDC vaccine, loaded with WT1 peptivator and able to stimulate autologous T-cells in an antigen-specific manner. Finally, these T-cells lysed primary pediatric AML demonstrating the competence of the CBDC vaccine strategy., (Copyright © 2020 Plantinga, Lo Presti, de Haar, Dünnebach, Madrigal, Lindemans, Boelens and Nierkens.)

Agronomic traits, oil content, fatty acid composition, antioxidant activity, and total phenolic content were studied on eight A. cruentus accessions cultivated in Southern Italy. A one-way ANOVA model was performed to compare accessions and the Principal Components Analysis was applied to identify patterns in our dataset and highlight similarities and differences. A. cruentus showed valuable seed yield (0.27 kg/m 2 , on average) comparable to the main tradition cereals used for animal feeding. Seed-oil composition showed significant differences among the accessions. Data showed a higher lipid content than most cereal grains (from 5.6 to 7.3%). Approximately 60% of fatty acids were unsaturated; linoleic fatty acid ranged from 19 to 34%, oleic acid from 29 to 36%, and alfa-linolenic fatty acid from 0.3 to 0.5%, respectively. The saturated/unsaturated fatty acid ratio ranged from 0.5 to 0.8, the hypocholesterolemic:hypercholesterolaemic ratio from 1.7 to 2.7, the Atherogenic Index from 0.38 to 0.66, the Thrombogenic Index from 0.85 to 1.48, the total phenolic content from 0.14 to 0.36 mg/g seeds, and the antioxidant activity (DPPH • ) from 0.30 to 0.50. The studied seed-oil composition evidenced A. cruentus as a healthy ingredient for animal feed and consequently, as a possible substitute for traditional cereals. Accessions from Mexico and Arizona emerged for their high qualitative traits.

Dietary partially destoned olive cake supplementation on performance, carcass traits and meat quality of intensively finished bulls was evaluated. Forty-five Limousin bulls, divided into three homogenous groups, received a diet with no supplementation (Control-CTR), 7.5% (Low Olive Cake-LOC), and 15.0% of olive cake supplementation (High Olive Cake-HOC). The trial was realized for 150 days; all bulls were individually weighed at the beginning, middle, and end of the trial, to calculate the individual average daily gain (ADG). At slaughtering, on each carcass, hot weight was recorded and, after 7 days, the pH and temperature were measured. On Longissimus lumborum muscle, color, cooking loss, and shear force of the cooked sample were determined. The chemical composition and the fatty acid content of muscle were determined. Olive cake inclusions (7.5% and 15.0%) increased ( p < 0.05) the body weight, ADG, slaughter traits and intramuscular fat content and influenced ( p < 0.05) the quality indices. The 15.0% of the inclusion reduced ( p < 0.05) the cooking loss and shear force, and increased the unsaturated fatty acid content. The olive cake can be considered as a functional component in beef production and, in substitution to a quote of cereals into the diet of bulls, could be an opportunity to improve agriculture sustainability.

Aim of the present study was to assess the effect of dietary dried partially destoned olive cake supplement on nutritional quality and sensory profile of milk and cheese produced by dairy cows. The experiment was carried out on eighty-four healthy dairy Friesian cows divided into two homogenous groups. The control group (CTR) received a conventional diet, whereas the experimental group (OC) received a conventional diet supplemented with olive cake as 15% of DM. The trial lasted five months. Monthly, on individual milk samples, yield and physical-chemical parameters were determined. Milk was used for the artisanal cheese production. On 10 samples of cheese for each group, physical-chemical and fatty acid profile were determined. Electronic nose analysis and sensory evaluation were performed. Data were analyzed by ANOVA. The diet affected ( p < 0.05) the milk yield, exclusively in September. Yield and quality of cheese of OC group after 60 d of ripening showed higher ( p < 0.05) yield, moisture and fat content, lower ( p < 0.05) pH, protein, salt and ash content, higher ( p < 0.01) MUFA and PUFA and CLA content, lower ( p < 0.05) SFA, higher ( p < 0.01) UFA/SFA and hypocholesterolemic/hypercholesterolemic ratios, better ( p < 0.01) atherogenic and thrombogenic indices. Data show dietary olive cake supplementation in lactating dairy cows improves nutritional and nutraceutical properties of cheese, volatile profile and level of assessors' acceptance.

