Detection of anti-HEV antibodies but no molecular positivity in dogs in the Campania region, southern Italy.

Paslahepevirus balayani genotypes 3 and 4 (HEV-3 and 4) have zoonotic potential and can be transmitted to humans and animals through the consumption of contaminated raw or undercooked meat. Although it has been demonstrated that dogs are susceptible to the infection and produce specific antibodies, the epidemiological role of this species is not yet well defined. This study aimed to evaluate the circulation of HEV at the serological and molecular level in the dog population of the Campania region, southern Italy. A total of 231 dogs were sampled, divided according to several variables (sex, age, origin, lifestyle, location, size, and breed), and tested for the presence of HEV antibodies using a commercial multi-species ELISA. A total of 197 blood samples and 170 stool samples were tested with two specific PCRs in order to detect viral RNA. A total of 19 out samples of 231 were seropositive, obtaining an exposure (8.2%) similar to that observed in other European countries. The univariate and multivariate analysis revealed a wide exposure to stray dogs and animals from the province of Salerno. All samples tested with molecular methods were negative. Defining the role of domestic carnivores continues to be a "one health" challenge, although it appears that they do not eliminate the virus and therefore do not pose a danger to humans. In the absence of other evidence, it is advisable to continue to carry out surveillance also for domestic animals, which, due to ethological characteristics or their position in the food chain, could be predisposed to being exposed to HEV.

Short communication: Seroprevalence of paratuberculosis in Italian water buffaloes (Bubalus bubalis) in the region of Campania.

Paratuberculosis is a chronic enteric disease affecting virtually all ruminants, but only anecdotal information is currently available about the occurrence of this disease in water buffaloes (Bubalus bubalis). We carried out a survey study aimed at determining the prevalence of paratuberculosis in 2 provinces in the region of Campania, Italy, where about half of all Italian buffaloes are reared. From May 2017 to December 2018, we collected 201,175 individual serum samples from 995 buffalo herds. The sera were collected from animals over 24 mo old and were tested using a commercial ELISA test. The herd-level apparent prevalence result was 54.7%, and the animal-level apparent prevalence was 1.8%. The herd-level true prevalence was estimated using a Bayesian approach, demonstrating a high herd-level prevalence of paratuberculosis in water buffaloes from the Campania area. These findings suggest that the urgent adoption of paratuberculosis herd-control programs for water buffaloes in this area would be beneficial.

Penile cancer: prognostic and predictive factors in clinical decision-making.

Penile cancer (PC) is a typical tumor of non-industrialized countries. The incidence is 20-30 times higher in Africa and South America, considering the elevated prevalence of sexually transmitted diseases. Histologically, PC includes squamous cell carcinoma (SCPC), the most frequent, and nonsquamous carcinoma (NSCPC). Early diagnosis is the goal, whereas later diagnosis relates to poor functional outcomes and worse prognosis. The 5-year survival rate is 85% for patients with histologically regional negative lymph nodes, compared to 29%-40% for those with histologically regional positive lymph nodes. To date no new drugs are approved, and there are few new data about molecular mechanisms underlying tumorigenesis. The SCPC remains a rare tumor and the current therapeutic algorithm is based principally on retrospective analysis and less on prospective trials. In this review article, biomarkers of prognosis and efficacy of current treatments are summarized with a focus on those that have the potential to affect treatment decision-making in SCPC.

Advanced/metastatic bladder cancer: current status and future directions.

In 2015 bladder cancer was the fourth most frequent malignancy and the eighth cause of death for cancer. At diagnosis, about 30% of bladder cancer (BC) patients present a muscle-invasive bladder cancer (MIBC) and 5% a metastatic bladder carcinoma (MBC). For fit MBC patients, combination chemotherapy (CC) is the standard of care for first-line treatment. CC includes both the treatment with methotrexate, vinblastine, doxorubicin, and cisplatin (MVAC) either the classical or the dose-dense MVAC regimen, and the doublet therapy with cisplatin and gemcitabine (CG). Median progression free survival (PFS) was 7 months and median overall survival (OS) was 15 months. The present review provides an update on the management of MBC, with focus on target therapies, immune checkpoint inhibition, looking for prognostic and predictive factors.

