Transcription profiling of sheep terminal ileum with 3 defined groups of MAP infection
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ABSTRACT: Comparative expression profiles of 3 defined groups of Mycobacterium avium subspecies paratuberculosis (MAP) infected terminal ileum from sheep. Defined by histopathlogy and IS900 real-time PCR as multibacillary, paucibacillary and asymptomatic sheep. Six sheep per group provided biological and technical replicates on the RIGUA custom array (Watkins et al., 2008).
Project description:Salmon pancreas disease, caused by salmonid alphavirus (SAV), is an economically important disease affecting farmed Atlantic salmon (Salmo salar L.) in Europe, with outbreaks reported in Scotland, Norway, and Ireland. A microarray-based study was performed to evaluate the host transcriptomic response during the early stages of an experimentally induced salmonid alphavirus 1 (SAV 1) infection in Atlantic salmon (Salmo salar L.) Head kidney was sampled from five fish PD infected Atlantic salmon parr and uninfected controls on days 1, 3 and 5 post injection (d.p.i). RNA from tissue samples was amplified and interrogated using the 17k TRAITS / SGP cDNA microarray, with results validated by SYBR green real-time PCR. The greatest number of significantly differentially expressed genes was recorded on day 3 p.i. These were found to be mainly associated with immune and defence mechanisms including genes involved in interferon I & II pathways and major histocompatibility complex class I & II responses. The expression of genes associated with apoptosis (BcL2 and caspase 3/7) and cellular stress (heat shock protein) were also found to differ significantly between infected and uninfected individuals as were genes involved in inhibiting viral attachment and replication, such as ubiquitin, serum myeloid, and and viperin.
Project description:Concerns have arisen recently over the possible environmental effects of human pharmaceuticals. Although acute toxicities are low, the continuous discharge of pharmaceuticals into the aquatic environment, coupled with the fact that such compounds are selected for use on the basis of a strong pharmacological effect, means that sublethal effects on non-target organisms need to be seriously considered. The juvenile stages of Atlantic salmon are present in many northern European rivers which are also used for the discharge of domestic wastewaters likely to contain pharmaceuticals. One year old salmon parr were exposed to an environmentally relevant concentration (5µg·/ L) of the antidepressant drug carbamazepine for five days and changes of mRNA expression in brain tissues were investigated by means of a custom 17k Atlantic salmon cDNA microarray. The TRAITS 17K Atlantic salmon cDNA microarray was employed. A dual-labelled experimental design was employed for the microarray hybridisations. Each experimental cDNA sample (Cy3 labeled) was competitively hybridised against a common pooled-reference sample (Cy5 labeled). The entire experiment comprised 10 hybridisations - 2 states (CBZ exposed / unexposed) × 1 time-point ( at 5 days) × 5 biological replicates (males only). Hybridisations were undertaken concurrently.
Project description:Infectious pancreatic necrosis (IPN) is a serious viral disease that causes significant economic losses in salmon aquaculture. To characterize the host-pathogen relationship in IPN, we analysed transcriptional profiles of salmon head kidney (SHK-1) cells infected with infectious pancreatic necrosis virus (IPNV) at three timepoints over six days (at 1, 3 & 6 days post infection. The transcriptome was investigated using the TRAITS / SGP 16950-feature Atlantic salmon cDNA microarray, which is enriched for genes with functions related to the immune response.
