Differential effects of frozen storage on the molecular detection of bacterial taxa that inhabit the nasopharynx

16S ribosomal RNA (rRNA) gene-based terminal restriction fragment length polymorphism (T-RFLP) analysis of nasopharyngeal (NP) swabs from twelve Gambian infants was employed. NP swabs were analysed within hours of collection and then after 30 days of storage at -70°C.There was substantial heterogeneity among subjects with respect to the effect of freezing on the number of operational taxonomic units (OTUs) detected. Nevertheless, the mean number of OTUs decreased after frozen storage and the relative abundance for 72% of the OTUs changed by less than 0.5% after deep frozen storage. There were differences in the odds of detection and relative abundance of OTUs matched with Moraxella sp., Haemophilus sp./Burkholderia sp., and Pseudomonas sp. A strong interaction between sex and the effect of freezing was found, whereby there was no significant change observed for males while the mean number of OTUs significantly declined among female infants following frozen storage.Although frozen storage of biological samples is often necessary for archiving and logistic purposes, the potential effects on the number of taxa (composition) detected in microbial community studies are significant and should not be overlooked. Moreover, genetic factors such as sex may influence the integrity of nucleic acids during the freezing process.Nasopharyngeal swabbing is an important diagnostic and epidemiological surveillance tool used to detect several pathogens in one of the most clinically relevant microbial reservoirs in the human body [1,2]. Although detection of microbes in nasopharyngeal (NP) swabs stored in skim milk-tryptone-glucose-glycerin (STGG) is often employed in epidemiology and diagnostic research [3], few studies have investigated the potential bias introduced during sample storage, which usually involves deep freezing at -20°C or -70°C [1,4-6]. Furthermore, these studies have focused on the effects of frozen storage on the culture-based detection of specific pathogens such as