Bayesian assessment of two competitive enzyme-linked immunosorbent assays for the detection of bovine viral diarrhoea virus antibodies in bovine sera.

Infections due to bovine viral diarrhoea virus (BVDV) are endemic in most cattleproducing countries throughout the world and bovine viral diarrhoea is considered a transboundary disease. The key elements of a BVDV control programme are vaccination, biosecurity, elimination of persistently infected (PI) animals and surveillance. The aim of this study was to assess the sensitivity (Se) and the specificity (Sp) of two commercial competitive enzyme-linked immunosorbent assays (ELISAs) based on selected immune-dominant BVDV proteins: the non-structural protein NS3 (p80) and the recombinant envelope glycoprotein E0(Erns). Both tests were used on individual serum samples from randomly sampled young bovines in southern Belgium in order to detect specific BVDV antibodies. The Se and Sp were assessed using a Bayesian approach and were estimated, respectively, at 97.2% (with 95% credibility interval [Cr I]: 95.1-99.8) and 98.7%(95% Cr I: 96.6-99.9) for the first test and 95.8% (95% Cr I: 91.1-99.7) and 96.1%(95% Cr I: 95.1-97.7) for the second test. The results obtained with the two tests were not significantly different. In addition, using both ELISAs, the current BVDV exposure among young bovines in southern Belgium was estimated at 23.3% (95%Cr I: 20.6-26.2). Combining virological testing of all newborns to detect PI animals with regular serological testing of young stock using ELISAs is recommended in the surveillance of BVDV.

Les infections par le virus de la diarrhée virale bovine (VDVB) sont endémiques dans la plupart des pays d'élevage du monde et la diarrhée virale bovine estune maladie transfrontalière. Les éléments d'un programme de contrôle de la VDVB sont la vaccination, la biosécurité, l'élimination d'animaux porteurs d'une infection persistante et la surveillance. L'objectif de cette étude était d'évaluer la sensibilité (Se) et la spécificité (Sp) de deux kits commerciaux ELISA (épreuve immuno-enzymatique) par compétition basés sur une sélection de protéines immunodominantes du VDVB, la protéine non structurale NS3(p80) et la glycoprotéine d'enveloppe recombinante E0 (Erns). Les deux kits étaient testés sur des échantillons individuels de sérum collectés demanière aléatoire chez de jeunes bovins dans le sud de la Belgique afin de détecter les anticorps VDVB spécifiques. L'analyse bayésienne montrait une Se de 97,2 % (intervalle de crédibilité de 95 % [ICr] de 95,1 à 99,8) et une Sp de 98,7 (ICr 95 % de 96,6 à 99,9) pour le premier kit et une Se de 95,8 (ICr 95 % de91,1 à 99,7) et une Sp de 96,1 (ICr 95 % de 95,1 à 97,7) pour le deuxième. Les différences n'étaient pas significatives. De même, l'application des deux ELISA montrait que l'exposition actuelle des jeunes bovins du sud de la Belgique au VDVB s'élevait à 23,3 % (ICr 95 % de 20,6 à 26,2). L'association de tests virologiques effectués chez tous les nouveau-nés afin de détecter des animaux à infection persistante et de tests sérologiques de routine par ELISA chez les jeunes animaux est recommandée pour la surveillance de VDVB.

Las infecciones por el virus de la diarrea viral bovina (BVDV), considerada enfermedad transfronteriza, son endémicas en la mayoría de los países del mundo que albergan producción bovina. Los principales elementos de todo programa de lucha contra este virus son la vacunación, la seguridad biológica, la eliminación de los animales con infección persistente y la vigilancia. Los autores describen un estudio encaminado a evaluar la sensibilidad y especificidad de dos ensayos inmunoenzimáticos (ELISA) comerciales basados en sendas proteínas inmunodominantes del virus: la proteína no estructural NS3 (p80) y la glucoproteína recombinante de envoltura E0 (Erns). Ambas pruebas fueron aplicadas a sueros procedentes de una muestra aleatoria de bovinos jóvenes del sur de Bélgica con el fin de detectar anticuerpos específicos contra el virus. Empleando un método de estadística bayesiana se calcularon la sensibilidad y la especificidad, que resultaron, respectivamente, de un 97,2% (intervalo de credibilidad [I Cr] al 95%: 95,1­99,8) y un 98,7% (I Cr 95%: 96,6­99,9) en el caso dela primera prueba y de un 95,8% (I Cr 95%: 91,1­99,7) y un 96,1% (I Cr 95%: 95,1­97,7) en el caso de la segunda. Los resultados obtenidos con una y otra prueba no diferían significativamente entre sí. Además, utilizando ambas técnicas ELISA se calculó que la exposición actual al virus de los bovinos jóvenes del sur de Bélgica se cifraba en un 23,3% (I Cr 95%: 20,6­26,2). Para las tareas de vigilancia del BVDV se recomienda combinar el análisis virológico de todos los recién nacidos ­ para detectar animales con infección persistente ­ con la realización periódica de pruebas serológicas en el ganado joven con la técnica ELISA.

