Assessment of extended-spectrum ß-lactamase, KPC carbapenemase and porin resistance mechanisms in clinical samples of Klebsiella pneumoniae and Enterobacter spp.

The emergence and spread of resistance mechanisms in Gram-negative bacilli has complicated the treatment of serious nosocomial infections. Current automated systems for detection of Klebsiella pneumoniae carbapenemase (KPC)-producing isolates are unreliable. One possible straightforward alternative method is evaluation of ertapenem resistance. However, the accuracy of this method is affected by other resistance mechanisms such as AmpC gene expression or extended-spectrum ß-lactamase production associated with porin loss. This study included 128 samples of K. pneumoniae and Enterobacter spp. that were non-susceptible to ertapenem. The disk diffusion and Etest method were applied to determine susceptibility to imipenem, meropenem and ertapenem. Isolates exhibiting intermediate or complete resistance to ertapenem were evaluated for resistance mechanisms. bla(TEM), bla(SHV), bla(CTX-M), bla(CTX-M-2) and bla(KPC) genes were tested for by PCR, and the presence of outer membrane protein was investigated by dot-blot assay. bla(TEM) was detected in 52.9% and 10.3%, bla(SHV) in 29.4% and 0.94%, bla(CTX-M) in 41.4% and 1.9% and bla(CTX-M-2) in 23.5% and 1.9% of K. pneumoniae and Enterobacter cloacae isolates, respectively. The bla(KPC) gene was present in 12.6% of Enterobacter spp. isolates. OmpC and OmpF were present in 6.6% of E. cloacae isolates. These results indicate that several resistance mechanisms contribute to potential therapeutic failure of carbapenem therapy and point to the need for better detection methods and surveillance strategies.

Avaliação preliminar da atividade antimicrobiana do extrato de erva-mate (Ilex paraguariensis A. St. - Hil.) obtido por extração com CO2 supercrítico / Preliminary evaluation of the antimicrobial activity of "erva mate" (Ilex paraguariensis A. St. - Hil.) extract obtained by using supercritical CO2

Ilex paraguariensis A. St. - Hil. (erva-mate) é uma árvore pertencente à família Aquifoleaceae, nativa da região sul da América do Sul e de grande importância cultural e econômica para os três estados da região sul do Brasil. O presente trabalho determinou o potencial antimicrobiano do extrato de erva-mate obtido por fluido supercrítico, frente a alguns microrganismos. As folhas de Ilex paraguariensis foram coletadas na região norte do RS. O extrato bruto da planta desidratada foi obtido pelo método de extração com fluido supercrítico, utilizando dióxido de carbono (CO2) como solvente. Foi obtida a solução mãe desse extrato (100 mg mL-1) que a seguir foi diluído (50; 25; 12,5; 6,25 mg mL-1) para avaliação da atividade antimicrobiana por meio da técnica de difusão do disco em ágar Müeller Hinton, sobre cepas de Acinetobacter baumannii (ATCC 19606), Escherichia coli (ATCC 25922), Pseudomonas aeruginosa (ATCC 27853), Staphylococcus aureus (ATCC 25923) e Enterococcus faecalis (ATCC 29212). As análises foram efetuadas em triplicata, utilizando como controle negativo disco estéril sem impregnação e como controle positivo discos de antibióticos comerciais variáveis de acordo com a espécie bacteriana. Após 16 a 24h de incubação a 35ºC os halos com significância foram medidos. Observou-se atividade antimicrobiana frente a S. aureus (halos de inibição variaram entre 7 e 18 mm) e para P. aeruginosa (halos entre 6 e 11 mm), ambos nas concentrações de 100 até 25 mg mL-1. O extrato obtido por extração em fluido supercrítico de Ilex paraguariensis apresentou atividade antimicrobiana frente a S. aureus e P. aeruginosa.

Ilex paraguariensis A. St. - Hil. ("erva mate") is a tree of the Aquifoliaceae family, native to the south region of South America and of great cultural and economic importance for the three states of the south region of Brazil. The present study aimed to determine the antimicrobial potential of "erva mate" extract obtained by supercritical fluid against some microorganism strains. The leaves of Ilex paraguariensis were collected in the north region of Rio Grande do Sul State. The crude extract of dehydrated plant was obtained by supercritical fluid extraction, using carbon dioxide (CO2) as solvent. The extract mother solution (100 mg mL-1) was diluted (50; 25; 12, 5; 6.25 mg mL-1) for antimicrobial activity evaluation, by means of disk diffusion method on Müeller-Hinton agar, against strains of Acinetobacter baumannii (ATCC 19606), Escherichia coli (ATCC 25922), Pseudomonas aeruginosa (ATCC 27853), Staphylococcus aureus (ATCC 25923) and Enterococcus faecalis (ATCC 29212). The analyses were done in triplicate, using as negative control sterile disk without impregnation and as positive control, commercial antibiotic disks according to the bacterial species. After 16-24h incubation at 35ºC, the diameters of inhibition zones were measured. Antimicrobial activity was observed against S. aureus (inhibition zones between 7 and 18 mm) and P. aeruginosa (inhibition zones between 6 and 11 mm), both at the concentrations of 100 mg mL-1 to 25 mg mL-1. The extract of Ilex paraguariensis obtained by supercritical fluid presented antimicrobial activity against S. aureus and P. aeruginosa.