Project description:Several methicillin resistance (SCCmec) clusters characteristic of hospital-associated methicillin-resistant Staphylococcus aureus (MRSA) strains harbor the psm-mec locus. In addition to encoding the cytolysin, phenol-soluble modulin (PSM) mec, this locus has been attributed gene regulatory functions. Here we employed genome-wide transcriptional profiling to define the regulatory function of the psm-mec locus. The immune evasion factor protein A emerged as the primary conserved and strongly regulated target of psm-mec, an effect we show is mediated by the psm-mec RNA. Furthermore, the psm-mec locus exerted regulatory effects that were more moderate in extent and possibly mediated by the PSM-mec peptide. For example, expression of PSM-mec limited expression of mecA, thereby decreasing methicillin resistance. Our study shows that the psm-mec locus has a rare dual regulatory RNA and encoded cytolysin function, both with the potential to enhance MRSA virulence. Furthermore, our findings reveal a specific mechanism underscoring the recently emerging concept that S. aureus strains balance pronounced virulence and high expression of antibiotic resistance.
Project description:Bacterial sepsis is a major killer in hospitalized patients. Coagulase-negative staphylococci (CNS) with the leading species Staphylococcus epidermidis are the most frequent causes of nosocomial sepsis, with most infectious isolates being methicillin resistant. However, which bacterial factors underlie the pathogenesis of CNS sepsis is unknown. While it has been commonly believed that invariant structures on the surface of CNS trigger sepsis by causing an over-reaction of the immune system, we show here that sepsis caused my methicillin-resistant S. epidermidis is to a large extent mediated by the methicillin resistance island-encoded peptide toxin, PSM-mec. PSM-mec contributed to bacterial survival in whole human blood and resistance to neutrophil-mediated killing, and caused significantly increased mortality and cytokine expression in a mouse sepsis model. Furthermore, we show that the PSM-mec peptide itself, rather than the regulatory RNA in which its gene is embedded, is responsible for the observed virulence phenotype. While toxins have never been clearly indicated in CNS infections, our study shows that an important type of infection caused by the predominant CNS species, S. epidermidis, is mediated to a large extent by a toxin. Of note, these findings suggest that CNS infections may be amenable to virulence-targeted drug development approaches. We used microarrays to detail the global gene expression between S. epidermidis strain Rp62A and S. epidermidis strain Rp62A isogenic Δpsm-mec deletion mutants
Project description:Comparing two subclones (Taiwan clone and Asian-Pacific clone) of CA-MRSA ST59. The Taiwan clone carries the Panton-Valentine leukocidin (PVL) genes, the staphylococcal chromosomal cassette mec (SCCmec) VT and is frequently isolated from patients with severe disease. The Asian-Pacific clone is PVL-negative, carries SCCmec IV, and is a frequent colonizer of healthy children.
Project description:The present work comprises the study of wound pathogenic bacteria as part of a community. It considers the interactions of two different S. aureus isolates with B. thuringiensis and K. oxytoca; all of them isolated from the same chronic wound of a patient with epidermolysis bullosa. Particular focus has been given on the interactions of S. aureus with other microbes due to its high prevalence among chronic wounds. During cultivation, no species performed as dominant or inhibited the growth of one another. Mass spectrometry was used to explore the inherent relationships between the staphylococcal strains and the coexisting bacteria exproteomes. The analysis showed an important reduction in the amount of staphylococcal cytoplasmic proteins when co-cultured with K. oxytoca and B. thuringiensis, this decrement did not occur with klebsiella and bacillus proteins. Interestingly, K. oxytoca and B. thuringiensis seemed to have a more evident response towards the presence of S. aureus in the culture, while the opposite was not observed with the staphylococcal isolates. Genomic analysis revealed isolate t13595 hypermutable characteristics, placing the interactions between staphylococcal isolates in the context of a chronic wound. Overall, the nature of the exoproteome variations among cultures suggests that adaptive mechanisms differ in all strains.
Project description:Mastitis is an inflammation of the mammary gland (MG), usually due to bacterial infection. Although considerable attention has been paid to this economically important disease, the early stages of the host response remain poorly defined. In particular, it is unclear how mammary epithelial cells (MEC), a first barrier against pathogens, respond to infection. Indeed, it is difficult to differentiate between the contributions of MEC and infiltrating immune cells to gene expression profiles of mammary tissue during mastitis. The current investigation examines the response of MEC at the early stage of infection using a non invasive RNA sampling method taking advantage of the presence of cytoplasmic crescents contained in milk fat globules. We have recently shown that, in goats, Milk Fat Globules (MFG) provide a unique source of RNA to study the in vivo regulation of gene expression in MEC. This non invasive RNA sampling method was used during the time course of an experimental intra mammary infection (IMI) with S. aureus. Experiments were performed using ovine microarrays (Agilent) to compare gene expression patterns before infection, at 12h, 18h and 24h post-infection (PI). Furthermore, we combined this approach with laser capture microdissection of MEC isolated from frozen slides of mammary tissue to study some specific genes at the late stage of infection (30h PI). We show that at 18h PI, before the burst of somatic cells in milk, MEC play an important role in the recruitment and activation of inflammatory cells through the IL-8 signaling pathways. Then, at the late stage of infection (30h PI), the contribution of MEC in immune response changes to produce different acute phase proteins, including SAA3, serpin A1 and PTX3. These cells also express factors that contribute directly to fighting infection, such as S100A12. In summary, we demonstrate for the first time in vivo how MEC orchestrate innate immune response during an IMI with S. aureus in the goat species. We report here new opportunities to assess the dynamics of gene expression in the mammary gland, thus providing significant advances in the understanding of MEC immune capacity. Furthermore, the production of some molecules by MEC, in the early stages of IMI, could provide sensitive biomarkers for early detection and therefore, treatment of mastitis.