Schmallenberg Virus in Belgium: Estimation of Impact in Cattle and Sheep Herds.

Schmallenberg virus (SBV) emerged during summer 2011. SBV induced an unspecific syndrome in cattle and congenital signs (abortions, stillbirths and malformations) in domestic ruminants. To study the impact of SBV in Belgium, a phone survey was conducted upon September 2012. Hereto two groups of cattle farmers (A and B) and two groups of sheep farmers (C and D) were randomly selected. Farms from groups A (n = 53) and C (n = 42) received SBV-positive result at RT-PCR in the Belgian National Reference Laboratory (NRL). Farms from groups B (n = 29) and D (n = 44) never sent suspected samples to NRL for SBV analysis but were however presumed seropositive for SBV after the survey. Questionnaires related to reproduction parameters and clinical signs observed in newborn and adult animals were designed and addressed to farmers. As calculated on a basis of farmers' observations, 4% of calves in group A and 0.5% in group B were reported aborted, stillborn or deformed due to SBV in 2011-2012. The impact as observed by sheep farmers was substantially higher with 19% of lambs in group C and 11% in group D that were reported aborted, stillborn or deformed due to SBV in 2011-2012. Interestingly, abortions or stillbirths were not clear consequences of SBV outbreak in cattle farms, and the birth of a deformed animal was an essential condition to suspect SBV presence in cattle and sheep farms. This study contributes to a better knowledge of the impact of the SBV epidemic. The results suggest that SBV impacted Belgian herds mostly by the birth of deformed calves, stillborn lambs and deformed lambs. This work also demonstrates that the birth of a deformed calf or lamb was a trigger for the farmer to suspect the presence of SBV and send samples to NRL for further analyses.

Resurgence of Schmallenberg Virus in Belgium after 3 Years of Epidemiological Silence.

In spring 2016, three years after the last reported outbreak of Schmallenberg virus (SBV) in Belgium, an abortion was notified in a two year old Holstein heifer that previously had not been vaccinated against SBV. The autopsy of the eight-month-old malformed foetus revealed hydrocephalus, torticollis and arthrogryposis. Foetal brain tissue and blood were found to be SBV-positive by RT-PCR and ELISA tests, respectively. Evidencing the circulation of SBV in Belgium in the autumn 2015 is important to anticipate future outbreaks and advise veterinarians about the risks associated with calving, as more bovine foetuses might have been infected.

Follow-up of the Schmallenberg Virus Seroprevalence in Belgian Cattle.

Schmallenberg virus (SBV), which emerged in Northwestern Europe in 2011, is an arthropod-borne virus affecting primarily ruminants. Based on the results of two cross-sectional studies conducted in the Belgian ruminant population during winter 2011-2012, we concluded that at the end of 2011, almost the whole population had already been infected by SBV. A second cross-sectional serological study was conducted in the Belgian cattle population during winter 2012-2013 to examine the situation after the 2012 transmission period and to analyse the change in immunity after 1 year. A total of 7130 blood samples collected between 1st January and 28 February 2013 in 188 herds were tested for the presence of SBV-specific antibodies. All sampled herds tested positive and within-herd seroprevalence was estimated at 65.66% (95% CI: 62.28-69.04). A statistically significant decrease was observed between the beginning and the end of 2012. On the other hand, age-cohort-specific seroprevalence stayed stable from 1 year to the other. During winter 2012-2013, calves between 6 and 12 months had a seroprevalence of 20.59% (95% CI: 15.34-25.83), which seems to be an indication that SBV was still circulating at least in some parts of Belgium during summer-early autumn 2012. Results showed that the level of immunity against SBV of the animals infected has not decreased and remained high after 1 year and that the spread of the virus has slowed down considerably during 2012. This study also indicated that in the coming years, there are likely to be age cohorts of unprotected animals.

Large-scale cross-sectional serological survey of Schmallenberg virus in Belgian cattle at the end of the first vector season.

