Transcription profiling by array of Staphylococcus aureus grown in cheese matrix in mixed culture with Lactococcus lactis
ABSTRACT: Effect of the presence of Lactococcus lactis on Staphylococcus aureus transcriptome in cheese matrix. S. aureus was co-cultured with L. lactis LD61 in cheese matrix during 7 days. RNA samples were extracted at different time points (6 h, 8 h, 10 h, 24 h and 7 days) in order to monitor the dynamic response of S. aureus MW2 in cheese matrix in presence of L. lactis
Project description:This SuperSeries is composed of the following subset Series: GSE23987: Transcriptomic profiles of six strains of Lactococcus lactis in ultrafiltration-cheese model GSE23990: Comparative genome hybridization profiles of six strains of Lactococcus lactis Refer to individual Series
Project description:The intra sub-species diversity of six strains of Lactococcus lactis subsp. lactis was investigated at the genomic level and in terms of phenotypic and transcriptomic profiles in UF-cheese model. Six strains were isolated from various sources, but all are exhibiting a dairy phenotype. Our results showed that, the six strains exhibited small phenotypic differences since similar behaviour in terms of growth was obtained during cheese ripening while only different acidification capability was detected. Even if all strains displayed high genomic similarities, sharing a high core genome of almost two thousands genes, the expression of this core genome directly in the cheese matrix revealed major strain-specific differences. This strains with the same dairy origin. Strains were cultured on skimmed raw milk ultrafiltration (UF) retentate. The UF retentate was pre-incubated overnight at 4 °C, then 45 minutes at 50 °C and homogenized during 1.5 minutes at 24 000 rpm with an ultra-turax (Imlab, France). After addition of rennet (0.3 µl ml-1), 400 g UF retentate was inoculated at 2 106 CFU/g with L. lactis subsp. lactis strains. After incubation for 8 hours at 30 °C, the cheeses were transferred at 12° C until 7 days for ripening simulation. At least three independent cultures of the six strains were performed. Total RNA was extracted from cells grown 24 hours in UF-cheese and radiolabelled cDNA were prepared and hybridized on nylon arrays. 1948 amplicons specific of Lactococcus lactis IL1403 genes were spotted twice on the array. 3 independent repetitions were performed.
Project description:The intra sub-species diversity of six strains of Lactococcus lactis subsp. lactis was investigated at the genomic level and in terms of phenotypic and transcriptomic profiles in UF-cheese model. Six strains were isolated from various sources, but all are exhibiting a dairy phenotype. Our results showed that, the six strains exhibited small phenotypic differences since similar behaviour in terms of growth was obtained during cheese ripening while only different acidification capability was detected. Even if all strains displayed high genomic similarities, sharing a high core genome of almost two thousands genes, the expression of this core genome directly in the cheese matrix revealed major strain-specific differences. This strains with the same dairy origin. Strains were cultured on M17. At least three independent cultures of the six strains were performed. Genomic DNA was extracted from cells grown overnight on M17 and radiolabelled cDNA were prepared and hybridized on nylon arrays. 1948 amplicons specific of Lactococcus lactis IL1403 genes were spotted twice on the array. 3 independent repetitions were performed.
Project description:Methicillin-resistant Staphylococcus aureus (MRSA) is problematic both in hospitals and the community. Currently, we have limited understanding of mechanisms of innate immune evasion used by S. aureus. To that end, we created an isogenic deletion mutant in strain MW2 (USA400) of the saeR/S two-component gene regulatory system and studied its role in mouse models of pathogenesis and during human neutrophil interaction. In this study, we demonstrate saeR/S plays a distinct role in S. aureus pathogenesis and is vital for virulence of MW2 in a mouse model of sepsis. Moreover, deletion of saeR/S significantly impaired survival of MW2 in human blood and after neutrophil phagocytosis. Microarray analysis of genes influenced by saeR/S demonstrated SaeR/S of MW2 influences a wide variety of genes with diverse biological functions. These data shed new insight into how virulence is regulated in S. aureus and associates a specific staphylococcal gene-regulatory system with invasive staphylococcal disease. Wild type control vs mutant at two different growth phases
Project description:The response of L. lactis to the presence of S. cerevisiae was analyzed during the exponential growth phase in fermentors in defined growth conditions. Although no growth kinetic difference was observed between the pure and mixed culture of L. lactis, the mRNA level of genes was significantly modified. More particularly, a strong reorientation of pyrimidine metabolism was observed when L. lactis was grown in the mixed culture. Keywords: microbial interaction, time course A pure culture of L. lactis was conducted in parallel with a co-culture of L. lactis and S. cerevisiae. For the transcriptomic analysis, the mixed culture (test condition, 2.5 hours of culture, B’) was compared to the pure culture (reference, 2.5 hours of culture, B) on the same slide at the same time. The mRNA level changes between 2 and 3 hours (A and C samples) flanking B sample (2.5 hours) in the pure culture were also determined on another slide. Total RNA was extracted from these samples and labelled cDNA (Cy3/Cy5) were prepared and hybridized on glass slides exhibiting 2004 amplicons specific of Lactococcus lactis IL1403 genes. 3 independent repetitions were performed.
