Schmallenberg virus detection in Culicoides biting midges in Spain: First laboratory evidence for highly efficient infection of Culicoides of the Obsoletus complex and Culicoides imicola.

Since Schmallenberg disease was discovered in 2011, the disease rapidly spread across Europe. Culicoides biting midges have been implicated as putative Schmallenberg vectors in Europe. The detection of Schmallenberg virus (SBV) in field collected Culicoides was evaluated through retrospective (2011-2012) collections and captures performed in 2013. This study represents the first detection of SBV in field collected Culicoides in Spain. Infectious midges were detected at the foothills of Pyrenees, Aramunt, in the summer 2012. All the specimens infected with Schmallenberg were of the species Culicoides obsoletus s.s. confirming its putative vector status in Spain. Experimental infection on field collected Culicoides provided evidence of atypical high efficiency for SBV vector infection and transmission potential in local populations of Culicoides imicola and in Culicoides of the Obsoletus complex. However, captured individuals of C. imicola were more susceptible to SBV infection than C. obsoletus s.l. (p < .001), with an infection ratio of 0.94 and 0.63, respectively. In contrast, a Culicoides nubeculosus colony appeared to be refractory to SBV infection.

Middle East respiratory syndrome coronavirus experimental transmission using a pig model.

Dromedary camels are the main reservoir of Middle East respiratory syndrome coronavirus (MERS-CoV), but other livestock species (i.e., alpacas, llamas, and pigs) are also susceptible to infection with MERS-CoV. Animal-to-animal transmission in alpacas was reported, but evidence for transmission in other species has not been proved. This study explored pig-to-pig MERS-CoV transmission experimentally. Virus was present in nasal swabs of infected animals, and limited amounts of viral RNA, but no infectious virus were detected in the direct contact pigs. No virus was detected in the indirect contact group. Furthermore, direct and indirect contact pigs did not develop specific antibodies against MERS-CoV. Therefore, the role of pigs as reservoir is probably negligible, although it deserves further confirmation.

A robust PCR for the differentiation of potential virulent strains of Haemophilus parasuis.

BACKGROUND: Haemophilus parasuis is the etiological agent of Glässer's disease in swine. H. parasuis comprises strains with heterogeneous virulence capacity, from non-virulent to highly virulent. Determination of the pathogenic potential of the strains is important for diagnosis and disease control. The virulence-associated trimeric autotransporters (vtaA) genes have been used to predict H. parasuis virulence by PCR amplification of their translocator domains. Here, we report a new and improved PCR designed to detect a different domain of the vtaA genes, the leader sequence (LS) as a diagnostic tool to predict virulence. METHODS: A collection of 360 H. parasuis strains was tested by PCR with LS specific primers. Results of the PCR were compared with the clinical origin of the strains and, for a subset of strains, with their phagocytosis and serum resistance using a Chi-square test. RESULTS: LS-PCR was specific to H. parasuis, and allowed the differential detection of the leader sequences found in clinical and non-clinical isolates. Significant correlation was observed between the results of the LS-PCR and the clinical origin (organ of isolation) of the strains, as well as with their phagocytosis and serum susceptibility, indicating that this PCR is a good predictor of the virulence of the strains. In addition, this new PCR showed a full correlation with the previously validated PCR based on the translocator domain. LS-PCR could be performed in a wide range of annealing temperatures without losing specificity. CONCLUSION: This newly described PCR based on the leader sequence of the vtaA genes, LS-PCR, is a robust test for the prediction of the virulence potential of H. parasuis strains.

Distribution and genetic characterization of Enterovirus G and Sapelovirus A in six Spanish swine herds.

The prevalence of Enterovirus G (EV-G) and Sapelovirus A (PSV-1) was investigated in Spanish swine herds by means of cross-sectional studies. Faecal samples from clinically healthy pigs were collected from six farms, and analysed by RT-PCR. The results indicated a high prevalence of EV-G detected in nearly all the animals older than 3 weeks of age. Otherwise, PSV-1 was only detected in 3-week-old piglets from one of the farms. Genetic analyses performed in the VP1 region of the EV-G indicated circulation of diverse strains in the same farm, related to genotypes G1, G2, G3, G4, G6, G9, G12, G13 and G14. Moreover, co-infection of several PSV-1 variants in the same animal was evident, typical of viral quasispecies. Evolutionary pressure analysis indicated that microevolution of PSV-1 seems to be driven by negative selection. This study gives further insights in the epidemiology of EV-G and PSV-1.

