Using multi-criteria risk ranking methodology to select case studies for a generic risk assessment framework for exotic disease incursion and spread through Europe.

We present a novel approach of using the multi-criteria pathogen prioritisation methodology as a basis for selecting the most appropriate case studies for a generic risk assessment framework. The approach uses selective criteria to rank exotic animal health pathogens according to the likelihood of introduction and the impact of an outbreak if it occurred in the European Union (EU). Pathogens were evaluated based on their impact on production at the EU level and international trade. A subsequent analysis included criteria of relevance to quantitative risk assessment case study selection, such as the availability of data for parameterisation, the need for further research and the desire for the case studies to cover different routes of transmission. The framework demonstrated is flexible with the ability to adjust both the criteria and their weightings to the user's requirements. A web based tool has been developed using the RStudio shiny apps software, to facilitate this.

A cross-sectional study to identify a set of risk factors for caprine herpesvirus 1 infection.

BACKGROUND: Caprine herpesvirus 1 (CpHV-1) causes neonatal mortality and reproductive failure in goats. Despite its impact on herd reproductive performance, few studies have investigated the risk factors associated with CpHV-1 infection. The aim of this cross-sectional study was to identify potential herd- and host-level risk factors associated with CpHV-1 prevalence in a goat population with heterogeneous seropositivity for CpHV-1. RESULTS: Blood samples and individual data from 4542 goats were collected from 255 herds in Piedmont, Italy. Enzyme-linked immunosorbent assay (ELISA) and serum neutralization tests were carried out to detect antibodies against CpHV-1. A mixed-effects model was applied to identify any statistical association between CpHV-1 seropositivity and a set of putative host-level and herd-level risk factors. A total of 630 samples tested were found positive by ELISA (prevalence = 13.9%; 95% confidence interval (CI) 12.9-14.9). Of the 255 tested herds, 85 were classified as positive for the presence of at least one gB-positive animal (herd prevalence 33.3%, 95% CI 27.5-39.2), with a within-herd prevalence between 0.7 and 100% (Q1 = 17.6%; median = 32.3%; Q3 = 50%) (Q = quartiles). The prevalence ratios showed a statistical association with the following risk factors: breeds other than Saanen, older age, larger herd size, meat and extensive herds, and co-existence of CAEV-infected animals. CONCLUSIONS: Results from this cross sectional study may help to elucidate the natural history of the infection and inform targeted strategies to control a disease with a potentially important impact on animal health and goat farming economy.

Histopathology as a simple and reliable method to detect 17ß-oestradiol illegal treatment in male calves.

17ß-Oestradiol is a steroid hormone banned as a growth promoter in food-producing animals all over Europe because of its carcinogenicity. Despite mandatory monitoring of illegal treatment all over Europe, official analytical methods in use test negative a few days after 17ß-oestradiol administration, requiring new sensitive tools to ensure a high level of protection for consumers. The aim of this work was the evaluation of the accuracy of histopathology and immunohistochemistry for progesterone receptor (PR) as a screening method for the detection of low-dosage illegal treatments with 17ß-oestradiol. Fresian male calves (153) were farmed under controlled conditions, and 89 of them were treated with 17ß-oestradiol (5 mg/animal once a week for 4 weeks). After 15 days of suspension, all animals were slaughtered and sexual accessory glands (prostate and bulbo-urethral glands) were sampled for histological examination and immunohistochemical staining with anti-PR antibody (clone hPRa 2). Microscopically 86 out of 89 bulbo-urethral glands showed mild to severe metaplasia, while mild metaplasia was observed only in 1 control. Eighteen out of 89 samples of prostate did not show metaplastic lesions. Immunopositivity for PR characterised all treated animals, while no signal was detected in controls. These findings show that metaplasia of the sexual accessory glands is a sensitive and specific parameter for illegal 17ß-oestradiol treatment in calves at the slaughterhouse, while the appliance of immunohistochemistry for PR can improve to 100% the accuracy of this highly reliable histological approach.

Molecular characterization of Pseudomonas fluorescens isolates involved in the Italian "blue mozzarella" event.