In poultry production, the intensive use of high-performing hybrid animals led to loss of genetic variability and a consequent lower response to climatic change and disease. Poultry biodiversity is seriously threatened, and its safeguard is a strong objective in developed countries. According to the FAO, which emphasized the importance of native breeds for its country of origin, the aim of this study was to present the first contribution on eggs quality for endangered the Siciliana chicken breed and deepen knowledge on the local Livorno breed. At 20 weeks of age, 108 laying hens (54 Siciliana breed and 54 Livorno breed) were divided into six homogeneous groups of 18 hens each and reared according to requirements imposed by the EC Regulation 889/08 for organic production. The production cycle was controlled over one year, and egg production was recorded daily by group. Eggs were collected, weighted, and measured. Physico-chemical parameter and fatty acids profile were analyzed and nutritional indexes calculated. The statistical model included the effects of breed (Siciliana, Livorno). Egg production was 190 egg/head for Siciliana and 180 for Livorno group. The results showed similar values for Siciliana and Livorno egg quality, highlighting several valuable quality traits from these breeds which might be taken into account for conservation programs.

The main objective of this study was to evaluate the efficacy of thymol in controlling environmental contamination in food processing facilities. The effect of thymol was tested as an agent to prevent planktonic and bacterial biofilm growth of twenty-five Listeria monocytogenes isolates from a variety of foods and five Escherichia coli isolates from a farm. The E. coli isolates were positive for extended spectrum β-lactamase (ESBL) genes. All isolates and reference strains were susceptible to thymol at Minimum inhibitory concentration (MIC) values ranging from 250 to 800 μg/mL. An interesting activity of interference with biofilm formation of L. monocytogenes and E. coli was found for thymol at sub-MIC concentrations of 200, 100, 75, and 50 μg/mL. Anti-biofilm activity ranging from 59.71% to 66.90% against pre-formed 24-h-old L. monocytogenes biofilms at concentrations of 500 or 800 µg/mL, corresponding to 2× MIC, was determined against free-living forms of six isolates chosen as the best or moderate biofilm producers among the tested strains. The property of thymol to attack L. monocytogenes biofilm formation was also observed at a concentration of 100 µg/mL, corresponding to 1/4 MIC, by using a stainless-steel model to simulate the surfaces in food industries. This study gives information on the use of thymol in food processing setting.

The aim of the study was to assess the inclusion of different levels of olive cake in pigs' diet as a strategy to replace conventional ingredients and to improve meat quality traits. Seventy-two Pietrain pigs, during the growing-finishing period (50-120 kg BW), were fed with three dietary treatments that contained or did not contain olive cake: 0% (Ctrl), 5% (Low), and 10% (High). The trial lasted 90 days. Weekly, individual body weight (BW) and feed intake (FI) were recorded to calculate average daily gain (ADG) and feed conversion ratio (FCR). At slaughter, the dressing percentage was calculated and carcass weight and backfat thickness were measured. On a section of Longissimus thoracis muscle (LT), pH, color, chemical, and fatty acid composition were determined. Fatty acid profile was also determined in backfat. The statistical model included the effects of diet (Ctrl, Low, and High). The inclusion of 5% of olive cake in the diet improved significantly ( p < 0.05) BW and FCR. Both levels of inclusion (5% and 10%) significantly reduced ( p < 0.05) backfat thickness and intramuscular fat and modified their fatty acid composition, increasing ( p < 0.05) the concentration of MUFA and PUFA and improving ( p < 0.05) quality indices. Results suggest that olive cake did not negatively affect the productive performances.

The effects of linseed feeding on the stearoyl-CoA desaturase (SCD) activity were evaluated on grazing dairy goats divided into two homogeneous groups (C, control, and L, treated) fed the same amount of concentrate which, for group L was supplemented with linseed. Milk yield was unaffected by the treatment. Group L showed significantly higher milk fat (4.10% vs 2.94%, p < 0.01) than group S. Within milk fatty acids, group C showed significantly higher levels of saturated fatty acids and lower values of mono-unsaturated and polyunsaturated fatty acids. In group L, total CLAs were higher than in group S (0.646% vs 0.311%; p < 0.01) mainly because of the differences in CLA cis9 trans 11 (0.623% vs 0.304%; p < 0.01). In treated animals, SCD activity, measured as cis9 C14:1/C14:0, was lower than in the control group, mainly in July and August.

