tRNA modification in Streptococci
tRNA modifications are central to bacterial translational control. Here, we integrate genetics, mass spectrometry, epitranscriptomics, and comparative genomics to comprehensively map the tRNA modification genes of the Gram-positive pathogens Streptococcus mutans and Streptococcus pneumoniae. Both species show a marked loss of modifications dependent on Fe–S enzymes, consistent with a broader trend of Fe–S enzyme reduction in Streptococcus central metabolism. We further uncover  differences in queuosine (Q) metabolism: while S. mutans synthesizes Q de novo, S. pneumoniae instead salvages preQ1 and accumulates the epoxy-Q precursor, a strategy shared with multiple other Streptococci as revealed by analysis of 1,499 genomes. Comparative essentiality profiling of modification genes revealed notable differences, including the essentiality of the N6-threonylcarbamoyladenosine (t6A) synthesis enzyme TsaE in S. pneumoniae but not in S. mutans that was confirmed by genetic studies., We found that suppressor mutations in asparaginyl-tRNA synthetase (AsnRS) restored viability to ?tsaE mutants, albeit with reduced growth, highlighting previously unrecognized roles for t6A in translation.