Combination of altered PBPs and expression of cloned extended-spectrum beta-lactamases confers cefotaxime resistance in Haemophilus influenzae.
BACKGROUND
Resistance to beta-lactams in Haemophilus influenzae is mostly due to the presence of TEM beta-lactamases. beta-Lactamase-negative ampicillin resistance (BLNAR) also occurs as a result of PBP3 modifications. BLNAR strains are particularly common in Japan and France, and strains with combined mechanisms of altered PBP3 and beta-lactamase (BLPACR) are emerging. Although the prevalence of beta-lactamase-positive strains is high, TEM-derived extended-spectrum beta-lactamase (ESBL)-mediated cephalosporin resistance, which is common in Enterobacteriaceae, has not been reported in H. influenzae. In this study, the ability of ESBLs TEM-3, -4 and -5 to confer beta-lactam resistance in H. influenzae strains with or without modified PBP3 was tested.


METHODS
The genes encoding TEM-1, -3, -4 and -5 were cloned into plasmid pLS88 and were used to transform H. influenzae Rd and BLNAR H. influenzae Rd strains with PBP3 modifications. MICs were determined using CLSI macrobroth dilution. The ability of amoxicillin/clavulanate pre-diffusion disc tests to discriminate between ESBL-producing strains and other strains was also tested.


RESULTS
The presence of ESBLs in BLNAR strains conferred cefotaxime resistance, but H. influenzae Rd strains expressing ESBLs and BLNAR strains expressing TEM-1 remained cefotaxime susceptible. The amoxicillin/clavulanate pre-diffusion tests were able to discriminate the ESBL-producing strains from the other strains.


CONCLUSIONS
The presence of altered PBP3 increases the effect of ESBLs in recombinant H. influenzae strains to produce cefotaxime resistance. Amoxicillin/clavulanate pre-diffusion disc tests can discriminate ESBL-producing strains from other strains.
Bozdogan
Bülent
B
Mikrobiyoloji ve Klinik Mikrobiyoloji, Adnan Menderes Universitesi Hastanesi, Aydin, Turkey. bbozdogan@adu.edu.tr
Tristram
Stephen
S
Appelbaum
Peter C
PC
D000900:Anti-Bacterial Agents / Q000494:pharmacology*; D002439:Cefotaxime / Q000494:pharmacology*; D018550:Cephalosporin Resistance; D003001:Cloning, Molecular; D006193:Haemophilus influenzae / Q000187:drug effects* / Q000201:enzymology / Q000235:genetics / Q000378:metabolism; D006801:Humans; D008826:Microbial Sensitivity Tests; D046915:Penicillin-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D011995:Recombination, Genetic; D001618:beta-Lactamases / Q000235:genetics / Q000378:metabolism*
D000900:Anti-Bacterial Agents; D046915:Penicillin-Binding Proteins; D001618:beta-Lactamases; D002439:Cefotaxime