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The present study assessed the efficacy of vaccination against genotype 1 porcine reproductive and respiratory syndrome virus (PRRSV) in terms of reduction of the transmission. Ninety-eight 3-week-old piglets were divided in two groups: V (n=40) and NV (n=58) that were housed separately. V animals were vaccinated with a commercial genotype 1 PRRSV vaccine while NV were kept as controls. On day 35 post-vaccination, 14 NV pigs were separated and inoculated intranasally with 2 ml of a heterologous genotype 1 PRRSV isolate ("seeder" pigs, SP). The other V and NV animals were distributed in groups of 5 pigs each. Two days later, one SP was introduced into each pen to expose V and NV to PRRSV. Sentinel pigs were allocated in adjacent pens. Follow-up was of 21 days. All NV (30/30) became viremic after contact with SP while only 53% of V pigs were detected so (21/40, p<0.05). Vaccination shortened viremia (12.2±4 versus 3.7±3.4 days in NV and V pigs, respectively, p<0.01). The 50% survival time for becoming infected (Kaplan-Meier) for V was 21 days (CI95%=14.1-27.9) compared to 7 days (CI95%=5.2-8.7) for NV animals (p<0.01). These differences were reflected in the R value as well: 2.78 (CI95%=2.13-3.43) for NV and 0.53 (CI95%=0.19-0.76) for V pigs (p<0.05). All sentinel pigs (10/10) in pens adjacent to NV+SP pens got infected compared to 1/4 sentinel pigs allocated contiguous to a V+SP pen. These data show that vaccination of piglets significantly decrease parameters related to PRRSV transmission., (Copyright © 2014 Elsevier B.V. All rights reserved.)

Bovine plasma was inoculated with porcine epidemic diarrhea virus (PEDV) at an average final titer of 4.2 log10 TCID50/mL to determine the effect of spray drying on viral inactivation. Using a laboratory scale drier, inoculated plasma was spray dried at 200 °C inlet temperature and either 70 or 80 °C throughout substance. Both liquid and dried samples were subjected to three passages on VERO cell monolayers to determine PEDV infectivity. Results indicated liquid samples contained infective virus, but none of the spray dried samples were infectious. Also, survivability of PEDV inoculated on spray dried bovine plasma (SDBP) and stored at 4, 12 or 22 °C was determined for 7, 14 and 21 days. Commercial SDBP powder was inoculated with PEDV to an average final titer of 2.8 log10 TCID50/g. Five samples per time and temperature conditions were subjected to three passages on VERO cell monolayers to determine PEDV infectivity. The virus was non-infectious for all samples stored at 22 °C at 7, 14 and 21 days. PEDV was infective in 1 out of 5 samples stored at 12 °C at 7 days, but none of the samples stored for 14 and 21 days were infectious in cell culture. For samples stored at 4 °C, 4 out of 5 samples were infectious at 7 days, 1 out of 5 samples were infectious at 14 days, but none were infectious at 21 days. In summary, PEDV was not infectious on cell culture within 7 days when stored at room temperature and within 21 days when stored at refrigerated temperature., (Copyright © 2014 Elsevier B.V. All rights reserved.)

Bluetongue (BT) is an infectious disease of wild and domestic ruminants caused by bluetongue virus (BTV). BTV-4 spread through southern Spain from 2004 to 2006, whereas a BTV-1 outbreak that started in southern Spain in 2007 is still ongoing. Vaccination and movement restriction regulations are applied to domestic ruminants to control BT, but the potential reservoir role of wild European ungulates has not been clarified so far. The aim of this study was to describe the epidemiology of BTV in the wild free-ranging red deer (Cervus elaphus) population of Cabañeros National Park (CNP) in central Spain during the BTV-4 and BTV-1 epizootics, assessing the potential role of this deer population as a BTV reservoir. Blood samples from 2885 (2542 adults, 208 calves and 135 undetermined) wild red deer were collected from 2005 to 2010 in CNP and surrounding hunting estates. All sera were tested for antibodies against the BTV VP7 protein by ELISA. Ninety-four of the ELISA-positive samples were analysed by serum neutralization to detect BTV-4 and BTV-1 specific antibodies, and 142 blood samples were analysed by RT-PCR for BTV RNA. A total of 371 (12.9%) out of the 2,885 deer (35/208 calves, 307/2,542 adults, and 29/135 undetermined) were positive for antibodies against BTV. Prevalence increased in adult deer from 2005-2006 to 2008-2009, declining thereafter. No positive samples for BTV-1 were found by serum neutralization, whereas 43 deer (38 adults, four calves and one undetermined) were positive for BTV-4 specific antibodies. No BTV RNA positive deer were found by RT-PCR. Antibody detection throughout the study period suggests a maintained circulation of BTV in red deer. However, the lack of BTV RNA detection suggests a minor transmission risk to livestock., (Copyright © 2012 Elsevier B.V. All rights reserved.)

