GONOCOCCAL RESISTANCE TO BETA-LACTAMS AND TETRACYCLINE INVOLVES MUTATION IN LOOP-3 OF THE PORIN ENCODED AT THE PENB LOCUS

penB is a chromosomal mutation that confers resistance to p-lactams an d tetracyclines and reduced susceptibility to quinolones in Neisseria gonorrhoeae, It is linked to the porin gene (por) and requires the inc reased expression of an efflux pump due to mtr. Transformation of a su sceptible gonococcus (strain H1) with chromosomal DNA from strain FA14 0 (penA mtr penB; porin serovar IB1) and conjugal transfer of a p-lact amase-expressing plasmid was used to produce isogenic strains for dete rmination of equilibrium periplasmic penicillin concentrations by the method of Zimmermann and Rosselet (W. Zimmermann and A. Rosselet, Anti microb, Agents Chemother, 12:368-372, 1977). In transformants with the Mtr and PenB phenotypes, equilibrium concentrations of penicillin wer e reduced. DNA sequence analysis of por from isogenic penB and penB(+) transformants revealed 14 sequence differences; nine of these differe nces resulted in amino acid changes. Three amino acid changes were fou nd in the putative gonococcal equivalent of the pore-constricting loop 3 of Escherichia coli OmpF. Two of these changes (Gly-101-Ala-102-->A sp-Asp) result in an increased negative charge at this position in por loop 3, PCR products comprising the complete por gene from strain FA1 40 were transformed into strain H1-2 (penA mtr; porin serovar IB-3), w ith the resulting transformants having the antibiotic susceptibility p henotype associated with penB, penB-like mutations were found in loop 3 of clinical isolates of gonococci with chromosomally mediated resist ance to penicillin. We conclude that penB is a mutation in loop 3 of p or that reduces porin permeability to hydrophilic antibiotics and play s an important role in the development of chromosomally mediated resis tance to penicillin and tetracycline in gonococci.