Characterization of Tn6001 in multi-drug resistant Pseudomonas aeruginosa (MDRPA) and efficacy of antibacterial photodynamic therapy against MDRPA with Toluidine Blue O
Date Issued
2008
Date
2008
Author(s)
Tseng, Sung-Pin
Abstract
The VIM-type metallo-β-lactamase (MBL) blaVIM-3 was found in multidrug- resistant (MDR) Pseudomonas aeruginosa clinical isolates from Taiwan. We identified a Tn501-related (class II) transposon, termed Tn6001, which carries a class 1 integron In450-associated multiple drug resistance genes including blaVIM-3. Tn6001 backbone which contains the tnpA, tnpR at the region of left-end and orfB, orfC, orfD at the region of right-end is very similar to that found in Tn1404* or Tn1403. In450 gene cassette contains the following six genes: the blaVIM-3 (resistance to carbapenems), orf2 (putative fosfomycin resistance gene), two copies of aacA4 (resistance to aminogylcoside), aadB (resistance to aminogylcoside), and another copy of aacA4. In450 is related to the In4-like family which contains the 5''-conserved segment (5''-CS), 3''-CS segment and one copy of IS6100. Sequence analysis revealed that In450 is integrated upstream of the tnpR gene in the site II of the res site and lacks the site I. This finding is unlike In4 and In28 with part of res site on each side and with a 5 bp duplication of backbone sequences. I-CeuI digested pulsed-field gel electrophoresis (PFGE) analysis revealed the blaVIM-3 gene is located on the chromosome. Further, hybridization with the blaVIM-3 revealed only one band difference between an imipenem susceptible and an imipenem resistant isolate recovered from the same patient, suggesting that the gene element might have transferred horizontally. ubsequently, we investigated the antibiotic and MBLs surveillance of 316 non-duplicate isolates of carbapenem intermediately-resistant or resistant P. aeruginosa at the National Taiwan University Hospital (NTUH) from 2000 to 2005 and 26 MDR P. aeruginosa isolates collected from different parts of Taiwan from March to August 2003. Overall, 6% (18/316) colistin-only- sensitive (COS) P. aeruginosa and 4% (13/316) pan-drug-resistant (PDR) P. aeruginosa were found in NTUH and 35% (9/26) MDR P. aeruginosa isolates from six hospitals were found colistin intermediately-resistant. 33% (6/18) COS P. aeruginosa and 38% (5/13) PDR P. aeruginosa isolates carried the Tn6001 which contained a blaVIM-3-harboring integron In450 were found in different genotypes at NTUH. These finding suggest that mobile element Tn6001 might have transferred horizontally and cause the resistant genes delivery in Taiwan.DR Pseudomonas aeruginosa infection is becoming a critical problem worldwide. Currently only limited therapeutic options are available for the treatment of infections caused by MDR P. aeruginosa, therefore the development of new alternative treatments may be needed. In this study, we evaluated the antibacterial photodynamic inactivation effect on MDR P. aeruginosa using toluidine blue O (TBO) as photosensitizer followed by red light irradiation. We demonstrated 4 to 6 logs of killing effect in MDR P. aeruginosa clinical isolates. We also found that TBO caused DNA fragmentation and protein carbonylation in both MDR and susceptible P. aeruginosa clinical isolates. These results suggest that TBO is able to permeate through the membrane and subsequently induces photodamages to DNA and proteins after red light irradiation. Although eleven strains of MDR P. aeruginosa contain an active efflux pump which can pump TBO out, the efficacy of PDT among these resistant strains did not differ from those susceptible strains as shown by minimum lethal drug concentrations (MLC) assay. In summary, antimicrobial photodynamic therapy with TBO showed the same efficacy between efflux pump-positive MDR P. aeruginosa strains and non-MDR strains.
Subjects
Tn6001
MDRPA
photodynamic therapy
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