Project description:The lactococcal phage p2 is a model for studying the Skunavirus genus, the most prevalent group of phages in cheese factories worldwide. It infects L. lactis MG1363, a model strain for the study of Gram-positive bacteria. The structural proteins of phage p2 have been thoroughly described. However, most of its non-structural proteins are still uncharacterized. Here, we developed an integrative approach, making use of structural biology, genomics, physiology, and proteomics to provide insights into the function of ORF47, the most conserved non-structural protein of unknown function among the Skunavirus genus. We found this small phage protein to have a major impact on the bacterial proteome and to be important to prevent bacterial resistance to phage infection.
Project description:Staphylococcus aureus causes a spectrum of human infection. Diagnostic delays and uncertainty lead to treatment delays and inappropriate antibiotic use. A growing literature suggests the host’s inflammatory response to the pathogen represents a potential tool to improve upon current diagnostics. The hypothesis of this study is that the host responds differently to S. aureus than to E. coli infection in a quantifiable way, providing a new diagnostic avenue. This study uses Bayesian sparse factor modeling and penalized binary regression to define peripheral blood gene-expression classifiers of murine and human S. aureus infection. The murine-derived classifier distinguished S. aureus infection from healthy controls and Escherichia coli-infected mice across a range of conditions (mouse and bacterial strain, time post infection) and was validated in outbred mice (AUC>0.97). A S. aureus classifier derived from a cohort of 95 human subjects distinguished S. aureus blood stream infection (BSI) from healthy subjects (AUC 0.99) and E. coli BSI (AUC 0.82). Murine and human responses to S. aureus infection share common biological pathways, allowing the murine model to classify S. aureus BSI in humans (AUC 0.84). Both murine and human S. aureus classifiers were validated in an independent human cohort (AUC 0.95 and 0.94, respectively). The approach described here lends insight into the conserved and disparate pathways utilized by mice and humans in response to these infections. Furthermore, this study advances our understanding of S. aureus infection; the host response to it; and identifies new diagnostic and therapeutic avenues. To create a host gene expression-based classifier for S. aureus infection, mice from a variety of experimental conditions were utilized. Seven different strains of inbred mice (n=187 total) were challenged with four different S. aureus strains via intraperitoneal inoculation and sacrificed at various time points as described in Methods. The comparator group for model derivation included 50 A/J or C57BL/6J mice inoculated with E. coli (O18:K1:H7) as well as 54 uninfected mice. Next, the murine S. aureus classifier was externally validated in outbred CD-1 mice with S. aureus infection (Sanger 476 or USA300), E. coli infection (O18:K1:H7), or uninfected controls (10 animals per condition). Method: All experiments were performed on mice 6-8 weeks old. For the murine S. aureus predictor, seven inbred mouse strains (3 mice/strain: 129S1/SvImJ, A/J, AKR/J, BALB/cByJ, C57BL/6J, C3H/HeJ, and NOD/LtJ) were IP inoculated with 107 CFU/g of S. aureus Sanger476, euthanized at 2h after injection, and bled. This was repeated using four different S. aureus strains (USA100, USA300, MW2, and Sanger476) in A/J mice (n=3 per S. aureus strain). For time series experiments, both A/J and C57BL/6J mouse strains were IP inoculated with S. aureus Sanger476 as above, and sacrificed at 2, 4, 6, and 12h after injection (n=5 per time point).
Project description:Comparison of L. lactis NZ9000 ΔlmrR versus L. lactis NZ9000 wild type Keywords: Transcription profiling Comparison between strain lacking transcriptional regulator LmrR of major Lactococcal mdr transporter LmrCD, and wild type parental strain.