Dataset Information

Specific capture and detection of Staphylococcus aureus with high-affinity modified aptamers to cell surface components.

ABSTRACT: Slow off-rate modified aptamer (SOMAmer) reagents were generated to several Staphylococcus aureus cell surface-associated proteins via SELEX with multiple modified DNA libraries using purified recombinant or native proteins. High-affinity binding agents with sub-nanomolar Kd 's were obtained for staphylococcal protein A (SpA), clumping factors (ClfA, ClfB), fibronectin-binding proteins (FnbA, FnbB) and iron-regulated surface determinants (Isd). Further screening revealed several SOMAmers that specifically bound to Staph. aureus cells from all strains that were tested, but not to other staphylococci or other bacteria. SpA and ClfA SOMAmers proved useful for the selective capture and enrichment of Staph. aureus cells, as shown by culture and PCR, leading to improved limits of detection and efficient removal of PCR inhibitors. Detection of Staph. aureus cells was enhanced by several orders of magnitude when the bacterial cell surface was coated with SOMAmers followed by qPCR of the SOMAmers. Furthermore, fluorescence-labelled SpA SOMAmers demonstrated their utility as direct detection agents in flow cytometry. Significance and impact of the study: Monitoring for microbial contamination of food, water, nonsterile products or the environment is typically based on culture, PCR or antibodies. Aptamers that bind with high specificity and affinity to well-conserved cell surface epitopes represent a promising novel type of reagents to detect bacterial cells without the need for culture or cell lysis, including for the capture and enrichment of bacteria present at low cell densities and for the direct detection via qPCR or fluorescent staining.

SUBMITTER: Baumstummler A

PROVIDER: S-EPMC4299505 | biostudies-literature | 2014 Oct

REPOSITORIES: biostudies-literature

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Specific capture and detection of Staphylococcus aureus with high-affinity modified aptamers to cell surface components.

Baumstummler A A Lehmann D D Janjic N N Ochsner U A UA

Letters in applied microbiology 20140702 4

Slow off-rate modified aptamer (SOMAmer) reagents were generated to several Staphylococcus aureus cell surface-associated proteins via SELEX with multiple modified DNA libraries using purified recombinant or native proteins. High-affinity binding agents with sub-nanomolar Kd 's were obtained for staphylococcal protein A (SpA), clumping factors (ClfA, ClfB), fibronectin-binding proteins (FnbA, FnbB) and iron-regulated surface determinants (Isd). Further screening revealed several SOMAmers that sp ...[more]

PMID: 24935714

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Project description:Metals are a limiting resource for pathogenic bacteria and must be scavenged from host proteins. Hemoglobin provides the most abundant source of iron in the human body and is required by several pathogens to cause invasive disease. However, the consequences of hemoglobin evolution for bacterial nutrient acquisition remain unclear. Here we show that the α- and β-globin genes exhibit strikingly parallel signatures of adaptive evolution across simian primates. Rapidly evolving sites in hemoglobin correspond to binding interfaces of IsdB, a bacterial hemoglobin receptor harbored by pathogenic Staphylococcus aureus Using an evolution-guided experimental approach, we demonstrate that the divergence between primates and staphylococcal isolates governs hemoglobin recognition and bacterial growth. The reintroduction of putative adaptive mutations in α- or β-globin proteins was sufficient to impair S. aureus binding, providing a mechanism for the evolution of disease resistance. These findings suggest that bacterial hemoprotein capture has driven repeated evolutionary conflicts with hemoglobin during primate descent.IMPORTANCE During infection, bacteria must steal metals, including iron, from the host tissue. Therefore, pathogenic bacteria have evolved metal acquisition systems to overcome the elaborate processes mammals use to withhold metal from pathogens. Staphylococcus aureus uses IsdB, a hemoglobin receptor, to thieve iron-containing heme from hemoglobin within human blood. We find evidence that primate hemoglobin has undergone rapid evolution at protein surfaces contacted by IsdB. Additionally, variation in the hemoglobin sequences among primates, or variation in IsdB of related staphylococci, reduces bacterial hemoglobin capture. Together, these data suggest that S. aureus has evolved to recognize human hemoglobin in the face of rapid evolution at the IsdB binding interface, consistent with repeated evolutionary conflicts in the battle for iron during host-pathogen interactions.

Dataset Information

Specific capture and detection of Staphylococcus aureus with high-affinity modified aptamers to cell surface components.

Publications

Specific capture and detection of Staphylococcus aureus with high-affinity modified aptamers to cell surface components.

Similar Datasets

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