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Vaccination with a UV-irradiated genetically attenuated mutant of Staphylococcus aureus provides protection against subsequent systemic infection.

ABSTRACT: Staphylococcus aureus are gram-positive bacteria that cause clinically significant infections in humans. Severe S. aureus infections are particularly problematic in hospitalized patients and reoccur despite therapeutic measures. The absence of natural protective immune responses and the lack of high-throughput approaches to identify S. aureus antigens have imposed constraints in the development of effective vaccines. Here, we showed that vaccination with the genetically attenuated S. aureus mutant, inactivated using UV irradiation rather than heat, significantly increased survival and diminished bacterial burden and kidney abscesses when mice were challenged with virulent methicillin-sensitive or methicillin-resistant S. aureus. Protection conferred by immunization could be transferred to the naive host and was not observed in B-cell-deficient mice. Using a novel S. aureus whole-proteome microarray, we show that immunoglobulin G antibody responses to 83 proteins were observed in the immunized mice. These results suggest that protection against S. aureus infections requires antibody responses to the wide repertoire of antigens/virulence factors. Vaccination using UV-irradiated genetically attenuated S. aureus induces humoral immunity and provides a vaccine strategy for pathogens that fail to induce protective immunity. We also describe a novel, high-throughput technology to easily identify S. aureus antigens for vaccine development.

SUBMITTER: Burnside K

PROVIDER: S-EPMC3488195 | biostudies-literature | 2012 Dec

REPOSITORIES: biostudies-literature

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Vaccination with a UV-irradiated genetically attenuated mutant of Staphylococcus aureus provides protection against subsequent systemic infection.

Burnside Kellie K Lembo Annalisa A Harrell Maria Isabel MI Klein Jessica Abbey JA Lopez-Guisa Jesus J Siegesmund Amy M AM Torgerson Troy R TR Oukka Mohamed M Molina Douglas M DM Rajagopal Lakshmi L

The Journal of infectious diseases 20120910 11

Staphylococcus aureus are gram-positive bacteria that cause clinically significant infections in humans. Severe S. aureus infections are particularly problematic in hospitalized patients and reoccur despite therapeutic measures. The absence of natural protective immune responses and the lack of high-throughput approaches to identify S. aureus antigens have imposed constraints in the development of effective vaccines. Here, we showed that vaccination with the genetically attenuated S. aureus muta ...[more]

PMID: 22966130

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Project description:The transcription level of a rex-deficient S. aureus mutant in comparison to its parental strain S. aureus SH1000 was analyzed using DNA microarrays. S. aureus N315 microarrays were purchased from Scienion (Scienion AG, Berlin, Germany) and were produced by spotting 2,338 PCR products of the 2,593 ORFs comprising annotated genome of S. aureus N315 [reference identification: NC_002745] on a glass slide. Each ORF is present in duplicate on the microarray (further details can be found at http://www.scienion.com), cDNA was synthesized from total RNA with the LabelStar Array Kit from QIAGEN using the QIAGEN protocol with slight modifications: Random hexamer primer were used (Invitrogen, Karlsruhe, Germany) and Cy3- and Cy5-dCTP were purchased from Perkin-Elmer (Rodgau - Juegesheim, Germany). As recommended by Scienion, 10 Âµg RNA from either SH1000 or AK1 were used for cDNA synthesis. After hybridisation for 72 h, the microarrays were washed as recommended by the manufacturer. Data analysis. The hybridized microarrays were scanned with a GenePix 4000B microarray scanner (MDS Analytical Technologies GmbH, Ismaning, Germany). A geometric raster was laid over the resulting microarray picture to distinguish the signals from the background. After localization of single spots, intensities and global background were calculated automatically. The hybridization patterns and intensities were quantitatively analyzed using the Imagene 6 software (BioDiscovery, El Segundo, CA). The replicates were averaged, and the spots identified by Imagene 6 (BioDiscovery) as flawed were omitted. The data set was normalized by application of the LOWESS algorithm. In a next step, the intensity values of all arrays for each time point as well as for all time points combined were used for t tests. Genes with a change of <0.5- or â?¥2.0-fold were characterized as having significantly differing amounts of transcripts based on t tests with a P value cut-off of at least 0.05. Gene functions were assigned to the respective accession numbers and annotations as compiled on DOGAN, a web page for S. aureus N315 (http://www.bio.nite.go.jp/dogan/MicroTop?GENOME_ID=n315G1). The parental strain SH1000 and the Rex deficient mutant AK1 were applied on full-genome microarrays to get a detailed view on the differences in the transcriptional profiles which are caused â?? directly or indirectly â?? by the introduced mutation. More specifically, expression levels were compared at five time points, covering different growth phases. To highlight the general changes in the expression profile between SH1000 and the rex mutant, the microarray data of all five time points were also analyzed in a combined way using standard statistical methods. In further experiments, we focused on those genes, which seemed to flag the general difference between the investigated strains.

Dataset Information

Vaccination with a UV-irradiated genetically attenuated mutant of Staphylococcus aureus provides protection against subsequent systemic infection.

Publications

Vaccination with a UV-irradiated genetically attenuated mutant of Staphylococcus aureus provides protection against subsequent systemic infection.

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