Developments in biologicals最新文献

In genome-wide association studies using single nucleotide polymorphisms (SNPs), typically thousands of SNPs are genotyped, whereas the number of phenotypes for which there is genomic information may be smaller. Atwo-step SNP (feature) selection method was developed, which consisted of filtering (using information gain), and wrapping (using naïve Bayesian classification). This was based on discretization of the continuous phenotypic values. The method was applied to chick early mortality rates (0-14 days of age) on progeny from 201 sires in a commercial broiler line, with the goal of identifying SNPs (over 5000) related to progeny mortality. Sires were clustered into two groups, low and high, according to two arbitrarily chosen mortality rate thresholds. By varying these thresholds, 11 different "case-control" samples were formed, and the SNP selection procedure was applied to each sample. To compare the 11 sets of chosen SNPs, predicted residual sum of squares (PRESS)from a linear model was used. Naive Bayesian classification accuracy was improved over the case without feature selection (from 50% to 90%). Seventeen SNPs in the best case-control group (with smallest PRESS) accounted for 31% of the variance among sire family mortality rates.

The pseudorabies virus (PrV), a porcine Alphaherpesvirus, is a good model for the study of virus-host cell dialog. As PrV has a strong tropism for mucous epithelial cells, we chose to follow in vitro the PrV time course-infection of porcine PK15 cells. The viral and cellular transcriptome modifications were simultaneously analysed using a combined SLA/PrV cDNA microarray, the porcine Qiagen-NRSP8 oligonucleotides microarray and real time quantitative PCR.Ahigh increase in viral gene expression was found from 4 h post-infection (PI), concomitantly to the first viral progeny and most viral genes were differentially expressed 12 h PI. No early global cellular shutoff was observed but many cellular genes were downregulated between 8 and 12 h PI, when UL41 transcripts encoding the virion shutoff protein, were first detected. Several genes involved in the MHC class I mediated antigenic pathway were downregulated including SLA-la, TAP1, TAP2, PSMB8 and PSMB9 genes. These results suggested that PrV prevents the viral antigen presentation by epithelial cells to cytotoxic T lymphocytes by decreasing transcription levels of SLA Ia mediated antigenic pathway genes. Other genes involved in the immune response, the apoptosis pathway, nucleic acid metabolism and cytoskeleton also appeared to be regulated during PrV infection. The combined approach will help to decipher host response evasion strategies developed by PrV and to study early cellular modifications.

Infectious diseases are an important cause of economic loss in the agri-food business. This study investigates indicators of bovine high (HR) and low (LR) immune response and their associated patterns of gene expression. Holstein cows were immunized to induce antibody (AMIR) and cell-mediated (CMIR) immune responses. Based on the results of enzyme-linked immunosorbent assay (ELISA) and delayed-type hypersensitivity (DTH), cows were ranked as HR, LR or average (AR) immune responders. For microarray analysis, phenotypic HR and LR status in both groups was confirmed and total RNA from blood mononuclear cells (BMCs) was obtained. RNA from a pool of AR cows was used as a common reference for hybridization to an in-house cDNAmicroarray. Results of microarray analysis showed transcriptional differences in several immune-related genes between the HR and LR groups. Genes identified as differentially expressed include transcription factors, cytokines, MHC, and TCR-related genes. These results can aid in the establishmentof selection programmes based on broad-based disease resistance, aimed at improving general health in cattle herds.

The Germplasm Evaluation (GPE) Project at the US Meat Animal Research Center (USMARC) is planned to produce about 3,000 calves per year in support of the following objectives: identification and validation of genetic polymorphisms related to economically relevant traits (ERT), estimation of breed and heterosis effects among 16 breeds for ERT, and estimation of genetic correlations among ERT and physiological indicator traits (PIT). Opportunities exist for collaboration in the development and collection of PIT phenotypes for disease resistance. Other areas of potential collaboration include detailed diagnosis (identification of disease causing organisms, etc.) of treated animals, collaborative development of epidemiological statistical models that would extract more information from the records of diagnoses and treatments, or pharmacogenetics. Concentrating a variety of different phenotypes and research approaches on the same population makes each component much more valuable than it would be individually.

Highly pathogenic (HP) avian influenza viruses (AIV) present an ongoing threat to the world poultry industry. In order to develop new AIV control strategies it is necessary to understand the underlying mechanism of viral infection at mucosal respiratory sites. Chicken and duck tracheal epithelial cells systems (TEC) were developed to study early host responses to AIV infection on TEC. Infection of 2 week-old chickens and ducks with the highly pathogenic AIV H5N1 Ck/Hong Kong/220/97 and Egret/Hong Kong/757.2/02 viruses together with TEC early responses to infection suggest the induction of differential innate immune responses. Growth curves indicated that although chicken and ducks TEC supported viral replication and re-infection, the capacity of the two viruses to replicate was not equal. A 42K probes chicken microarray system used to characterize differences in gene expression between chicken tracheal epithelial cells infected with these two highly pathogenic AIV identified expression of virus-specific molecular markers. The existence of dissimilar patterns of host gene expression as early as six hours post infection suggests that the differential growth characteristics of the two highly pathogenic AIV in tracheal epithelial cells is preceded by distinct host responses.