Mesothelioma is a tumor of the serosal membranes described both in human and veterinary medicine. While in humans the relationship between mesothelioma and exposure to asbestos and some other asbestiform minerals is well known, in animals it is still difficult to establish. In this paper a case of malignant peritoneal mesothelioma probably related to asbestos exposure in a wild boar is described. At post-mortem evaluation the peritoneum, diaphragm and serosal surface of liver and kidneys showed isolated to coalescent multiple nodular lesions. Samples from diaphragm, liver and lung were collected to perform microbiological and histological investigations. To assess the presence of asbestos and/or other asbestiform minerals, SEM-EDS investigations were performed on organs and soil samples collected from the area where the wild boar lived. Microbiological investigations were negative for Mycobacterium species. Gross and histological examination were compatible with a biphasic mesothelioma, with nodules composed of epithelioid and sarcomatoid elements with high pleomorphism. Immunohistochemistry revealed only multifocal scattered positivity for WT-1 and D2-40. Asbestos fibres were detected in all samples (organs and soil) by SEM-EDS, demonstrating a potential relationship between the neoplasia and the exposure to naturally occurring asbestos (NOA). In conclusion, the results of the present study are further confirmation that wild animals, such as the boar, are suitable sentinels to indicate the risk of environmental exposure to asbestos for human populations., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Two different diets characterized by the absence of cereals or by the presence of conventional cereals were evaluated on the nutrient digestibility and faecal characteristics and faecal fermentative end-product concentrations of 8 neutered adult Labrador retrievers housed at the Regional Centre Helen Keller (Messina, Italy) during the training work for the service guide for the blind. Dogs (age = 17 ± 1 months, initial body weight [BW] = 26.3 ± 1 kg, and body condition score [BCS] = 4.5 ± 0.11) were divided into 2 homogeneous groups for sex (half males and half females). Dogs in the grain free (GF) group were fed a commercial diet characterized by the absence of grain cereals, and dogs in the control (CTR) group were fed a super-premium pet food characterized by conventional grains as the carbohydrate source. The trial lasted 84 d, preceded by a 7-d of adaption period. Physical examination, digestibility, and faecal characteristics were studied. The statistical model included the effects of diet (GF vs. CTR), time (from d 0 to 84, end of the trial) and the interaction (diet × time). The high-protein, low-carbohydrate dry diet (GF) offered higher apparent nutrient digestibility of protein (+10%; P  = 0.002) and fat (+7%; P  < 0.001) and more stable large intestinal fermentation of carbohydrate compared to the commercial high-carbohydrate dry diet, enabling dogs to use nutrients from the diet more efficiently and thus requiring less food (-13%) to satisfy their nutrient requirements, producing less excrement (-33%; P  = 0.033), and reaching a higher final BW (+8%; P  < 0.0001) and a higher final BCS (+15%; P  = 0.003). Therefore, the GF diet appears the nutritional plan most suitable for these animals taking due account not only of the training work done by animals with their increased nutrient and energy needs, but also of the gastrointestinal disorders consequent to stress coming from work and life in kennels, which cause in the Labrador retrievers an unusual weight loss.

Despite some researchers reporting clinical signs in cattle associated with Trypanosoma theileri, its role as a pathogen is still unclear. We describe here the isolation of Trypanosoma theileri during a routine laboratory investigation. Mature and immature vital parasitic forms were observed within hematopoietic cell cultures from the bone marrow of one cow for monocyte isolation. The animal was submitted to clinical examination and blood sample counting (CBC). Postmortem analysis included gross and histological examination and PCR in the liver, spleen, brain, lymph nodes, and lungs. PCR and Giemsa staining were used for parasite identification. A second cow belonging to the same farm was positive for Trypanosoma theileri by PCR performed on blood sample. In this case, the postmortem analysis included also testis. Clinical examination showed only a reduction in body weight in both cases. The CBC revealed an increase of lymphocytes and neutrophils while red blood cells were within the normal range. Spleen was slightly increased in volume and the histology revealed a proliferative activity of the white and red pulp. The biomolecular analysis identified the parasite as Trypanosoma theileri and its DNA was detected in the bone marrow, testis, and brain. The unusual finding of parasite in the brain, testis, and bone marrow raises new clinical implication on disease course and also possible sexual transmission.

