Dataset Information

Utilisation of Galleria mellonella larvae to characterise development of Staphylococcus aureus infection

ABSTRACT: Staphylococcus aureus is a common human opportunistic pathogen which causes a wide range of infections from superficial skin infections to invasive infections including pneumonia and bloodstream infections. Here we describe how an inoculum of S. aureus activates the cellular and humoral response of Galleria mellonella larvae while growing and disseminating throughout the host, forming nodules and ultimately killing the host. An inoculum of S. aureus (2×106/ larva) decreased larval viability at 24 (80 ± 5.77%), 48 (55.93 ± 5.55%) and 72 (10.23 ± 2.97%) hours and this was accompanied by significant proliferation and disseminated of S. aureus between 6 hours to 48 hours and the formation of nodules in the host. The hemocyte (immune cell) densities increased between 4 and 24 hours and cells isolated from larvae after 6 and 24 hours exposure to heat killed S. aureus (2×106/ larva) showed altered killing kinetics as compared to control larvae. Alterations in the humoral immune response of larvae 6 and 24 hours post infection was also determined by quantitative shotgun proteomics. The proteome of 6 hour infected larvae was enriched for antimicrobial proteins, members of the prophenoloxidase cascade and a range of peptidoglycan recognition proteins. By 24 hours there was a significant increase in a range of antimicrobial peptides with anti-staphylococcal activity and proteins associated with nodule formation. The results presented here find S. aureus interacts with the larval host immune response, induces its expression and also forms nodules which are a hallmark of soft tissue infections during human infection.

INSTRUMENT(S): Q Exactive

ORGANISM(S): Galleria Mellonella

SUBMITTER: Gerard Sheehan

LAB HEAD: Kevin Kavanagh

PROVIDER: PXD012766 | Pride | 2019-05-21

REPOSITORIES: Pride

ACCESS DATA

Dataset's files

Source:

Items per page:

1 - 5 of 13

Publications

Utilization of Galleria mellonella larvae to characterize the development of Staphylococcus aureus infection.

Sheehan Gerard G Dixon Amy A Kavanagh Kevin K

Microbiology (Reading, England) 20190520 8

Staphylococcus aureus is a human opportunistic pathogen that causes a wide range of superficial and systemic infections in susceptible patients. Here we describe how an inoculum of S. aureus activates the cellular and humoral response of Galleria mellonella larvae while growing and disseminating throughout the host, forming nodules and ultimately killing the host. An inoculum of S. aureus (2×10<sup>6</sup> larva<sup>- 1</sup> ) decreased larval viability at 24 (80±5.77 %), 48 (55.93±5.55 %) and ...[more]

PMID: 31107207

Similar Datasets

Project description:Polymicrobial infections are intrinsically less susceptible to antimicrobial therapy, are more invasive and can induce enhanced inflammatory responses leading to negative clinical outcomes. The importance of studying the interaction between microbes themselves and their interactions with their host in combination has only been realised in recent years. However, there is a lack of simple, effective and low cost in vivo models to study these interactions with the host immune response. This study details the responses of Galleria mellonella larvae to disseminated combination infection by C. albicans and S. aureus and demonstrates the efficacy of antimicrobial chemotherapy on treating co-infected larvae. Doses of C. albicans (1 × 105 larva-1) and S. aureus (1 × 104 larva-1) which were non-lethal in mono-infection, when combined significantly lowered larval survival at (70 ± 3.33% [p < 0.01]), 48 (10 ± 3.33% [p < 0.0001]) and 72h (5 ± 6.66% [p < 0.0001]) h relative to larvae receiving S. aureus 2 × 104 larva-1 mono-infection. Co-infected larvae with S. aureus 2 × 104 larva-1 resulted in an increase in the density of circulating hemocytes relative to S. aureus mono-infected larvae. Co-infected larvae displayed a significantly higher overall S. aureus c.f.u larva-1 compared to larvae only infected with S. aureus. Co-infection resulted in dissemination throughout the host and the appearance of large nodules. Proteomic analysis of co-infected larval hemolymph revealed specific immune responses to bacterial and fungal infection by the increased abundance of a range of antimicrobial peptides, peptidoglycan, lipopolysaccharide and β-glucan recognition proteins and proteins associated with tissue invasion and nodule formation. The in vitro and in vivo efficacy of empirical antimicrobial drugs (amphotericin B & meropenem) was examined against C. albicans and S. aureus was also examined. meropenem was more effective than amphotericin B but a combination was most effective at improving survival of co-infected larvae. The effect of antimicrobial therapy on symptoms associated with co-infection was also assessed using Cryo-imaging. The processes of polymicrobial co-infection in G. mellonella are compared to those in mice models and larvae provide an easy to use and ethically acceptable in vivo system to assess polymicrobial interactions with their host.