[Selective Laser Trabeculoplasty in the treatment of chronic open-angle glaucoma: retrospective analysis 12 years after treatment in a cohort of 28 patients]. / Trabéculoplastie sélective au Laser Selecta (SLT) dans le traitement du glaucome chronique à angle ouvert : analyse rétrospective 12 ans après traitement d'une cohorte de 28 patients.

PURPOSE: To determine long-term efficacy of selective Laser Trabeculoplasty (SLT) over 12 years in chronic open-angle glaucoma (OAG) patients. METHODS: In this retrospective study, all patients treated by SLT between 1997 and 1999 for OAG were included and followed up every 6 months. The procedure was performed with a Coherent Selecta 7000 Nd:YAG with 100 ± 10 non overlapping 400 µm spots over 360 degrees centered on the trabecular meshwork. Patients were excluded in the case of prior filtration surgery or Argon laser trabeculoplasty. Our primary study parameter was the number of patients requiring filtration surgery within the follow-up period. Our secondary parameters were intraocular pressure (IOP) and SLT-related complications. RESULTS: We included 46 eyes of 28 patients. The 12-year success rate was 26.1%. Thirty-nine percent of all eyes underwent filtration surgery (failure) during the follow-up period, and 34.8% were lost to follow-up. In the pigmentary glaucoma (PG) subgroup, the 12-year success rate was 16%, while it was 37.5% in the Primary OAG subgroup. The overall mean IOP was 22.8 mm Hg (D.S. 3.78) prior to laser, 16.08 mm Hg (D.S. 2.7) at 1 year and 15 mm Hg (D.S. 1.8) at 12 years. The mean number of medications was 1.6 (D.S. 0.8) prior to SLT, 1.36 (D.S. 0.8) at 1 year, and 1.3 (D.S. 1.2), 12 years after SLT respectively. No patients had a second SLT treatment. No significant complications occurred during follow-up. CONCLUSION: Selective laser trabeculoplasty may at times be a useful resource to lower IOP in patients with OAG. Nonetheless, the failure rate is significant especially in PG, which requires confirmation by larger prospective studies.

Microarray analysis of mediastinal lymph node of pigs naturally affected by postweaning multisystemic wasting syndrome.

Postweaning multisystemic wasting syndrome (PMWS) is one of the pig diseases with major economic impact worldwide. Clinical, pathologic and some immunologic aspects of this disease are relatively well-known, but the molecular mechanisms underlying the pathogenic mechanisms of the disease are still poorly understood. The objective of the present study was to investigate the global transcriptome changes in the mediastinal lymph nodes from pigs naturally affected by PMWS, as well as healthy counterparts, using the Affymetrix Porcine Genechip(®). From 366 transcripts showing significant differential abundance in the PMWS group of pigs relative to healthy animals, 229 showed higher and 137 lower abundance. A relative increased abundance of mRNAs coded by a large set of genes involved in the inflammatory responses (e.g. cytokines, acute phase proteins, and respiratory burst) was observed in PMWS affected pigs. The Gpnmb and Lgals3 genes, which have antagonistic functions in regulation of inflammatory processes, showed high mRNA levels in diseased pigs. The complement system was altered by PMWS, notably by the lower levels of Cr1 mRNA, which might favour both complement deposition and secondary infections by impairing phagocytosis. Decreased mRNA abundance of several genes involved in lymphocyte activation/differentiation, such as Cd79b, Cd19, Cd21 and MybL1, and the high level of Vsig4 mRNA, which can compromise the activation of residing T-cells, pointed towards a defective adaptive immunity. This is the first study on gene expression in pigs naturally affected by PMWS. The present results allowed identifying potential mechanisms underlying the inflammation and lymphocyte depletion in lymphoid tissues by complement mediated damage and immunosuppression, which are key features of PMWS.