Project description:The altitude gradient limits the growth and distribution of alpine plants.Alpine plants have developed strategies to survive the extremely cold conditions prevailing at high altitudes; however, the mechanism underlying the evolution of these strategies remains unknown. The alpine plant Potentilla saundersiana is widespread in the Northwestern Tibetan Plateau. In this study, we conducted a comparative proteomics analysis to investigate the dynamic patterns of protein expression of P. saundersiana located at five different altitudes. We detected and functionally characterized 118 differentially expressed proteins. Our study confirmed that increasing levels of antioxidant proteins, and their respective activities, and accumulation of primary metabolites, such as proline and sugar, confer tolerance to the alpine environment in P. saundersiana. Proteins species associated with the epigenetic regulation of DNA stability and post-translational protein degradation were also involved in this process. Furthermore, our results showed that P. saundersiana modulated the root architecture and leaf phenotype to enhance adaptation to alpine environmental stress through mechanisms that involved hormone synthesis and signal transduction, particularly the cross-talk between auxin and strictosidine. Based on these findings, we conclude that P. saundersiana uses multiple strategies to adapt to the high-altitude environment of the Northwestern Tibetan Plateau.
Project description:There is an increasing drive to replace fish oil (FO) in finfish aquaculture diets with vegetable oils (VO), driven by the short supply of FO derived from wild fish stocks. Little is known of the consequences for fish health after such substitution. The effect of dietary VO on hepatic gene expression was determined in Atlantic salmon (Salmo salar) byg a cDNA microarray analysis. Post-smolt farmed salmon were reared for x weeks on diets where the FO component of the feed was replaced with one of three different VOs - rapeseed (RO), soybean (SO) or linseed (LO). RNA from five fish fed on each diet was extracted. A total of 20 cDNA microarray hybridisations - TRAITS / SGP Atlantic salmon 17k feature cDNA microarray - were performed - 4 diets (three VO + FO control) x 5 individuals - using a common pooled reference control design. Data were obtained from 19 of the 20 hybridisations.
Project description:In patients undergoing thyroidectomy one of the goals of post-surgical follow-up is the correct maintenance of thyroid hormone (TH) levels by replacement therapy with levo-thyroxine (LT4). Nevertheless, some patients complain of symptoms of hypothyroidism (memory loss, weight gain, fatigue, depression, and reduced quality of life) despite values of thyroid hormones get the physiological range. Since this recurrent issue, we aim to compare serum proteomic profiles of LT4-euthyroid thyroidectomized patients with stable or reduced post-operative quality of life. Proteomic analysis highlights a different protein profile in serum of patients with reduced quality of life compared to the other although both resulted euthyroid. Differential proteins are involved in coagulation processes, complement system activation and in lipoprotein particles remodeling, that together lead to a pro-inflammatory response. Moreover, a particular protein isoform pattern of apolipoprotein A1 and alpha 1 antitrypsin, was also detected. This study suggests that LT4 replacement therapy might restore euthyroid conditions in thyroidectomized patients but in some cases without re-establish body tissues euthyroidism. This condition is reflected by the serum protein profile that shows a behavior similar to overt hypothyroidism.
Project description:Data analysis. Spot detection and matching were performed with a comparative cross analysis of all the gels using DeCyder software v.6.5 (GE Healthcare). 178 spots were selected based on 1.15-fold for protein ratio cut-off, allowing for the appearance of the spots in 23 out of 28 gels (69 out of 84 total images). Data from 95 spots were submitted. 93 spots were identified with high confidence. Spot picking and Trypsin digestion. The spots of interest were picked up by Ettan Spot Picker (GE Healthcare) based on the in-gel analysis and spot picking design by DeCyder software. The gel spots were washed a few times then digested in-gel with modified porcine trypsin protease (Promega, Fitchburg, WI). The digested tryptic peptides were desalted using a Zip-tip C18 (Millipore, Billerica, MA). Peptides were eluted from the Zip-tip with 0.5 uL of matrix solution (alpha-cyano-4-hydroxycinnamic acid, 5 mg/mL in 50% acetonitrile, 0.1% trifluoroacetic acid, 25mM ammonium bicarbonate) and spotted on a MALDI plate. Mass Spectrometry. MALDI-TOF MS and TOF/TOF tandem MS/MS were performed on AB SCIEX TOF/TOF 5800 System (AB SCIEX). MALDI-TOF mass spectra were acquired in reflectron positive ion mode, averaging 4000 laser shots per spectrum. TOF/TOF tandem MS fragmentation spectra were acquired for each sample, averaging 4000 laser shots per fragmentation spectrum on each of the 7-10 most abundant ions present in each sample (excluding trypsin autolytic peptides and other known background ions). Database search. Both the resulting peptide mass and the associated fragmentation spectra were submitted to GPS Explorer workstation equipped with MASCOT search engine (Matrix Science, Boston, MA) to search the Swiss-Prot database. Searches were performed without constraining protein molecular weight or isoelectric point, with variable carbamidomethylation of cysteine and oxidation of methionine residues, and with one missed cleavage also allowed in the search parameters. Candidates with either protein score C.I.% or Ion C.I.% greater than 95 were considered significant. When multiple IDs were significant for a given spot, the selection was made by evaluating apparent molecular weight, isoelectric point, the location of the spot in the gel, and the presence of strips of multiple protein isoforms in the adjacent spots.