Prognostic and predictive factors in testicular cancer.

OBJECTIVE: Testicular cancer is a relatively rare neoplasia, with an incidence of about 1,5% among male malignancies, usually in the third and fourth decade of life. Although several histological variants are known, with some histotypes affecting older patients (e.g., spermatocytic seminoma), there is a clear predominance (90-95%) of germ cell tumors among young adults patients1. Testicular Germ Cell Tumor (TGCT), undoubtedly the seminoma histological variant more than non-seminoma one, is definitely a highly curable disease, with a distinctive sensitivity to cisplatin-based therapy (and for seminomas to radiotherapy) and an outstanding cure rate of nearly 80% even for patients with advanced disease. So far, clinical and pathohistological features supported our efforts to choose the best treatment option for patients suffering from this malignancy, but we don't clearly enough know molecular and pathological features underlying different clinical behaviors, mostly in early-stage disease: by improving this knowledge, we should better "shape" therapeutic or surveillance programs for each patient, also in order to avoid unnecessary, if not harmful, treatments.

Health-related quality of life and psychosocial implications in testicular cancer survivors. A literature review.

OBJECTIVE: In this review, we focused our attention on Quality of Life (QoL) of testicular cancer survivors (TCSs), in general and in the most relevant areas. Several key findings have been highlighted in our review. MATERIALS AND METHODS: PubMed, MEDLINE and PsycINFO databases were consulted to find published studies, from 1980 to May 2017, that met our inclusion criteria. RESULTS: The majority of studies investigated older adult TCSs, while few studies on adolescent and young adult patients were available. Many studies indicate that health-related QoL (HRQoL) is similar among the TCSs and the general population. Even if QoL deteriorates so clear at the time of diagnosis and throughout treatment, afterward returns to normal levels, as defined by the matched controls. However, there are numerous chronic conditions consequent to diagnosis and treatment of testicular cancer that plague survivors and affect QoL, like Raynaud-like phenomena, peripheral neuropathy, fatigue, anxiety, sexual, fertility and body image problems. Even if these problems can have no effects on the measures of global QoL, they have an impact on the quality of life. Differences between TCSs with and without a partner bring to different outcomes in the adjustments to cancer. CONCLUSIONS: It is necessary to identify TCSs with higher risks of poorer QoL outcomes, to focus interventions on the areas with the greatest impairments. Further researches should consider the effects of testicular cancer on the impaired areas, collecting more data to better identify survivor's needs and consequent interventions, with a special focus on adolescent and young adult TCSs. Other works are requested on therapies, preventive and ameliorative, to reduce chronic side effects of testicular cancer treatment.

Rectal/urinary toxicity after hypofractionated vs. conventional radiotherapy in high risk prostate cancer: systematic review and meta analysis.