Infectious bronchitis (IB) is a worldwide disease affecting chickens of all ages and causing important economic losses in poultry industry. Despite being one of the predominant IB virus (IBV) serotype in several European countries, slightly is known about pathogenesis and pathogenicity of Italy 02 serotype. In this study chicks and old hens were infected by oculo-nasal route with Italy 02 serotype. Clinical signs, gross and microscopic findings were evaluated, viral nucleic acid detection was assessed by in situ hybridization (ISH) in several tissues and viral RNA was detected by RT-PCR in trachea, kidney and nasal and cloacal swabs. Italy 02 serotype was demonstrated to cause severe respiratory and renal damage in one-day old chicks but not in adult hens in which only respiratory disease and drop in egg production was observed. The use of ISH technique demonstrated the presence of viral RNA in nasal turbinates prior to trachea, but more consistent and longer replication periods in enterocytes of lower gastrointestinal tract. The detection of viral nucleic acid in gut by RT-PCR was consistent and more persistent viral shedding was detected in faeces than in nasal exudates. We describe a complete update of IBV distribution in tissues by the use of molecular techniques and we also provide and in-depth pathological characterization of the new Italy 02 IBV serotype. Furthermore, new data about IBV pathogenesis essential in field control is afforded., (Copyright © 2011 Elsevier B.V. All rights reserved.)

Red deer (Cervus elaphus) is a widespread and abundant species susceptible to bluetongue virus (BTV) infection. Inclusion of red deer vaccination among BTV control measures should be considered. Four out of twelve BTV antibody negative deer were vaccinated against serotype 1 (BTV-1), and four against serotype 8 (BTV-8). The remaining four deer acted as unvaccinated controls. Forty-two days after vaccination (dpv), all deer were inoculated with a low cell passage of the corresponding BTV strains. Serological and virological responses were analyzed from vaccination until 28 days after inoculation (dpi). The vaccinated deer reached statistically significant (P<0.05) higher specific antibody levels than the non vaccinated deer from 34 (BTV-8) and 42 (BTV-1) dpv, maintaining stable neutralizing antibodies until 28 dpi. The non vaccinated deer remained seronegative until challenge, showing neutralizing antibodies from 7 dpi. BTV RNA was detected in the blood of the non vaccinated deer from 2 to 28 dpi, whereas no BTV RNA was found in the vaccinated deer. BTV was isolated from the blood of non vaccinated deer from 7 to 28 dpi (BTV-1) and from 9 to 11 dpi (BTV-8). BTV RNA could be identified by RT-PCR at 28 dpi in spleen and lymph nodes, but BTV could not be isolated from these samples. BT-compatible clinical signs were inapparent and no gross lesions were found at necropsy. The results obtained in the present study confirm that monovalent BTV-1 and BTV-8 vaccines are safe and effective to prevent BTV infection in red deer. This finding indicates that vaccination programs on farmed or translocated red deer could be a useful tool to control BTV., (Copyright © 2011 Elsevier B.V. All rights reserved.)

Genetic diversity of porcine reproductive and respiratory syndrome virus (PRRSV) has been based on ORF5/GP5 and ORF7/N protein variations. Complete viral genome studies are limited and focused on a single or a few set of strains. Moreover, there is a general tendency to extrapolate results obtained from a single isolate to the overall PRRSV population. In the present study, six genotype-I isolates of PRRSV were sequenced from ORF1a to ORF7. Phylogenetic comparisons and the variability degree of known linear B-epitopes were done considering other available full-length genotype-I sequences. Cytokine induction of all strains was also evaluated in different cellular systems. Non structural protein 2 (nsp2) was the most variable part of the virus with 2 out of 6 strains harboring a 74 aa deletion. Deletions were also found in ORF3 and ORF4. Phylogenetic analyses showed that isolates could be grouped differently depending on the ORF examined and the highest similarity with the full genome cluster was found for the nsp9. Interestingly, most of predicted linear B-epitopes in the literature, particularly in nsp2 and GP4 regions, were found deleted or varied in some of our isolates. Moreover, 4 strains, those with deletions in nsp2, induced TNF-α and 3 induced IL-10. These results underline the high genetic diversity of PRRSV mainly in nsp1, nsp2 and ORFs 3 and 4. This variability also affects most of the known linear B-epitopes of the virus. Accordingly, different PRRSV strains might have substantially different immunobiological properties. These data can contribute to the understanding of PRRSV complexity., (Copyright © 2011 Elsevier B.V. All rights reserved.)

