Mutations and horizontal transmission have contributed to sulfonamide resistance in Streptococcus pyogenes

Two variants of dihydropteroate synthase (DHPS) were found among sulfonamide-resistant Streptococcus pyogenes, one of which was characterized by a 2-amino-acid addition in a conserved part of the enzyme. The enzyme kinetics of both variants was compared with the kinetics of DHPS from a sulfonamide-susceptible S. pyogenes. The most striking difference was a substantially elevated Ki for both variants, but variations in Km for both of its substrates p-aminobenzoic acid (p-AB) and dihydropteridine-pyrophosphate (pteridine) were also found. In the resistance variant lacking additions, the amino acid at position 213 was changed by site-directed mutagenesis from a Gly to an Arg, which resulted in a lower Ki. The corresponding change from an Arg to a Gly in the DHPS from a susceptible isolate led to a substantially increased Ki, confirming the importance of this amino acid difference for the resistance. Nucleotide sequence determinations of the complete folate operon revealed in some isolates a mosaic pattern of differences compared to the wild type, not only in the genes coding for DHPS and GTP cyclohydrolase (GTPCH) noted earlier but also in genes coding for dihydroneopterin aldolase (DHNA) and hydroxymethylpterin pyrophosphokinase (HPPK). Regions of sequence differences were interspersed with regions of complete identity in a mosaic pattern, indicating a dispersed pattern of uptake of foreign DNA in the resistant isolates.