OBJECTIVE: The aim of our report was to review the literature concerning the toxicity of radiation therapy in patients treated for high-risk prostate cancer, and to evaluate the differences in toxicity between conventional fractionation and hypofractionated treatments, in view of different techniques used in high-risk prostate cancer patients. MATERIALS AND METHODS: PubMed database has been explored for studies concerning acute and late urinary/gastrointestinal toxicity in high-risk prostate cancer patients treated with radiotherapy. Prospective studies, concerning potential relationship between acute/late genitourinary (GU)/gastrointestinal (GI) toxicity and prostate radiotherapy in patients with high-risk prostate cancer, were included in the final analysis. Data collected from single arm, phase II non-randomized and randomized studies have been evaluated to perform odds ratio for toxicity risk. Furthermore, meta-analysis randomized prospective trials were considered suitable because they had recruited high-risk prostate cancer patients who didn't undergo surgery, with available data on ≥ G2 toxicity frequency. RESULTS: The initial search provided 606 results, but only 35 manuscripts met all eligibility requirements and were included in this report. In order to perform odds ratio we observed a decrease in late gastrointestinal toxicity for patients treated with hypofractionated schemes compared to CV treated ones. Among patients who underwent conventional treatment, SIB seemed to decrease acute genitourinary side effects; SIB-Hypo treated patients suffered less toxicity than patients treated with hypofractionated- sequential boost schemes. Hypo-SIB schemes would seem less toxic in terms of acute gastrointestinal and late genitourinary side effects than CV-SIB. Therefore, our focus shifted to 6 clinical trials evaluating genitourinary and gastrointestinal toxicity in patients who had been randomized to receive conventional fractionation or hypofractionated treatment, in both cases with IMRT technology. Our meta-analysis of these randomized trials involving patients with high-risk prostate cancer showed a statistically significant increase in late genitourinary toxicity for hypo-treated patients; no difference was observed in acute genitourinary/gastrointestinal toxicity, and in late gastrointestinal toxicity. CONCLUSIONS: Our analysis doesn't want to establish a definitive truth; very few trials assessed only high risk-class patients. Our purpose is to stimulate further randomized prospective trials focusing both on the effectiveness and toxicity profile (toxicity/effectiveness ratio), taking into account the use of different technologies and doses.

Prevalence and antimicrobial susceptibility of salmonella in European wild boar (Sus scrofa); Latium Region - Italy.

The prevalence of Salmonella spp. infection was determined in 499 wild boars harvested during the 2010-2011 and 2011-2012 hunting seasons in the Latium Region of Italy. We conducted a microbiological assessment on faeces collected at slaughter and we examined serum samples for the presence of antibodies to Salmonella spp. by ELISA assay. Out of 383 serum samples examined, 255 (66.5%) were positive for Salmonella spp. antibodies. Overall, 10.8% (54/499) of the animals were positive by microbiological assessment. The Salmonellae most frequently isolated were S. enterica subsp. salamae II (24%), S. enterica subsp. Diarizonae III b (12.9%), S. enterica subsp. houtenae IV (11.1%) and S. Fischerhuette (7.4%); less common Salmonella isolates included S. Veneziana (5.5%), S. Napoli (5.5%), S. Kottbus (5.5%), S. Thompson (5.5%), S. enterica subsp. arizonae III a (3.7%), S. Toulon (3.7%), S. Burgas (1.8%), S. Tennelhone (1.8%), S. Ferruch (1.8%), S. choleraesuis (1.8%), S. Paratyphi (1.8%), S. Stanleyville (1.8%), S. Typhimurium (1.8%) and S. enterica subsp. enterica 4,5,12:1:- (1.8%). These isolates were tested against 16 antimicrobial agents and exhibited resistance to sulphonamides (92.5%), sulphonamides and thrimetroprim (14.8%), colistin (14.8%), streptomycin (18.5%), gentamycin (5.5%), tetracycline (5.5%), ceftiofur (3.7%), cefazoline (1.8%), cefotaxime (1.8%), nalidixic acid (1.8%), amoxicillin and clavulanic acid (1.8%) and ampicillin (3.7%). Our data, the first collected on this species in Italy, suggest that European wild boars are frequent carriers of antimicrobial-resistant Salmonellae and are likely involved in the transmission of antimicrobial resistance throughout the environment.

Characterization of enterotoxigenic E. coli (ETEC), Shiga-toxin producing E. coli (STEC) and necrotoxigenic E. coli (NTEC) isolated from diarrhoeic Mediterranean water buffalo calves (Bubalus bubalis).

Two hundred and twenty Escherichia coli isolates from 314 Mediterranean water buffalo calves less than 4 weeks old affected by severe diarrhoea with a lethal outcome were characterized for the presence of the virulence factors LT, ST, Stx1, Stx2, haemolysins, intimin, CNF1, CNF2, CDT-I, CDT-II, CDT-III, CDT-IV, and F17-related fimbriae (F17a, F17b, F17c, F17d). The prevalence of ETEC, STEC and NTEC were 1.8%, 6.8% and 20.9%, respectively. The ETEC isolates were all LT-positive and ST-negative. The STEC isolates were all Stx and intimin-positive, with Stx1 (80%) more frequent than Stx2 (27%). The NTEC isolates were all CNF and Hly-positive, with CNF2 (83%) more frequent than CNF1 (22%). Susceptibility assays to 11 antimicrobials displayed high rates of resistance (>30%) to antimicrobials tested. These data show that the most prevalent strains in diarrhoeic water buffalo calves were NTEC, mostly CNF2 and HlyA-positive, with strong associations CNF2/CDT-III and CNF2/F17c.