Bovine tuberculosis (bTB) caused by Mycobacterium bovis is an important re-emerging disease affecting livestock, wildlife and humans. Epidemiological studies are crucial to identifying the source of bTB infection, and its transmission dynamics and host preference, and thus to the implementation of effective strategies to contain it. In this study, we typed M. bovis isolates from livestock, and investigated their genetic diversity and distribution. A total of 204 M. bovis isolates were collected from cattle (n = 164) and Sicilian black pigs (n = 40) reared in a limited area of the province of Messina, northeastern Sicily, an area that had previously been identified as having the highest incidence of bTB in livestock on the island. All M. bovis isolates were typed by both spoligotyping and 12-loci MIRU-VNTR analysis. Results from both methods were then combined in order to improve the discriminatory power of M. bovis typing. We identified 73 combined genetic profiles. Thirty-five point six percent of the profiles were common to at least two animals, whereas 64.4% of profiles occurred in only one animal. A number of genetic profiles were predominant in either cattle or black pigs. We identified common genetic patterns in M. bovis isolates originating not only from neighboring districts, but also from non-neighboring districts. Our findings suggest that bTB is widespread in our setting, and is caused by a large number of genetically diverse M. bovis strains. The ecology and farming practices characteristic of the area may explain the substantial M. bovis heterogeneity observed, and could represent obstacles to bTB eradication., Competing Interests: The authors have declared that no competing interests exist.

We report a case of a 32-year old woman with severe pelvic abscess formation who was readmitted to the hospital after ten days of postoperative caesarean course. The patient had undergone emergency caesarean section (CS) for acute foetal sufferance during expulsive period and following three days had been discharged regularly. Enterococcus Faecalis was isolated from purulent material. A prompt antibiotic therapy was carried out, therefore clinic condition did not improve. A percutaneous Computed Tomography (CT)-guided drainage of the abscess was planned. Under local anesthesia abscess was drained. The hospitalization stay allowed monitoring the patient's clinical condition and laboratory blood panel until normalization of all parameters.

We present a very rare case of a 49-year old woman suffering from Nuck canal cyst reaching and compressing femoral vein. Nuck canal cyst is very uncommon event because the pouch accompanying the gubernaculum during intrauterine descent of ovaries usually obliterates, whereas when it persists a cystic cavity containing citrine fluid develops. A gravid 0 para 0 49 old woman was admitted to Catania University Surgery Department owing to suspected lymphatic tumor compressing right femoral vein and causing groin pain with ipsilateral leg partial stasis. Patient believed right venous stasis was due to fibromatous uterus. Ultrasounds and computed tomography (CT) scan defined size (7.1 × 4.2 × 1.5 cm), structure (cystic) of mass and its relation with femoral vein, although they were not diriment for diagnosing its nature. Color Doppler detected circulatory function of compressed femoral vein. Surgery was challenging and Nuck cyst was removed after accurate separation from the right femoral venous walls. A case of Nuck cyst involving femoral vein has never been reported so far.

Background: Polymorphisms of the prion protein gene may influence scrapie susceptibility in small ruminants through modified protein conformation. At least 47 amino acid substitutions and 19 silent polymorphisms have been described in goat PRNP reported from several countries. The objective of this study was to investigate PRNP polymorphisms of native Ethiopian goat breeds and compare the results with other goat breeds., Results: The analysis of the prion protein gene PRNP in 229 goats belonging to three of the main Ethiopian native goat breeds showed a remarkably high frequency (> 34.6%) of p.(Asn146Ser) in these breeds, a variant involved in scrapie resistance in Cyprus. In addition, two novel amino-acid substitutions p.(Gly127Ala) and p.(Thr193Ile), with frequencies ranging from 1.5 to 7.3% were detected. Both amino acids are well conserved in prion proteins (PrP) of most species and these changes have never been reported before in goats worldwide. Residue 127 is within the N-terminal domain of PrP and is probably involved in the recruitment of neural cell adhesion molecules (NCAM). Residue 193 is within the highly conserved string of 4 threonines that plays a role in determining the efficiency of prion protein conversion towards its pathological form., Conclusion: Two novel coding polymorphisms and a high frequency of a scrapie protective variant indicate a high level of genetic diversity in PRNP of Ethiopian goats. This finding increases the interest in exploring PRNP polymorphisms of native goat breeds in areas where cross breeding with foreign goats has rarely occurred.