Porcine reproductive and respiratory syndrome virus (PRRSV) is a major pathogen of swine worldwide and causes considerable economic loss. The main target of infection is the porcine alveolar macrophage (PAM). Infection of PAMs by PRRSV causes significant changes in their function by mechanisms that are not understood. We have employed Serial Analysis of Gene Expression (SAGE) to examine the global expression of genes in PRRSV-infected PAMs. Total cellular RNAwas prepared from in vitro mock-infected and PRRSV strain VR-2332-infected PAMs at 0, 6, 12, 16 and 24 hours after infection, and subjected to SAGE analysis to obtain > 100,000 tags per time point. These sequences were processed to account for sequencing error before generating tag:count lists. These lists were deposited into a modified Identitag database for mapping to porcine and PRRSV genes. Identified unique mRNAtags were analyzed for their identity and relative abundance. Examination of the SAGE data indicated that there were changes in gene expression occurring in the PRRSV-infected PAMs over time post-infection. More than 400 unique tags with significantly altered expression levels were identified (p < 0.01 with Bonferroni correction). The validity and kinetics of expression of SAGE identified genes were evaluated using real-time RT-PCR.

Oral vaccination programmes conducted in rabies infected countries from Eastern Europe and Eurasia should not be restricted to foxes but should target other major rabies vectors such as dogs and raccoon dogs as well. The objective of this experimental trial was to assess the protection induced by the vaccine by challenging these different species, which had been previously vaccinated intramuscularly with the square V-RG baits (produced in the US). Different parameters were evaluated such as attractiveness of the baits and induction of neutralising antibodies as an indicator for immunogenicity and protection after rabies challenge. The acceptability of the square bait was satisfactory in dogs, foxes and raccoon dogs, confirming previous laboratory and field studies conducted with the rectangular baits. Only one vaccinated dog out of nine seroconverted after vaccination and among them one dog died of rabies. Eight of ten vaccinated foxes seroconverted after vaccination and survived the rabies challenge. All vaccinated raccoon dogs seroconverted after challenge and all survived the challenge. These trials demonstrated that the square presentation of the V-RG vaccine was attractive, immunogenic and efficacious.

In response to a Commission request, EFSA has carried out a quantitative assessment of the risk of rabies introduction into the UK, Ireland, Sweden, and Malta due to the movement of pets incubating rabies at the time of movement. The risk that a pet is incubating rabies at the time of first vaccination is equal to the prevalence of rabies-incubating pets in the population of origin. Following induction of protective immunity by vaccination, animals already incubating rabies will still develop clinical disease as a function of time after vaccination (termed type A risk). A waiting period will reduce this risk. Afew animals may not be protected after single-shot primary vaccination. Such animals may become infected during the waiting period after vaccination. The risk of becoming infected after the first vaccination (termed type B risk) depends on the prevalence and efficiency of vaccination. Serological testing can be used to identify non-immune pets (depending on test specificity) and will therefore reduce this risk accordingly. The type A and B risks were modelled as a function of the waiting period after vaccination and fitted to a non-linear model incorporating vaccination efficiency and test specificity. The model can be used to quantify the risk of moving pets from rabies infected areas and also to investigate the effect of different control measures. In quantitative terms, the type A risk constitutes by far the major risk. Therefore, a waiting period (defined as the time spent between vaccination and pet movement to the destined country) is the major effective measure to mitigate the risk of rabies introduction due to an animal being infected before primo-vaccination. Serological testing will only add significantly to risk reduction when waiting periods exceed 100 days. Within the EU, the rabies prevalence in most countries is so low that the risk can be considered negligible. However, for some countries the risk is non-negligible.

Rabies virus causes severe encephalitis that is invariably fatal for the victim. However, the contribution of the virus and the host to damage of the CNS is unclear. In order to investigate this we studied the neuropathology and CNS gene expression patterns in a murine model of rabies using a 'street' isolate RV61. This virus was derived from a human case of disease. In this model, infection of the CNS progresses rapidly following inoculation in the periphery, leading to extensive virus replication in the brain and the development of disease. However, previous studies have found little evidence of inflammation and lymphocyte infiltration in many regions of the CNS of infected mice. During the current study virus replication was detected in the dorsal root ganglia (DRG), spinal cord, brain and salivary gland at 11 days postinfection (dpi). Mononuclear cell infiltration was observed in the DRG and to a lesser extent, the spinal cord. Immunolabelling demonstrated that T-lymphocytes were the dominant population of infiltrating cells. Murine innate immune response gene transcripts were detected in the brain as early as 5 dpi. At 11 dpi, coincidentwith the onset of disease, elevated levels of mRNA transcripts were recorded for type-1 (alpha and beta) and type-2 interferon (gamma) and certain chemokines (CCL5 and CXCL10) with chemotactic properties for T-cells. We suggest that damage to the DRG and spinal cord could be due to a combination of both virus infection and the infiltration of T-cells.

The objective of this study was to obtain transgenic maize expressing the rabies virus glycoprotein (G) of the Vnukovo strain and to evaluate its immunogenicity in mice, by the oral route. The ubiquitin maize promoter fused to the whole coding region of the rabies virus G gene, and a constitutive promoter from cauliflowermosaic virus (CaMV)were used. Maize embryogenic callus were transformed with the above construct by biolistics. Regenerated maize plants were recovered and grown in a greenhouse. The presence of the G gene and its product was detected by PCR and western blot, respectively. The amount of G protein detected in the grains was approximately 1% of the total soluble plant protein. Transformed kernels containing 50 microg of G protein were given once by the oral route in adult mice (BALB-C strain). Challenge was undertaken at 90-days post-vaccination using a lethal dose of a vampire bat rabies virus (100 LD 50% in mice); vampire bats are one of the main reservoirs in Latin America. The edible vaccine induced viral neutralizing antibodies (VNA) which, protected mice 100% against challenge. The control group did not survive. The G protein of the Vnukovo strain expressed in transgenic maize may be considered as an oral immunogen against rabies, conferring cross-protection.