Exploratory study on the transcriptional profile of pigs subclinically infected with porcine circovirus type 2.

The aim of this exploratory work was to use a microarray-based approach to study the global transcriptome profile of caesarean-derived, colostrum-deprived (CDCD) piglets experimentally infected with porcine circovirus type 2 (PCV2). PCV2-inoculated piglets developed a subclinical infection, as confirmed by serology, in situ hybridization and quantitative PCR. Total RNA from mesenteric lymph nodes and lungs was obtained by duplicate from 2 control and 2 PCV2-inoculated piglets and was hybridized to Affymetrix Porcine GeneChip. Among the 24,123 probesets studied, 25 and 33 were found to be significantly differentially expressed (DE) between control and PCV2 groups for mesenteric lymph node and lung, respectively. Most up-regulated genes in PCV2 group were closely related to the immune response, such as cytokines (CCL4L, CXCL9, CXCL11), MHC binding molecules (TCRalpha, CD8alpha), immunoglubulins (IgG) and T cell activation (LCK, KLRK1, RASSF2, GBP2). Quantitative real-time PCR was performed to verify the microarray results. Therefore, from a transcriptional point of view, PCV2-inoculated pigs were apparently able to activate a cell-mediated response and develop PCV2-specific antibodies, which probably led to a subclinical infection. The results from this study indicate that a microarray based approach is a helpful tool in order to better understand the pathogenesis of PCV2 infection.

[Postoperative pain in retinal detachment surgery]. / Douleur postopératoire dans la chirurgie de décollement de rétine.

Postoperative pain in retinal detachment surgery is frequent but it is often underestimated. The aim of this study was to determine the incidence of postoperative pain after retinal detachment surgery and to identify its predictive factors in a longitudinal study. We included 106 patients operated for retinal detachment surgery using an endo-ocular or exo-ocular approach with general anesthesia. Postoperative monitoring for 24 h evaluated the intensity of pain using a numerical scale. The possible predictive factors of this pain were studied: ocular antecedents, premedication, total amount of morphine used, type of surgery, duration of surgery, and vomiting. The incidence of postoperative pain was 57.5%, 56% of which was intense pain. Postoperative pain was greatest during the first 4 h. The predictive factors of this pain revealed by bivariate analysis of the data were the type of surgery and vomiting. The incidence and intensity of postoperative pain after retinal detachment surgery remain high. Pain management requires postoperative treatment of vomiting as well as the development of the endo-ocular surgery and locoregional anesthesia techniques.

[Emphysematous pyelonephritis: a case report]. / La pyélonéphrite emphysémateuse: à propos d'un cas guéri.

Diabetic patients with urinary tract infections had a certain risk of developping emphysematous pyelonephritis with gas producting bacteria. This disease was accompagned with high mortalité. We will report one case who was treated successfully by high dose antibiotic regim and emergency nephrectomy.

IFN-gamma as an indicator of successful immunization of goats vaccinated with a killed Cowdria ruminantium vaccine.

Cowdria ruminantium-induced production of IFN-gamma was measured by ELISA on a weekly basis during the course of vaccination with killed organisms emulsified in ISA50. Upon challenge, all (3/3) vaccinated animals that gave the lowest IFN-gamma response died of peracute cowdriosis. On the other hand, only one of three animals showing high IFN-gamma responses to vaccination died, but with a delay of 4 days in comparison with naïve controls. Thus, there seems to be a threshold level of IFN-gamma below which the probability for vaccinated animals to survive a lethal challenge is very low. During challenge, a much lower, but still physiologically meaningful production of IFN-gamma was detected using the 24-hour whole blood assay on day 5 after infection in animals controlling the infection. In contrast, IFN-gamma production was absent or negligible in naïve and vaccinated animals that died within 8-10 days after infection. Although these results need to be validated on a larger number of animals, they strongly suggest that IFN-gamma is a useful indicator of protective immunity in animals immunized with killed COWDRIA:

Cowdria ruminantium antigens of around 15 kDa are potent inducers of IFN-gamma.

