Expression analysis of 6 serotypes of Actinobacillus pleuropneumonia during iron deprivation
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ABSTRACT: Investigation of whole genome changes in six serotypes of Actinobacillus pleuropneumoniae in control cultures compared to bacteria grown in media containg the iron chelator 2,2'-dipyridyl. A 48 chip study using total RNA recovered from four different control cultures and four different iron depleted cultures of Actinobacillus pleuropneumoniea serotype 1 (4074), serotype 2 (4226), serotype 3 (1421), serotype 5b(L209, serotype 6 (7712640) and serotype 7 (WF87).
ORGANISM(S): Actinobacillus pleuropneumoniae serovar 2 str. 4226
Project description:G. sulfurreducens was cultured on a variety of different iron concentrations and differential expression analysis was used to identify the iron stimulon. Wild type G. sulfurreducens was cultured on fresh water media with trace concentrations of iron (iron sufficient condition) as well as no trace iron (iron deficient condition). It was also cultured in ferric citrate (iron excess condition) media
Project description:Investigation of whole genome gene expression level changes in a Nitrosomonas europaea (ATCC 19718) wildtype and pFur::Kan mutant [kanamycin resistance cassette insertion in the promoter region of the fur gene (NE0616)] strains grown in Fe-replete and Fe-limited media. The Nitrosomonas europaea (ATCC 19718) wiltype cells grown in Fe-limited media were compared to cells grown in Fe-replete media to gain a better understanding of the metabolic changes occurring in response to iron stress. The Nitrosomonas europaea (ATCC 19718) pFur::Kan mutant strain grown in Fe-replete & Fe-limited media were compared to wildtype cells grown in Fe=replete & Fe-limited media to gain a better understanding of the role Fur (NE0616) plays in iron homeostasis control. A 4-plex 3 chip study using total RNA recovered from three separate wild-type cultures each of N. europaea grown in Fe-replete media and Fe-limited media and three seperate cultures each of N. europaea pFur::Kan mutant strain grown in Fe-replete and Fe-limited media. Each chip measures the expression level of 2368 genes from Nitrosomonas europaea (ATCC19718) with 4 X 72,000 60-mer 14 probe pairs per gene, with two-fold technical redundancy.
Project description:The transcriptional profile of the porcine lung pathogen, Actinobacillus pleuropneumoniae, was monitored during the acute phase of infection in its natural host. Bacterial expression profiles of A. pleuropneumoniae isolated from lung lesions of 25 infected pigs were compared in samples taken 6, 12, 24 and 48 hours post infection. A 84 chip study using total RNA recovered from A. pleuropneumoniae serotype 2 (4226) and serotype 6 (7712640) isolated from infected pig lung tissue during the first 48 of infection. Samples were taken 6, 12, 24 and 48 hours post infection, respectively. Before hybridization the samples were enriched for bacterial RNA and submitted to linear amplification. Each chip measures the expression level of 4,876 target genes from A. pleuropneumoniae with each gene covered by an average of 26.7 probes. Three biological replicates per sample. Microarray data from 3 pigs (no. 39, no. 43 and no. 72) were omitted from the final analysis due to poor signal intensity. A total of 75 microarrays were included in the final analysis.
Project description:In Neisseria meningitidis iron responsive gene regulation is mediated primarily by the Ferric Uptake Regulator (Fur) protein. When complexed with iron, Fur represses gene expression by preventing transcription initiation. Fur can also indirectly activate gene expression via the repression of regulatory small RNAs (sRNA). One such Fur-and iron-regulated sRNA, NrrF, was previously identified in N. meningitidis and shown to repress expression of the sdhA and sdhC genes encoding subunits of the succinate dehydrogenase complex. In the majority of Gram-negative bacteria sRNA-mediated regulation requires a cofactor RNA-binding protein (Hfq) for proper gene regulation and stabilization. In this study we examined the role of Hfq in NrrF-mediated regulation of the succinate dehydrogenase genes in N. meningitidis and the effect of an hfq- mutation on iron-responsive gene regulation more broadly. We first demonstrated that the stability of Nrrf as well as the regulation of sdhC and sdhA in vivo was unaltered in the hfq- mutant. Secondly, we established that iron responsive gene regulation of the Fur-regulated sodB gene was dependent on Hfq. Finally, we demonstrate that in N. meningitidis Hfq functions to control expression of both ORFs and intergenic regions via iron independent mechanisms. Collectively these studies demonstrate that in N. meningitidis iron and NrrF mediated regulation of sdhC and sdhA can occur independently of Hfq, although Hfq functions more globally to control regulation of other N. meningitidis genes primarily by iron-independent mechanisms. RNA was isolated from wild-type MC58 Neisseria meningitidis, from an hfq- mutant, and from a complemented hfq- mutant under both iron-replete and iron-deplete conditions. Three biological replicates were analyzed for each strain and condition were analyzed.
