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O055 - MAGNETIC BEAD BASED PURIFICATION OF PNEUMOCOCCAL STRAINS (ID 775)
Abstract
Background
Studies have demonstrated the use of saliva for detection of pneumococcal carriage. However, isolating pneumococci from saliva is challenging due to the density and diversity of bacteria present in the oral cavity, and this is particularly challenging in individuals with colonized at a low density. Furthermore, co-colonisation with multiple pneumococcal serotypes is seldom detected by the gold standard culture-based methods, and often only the majority serotype is identified. We therefore developed a novel assay to enrich for pneumococcus in a serotype-specific manner from a complex sample such as saliva.
Methods
Serotype-specific antisera/antibodies were incubated in a mixed sample containing two pneumococcal serotypes (majority and minority concentrations), and in culture-enriched saliva spiked with varying concentrations of pneumococcus. Magnetic IgG microbeads (Miltenyi Biotech) were used to purify the antibody-bound target strain using a Kingfisher™ Flex, and the eluate were plated on blood agar for counting and isolation.
Results
For all serotypes investigated, the minority serotype was enriched from ~0.1% of the initial sample, to between 13% and 90% in the final sample (corresponding to a 100-to-900-fold enrichment). Even at concentrations as low as 50 CFU/mL pneumococcus can be enriched from a saliva sample, to an extent that allows easy isolation of individual colonies.
Conclusions
This method is an important development for isolation of pneumococci from clinical samples and may be particularly useful for isolation of minority serotypes in co-colonization studies. This technique will also be a useful in a laboratory setting where mixed samples need to be separated (e.g. transformation experiments).