Project description:The precise mechanism and effects of antibiotics in host gene expression and immunomodulation in MRSA infection is unknown. Using a well characterized Methicillin Resistant Staphylococcus aureus (MRSA) isolate USA300 in a murine model of infection, we determined that linezolid and vancomycin induced differential production of bacterial toxins and host cytokines, differences in host gene expression, and differences in immunomodulators during MRSA bloodstream infection. A total of 35 A/J mice, categorized into seven groups (no infection; no infection with linezolid; no infection with vancomycin; 2 hour post-infection (hpi) S. aureus; 24 hpi S. aureus; 24 hpi S. aureus with linezolid; and 24 hpi S. aureus with vancomycin), were used in this study. Mice were injected with USA300 (6 x 106 CFU/g via i.p. route), then intravenously treated with linezolid (25 mg/kg) or vancomycin (25 mg/kg) at 2 hpi. Control and S. aureus infected mice were euthanized at each time point (2 h or 24h) following injection. Whole blood RNA was used for microarray; three cytokines and two S. aureus toxins [PantonValentine Leukocidin (PVL) and alpha hemolysin] were quantified in mouse serum by ELISA. S. aureus CFUs were significantly reduced in blood and kidney after linezolid or vancomycin treatment in S. aureus-infected mice. In vivo IL-1β in mouse serum was significantly reduced in both linezolid (p=0.001) and vancomycin (p=0.006) treated mice compared to untreated ones. IL-6 was significantly reduced only in linezolid treated (p<0.001) but not in vancomycin treated mice. However, another proinflammatory cytokine, TNF-α, did not exhibit altered levels in either linezolid or vancomycin treated mice (p=0.3 and p=0.51 respectively). In vivo level of bacterial toxin, Panton-Valentine leukocidin, in mouse serum was significantly reduced only in linezolid treated mice (p=0.02) but not in vancomycin treated mice. There was no significant effect of either treatment in in vivo level of alpha hemolysin production. Unsupervised hierarchical clustering using the gene expression data from 35 microarrays revealed distinct clustering based on infection status and treatment group. Study of the antibiotic-specific difference in gene expression identified the number of genes uniquely expressed in response to S. aureus infection, infection with linezolid treatment, and infection with vancomycin treatment. Pathway associations study for the differentially expressed genes in each comparison group (Control vs. 24 h S. aureus infection, 24 h S. aureus infection vs. 24 h S. aureus linezolid, and 24 h S. aureus infection vs. 24 h S. aureus vancomycin) in mice using Kyoto Encyclopedia of Genes and Genomes (KEGG) identified toll-like receptor signaling pathway to be common to every comparison groups studied. Glycerolipid metabolism pathway was uniquely associated only with linezolid treatment comparison group. The findings of this study provide the evidence that protein synthesis inhibitor like linezolid does a better job in treating MRSA sepsis compared to cell wall acting antibiotics like vancomycin.

