Characterization of glycopeptides, aminoglycosides and macrolide resistance among Enterococcus faecalis and Enterococcus faecium isolates from hospitals in Tehran

The prevalence of glycopeptides, aminoglycosides and erythromycin resistance among Enterococcus faecalis and Enterococcus faecium was investigated. The susceptibility of 326 enterococcal hospital isolates to amikacin, kanamycin, netilmicin and tobramycin were determined using disk diffusion method. The minimum inhibitory concentration (MIC) of vancomycin, teicoplanin, gentamicin, streptomycin, and erythromycin were determined by microbroth dilution method. The genes encoding aminoglycoside modifying enzymes described as AMEs genes, erythromycin-resistant methylase (erm) and vancomycin-resistant were targeted by multiplex-PCR reaction. High level resistance (HLR) to gentamicin and streptomycin among enterococci isolates were 52% and 72% respectively. The most prevalent of AMEs genes were aac (6')-Ie aph (2") (63%) followed by aph (3')-IIIa (37%). The erythromycin resistance was 45% and 41% of isolates were positive for ermB gene. The ermA gene was found in 5% of isolates whereas the ermC gene was not detected in any isolates. The prevalence of vancomycin resistant enterococci (VRE) was 12% consisting of E. faecalis (6%) and E. faecium (22%) and all of them were VanA Phenotype. The results demonstrated that AMEs, erm and van genes are common in enterococci isolated in Tehran. Furthermore our results show an increase in the rate of vancomycin resistance among enterococci isolates in Iran.     

Kinetic and physical characterisation of recombinant wild-type and mutant human protoporphyrinogen oxidases

The effects of various protoporphyrinogen oxidase (PPOX) mutations responsible for variegate porphyria (VP), the roles of the arginine-59 residue and the glycines in the conserved flavin binding site, in catalysis and/or cofactor binding, were examined. Wild-type recombinant human PPOX and a selection of mutants were generated, expressed, purified and partially characterised. All mutants had reduced PPOX activity to varying degrees. However, the activity data did not correlate with the ability/inability to bind flavin. The positive charge at arginine-59 appears to be directly involved in catalysis and not in flavin-cofactor binding alone. The K(m)s for the arginine-59 mutants suggested a substrate-binding problem. T(1/2) indicated that arginine-59 is required for the integrity of the active site. The dominant alpha-helical content was decreased in the mutants. The degree of alpha-helix did not correlate linearly with T(1/2) nor T(m) values, supporting the suggestion that arginine-59 is important for catalysis at the active site. Examination of the conserved dinucleotide-binding sequence showed that substitution of glycine in codon 14 was less disruptive than substitutions in codons 9 and 11. Ultraviolet melting curves generally showed a two-state transition suggesting formation of a multi-domain structure. All mutants studied were more resistant to thermal denaturation compared to wild type, except for R168C.