Novel Genes Required for the Fitness of Streptococcus pyogenes in Human Saliva

Abstract

The human bacterial pathogen Streptococcus pyogenes (group A streptococcus [GAS]) causes more than 600 million cases of pharyngitis annually worldwide, 15 million of which occur in the United States. The human oropharynx is the primary anatomic site for GAS colonization and infection, and saliva is the first material encountered. Using a genome-wide transposon mutant screen, we identified 92 GAS genes required for wild-type fitness in human saliva. Many of the identified genes are involved in carbohydrate transport/metabolism, amino acid transport/metabolism, and inorganic ion transport/metabolism. The new information is potentially valuable for developing novel GAS therapeutics and vaccine research.

Genome-Wide Assessment of Streptococcus agalactiae Genes Required for Survival in Human Whole Blood and Plasma

“…CNCTC 10/84 is considered a hypervirulent strain, in part due to its enhanced betahemolytic and cytolytic activities (30). Using the transposon plasmid pGh9:ISS1 (60, 61), we successfully generated a dense transposon mutant library from CNCTC 10/84 via a technique described previously (60,62,63). This mutant library contained 193,313 unique insertions, which corresponded to an average distance between insertions of 10.4 nucleotides.…”

“…1C). TraDIS was used to identify genes with a significantly altered mutant frequency in the output mutant pools relative to the input pools, as described previously (62)(63)(64). Genes with a significantly decreased mutant frequency (log 2 fold change of Ͼ1, q value of Ͻ0.1) in the output mutant pools were regarded as important for growth and persistence in whole blood (Fig.…”

“…Generation of GBS transposon insertion mutant library. A transposon mutant library was generated in strain CNCTC 10/84 using a recently described protocol (62). Briefly, transposon plasmid pGh9:ISS1 was transformed into strain CNCTC 10/84 by electroporation.…”

“…DNA preparation and massive parallel sequencing. Preparation of genomic DNA and sequencing of the TraDIS libraries were performed as previously described (60,62). Briefly, genomic DNA of mutant pools collected at each time point was isolated using the DNeasy blood and tissue kit (Qiagen).…”

“…Processing of TraDIS sequencing reads and data analysis. Processing of TraDIS sequencing reads and data analysis were performed as previously described (62)(63)(64). Sequencing reads were analyzed with the TraDIS toolkit (24).…”