Between June and September 2010, widespread Italian consumer reports of unusual blue spoilage on fresh dairy products were publicized, resulting in the so-called blue mozzarella event. An inordinately high number of samples from mozzarella and whey cheese products of Italian and German production subsequently tested positive for Pseudomonas fluorescens. The aim of this study was to verify whether a selected P. fluorescens strain was responsible for this apparently unusual event. Molecular characterization of 181 isolated P. fluorescens strains was conducted using a newly optimized pulsed-field gel electrophoresis protocol. Although a high number of pulsotypes was found (132), only four pulsotypes were associated with more than one production plant, and only one German isolate had the same pulsotype as was detected in two Italian plants. This is the only evidence of possible cross-contamination among cheeses from the two countries. The overall results did not support the spread of contamination from German to Italian plants or the presence of one environmental strain that spread in both countries.

Proteinuria in canine patients with lymphoma.

OBJECTIVES: To determine if proteinuria is more common in dogs with lymphoma when compared with healthy dogs and to assess the severity and frequency of proteinuria in dogs with lymphoma. METHODS: Determination of urine protein:creatinine ratio in 32 dogs with lymphoma compared with 30 healthy dogs. RESULTS: Canine patients with lymphoma are more likely to be proteinuric compared with healthy dogs. Proteinuria is common in dogs with lymphoma, although in most cases it is not severe. The presence of proteinuria is not linked with the stage or substage of lymphoma. CLINICAL SIGNIFICANCE: Mild proteinuria is a common finding in dogs with lymphoma. The clinical impact of the proteinuria is probably low.

Mosquito surveillance in northwestern Italy to monitor the occurrence of tropical vector-borne diseases.

Mosquito-borne arboviruses (MBV) represent an important health problem, causing diseases and deaths both in human and animals mainly in tropical and subtropical countries. In recent years, they have emerged also in temperate regions where they have caused epidemics. Of mounting concern among public health authorities in Europe are zoonotic mosquito-borne viruses belonging to the Flavivirus genus. The aim of this study was to carry out active surveillance on mosquitoes in two regions of northwestern Italy (Liguria and Piedmont) to gain a better knowledge of the mosquito populations by identifying potential vectors of arboviruses and to investigate arbovirus infection. A network of 61 CO2 CDC traps was placed in the study area; sampling was conducted from May to October 2011. A total of 46,677 mosquitoes was collected, identified to species level, and classified according to their vector competence. Mosquitoes collected from 16 traps, selected according to risk-based factors, were tested by biomolecular analysis to detect flavivirus infection. This study highlights the importance of entomological surveillance in northwestern Italy because most of the mosquitoes collected were found to have high vector competence. Moreover, the risk-based virological surveillance allowed to detect the presence of mosquito flavivirus RNA, phylogenetically closely related to the MMV Spanish isolate, in three pools and USUV RNA in one pool in new areas where it has not been reported previously. The availability of continuous data on mosquito populations provides invaluable information for use in cases of an epidemic emergency. Maintenance of this integrated system for the next years will provide stronger data that can inform the design of a risk-based surveillance for the early detection of the occurrence of outbreaks of tropical MBDs.

Sepsis and bacterial suppurative meningitis-meningoencephalitis in critically ill neonatal Piedmontese calves: clinical approach and laboratory findings.

Sepsis (S) and bacterial suppurative meningitis-meningoencephalitis (M-ME) are common causes of death in bovine neonates. The aim of this prospective study was to evaluate the prevalence of S and M-ME in critically ill neonatal Piedmontese calves. Critically ill animals up to 15 days old referred by practitioners were registered according to their status and subsequently assigned to clinical standardized score. Calves with a clinical score > = 5 were further assessed under a clinical and clinical-pathological protocol to strengthen the suspicion of S and M-ME. Critically ill neonatal calves sent for necropsy were included in the study as well. Fifty-nine calves were investigated, 26 of which referred alive and 33 dead. Ten out of the 26 clinically evaluated calves were classified as suspicious of S on the basis of the clinical and clinical-pathological protocols. S was confirmed by positive bacteriologic culture in 7 cases and in 3 cases on the basis of necroptic lesions. Concomitant suppurative M-ME suspected in 6 of these 10 calves was subsequently confirmed by CSF analysis or histological findings. Of the 33 calves examined only post-mortem, 20 showed pathognomonic findings of S and 14 signs of M-ME. The prevalence of S and M-ME was 46 and 36 %, respectively. Clinical signs of S were confirmed to be vague and overlapping with other diseases. The developed protocol was highly accurate in predicting S in these neonatal calves.