Pulmonary arteriovenous malformations (PAVMs) are anatomical abnormalities consisting in a direct connection between pulmonary arteries and veins. Most of PAVMs are related to Hereditary Hemorrhagic Teleangiectasia, whereas only 10 to 20% are isolated sporadic cases. PAVMs tend to increase in size naturally; however, several factors can influence their growth such as pulmonary arterial hypertension, puberty, and pregnancy. Clinical manifestations are related to the right-to-left shunting and include dyspnoea, hypoxia, and pulmonary hypertension. The presence of PAVMs during pregnancy is associated with an increased risk of complications such as their rupture, haemothorax, and hypovolemic shock. The treatment reserved to PAVMs was the surgical resection of the lung lobe involving the malformation. Due to the worldwide acceptance of endovascular technique, the transcatheter embolization (TCE) is today considered as the mainstay of treatment. Recent studies reported the safeness of the TCE during pregnancy if performed by an experienced radiologist, at second or third trimesters when radiation exposure is believed to have minimal effect on the foetus. However, although the TCE during pregnancy represents an option, the treatment prior to pregnancy has to be considered the auspicial solution. Our study reports the case of a dyspnoeic pregnant woman with unknown pAVM causing hemothorax and simultaneously treated for pAVM reparation, left lower lobe resection, and hysterectomy. Postoperative treatment of embolization was performed to definitively close the pAVM.

During a routine abattoir inspection of pig carcasses and control activities of hunted wildlife in 2013, 118 large white pigs Sus scrofa domesticus, 474 Nebrodi black pigs and 135 wild boars Sus scrofa scrofa were submitted to anatomopathological examination to evaluate the presence of tuberculosis-like lesions. Localized and generalized granulomatous lesions were detected with a prevalence of about 25% in large white pigs, 13% in Nebrodi black pigs and 8.15% in wild boars. Localized lesions involved mainly the submandibular lymph nodes, but when the disease was spread throughout the body, the inner organs and, also, in some cases, udders and/or bones were injured. The highest prevalence of generalized lesions (15/30) was observed in large white pigs, the only ones in which tuberculous granuloma affected also the spine. The bovine tuberculous spondylitis cases observed showed some similarities with Pott disease in humans regarding aspect and localization of lesions and age of the affected animals.Tissue samples of the positive animals were collected and submitted to bacteriological analysis, and the bone samples were also subjected to histological and immunohistochemistry analysis. M. bovis was isolated in all the analysed samples, and the granuloma encapsulation was found often incomplete indicating that the disease was in an active phase. The presence of lesions associated to tuberculous spondylitis in pigs suggests the possibility to use this animal species as model for the study of Pott disease in humans.

Introduction: Hematopoietic cell transplantation is a potentially lifesaving procedure for patients with hematological malignancies who are refractory to conventional chemotherapy and/or irradiation treatment. Umbilical cord blood (CB) transplantation, as a hematopoietic stem and progenitor cell (HSPC) source, has several advantages over bone marrow transplantation with respect to matching and prompt availability for transplantation. Additionally, CB has some inherent features, such as rapid expansion of T cells, lower prevalence of graft-versus-host disease and higher graft versus tumor efficacy that make this HSPC cell source more favorable over other HSPC sources. Areas covered: This review summarizes the current CB and CB derived T cell applications aiming to better disease control for hematological malignancies and discusses future directions to more effective therapies. Expert commentary: CB transplantation could be used as a platform to extract cord blood derived T cells for ex vivo expansion and/or gene modification to improve cellular immunotherapies. In addition, combining cord blood gene-engineered T cell products with vaccination strategies, such as cord blood derived dendritic cell based vaccines, may provide synergistic immunotherapies with enhanced anti-tumor effects.