A cross-sectional survey was conducted in the Belgian cattle population after the first period of infection of the emerging Schmallenberg virus. A total number of 11 635 cattle from 422 herds sampled between 2 January and 7 March 2012 were tested for the presence of Schmallenberg-specific antibodies using an ELISA kit. Between-herd seroprevalence in cattle was estimated at 99.76% (95% CI: 98.34-99.97) and within-herd seroprevalence at 86.3% (95% CI: 84.75-87.71). An Intraclass Correlation Coefficient of 0.3 (P < 0.001) was found, indicating that the correlation between two animals within a herd with respect to their serological status was high. Those results corroborate the conclusion that the Schmallenberg virus was widespread in Belgium during winter 2011. Seroprevalence was shown to be statistically associated to the animal's age (P < 0.0001): with 64.9% (95% CI: 61.34-68.3) estimated for the 6-12 months of age, 86.79% (95% CI: 84.43-88.85) for the 12-24 months of age and 94.4% (95% CI: 93.14-95.44) for the animals older than 24 months. Based on the results of the described serological survey, we can conclude that after the first Schmallenberg virus episode, almost every Belgian cattle has already been in contact with the virus. In consequence, the vast majority of the host animals should have developed post infection protective immunity against the virus.

Drastic decrease of Salmonella Enteritidis isolated from humans in Belgium in 2005, shift in phage types and influence on foodborne outbreaks.

In Belgium, non-typhoidal salmonellosis and campylobacteriosis are the two most frequently reported foodborne illnesses. During 2005, a 71% decrease of Salmonella Enteritidis infections compared with the average annual number cases in the period 2000-2004 was recorded by the Belgian National Reference Centre for Salmonella and Shigella. After the peak of 1999, the total number of salmonellosis cases decreased gradually, with the exception of 2003 when an increase was again recorded due to the rise of isolates belonging to the serotype Enteritidis. PT4, the predominant phage type of serotype Enteriditis over recent years (except in 2003), became the second most prevalent phage type in 2005 after PT21. We present in this paper the epidemiology (incidence and trends) of human salmonellosis in Belgium and assess the role of the vaccination programme in layer flocks on the decline of the incidence of human salmonellosis and foodborne outbreaks due to S. Enteritidis.

Generation of immature autologous clinical grade dendritic cells for vaccination of cancer patients.

BACKGROUND: Dendritic cell (DC)-based vaccine is a promising approach for cancer therapy. Pioneer trials have been conducted using DC generated in research conditions. There is now a need for generating DC in clinical grade conditions, including the use of closed systems, avoidance of FCS and respect of good manufacturing practices (GMP). METHODS: DC were generated from 84 leukapheresis products of 27 cancer patients enrolled in two Phase I/II trials of vaccination of either MAGE+tumors (n = 24) or prostate cancer (n = 3). Monocytes were seeded in culture bags in a serum-free medium supplemented with IL-4 and GM-CSF. After a 7 day culture, DC were collected and most were pulsed with various MAGE-derived peptides. RESULTS: After a short leukapheresis (mean time: 66 min; mean processed blood: 5 L), a mean of 6 x 10(9) WBC were collected, from which 2.25 x 10(9) were seeded. The culture procedure yielded a large number of DC (mean: 62 x 10(6) DC) harboring the expected phenotype of immature DC (CD1a(+) CD14(-) HLA-DR(+) CD80(+) CD86(+) CD83(-)). This phenotype was not altered by peptide loading. These DC, either fresh or thawed, were functionally effective invitro. Their s.c. and i.v. injections were devoid of any short-term side effect and associated with the induction of immune responses in the patients. DISCUSSION: Large numbers of functional immature clinical grade DC can be generated in a closed system from leukapheresis products in cancer patients. These results provide the basis for large-scale studies of cancer immunotherapy under improved safety conditions.

[Novel autograft method using positive selection of CD34 stem cells]. / Méthode nouvelle d'autogreffe par sélection positive de cellules souches sanguines CD34.

High dose chemotherapy with autologous blood stem cell rescue becomes widely used for patients with hematologic malignancies and solid tumors. Recently, it has been demonstrated that stem cells characterized by the CD34 antigenic marker could be positively selected using an anti CD34 monoclonal antibody and an avidin biotin immunoabsorption device. We report our experience of twelve selections and ten grafts. A CD34+ cells enrichment of 1.9 log (purity: 72%) and a CFU-GM cells concentration of 1.6 log have been obtained. In ten transplanted patients, the hematological recovery was similar to that obtained with non selected blood stem cells. The CD34+ cells purification allows mini graft infusion and purge of residual tumor cells implicated in relapse after autologous stem cells transplantation.