Cellular responses induced in two Creole goats by vaccination with killed Cowdria ruminantium (Cowdria) were confirmed by IFN-gamma production and interleukin-2 receptor (IL-2R) expression. Both CD4+ and CD8+ but not WC1+ T cells showed a substantial increase in cell surface expression of IL-2R molecules in response to whole Cowdria lysate. Cowdria (Welgevonden strain) proteins were fractionated using continuous-flow electrophoresis and tested for their ability to induce IFN-gamma production by PBMC collected three weeks after the first inoculation and one week after the booster injection. Pooled fractions of around 15, 22, and 24 kDa were found to induce significant IFN-gamma production in both vaccinated animals on one of the two occasions. Antigens of around 15 kDa induced substantially higher IFN-gamma production than any other fractions in both animals. These pilot experiments pave the way towards the identification of proteins/genes that have potential for the development of a recombinant vaccine against heartwater.

[Malignant non-Hodgkin's lymphoma presenting with cardiac tamponade]. / Tamponnade cardiaque révélatrice d'un lymphome malin non hodgkinien.

Malignant non-Hodgkin's lymphoma rarely involves the heart. However, all cardiac structures may be invaded, although the myocardium and pericardium are affected more often than the valvular structures. Diagnosis is important because treatment must be adapted. The cardiac signs are usually non-specific and lymphomas presenting in this way have only occasionally been reported. The authors report two cases in which malignant non-Hodgkin's lymphoma presented with cardiac tamponade.

Ehrlichia ruminantium major antigenic protein gene (map1) variants are not geographically constrained and show no evidence of having evolved under positive selection pressure.

In a search for tools to distinguish antigenic variants of Ehrlichia ruminantium, we sequenced the major antigenic protein genes (map1 genes) of 21 different isolates and found that the sequence polymorphisms were too great to permit the design of probes which could be used as markers for immunogenicity. Phylogenetic comparison of the 21 deduced MAP1 sequences plus another 9 sequences which had been previously published did not reveal any geographic clustering among the isolates. Maximum likelihood analysis of codon and amino acid changes over the phylogeny provided no statistical evidence that the gene is under positive selection pressure, suggesting that it may not be important for the evasion of host immune responses.

Daily delivery of dietary nitrogen to the periphery is stable in rats adapted to increased protein intake.

Dietary nitrogen was traced in rats adapted to a 50% protein diet and given a meal containing 1.50 g (15)N-labeled protein (HP-50 group). This group was compared with rats usually consuming a 14% protein diet and fed a meal containing either 0.42 g (AP-14 group) or 1.50 g (AP-50 group) of (15)N-labeled protein. In the HP group, the muscle nonprotein nitrogen pool was doubled when compared with the AP group. The main adaptation was the enhancement of dietary nitrogen transferred to urea (2.2 +/- 0.5 vs. 1.3 +/- 0.1 mmol N/100 g body wt in the HP-50 and AP-50 groups, respectively). All amino acids reaching the periphery except arginine and the branched-chain amino acids were depressed. Consequently, dietary nitrogen incorporation into muscle protein was paradoxically reduced in the HP-50 group, whereas more dietary nitrogen was accumulated in the free nitrogen pool. These results underline the important role played by splanchnic catabolism in adaptation to a high-protein diet, in contrast to muscle tissue. Digestive kinetics and splanchnic anabolism participate to a lesser extent in the regulation processes.

The use of the tetrazolium salt XTT for the estimation of biological activity of activated sludge cultivated under steady-state and transient regimes.

The tetrazolium salt 3'-¿1-[(phenylamino)-carbonyl]-3, 4-tetrazolium¿-bis (4-methoxy-6-nitro) benzenesulfonic acid hydrate (XTT) was used as a tool for estimating the activity of the electron transport system (ETS) in activated sludge cultivated under steady-state and transient regimes in chemostat culture. Production of formazan by reduction of XTT depended on the initial concentration of the XTT following a saturation law and was proportional to live cell biomass. Addition of cyanide (KCN) to activated sludge gave an initial 1.5-fold increase in XTT reduction, while addition of 3,5-dichlorophenol (3,5-DCP) reduced this value drastically. At steady-state and transient regimes of an activated sludge chemostat, oxygen uptake rate (OUR) and XTT reduction rate were highly correlated and indicated significant variations depending on the growth conditions.