Staphylococcus aureus is the major cause of foodborne diseases worldwide. In this retrospective study, 84 S. aureus strains were characterized. The collection comprises 78 strains isolated during 1998 and 2014 from dairy products and tissue samples from livestock bred for dairy production in Sicily. One isolate was obtained from a pet (dog), one from an exotic animal (a circus elephant), and four human isolates were obtained during a severe food poisoning outbreak that occurred in Sicily in 2015. All the strains were characterized by pulsed-field gel electrophoresis (PFGE), for antibiotic resistance and presence of toxin genes. PFGE results showed 10 different pulsotypes, with three relatively frequent and three unique. The antibiotic resistance profiling showed that penicillin G (35.7%) and tetracycline (20.2%) resistance is largely spread. Most isolates contained at least one toxin gene making them a potential threat for public health. Enterotoxin sec gene was observed in 28.6% and seg in 23.8% of the strains, respectively; the human isolates were the only ones to concurrently harbor both seg and sei genes. In addition, 24 isolates were randomly selected and analyzed by multilocus sequence typing. Interestingly, the analysis showed the presence of 12 sequence types (STs), of which 6 were novel. One of them, ST700, was detected in 29% of the isolates and was found to be spread throughout Sicily. ST700 has been present in the island for almost 16 years (1998-2014) and it shows no host preference since it was isolated from different ruminant species. Four human isolates shared both the pulsotype (PT10) and the sequence type (ST9), as well as the virulence genes (seg-sei); this observation suggests that the isolates originated from a single clone, although they were obtained from two different individuals.

The interferon-gamma (IFN-γ) test measures cell mediated immune response (CMI) during the early stages of tuberculosis infection. Although Bovine Tuberculosis (BT) spread in feral pigs is widely documented in literature, the effectiveness of IFN-γ in this species has been only recently reported. One of the major obstacle of this assay is that whole blood samples should be stimulated with purified protein derivative (PPD) cocktail within 8 h from the blood sampling. This study set up a defined broth culture in which lymphocytes, the cell population predominantly responsible for IFN-γ production, are maintained in a steady-state and their vitality is preserved. The IFN-γ production measured from the samples added with the maintenance medium and stored at 4 °C was similar to the enzyme-linked immunosorbent assay (ELISA) optical density values obtained from the same assay performed within 8 h from sampling., (Copyright © 2017. Published by Elsevier Ltd.)

Purpose: Alternatives to antibiotic therapy for mastitis in ruminants are needed. We present an evaluation, in two trials, of the efficacy of an intramammary infusion of a live culture of Lactococcus lactis for the treatment of subclinical and clinical mastitis in ewes., Methodology: In total, 67 animals were enrolled: 19 lactating ewes (study 1), including healthy (N=6) and coagulase-negative staphylococci (CNS)-infected ewes (N=13); and 48 lactating ewes (study 2) with either CNS mastitis (N=32), or Staphylococcus aureus mastitis (N=16), for a total of 123 mammary glands. Intramammary infusions were performed with either L. lactis or PBS for 3 (study 1) or 7 (study 2) consecutive days. Antibiotic-treated and untreated control glands were included. Milk samples for microbiology, somatic cell analysis and milk production were collected before and after treatment.Results/Key findings.L. lactis rapidly activated the mammary glands' innate immune response and initiated an inflammatory response as evidenced by the recruitment of polymorphonuclear neutrophils and increased somatic cell counts. But while leading to a transient clearance of CNS in the gland, this response caused mild to moderate clinical cases of mastitis characterized by abnormal milk secretions and udder inflammation. Moreover, S. aureus infections did not improve, and CNS infections tended to relapse., Conclusion: Under our experimental conditions, the L. lactis treatment led to a transient clearance of the pathogen in the gland, but also caused mild to moderate clinical cases of mastitis. We believe it is still early to implement bacterial formulations as alternatives in treating mastitis in ruminants and further experimentation is needed.

