Transplacental transmission of the Italian Bluetongue virus serotype 2 in sheep

In order to study the capability of a Bluetongue virus serotype 2 (BTV­2) field isolate to cross the placental barrier, 2 groups of 5 pregnant ewes were infected with a field BTV­2 Italian strain (Group A) or with the same strain passaged once in Culicoides cells (Kc) (Group B). Following infection, EDTA­blood and serum samples were collected weekly and tested for the presence of BTV RNA/infectious virus and anti­BTV­2 antibodies, respectively. At lambing, precolostral EDTA­blood and serum samples were collected from lambs and tested as before. The lambs were then sampled as scheduled for the dams. All sheep seroconverted on day 12 post­infection (pi) and remained seropositive throughout the sampling period (day 68 pi). BTV was isolated from day 7 pi to day 14 pi in animals of Group A and from day 5 pi to day 12 pi in animals of Group B. None of the 14 lambs born had pre­colostral antibodies. Three lambs born from two ewes of Group B were viraemic at birth and in one lamb infectious virus was isolated from blood up to 11 days of age. This study proved for the first time that a single passage of BTV­2 field strain in Kc cells is able to give to BTV the ability to cross the placenta barrier and infect foetal tissues.

Exploiting serological data to understand the epidemiology of bluetongue virus serotypes circulating in Libya.

The epidemiological patterns of Bluetongue (BT) in North Africa and Mediterranean Basin (MB) dramatically changed by emergence of subsequent episodes of novel bluetongue virus (BTV) serotypes with highly pathogenic indexes and socio-economic impacts. The objective of the study was to investigate the sero-prevalence and serotype distribution of BTV in Libya. During 2015-2016, a total of 826 serum samples were collected from domestic ruminants in Libya. All sera were assayed by competitive enzyme-linked immunosorbent assays (c-ELISA). C-Elisa-positive samples (43.3%; 173/400) were further analyzed by virus neutralization assay to identify BTV serotypes and determine the antibody titre of positive samples. An overall BTV sero-prevalence was 48.4% (95% CI: 45.0%-51.8%). Neutralizing antibodies were detected against the following BTV serotypes namely: BTV-1, BTV-2, BTV-3, BTV-4, BTV-9 and BTV-26. While BTV-1, BTV-2, BTV-4 and BTV-9 circulation was unsurprising as they have been responsible of the last year outbreaks in Northern African Countries, the detection of BTV-3 and BTV-26 was definitely new and concerning for the animal health of the countries facing the Mediterranean Basin. It is crucial that European and Northern African authorities collaborate in organizing common surveillance programmes to early detect novel strains or emerging serotypes in order to set up proper preventive measures, and, in case, develop specific vaccines and plan coordinated vaccination campaigns.

Prevalence and risk factors for bluetongue in the State of São Paulo, Brazil.

Bluetongue (BT), caused by Bluetongue virus (BTV), is a disease that affects ruminants such as cattle, sheep, goats and deer. BTV is transmitted by female midges of the genus Culicoides. In Brazil, information on the prevalence of BTV in cattle is limited, so the objective of this work was to identify BTV serotypes in cattle. The State of São Paulo was divided into seven cattle-producing regions, and in each of them, 300 cattle farms were randomly selected. One animal from each farm (out of a total of 1,598 farms) was selected and its sera tested by virus neutralization technique against BTV serotypes (1-24 and 26) for determining antibody titre. Moreover, for each sampled farm, an epidemiological questionnaire was submitted to verify the type of cattle production and the zootechnical and sanitary practices carried out, which could be associated with a higher risk of BTV infection. In this study, antibodies (percentage, [95% confidence interval]) were identified against 11 serotypes: BTV-1 (22.15%, [15.72-27.92]), BTV-2 (31.03%, [26.65-37.98]), BTV-3 (18.96%, [12.42-24.90]), BTV-4 (24.90% [19.41-29.12]), BTV-9 (6.82%, [1.45-11.72]), BTV-12 (7.50%, [2.82-12.51]), BTV-17 (23.90%, [17.35-29.35]), BTV-19 (10.20%, [4.62-5.56]), BTV-21 (30.66%, [25.00-36.00]), BTV-22 (12.14%, [5.91-18.55]), BTV-26 (57.00%, [51.41-63.59]). In this study, for the first time in Brazil serological evidence of the presence of serotypes BTV-2, BTV-9, BTV-21 and BTV-26 is reported. The variable 'new cattle entering herd' was considered a risk factor for the occurrence of infection (OR = 2.183, 95% CI = 1.6-2.9).

Analysis of bluetongue serotype 3 spread in Tunisia and discovery of a novel strain related to the bluetongue virus isolated from a commercial sheep pox vaccine.