Ten years of BSE surveillance in Italy: neuropathological findings in clinically suspected cases.

Between 2001 and 2010, 244 clinically suspected cases of bovine spongiform encephalopathy (BSE) were reported in Italy. This report summarizes the neuropathological findings in cattle displaying clinical signs consistent with a diagnosis of BSE. All animal specimens were submitted for confirmatory testing; samples testing negative underwent neuropathological examination to establish the differential diagnosis. Immunohistochemistry for scrapie prion protein (PrPSc) at the level of frontal cortex was carried out to exclude atypical BSE. Neuropathological changes were detected in 34.9% of cases; no histological lesions were found in 52.3% of subjects; 12.8% of samples were found unsuitable for analysis. BSE was detected in one case, but no cases of atypical BSE were observed. This study identified the diseases most commonly encountered in the differential diagnosis of BSE; furthermore, it demonstrated that the surveillance system is necessary for monitoring neuropathological disease in cattle and for the detection of BSE cases.

Development of an enhanced histopathological approach to detect low-dose dexamethasone illicit treatment in veal calves.

Dexamethasone is one of a number of synthetic corticosteroids illegally used to promote growth in food-producing animals. Since these low-level drug cocktails evade detection by currently available chemical methods, simple biological indicators that can aid in laboratory analysis are needed. In an attempt to devise an accurate biological method that could detect illicit drug treatment in food-producing animals, we characterized microscopic morphologic alterations of the thymus in veal calves administered low-dose dexamethasone versus control animals. For this purpose, 122 male calves were farmed for 6 months in controlled condition: 81 animals were orally administered dexamethasone (0.4 mg day(-1)) for 20 days during the sixth month and the remaining 41 were kept as control. Urine samples were collected systematically during the treatment period, the suspension period and at the slaughterhouse. All animals were slaughtered 10 per day starting from 10 days after the last dexamethasone administration and the thymus was sampled for histological examination. The difference between the two animal groups was evaluated by means of a non-parametric test of hypothesis. No residues were detected in the urines collected since the third day after the last administration, whereas morphometric analysis of the thoracic thymus revealed a significant decrease in the cortex:medulla ratio in the treated animals (p<0.0005). We can conclude that this histological approach offers encouraging prospects as a screening method to overcome current limitations in controlling growth promoter abuse.

Heat-related mortality in dairy cattle: A case crossover study.

The relationship between mortality and stressful weather is not only a complex health, social and environmental issue, but also becomes an economical problem when considering livestock. Future-climate scenarios suggest that higher global mean temperatures could result in marked changes in the frequency of extreme temperatures. The relationship between high temperature and mortality is well known in people living in urban areas, but it has been poorly investigated in livestock such as dairy cows. The aim of this study was to quantify the relationship between heat and mortality from all causes among dairy cows in Italy. We studied 6697 adult (≥24 months) dairy cows that died during the summer in the 5-year period 2002-2006 in three Italian districts (Brescia, Cuneo, Rome) which were selected because of the high livestock production and the availability of weather data for these geographical areas. A case-crossover design was applied using the temperature-humidity index (THI) as exposure of interest. Pooled odds ratio (OR) of mortality at high exposure values versus low exposure values was estimated. An exposure over the THI cut-off increased the risk of mortality by 1.6 times, with a 4% increase in mortality for each THI degree increase above the threshold. We found no effect modification for age-class, herd size or breed, but a certain degree of geographical heterogeneity was identified. The increased dairy cow mortality highlighted in this study should be considered when adopting animal welfare and farm management measures. This phenomenon can be prevented by adopting appropriate interventions to mitigate environmental thermal challenges.