Background: Typing of Toxoplasma gondii strains is important in epidemiological surveys, to understand the distribution and virulence of different clones of the parasite among human and animal populations. Stray dogs can be consider sentinel animals for contaminated environments playing an important but probably under- evaluated role in the epidemiology of T. gondii. We reported a rare case of acute toxoplasmosis in a stray dog due to clonal type I infection. The clonal type I, sporadic in Europe, is frequently associated with severe toxoplasmosis in humans and the control of its circulation is particularly relevant for public health. The symptomatology suggested a potential infection with the high similar parasite Neospora caninum but differential diagnosis showed that only T. gondii was involved highlighting the importance of multiple diagnostic methods beyond the clinical signs., Case Presentation: A female stray dog approximately six-month of age presented muscular atrophy of the femoral region and hyperextension of hind limbs. Body condition score (BCS) was 20% below ideal weight, ribs had almost no fat and the sensor state was depressed. Haematological values were normal and the dog did not show any neurological abnormalities. Serological analysis showed a positive response for T. gondii immunoglobulin G (IgG) antibodies and exclude N. caninum infection. To confirm T. gondii infection, a muscle biopsy was performed and genomic DNA was extracted. PCR analysis resulted positive to T. gondii and strain genotyping reveals clonal type I infection. The dog recovered after 4 weeks of treatment with clindamycin hydrochloride and aquatic physiotherapy., Conclusions: Our study reports a rare and severe case of T. gondii clonal type I infection in a stray dog feeding in garbage containers. The data confirm the importance of an in vivo early diagnosis for toxoplasmosis in dog. Clinical signs are often related to specific T. gondii genotype and parasite genotyping is important in the epidemiological survey of toxoplasmosis in public health. The detection of parasitic DNA in the tissue could be an useful diagnostic method in facilitating early treatment of the disease, which is important for a timely clinical recovery.

Purpose. Highly discriminatory genotyping strategies are essential in molecular epidemiological studies of tuberculosis. In this study we evaluated, for the first time, the efficacy of the repetitive sequence-based PCR (rep-PCR) DiversiLab Mycobacterium typing kit over spoligotyping, 12-locus mycobacterial interspersed repetitive unit-variable number tandem repeat (MIRU-VNTR) typing and embB single nucleotide polymorphism (SNP) analysis for Mycobacterium bovis typing. Methodology. A total of 49 M. bovis animal isolates were used. DNA was extracted and genomic DNA was amplified using the DiversiLab Mycobacterium typing kit. The amplified fragments were separated and detected using a microfluidics chip with Agilent 2100. The resulting rep-PCR-based DNA fingerprints were uploaded to and analysed using web-based DiversiLab software through Pearson's correlation coefficient. Results. Rep-PCR DiversiLab grouped M. bovis isolates into ten different clusters. Most isolates sharing identical spoligotype, MIRU-VNTR profile or embB gene polymorphism were grouped into different rep-PCR clusters. Rep-PCR DiversiLab displayed greater discriminatory power than spoligotyping and embB SNP analysis but a lower resolution power than the 12-locus MIRU-VNTR analysis. MIRU-VNTR confirmed that it is superior to the other PCR-based methods tested here. Conclusion. In combination with spoligotyping and 12-locus MIRU-VNTR analysis, rep-PCR improved the discriminatory power for M. bovis typing.

Bovine tuberculosis, a reemerging zoonosis in diverse ecological scenarios, has been reported in the autochthonous Nebrodi black pig breed population used for meat production in Italy. During a routine abattoir inspection in 2013, 24 of 299 carcasses (8%) of Nebrodi black pigs presented tuberculosis-like lesions at pathologic examination. Mycobacterium bovis was isolated from 23 animals and M. caprae from a 3-year-old sow. The sow showed severe diffuse lesions involving the visceral organs, right coxofemoral joint, and mammary glands. Isolation of M. caprae from mammary glands is uncommon, with only one other case involving a sow reported so far; however, Mycobacteria infection of the mammary glands may be transmitted from lactating sows to piglets, contributing to the spread and maintenance of bovine tuberculosis in swine. Genotyping analysis showed M. caprae spoligotype SB0866 and profile 4,1,5,4,4,11,4,2,4,3,8,7 MIRU-VNTR (mycobacterial interspersed repetitive units-variable number of tandem repeats). The worldwide prevalence of this spoligotype is very low. The finding of severe, diffuse tuberculous lesions strongly suggests that Nebrodi black pigs are susceptible for Mycobacterium spp. and that they might act as a distributor for these microorganisms. Since natural ecosystems with multiple contacts among different livestock species and wild animals are very common in Mediterranean regions, current surveillance and eradication plans for bovine tuberculosis will need to be extended to other potential reservoir species in regions where extensive and traditional breeding systems are operated., (Copyright © 2017 The Authors. Published by Elsevier B.V. All rights reserved.)