A survey of dioxins (PCDDs and PCDFs) and dioxin-like PCBs in sheep and goat milk from Campania, Italy.

In 2008, after the crisis of buffalo dairy fields in Campania, Italy, an assessment of the contamination of polychlorinated dibenzo-p-dioxins and polychlorinated dibenzofurans (PCDD/Fs) and dioxin-like polychlorinated biphenyls (dl-PCBs) was also necessary for other animal species bred in the region. The contents of PCDDs, polychlorinated dibenzofurans (PCDFs), and dl-PCBs were determined by high-resolution gas chromatography/mass spectrometry (HR-GC/MS) (according to USEPA method 1613) in 69 sheep and goat milk samples from 63 farms. In eleven samples from six sheep farms, the PCDD/Fs levels exceeded the maximum limit of 3.0 pg g(-1) fat established by the European Commission, in particular the concentrations ranged between 3.89 and 12.90 pg g(-1) fat. Statistical treatment of the results for the congener profiles of the non-compliant and compliant samples has been used to identify the sources of contamination.

Analysis of apoptosis induced by Caprine Herpesvirus 1 in vitro.

It is known that Caprine Herpesvirus 1 (CpHV-1) causes apoptosis in mitogen-stimulated as well as not stimulated caprine peripheral blood mononuclear cells (PBMC). Initial experiments in Madin Darby bovine kidney (MDBK) cells revealed that CpHV-1 infection induced apoptotic features like chromatin condensation and DNA laddering. Thus, to characterize in more detail this apoptotic process, activation of caspase-8, -9 and -3 in MDBK cells CpHV-1 infected was investigated and demonstrated. In addition, CpHV-1 infection resulted in disruption of mitochondrial membrane potential, cytochrome c release and alterations in the pro- and anti-apoptotic proteins of Bcl-2 family. Proteolytic cleavage of poly(ADP-ribose) polymerases (PARP), confirming the activation of downstream caspases, was also observed. Our data indicated that a "cross-talk" between the death-receptor (extrinsic) pathway and the mitochondrial (intrinsic) pathway occurred in CpHV-1-induced apoptosis in vitro.

Bovine papillomavirus type 1 DNA and E5 oncoprotein expression in water buffalo fibropapillomas.

Papillomas and fibropapillomas may occur in the skin and in different organs in animals. Ten different genotypes of bovine papillomavirus (BPV) have been identified. BPV-1 through BPV-10 are all strictly species-specific, but BPV-1/2 may also infect other species such as equids, inducing fibroblastic tumors. BPV-1 and BPV-2 are associated with fibropapillomas in cattle; these tumors are formed by excessive proliferation of virus-infected dermal fibroblasts and epidermal keratinocytes. Nine water buffalo (Bubalus bubalis) were examined for the presence of multiple cutaneous and perivulvar tumors. Cutaneous and perivulvar fibropapillomatosis were confirmed histologically. Negative-stain transmission electron microscopic examination revealed papillomavirus-like particles in the fibropapillomas, and papillomaviral DNA was also detected by the polymerase chain reaction. The amplified long control region (LCR) DNA sequence was identical to that of BPV-1. The BPV-1 E5 oncoprotein was strongly expressed in the tumor cells thus confirming a causal role of the virus. This article represents the first report of cutaneous, perivulvar, and vulvar fibropapilloma associated with BPV-1 infection in the water buffalo and describes another example of cross-species infection by BPV-1.

Detection of Coxiella burnetii in buffaloes aborted fetuses by IS111 DNA amplification: a preliminary report.

