To some extent, our understanding is limited by the methods used to detect and characterize multiple carriage. Ideally, a new method should detect multiple serotypes directly from the specimen (i.e., without a culture step which may alter the relative proportions of various strains) without

false positive reactions, and be quantitative, affordable, practical and capable of detecting all known serotypes. Although many potential methods have recently been developed they have not been sufficiently validated. The PneuCarriage project has compared 20 serotyping methods from 15 research groups, including their ability to detect multiple serotype carriage, using a well-characterized reference find more bank of samples (Satzke et al., manuscript in preparation). This project will provide further information on suitable methods for detecting multiple serotype carriage with high sensitivity and specificity. Current methods routinely underestimate the prevalence of multiple serotype carriage. Although many new techniques are in development, there is insufficient evidence to make a recommendation. For studies where multiple carriage is relevant, we Z-VAD-FMK ic50 recommend retaining the original STGG specimens

for future assessment when optimal methods are defined. A thorough comparison of methods to detect NP carriage of multiple pneumococcal serotypes from pneumococcal cultures and directly from specimens is needed. The clinical and public health importance of multiple serotype carriage needs to be determined. Several storage methods, such as lyophilization, or ULT storage on commercially available

chemically-treated beads, are appropriate for long-term storage of pure pneumococcal isolates. However, our recommendations for storage of pneumococcal isolates in STGG media are consistent with the 2003 methods [1], but with some minor amendments to reflect the breadth of consensus practice. The storage of at least one tube of each pneumococcal isolate is recommended. To do this inoculate (using a swab or loop) a fresh, overnight, pure lawn culture into suitable media, such as STGG, under aseptic conditions. After ensuring the growth is homogenized, for example by a short vortex step, freeze at ULT. Short-term storage (<12 months) of these high-titer stocks at −20 °C in a non-defrosting freezer is acceptable, these although survival will decrease over this time [33] and [37]. To recover the isolate, a small amount of frozen material can be scraped from the surface of the STGG medium, or the entire volume thawed and an aliquot taken. The scraping or aliquot is then usually inoculated onto solid medium to check for purity of the isolate. Recovery of isolates should be undertaken aseptically, with a view to minimizing temperature fluctuations of the stored isolate by, for example, keeping tubes on dry-ice (or if necessary, and for short periods, wet ice) when handling them, and only processing a few tubes at a time.