Analysis of cerebrospinal fluid from 20 calves after storage for 24 hours.

Samples of CSF collected from 20 normal healthy calves were analysed either immediately or after having been stored for 24 hours at 4 degrees C in the presence of 11 per cent autologous serum. There were no significant differences between the total and differential cells counts of the fresh and stored samples, but there was a positive linear correlation between them. There were some morphological changes to the nuclei of the mononuclear cells in the stored samples.

Neuropathology of italian cats in feline spongiform encephalopathy surveillance.

Feline spongiform encephalopathy (FSE) is a transmissible spongiform encephalopathy associated with the consumption of feedstuffs contaminated with tissue from bovine spongiform encephalopathy-affected cattle and characterized by the accumulation in the central nervous system of an abnormal isoform of the prion protein (PrP(sc)). Clinically, it presents as a progressive fatal neurologic syndrome that is not easily distinguished from other feline neurologic conditions. Most cases of FSE have been reported in England, where it was first detected in 1990, but a few cases have been reported from other European countries. To identify possible cases of FSE in Italy, the Italian Ministry of Health funded a 2-year surveillance project during which the brains from 110 domestic cats with neurologic signs were evaluated histologically for spongiform encephalopathy and immunohistochemically to detect PrP(sc). Although no cases of FSE were found, the study proved useful in monitoring the Italian cat population for other neurologic diseases: neoplasia (21.8%), toxic-metabolic encephalopathy (18.2%), granulomatous encephalitis (15.5%), suppurative encephalitis (4.6%), trauma (3.6%), circulatory disorders (3.6%), degeneration (2.7%), nonsuppurative encephalitis (2.7%), and neuromuscular diseases (1.8%). No histologic lesions were found in 20% of the brains, and samples from 5.5% of the cats were rejected as unsuitable.

Epidemiological study of the decline of BSE in Italy.

In this paper, data derived from the national database of the Italian surveillance system for bovine spongiform encephalopathy (BSE) are used to describe the Italian epidemic of bse. Two data flows were established to collect the results of active and passive surveillance, and 25 regional laboratories were involved. The National Reference Centre (CEA) was in charge of the data analysis. Crude and age-standardised estimates of the prevalence and incidence of bse were obtained to describe the distribution of the disease in terms of the main risk factors (age, breed and herd size), year of birth, time of diagnosis and geographical location. The increased risk was calculated in terms of the incidence rate ratio. During the five years since January 2001, 128 cases of bse were identified in domestic cows and four were identified in imported cattle. All but one of the cases were detected through active surveillance. The risk of the disease was highest in dairy stock and in large herds. The northern regions of Italy had an incidence of bse 2.6 times higher than the southern regions. There was a clear decline in the age-standardised prevalence, from one positive case per 10,000 tests in 2001 to one per 100,000 tests in 2005.

Evaluation of an enzyme immunoassay for the detection of central nervous system tissue contamination at the slaughterhouse.

To protect public health from bovine spongiform encephalopathy, European Commission Regulation EC 1139/2003 on monitoring programs and specified risk material requires that as of 1 October 2003, each member state has in place a sampling plan with an appropriate laboratory test to detect central nervous system (CNS) tissue in bovine head meat harvested at slaughterhouses or cutting plants. With this study, we wanted to evaluate the accuracy and reliability of an enzyme immunoassay, the RIDASCREEN Risk Material 10/5, in targeting a CNS-specific marker, the glial fibrillary acidic protein. A receiver operating characteristics curve was plotted to identify the best cutoff of CNS concentration. Reliability was calculated by Cohen's kappa on data from two diagnostic sessions. Test performance showed high sensitivity and specificity (97.9 and 97.4%, respectively) for a cutoff value between positive and negative at a CNS concentration of 0.049%; reliability of test precision was also very good. When these criteria are applied, the RIDASCREEN Risk Material 10/5 test appears to be a reliable tool for monitoring CNS tissue contamination in meat. This diagnostic procedure should therefore be recommended for national application in monitoring programs.