The aim of this study was to evaluate by PCR analyses the presence of Coxiella burnetii infection in fetuses of water buffalo (Bubalus bubalis) in the Campania region (Southern Italy). Samples were collected only from aborted fetuses and C. burnetii presence was evaluated by one-tube nested PCR amplification of the IS111 repetitive element. Of the 164 fetuses examined 14 (17.5%) were positive after DNA amplification, showing that C. burnetii occurs in this population of water buffaloes. However, more extensive prevalence studies need to be carried out to define the role of buffaloes as reservoirs for this pathogen and also the role of C. burnetii as an abortive agent in this animal.

Feline herpesvirus-1 down-regulates MHC class I expression in an homologous cell system.

Cytotoxic T lymphocytes (CTLs) are an essential component of the immune defense against many virus infections. CTLs recognize viral peptides in the context of the major histocompatibility complex (MHC) class I molecules on the surface of infected cells. Many viruses have evolved mechanisms to interfere with MHC class I expression as a means of evading the host immune response. In the present research we have studied the effect of in vitro Feline Herpesvirus 1 (FeHV-1) infection on MHC class I expression. The results of this study demonstrate that FeHV-1 down regulates surface expression of MHC class I molecules on infected cells, presumably to evade cytotoxic T-cell recognition and, perhaps, attenuate induction of immunity. Sensitivity to UV irradiation and insensitivity to a viral DNA synthesis inhibitor, like phosphonacetic acid, revealed that immediate early or early viral gene(s) are responsible. Use of the protein translation inhibitor cycloheximide confirmed that an early gene is primarily responsible.

Evaluation of molecular methods for the detection of Brucella species in water buffalo milk.

Brucellosis is a highly infectious disease affecting both animals and humans. The current standard tools for the diagnosis of this bacterial infection are serological and microbiological. In this study, we evaluated the feasibility of molecular assays as diagnostic tools for the detection of Brucella spp. in water buffalo milk. For this purpose, we first compared different DNA extraction protocols and PCR methods on artificially spiked milk samples. The most sensitive methods were then used to examine milk from serologically positive and negative water buffaloes. Molecular results were compared with serological and bacteriological test results. Milk samples from 53 Brucella seropositive buffaloes (by either rose Bengal or complement fixation test) were positive by ELISA, 37 were positive by culture, 33 were positive by PCR, and 35 were positive by real-time PCR. Of the 37 culture-positive samples, a total of 25 and 26 were positive by PCR and real-time PCR, respectively. Of the 16 culture-negative samples, 8 were positive by PCR and 9 by real-time PCR. Thus, although culture showed greater sensitivity than PCR, some animals found positive by serological methods and PCR tested negative by milk culture. The combined use of bacteriological and molecular tools increased the number of positive samples to 46. In conclusion, these results suggest that the simultaneous application of these 2 direct detection methods (culture and PCR) could be more useful than one test alone for the diagnosis of Brucella spp. in buffalo milk.

Evaluation of a fluorescence polarization assay for the detection of serum antibodies to Brucella abortus in water buffalo (Bubalus bubalis).

The fluorescence polarization assay (FPA) was evaluated for the serological diagnosis of brucellosis in water buffalo (Bubalus bubalis) in southern Italy. This assay uses O-polysaccharide prepared from Brucella abortus lipopolysaccharide conjugated with fluorescein isothiocyanate as a tracer. It has many methodological advantages over older, more established tests and can be performed in a fraction of the time. Sera from 890 buffalos from the Campania Region - 526 positive sera and 364 negative sera according to the complement fixation test (CFT) - were evaluated in this study. All samples were tested with the Rose Bengal test (RBT), CFT, and FPA in parallel and in blind fashion. Sensitivities (Sn) were 84.5% and 92.6%, and specificities (Sp) were 93.1% and 91.2% for RBT and FPA, respectively, relative to CFT. Finally, receiver operating characteristic (ROC) analysis suggested a cut-off value of 117 millipolarization (mP) units. On the whole, these results suggested that FPA might replace RBT in the diagnosis of buffalo brucellosis for its better performance relative to CFT, its adjustable cut-off useful in different epidemiological situations, its reliability, ease of performance, and for its potential application in field and high-throughput laboratories.