Project description:The precise mechanism and effects of antibiotics in host gene expression and immunomodulation in MRSA infection is unknown. Using a well characterized Methicillin Resistant Staphylococcus aureus (MRSA) isolate USA300 in a murine model of infection, we determined that linezolid and vancomycin induced differential production of bacterial toxins and host cytokines, differences in host gene expression, and differences in immunomodulators during MRSA bloodstream infection. A total of 35 A/J mice, categorized into seven groups (no infection; no infection with linezolid; no infection with vancomycin; 2 hour post-infection (hpi) S. aureus; 24 hpi S. aureus; 24 hpi S. aureus with linezolid; and 24 hpi S. aureus with vancomycin), were used in this study. Mice were injected with USA300 (6 x 106 CFU/g via i.p. route), then intravenously treated with linezolid (25 mg/kg) or vancomycin (25 mg/kg) at 2 hpi. Control and S. aureus infected mice were euthanized at each time point (2 h or 24h) following injection. Whole blood RNA was used for microarray; three cytokines and two S. aureus toxins [PantonValentine Leukocidin (PVL) and alpha hemolysin] were quantified in mouse serum by ELISA. S. aureus CFUs were significantly reduced in blood and kidney after linezolid or vancomycin treatment in S. aureus-infected mice. In vivo IL-1M-NM-2 in mouse serum was significantly reduced in both linezolid (p=0.001) and vancomycin (p=0.006) treated mice compared to untreated ones. IL-6 was significantly reduced only in linezolid treated (p<0.001) but not in vancomycin treated mice. However, another proinflammatory cytokine, TNF-M-NM-1, did not exhibit altered levels in either linezolid or vancomycin treated mice (p=0.3 and p=0.51 respectively). In vivo level of bacterial toxin, Panton-Valentine leukocidin, in mouse serum was significantly reduced only in linezolid treated mice (p=0.02) but not in vancomycin treated mice. There was no significant effect of either treatment in in vivo level of alpha hemolysin production. Unsupervised hierarchical clustering using the gene expression data from 35 microarrays revealed distinct clustering based on infection status and treatment group. Study of the antibiotic-specific difference in gene expression identified the number of genes uniquely expressed in response to S. aureus infection, infection with linezolid treatment, and infection with vancomycin treatment. Pathway associations study for the differentially expressed genes in each comparison group (Control vs. 24 h S. aureus infection, 24 h S. aureus infection vs. 24 h S. aureus linezolid, and 24 h S. aureus infection vs. 24 h S. aureus vancomycin) in mice using Kyoto Encyclopedia of Genes and Genomes (KEGG) identified toll-like receptor signaling pathway to be common to every comparison groups studied. Glycerolipid metabolism pathway was uniquely associated only with linezolid treatment comparison group. The findings of this study provide the evidence that protein synthesis inhibitor like linezolid does a better job in treating MRSA sepsis compared to cell wall acting antibiotics like vancomycin. To identify differences in host gene expression in a murine sepsis model treated with a) linezolid and b) vancomycin, we used whole blood gene expression (RNA) signatures from A/J inbred mice infected with USA 300 MRSA to evaluate differences in host gene expression among mice treated with linezolid and vancomycin. We used 5 RNA samples from MRSA-infected, linezolid- or vancomycin-treated mice. A total of 7 experimental groups have been employed: 1) Uninfected control group: (negative controls). 2) Uninfected, linezolid-treated group: Uninfected, linezolid-treated mice. 3) Uninfected vancomycin-treated group: Uninfected, vancomycin-treated mice. 4) Infected control group (positive control 2 h) MRSA-infected, untreated mice. 5) Infected control group (positive control 24 h): MRSA-infected, untreated mice. 6) Infected linezolid group: MRSA-infected, linezolid-treated mice. 7) Infected vancomycin group: MRSA-infected, vancomycin-treated mice.

			Action	DRS
	MaxQuantOutput.zip	Other
	SA1.raw	Raw
	SA10.raw	Raw
	SA11.raw	Raw
	SA12.raw	Raw