Project description:Staphylococcus aureus is recognized worldwide as a major pathogen causing clinical or subclinical intramammary infections in all the dairy species (sheep, goats and cows). The present study was designed to comparatively investigate 65 S. aureus isolates recovered from dairy sheep and S. aureus suclinical mastitis from cows (n=21) and goats (n=22), for the presence of 190 putative virulence determinants with a single-dye DNA microarray and PCR. The probes (65 mer) were mainly designed from the S. aureus Mu50. The extracted DNA of each strain was labelled with Cy5. The microarray results were validated with PCR.The genomic comparative study with the DNA microarrays showed lineage and species specificity genes leading to the host-specific pathogenic traits of S. aureus in dairy species.
Project description:The combination of increasing consumption rates and limited elimination under conventional waste water treatment practices of many pharmaceutical compounds has now led to their detection in aquatic environments. Three of the most frequently detected pharmaceuticals in the environment are Acetaminophen (APAP), Atenolol (AT) and Carbamazepine (CBZ). Atlantic salmon (parr) was exposed to environmentally relevant levels of Acetaminophen (APAP) (54.77 ± 34.67 µg·L-1), Atenolol (AT) (11.08 ± 7.98 µg·L-1) and Carbamzepine (CBZ) (7.85 ± 0.13 µg·L-1). Gene expression was analyzed in liver tissues using a 16K GRASP (University of Victoria, Canada) cDNA microarray. GRASP 16K v.2 cDNA microarrays were used for this study (Accession # A-GEOD-2716). A dual-labelled experimental design was employed for the microarray hybridisations. Each experimental cDNA sample (Cy3 labeled) was competitively hybridised against a common pooled-reference sample (Cy5 labeled). The entire experiment comprised 20 hybridisations - 4 states (APAP, AT, CBZ, control) × 1 time-point ( at 5 days) × 5 biological replicates (males only). Hybridisations were undertaken concurrently.
Project description:As part of a study investigating the effects of genotype on responses to sustainable feeds in Atlantic salmon, a microarray analysis of the liver transcriptome of two family groups, identified as 'Lean' or 'Fat' (based on flesh lipid content), which were fed a diet containing either 100% fish oil (FO) or 100% vegetable oil (VO) was undertaken. Cholesterol and lipoprotein metabolism pathways that were differentially affected by diet depending on the genetic background of the fish were identified.<br><br>The TRAITS/SGP (v.2.1) salmon 17k cDNA microarray was used in this experiment. A dual-labelled experimental design was employed for the microarray hybridisations. aRNA from each experimental sample (Cy3 labelled) was competitively hybridised against a common pooled-reference sample (Cy5 labelled), which comprised equal amounts of aRNA from each of the samples used in the study. This design permits valid statistical comparisons across all treatments to be made. The entire experiment comprised 24 hybridisations - 2 lipid phenotypes (Lean/Fat) × 2 diets (FO/VO) × 6 biological replicates.