Project description:To identify specific predictive biomarkers for septic arthritis, transcriptome sequencing analysis was performed on Staphylococcal aureus infected mouse RNA samples. We then evaluated differentially expressed genes from RNA-seq of Day 0, Day 2 and Day 10 samples of arthritic and non-arthritic mice.
Project description:Mastitis is an inflammation of the mammary gland (MG), usually due to bacterial infection. Although considerable attention has been paid to this economically important disease, the early stages of the host response remain poorly defined. In particular, it is unclear how mammary epithelial cells (MEC), a first barrier against pathogens, respond to infection. Indeed, it is difficult to differentiate between the contributions of MEC and infiltrating immune cells to gene expression profiles of mammary tissue during mastitis. The current investigation examines the response of MEC at the early stage of infection using a non invasive RNA sampling method taking advantage of the presence of cytoplasmic crescents contained in milk fat globules. We have recently shown that, in goats, Milk Fat Globules (MFG) provide a unique source of RNA to study the in vivo regulation of gene expression in MEC. This non invasive RNA sampling method was used during the time course of an experimental intra mammary infection (IMI) with S. aureus. Experiments were performed using ovine microarrays (Agilent) to compare gene expression patterns before infection, at 12h, 18h and 24h post-infection (PI). Furthermore, we combined this approach with laser capture microdissection of MEC isolated from frozen slides of mammary tissue to study some specific genes at the late stage of infection (30h PI). We show that at 18h PI, before the burst of somatic cells in milk, MEC play an important role in the recruitment and activation of inflammatory cells through the IL-8 signaling pathways. Then, at the late stage of infection (30h PI), the contribution of MEC in immune response changes to produce different acute phase proteins, including SAA3, serpin A1 and PTX3. These cells also express factors that contribute directly to fighting infection, such as S100A12. In summary, we demonstrate for the first time in vivo how MEC orchestrate innate immune response during an IMI with S. aureus in the goat species. We report here new opportunities to assess the dynamics of gene expression in the mammary gland, thus providing significant advances in the understanding of MEC immune capacity.M-bM-^@M-^BFurthermore, the production of some molecules by MEC, in the early stages of IMI, could provide sensitive biomarkers for early detection and therefore, treatment of mastitis. Experiments were performed using ovine microarrays (Agilent) to compare gene expression patterns before infection, at 12h, 18h and 24h post-infection (PI). 20 sample records; mono-colour experimental design
Project description:The goal of our study was to perform RNA sequencing of whole transcripts to determine and compare the real length of 3’UTRs among closely-related staphylococcal species. We found that most of the mRNAs encoding orthologous genes in staphylococcal species have 3’UTRs with different lengths in addition to sequence variation.
Project description:In this study, we performed bulk RNA-seq of pluripotent stem cells (PSCs) and induced pre-somitic mesoderm (PSM) cells of six different mammalian species. The species studied are: mouse (Mus musculus), marmoset (Callithrix jacchus), rabbit (Oryctolagus cuniculus), human (Homo sapiens), cattle (Bos taurus) and southern white rhinoceros (Ceratotherium simum). We used mouse ESCs, marmoset iPSCs, rabbit ESCs, human iPSCs, bovine ESCs and rhinoceros ESCs to induce PSM-like cells from these species following protocols already described. PSC samples were extracted under maintenance conditions. PSM samples were extracted on the day when the differentiation efficiency was higher based on the percentage of cells expressing the PSM marker TBX6. We used identical culture conditions when extracting the induced PSM cells to minimize the effect of external factors on our results. Two replicates per each cell type and species were collected for a total of 24 samples. We compared the expression levels of more than 10,000 orthologous protein-coding genes across the six species. With this, we determined that the species-specific segmentation clock periods might be derived from species-specific gene expression profiles controlling basic biological processes.
Project description:S. aureus and S. epidermidis were challenged with D-sphingosine, an antimicrobial lipid similar to sphingosines found in the major staphylococcal niche- human skin. Comparison of responses was used to identify resistance mechanisms and likely mode of action