Comparative Experimental Evolution of Gene Essentiality in Streptococcus sanguinis and S. mutans

Title: Comparative Experimental Evolution of Gene Essentiality in Streptococcus sanguinis and S. mutans
Project Summary/Abstract
Streptococcus sanguinis and S. mutans are important oral microbiota. Despite both belonging to the
Streptococcal genus, these bacteria exhibit distinct roles in oral health, with S. sanguinis serving as a pioneer
colonizer and commensal on teeth, while S. mutans is notorious for its major pathogenic role in dental caries
development. The contrasting virulence in dental caries is underscored by the similarities and differences in
their essential genomes. S. sanguinis SK36 possesses 218 essential genes (EGs), whereas S. mutans UA159
has 295. As key regulators for fitness and viability, EGs control the pathogenesis of these bacteria. However,
our understanding of gene essentiality has been hindered by the traditional belief that generating EG-deletion
mutants is impractical due to the indispensable nature of these genes for survival. Recognizing the quantitative
characteristics of gene essentiality, we have recently developed a high-throughput method that enables the
generation of dozens of EG deletion mutants in S. sanguinis SK36. We categorized the 218 EGs into three
groups:186 “non-evolvable EGs”, 23 “evolvable EGs” and 9 “conditional EGs”, marking a significant
advancement across the entire Streptococcus species. We hypothesized that A) the list of “evolvable EGs” and
“non-evolvable EGs” and B) the profile of suppressor genes for the same “evolvable EGs” are different
between S. mutans and S. sanguinis. Using an experimental evolution approach combined with whole-genome
sequencing, we have identified >1000 suppressor mutation in evolved populations deled of “evolvable EGs” of
S. sanguinis SK36. These suppressors were further be classified into three categories: “activation,” “inhibition,”
and “unknown.” in the previous study. Our primary objective is to categorize the 295 EGs in S. mutans UA159,
identify and classify corresponding suppressor mutations for its evolvable EGs. By examining essential gene
mutants and their corresponding suppressors, we aim to construct a comprehensive functional network of gene
essentiality. Through a comparative analysis of suppressor mutation profiles evolved in essential-gene mutants
between S. sanguinis and S. mutans, our goal is to comprehend gene essentiality of these two Streptococcus
species. This thorough examination will provide valuable insights into the conservation and diversity of
essential gene functions within the broader context of Streptococcus biology. Ultimately, these findings will be
instrumental in rational design of targeting S. mutans-specific anti-caries strategies by targeting EGs whose
deletion is lethal in S. mutans, while leaving S. sanguinis unaffected or allowing for compensatory mechanisms
through suppressor mutations.

Grant Number: 1R03DE034511-01A1
NIH Institute/Center: NIH
Principal Investigator: Liang Bao