Retrospective study of bacterial isolates and their antimicrobial susceptibilities in equine uteri during fertility problems.

Bacterial pathogens are a potential cause when a mare fails to conceive to a fertile stallion on a well-managed breeding farm on one or more cycles in the same season. Furthermore, emerging bacterial resistance to commonly used (topical) antibiotics has been demonstrated. In this study, a total of 586 uterine swabs from mares with fertility problems were evaluated and the bacterial isolates were identified and measured for resistance to 10 antibiotics most commonly used during bacterial equine infection. Forty-nine percent of the examined mares were positive at bacteriological investigations. Amongst 347 successful isolations, 31.7% were Streptococcus group C and 18.4% Escherichia (E.) coli, both considered frequently associated with fertility problems. Determination of the antibiotic susceptibility pattern of Streptococcus group C (110 organisms) revealed that only the amoxicillin/clavulanic acid was highly active with 82.7% of the isolates being inhibited. For E. coli, a major number of drugs displayed a high potency.

2,3,7,8-Tetrachlorodibenzo-p-dioxin increases Bovine Herpesvirus type-1 (BHV-1) replication in Madin-Darby Bovine Kidney (MDBK) cells in vitro.

Dioxin-2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) is a common environmental toxin of current interest. In the last years, higher levels of TCDD than those permitted in UE [European Commission. 2002. European Commission Recommendation 2002/201/CE. Official Gazette, L 67/69] were detected in milk samples from cow, water buffalo, goat, and sheep raised on some areas of Campania Region (South Italy). Dioxin often causes immunosuppression and might render the animal liable to viral infections. In addition, viral infections are able to alter the pattern of dioxin distribution in different organs of the exposed animals. Bovine Herpesvirus type-1 (BHV-1) is a widespread pathogen, which causes infectious rhinotracheitis and infectious pustular vulvovaginitis in cattle. Herein, we have studied the effects of TCDD and BHV-1 infection, in Madin-Darby Bovine Kidney (MDBK) cells, alone as well as in association, so as cellular proliferation, apoptosis, and virus replication. We have observed an increase in cell viability of confluent monolayers at low TCDD concentrations. TCDD treated cells demonstrated increased viability compared to controls as evaluated by MTT test. TCDD exposure increased cell proliferation but induced no changes on apoptosis. Cells exposed to TCDD along with BHV-1 showed a dose-dependent increase in cytopathy, represented by ample syncytia formation with the elimination of the cellular sheets and increased viral titer. These results suggest that TCDD increases viral replication in MDBK cells while BHV-1 further decreases viability of TCDD exposed cells. Since very low concentrations (0.01 pg/ml) are sufficient to augment BHV-1 titer, TCDD may contribute to reactivate BHV-1 from latency, leading to recurrent disease and increase virus transmission.

Survey of Neospora caninum and bovine herpes virus 1 coinfection in cattle.

A seroprevalence survey of Neospora caninum and bovine herpesvirus 1 (BHV-1) was conducted in cattle pasturing in an area of the southern Italian Apennines to investigate the coinfection of these two pathogens. Blood samples were collected from 948 pastured cattle raised on 81 farms. Sera were tested for antibodies to N. caninum and to BHV-1 using an ELISA assay and a neutralization test, respectively. Out of the 81 farms sampled, 63 (77.8%) were positive for N. caninum and 80 (98.8%) for BHV-1. Coinfection was found in 62 (76.5%) farms. Out of the 948 bovine sera samples, 303 (32.0%) had antibodies to N. caninum and 735 (77.5%) to BHV-1. The copresence of antibodies to N. caninum and BHV-1 was found in 256 (27.0%) cattle. The logistic regression results indicated that seropositivity for BHV-1 was a risk factor for N. caninum seropositivity and seropositivity for N. caninum was a risk factor for BHV-1 seropositivity.