Project description:Differential expression of electron transfer genes during growth with insoluble iron provided as an electron acceptor compared to soluble iron. A four chip study using total RNA recovered from two separate cultures of Ferroglobus placidus DSM 10642 grown with 10 mM acetate provided as electron donor and insoluble iron hydroxide provided as electron acceptor (experimental condition) and two separate cultures of Ferroglobus placidus DSM 10642 grown on 10 mM acetate with soluble iron citrate provided as electron acceptor (control condition). Each chip measures the expression level of 2613 genes from Ferroglobus placidus DSM 10642 with nine 45-60-mer probe pairs (PM/MM) per gene, with three-fold technical redundancy.
Project description:Investigation of whole genome changes in six serotypes of Actinobacillus pleuropneumoniae in control cultures compared to bacteria grown in media containg the iron chelator 2,2'-dipyridyl.
Project description:Actinobacillus pleuropneumoniae is the etiological agent of porcine pleuropneumonia, a respiratory disease which causes great economic losses worldwide. Many virulence factors are involved in the pathogenesis, namely capsular polysaccharides, RTX toxins, LPS and many iron acquisition systems. In order to identify genes that are expressed in vivo during a natural infection, we undertook transcript profiling experiments with an A. pleuropneumoniae DNA microarray, after recovery of bacterial mRNAs from serotype 5b-infected porcine lungs. Comparative Genomic Hybridizations between Actinobacillus pleuropneumoniae serotype 5b strain L20 (ref) and serotype 5b fresh field isolate 896-07, recovered from infected pig lung tissues following natural acute infection. Two condition transcript profiling experiments : infectious 5b field strain isolated directly from lungs of naturally deceased pigs after acute infection vs infectious 5b field strain grown in BHI broth to an OD600 of 0.300.
Project description:To identify genes involved in the iron-limited stress response and the induction of programmed cell death, gene expression analaysis using whole-genome microarrays was performed on cultures of T. pseudonana growing in replete or iron-limiting conditions Samples were collected at day 3 and day 5 from duplicate cultures; RNA was extracted and microarray analysis was performed Duplicate biological replicates were analyzed from replete growing or iron-limited cultures; Analysis was performed on cultures harvested in late exponential phase (day 3) and stationary phase (day 5)
Project description:The goal of this analysis is to define the aerobic and anaerobic regulons for the transcription factor Mlc (Makes Large Colonies) in the periodontal pathogen Aggregatibacter actinomycetemcomitans. The objectives were met by comparing the mRNA profile of a wild type strain, JP2, to that of an isogenic mutant deleted of the Mlc gene. Both strains were grown aerobically and anaerobically. The results show that Mlc is involved in the regulation of dozens of genes in both growth conditions. Half of the study (anaerobic samples) was a four biological replicates study using total RNA recovered from four separate anaerobic cultures of wild-type Aggregatibacter (formerly Actinobacillus) actinomycetemcomitans strain JP2 and four separate anaerobic cultures of strain AAM155, an isogenic deletion mutant in which the gene for Mlc (Makes Large Colonies) has been replaced by a spectinomycin resistance gene cassette. The cells were grown anaerobically in pairs (wild type paired with mutant). The second half of the study (aerobic samples) was a four biological replicates study using total RNA recovered from four separate aerobic cultures of wild-type Aggregatibacter actinomycetemcomitans strain JP2 and four separate anaerobic cultures of strain AAM155, an isogenic deletion mutant of Mlc. The cells were grown aerobically in pairs (wild type paired with mutant). Each chip measures the expression level of 2,116 genes from A. actinomycetemcomitans strain HK1651. There were 11 different 60-mer probes /gene for 94% of the genes, and the entire probe set (22,537 probes) was replicated (technical replicates) three times on each array. Each probe set is contained on a quarter of a chip and there were also 4,426 random probes per array. A total of four chips were used in these experiments.
Project description:Actinobacillus pleuropneumoniae is the etiological agent of porcine pleuropneumonia, a respiratory disease which causes great economic losses worldwide. Many virulence factors are involved in the pathogenesis, namely capsular polysaccharides, RTX toxins, LPS and many iron acquisition systems. In order to identify genes that are expressed in vivo during a natural infection, we undertook transcript profiling experiments with an A. pleuropneumoniae DNA microarray, after recovery of bacterial mRNAs from serotype 5b-infected porcine lungs.