Bluetongue (BT), is one of the OIE-listed major diseases of ruminants. Following the official report of BT virus serotype 3 (BTV-3) in a sheep in Cap Bon (Tunisia), blood and serum samples of ruminants were collected from some areas of Tunisia to further investigate the presence of this virus in the country. A quantitative real time RT-PCR has been first developed for the detection and quantitation of BTV-3 RNA from field specimens. Out of 62 collected blood samples, 23 were shown to be positive for BTV-3 RNA. Isolation on cell cultures was also possible from six samples. Genome sequencing revealed the circulation of two unrelated western strains of BTV-3, one circulating in Cap Bon and neighboring areas, and the other circulating nearby the border with Libya. The presence of a putative novel BTV serotype (BTV-Y TUN2017) in sheep introduced from Libya to Tunisia, genomically related to the BTV strain contaminating a commercially-available sheep pox vaccine and to BTV-26, has been also demonstrated. This finding highlights the pressing need for a prompt production and release of a novel inactivated BTV-3 vaccine to be used in case of emergence or proactively in the areas of Southern Europe at major risk of BTV introduction. The assessment of a novel vaccine will certainly exalt the role and importance of surveillance activities and collaboration with Northern African countries.

Competitive enzyme-linked immunosorbent assay using baculovirus-expressed VP7 for detection of epizootic haemorrhagic disease virus (EHDV) antibodies.

Epizootic haemorrhagic disease (EHD) is a vector-borne infectious viral disease of domestic and wild ruminants. EHD could spread from infected northern African countries in free territories like the EU; therefore, the availability of diagnostic assays would represent key components for adequate surveillance and control programs. In this study, the gene encoding the VP7 protein of EHD virus (EHDV) was expressed into a baculovirus-infected insect cell system. With this unpurified protein we developed a home-made competitive ELISA (cELISA) and a total number of 275 serum samples, originating from domestic and wild ruminants, were tested. 74/275 were previously shown to be positive for EHDV antibodies by a commercially available ELISA kit. A "very good" agreement was demonstrated when compared to a commercial ELISA kit (Cohen's kappa value=0.832). Samples which caused disagreement between the two assays originated from wildlife which highlights the need for further validation by using serum samples from wild animals.

Whole-Genome Sequence of a Suid Herpesvirus-1 Strain Isolated from the Brain of a Hunting Dog in Italy.

Suid herpesvirus-1 (SHV-1), a DNA virus of the family Herpesviridae, causes a severe and fatal disease in a wide range of mammals. Here, we report the whole-genome sequence of an SHV-1 isolated in Italy in 2014 from the brain of a hunting dog that suffered from an acute and severe disease.

Bluetongue virus surveillance in the Islamic Republic of Mauritania: Is serotype 26 circulating among cattle and dromedaries?

In March 2013, EDTA-blood and serum samples were collected from 119 cattle and 159 dromedaries at the slaughterhouse of Nouakchott, the capital city of the Islamic Republic of Mauritania. Serum samples were screened for the presence of Bluetongue (BT) antibodies by competitive ELISA (cELISA). Positive samples were then tested by serum-neutralization (SN) to determine BTV serotype. RNA from blood samples was first tested by a genus-specific quantitative RT-PCR assay which is able to detect all 27 existing BTV serotypes (RT-qPCR1-27). Positive samples were further screened by a RT-qPCR assay which, instead, is able to detect the classical 24 BTV serotypes only (RT-qPCR1-24). Of the 278 serum samples tested, 177 (mean=63.7%; 95% CI: 57.9%-69.1%) resulted positive by cELISA. Of these, 69 were from cattle (mean=58.0%; 95% CI: 49.0%-66.5%) and 108 from dromedaries (mean=67.9%; 95% CI: 60.3%-74.7%). BTV-26 neutralizing antibodies were by far the most frequently found as they were detected in 146 animals with titres ranging from 1:10 to 1:80. Out of 278 blood samples, 25 (mean=9.0%; 95% CI: 6.2%-12.9%) were found positive for BTV by RT-qPCR1-27, 20 (mean=16.8%; 95% CI: 11.2%-24.6%) were from cattle and 5 (mean=3.1%; 95% CI: 1.4%-7.1%) from dromedaries. When tested by RT-qPCR1-24 the 25 BTV positive samples were negative. Unfortunately, no genetic information by molecular typing or by next generation sequencing has been obtained as for the very low levels of RNA in the blood samples.

Innocuity of a commercial live attenuated vaccine for epizootic hemorrhagic disease virus serotype 2 in late-term pregnant cows.

Epizootic hemorrhagic disease (EHD) is an arthropod-borne infectious viral disease sustained by the epizootic hemorrhagic disease virus (EHDV). The only commercially available and currently used vaccines are manufactured for EHDV-2 in Japan, either live or inactivated vaccines. In this study we tested the innocuity for fetuses of the live attenuated EHDV-2 vaccine in five late-term pregnant cows. Whole blood and serum samples were collected from dams and screened for the presence of EHDV-2 RNA, infectious virus and antibodies. After calving, whole blood and serum samples collected from calves, before and after colostrum intake, were also tested for antibodies and for virus detection. In dams, neither fever nor clinical signs were observed. All of them seroconverted and a strong humoral response was detected throughout the sampling period. All blood samples tested negative for EHDV-2 except for one sample collected from a dam 11 days post-vaccination which tested positive at virus isolation at the third cell passage following two rounds of blind passages. Although they had free access to colostrum, calves tested serologically negative for EHDV-2 during the entire course of the experiment. Overall, the tested live attenuated vaccine can be safely administered to late-term pregnant cows as it was not demonstrated to cross the placental barrier. The safety of the live-attenuated vaccine is further confirmed by the emergence of Ibaraki virus in 2013 in Japan which is apparently not related to the spread of the vaccine strain currently used in Japan.