A cross-sectional study was carried out to assess the prevalence and circulation of bluetongue virus (BTV) in Spanish ibexes (Capra pyrenaica hispanica). A total of 770 sera samples, 380 blood samples and 34 spleen samples were collected between 2006 and 2009 in Andalusia (southern Spain), a region and time period with a wide circulation of BTV in livestock. Thirty-one out of 770 (4.0%; CI(95%): 2.6-5.4) sera samples analyzed by ELISA showed antibodies against BTV. Twenty-four out of 31 seropositive samples were tested against BTV serotypes 1, 4 and 8 by serum neutralization test (SNT). Neutralizing antibodies against BTV-1 and BTV-4 were detected in seven and ten animals, respectively, four of them showed neutralizing antibodies to both serotypes. The animals seropositive to BTV-4 were sampled between 2006 and 2008, while BTV-1 circulation was confirmed in ibexes sampled between 2007 and 2009. None of the ibexes presented neutralizing antibodies against BTV-8. Statistically significant differences were found among regions and years, which is in coincidence with what occurred in domestic ruminants. There were no statistically significant differences between sexes, age classes and habitats (captivity vs. free-living). BTV RNA was not found in any of the 380 blood samples analyzed. However, BTV-1 RNA was detected from spleen in one Spanish ibex from Málaga province in August 2008. This finding evidences the presence of BTV-1 in Spanish ibex in a municipality where BT outbreaks were not detected in domestic ruminants during that period. Results of the present study show that Spanish ibexes were exposed and responded serologically to both BTV-1 and BTV-4. The low seroprevalence obtained suggests that Spanish ibex is not a relevant species in the dissemination of BT. However, the detection of BTV-1 RNA and the presence of seropositive ibexes in areas where BT outbreaks were not detected in livestock, could not exclude a significant role in the epidemiology of BTV in certain areas., (Copyright © 2010 Elsevier B.V. All rights reserved.)

The objective of the present work was to ascertain the date in which hepatitis E virus (HEV) was introduced in the Spanish pig population. For this, a serological retrospective study was carried out using archived sera. A total of 2871 serum samples gathered between 1985 and 1997 and collected in 208 farms of Spain were tested for anti-HEV IgG by an in-house ELISA. Of the 2871 sera analyzed by ELISA, 1390 were positive for anti-HEV antibodies (48.4%, 95% CI: 46.9-49.9%) and that corresponded to 204/208 farms (98%, 95% CI: 96.1-99.9%) having at least one positive pig. Our results show that HEV was present and widespread in Spanish swine farms at least since 1985. Any significant changes in prevalence were detected from 1 year to another and therefore, HEV infection in swine should be considered endemic in Spain.

In March of 1991, a disease that affected pregnant sows and caused a high mortality in unweaned piglets was detected in Spain. Based on the clinical signs observed, mystery swine disease, which had been described recently in Germany, Holland and Belgium, was suspected. From the samples obtained from the affected farm, a filtrable agent (0.22 micron) was isolated on cell culture. It produced cytopathic effects, its replication was intracytoplasmic, it was sensitive to chloroform, and cross-reacted with a Lelystad reference serum. When inoculated into pregnant sows, the agent produced inappetence for 2-4 days, without hyperthermia. One of the sows aborted at 100 days of gestation; the two others had delayed parturitions (days 115 and 116). There was a mixture of healthy piglets, mummified fetuses, stillbirths and weak piglets. Microscopic examination of the lungs of healthy piglets killed at 8 and 12 days of life revealed the presence of interstitial pneumonia. The sera from the three sows at 39 days after infection cross-reacted with the Lelystad virus (titres > or = 1/640), whereas pre-inoculation sera did not recognize it (titres < or = 1/10). This is the first report from Spain of the isolation of an agent (antigenically related to the Lelystad virus), capable of reproducing the disease previously designated as mystery swine disease.