Recently, polymerase chain reaction (PCR)-based methods have been used to reclassify Ureaplasma urealyticum into two independent species (spp.), designating U. parvum and U. urealyticum. In the current study, we aim to reclassify U. urealyticum and to analyze the correlation between the presence of a genetic marker and an antibiotic resistance of U. urealyticum. Susceptibility test against tetracycline, levofloxain or moxifloxacin was performed by broth microdilution method. The presence of tet(M) gene and the mutations of quinolone resistance-determining regions (QRDRs) were analyzed by PCR and sequencing. Among fourteen Ureaplasma isolates, three were identified as U. parvum and eleven were identified as U. urealyticum, and this is first report showing that two independent spp. of U. urealyticum isolated from Korean are present. The minimum inhibitory concentration (MIC) ranges for Ureaplasma isolates were as follows: tetracycline 0.25~128 microg/ml, levofloxacin 1~8 microg/ml, and moxifloxacin 0.5~4 microg/ml. The tet(M) determinant was found in 3 among 14 Ureaplasma isolates with tetracycline MIC of >16 microg/ml, suggesting that the presence of the tet(M) determinant is associated with tetracycline resistance. Mutations of gyrA, gyrB, parC, and parE genes in the QRDRs were found in 3 among 14 Ureaplasma isolates, exhibiting only parE gene mutation is associated with fluoroquinolone resistance.
Aza Compounds ; Drug Resistance, Microbial ; Fluoroquinolones ; Genetic Markers ; Microbial Sensitivity Tests ; Ofloxacin ; Polymerase Chain Reaction ; Quinolines ; Tetracycline ; Tetracycline Resistance ; Ureaplasma ; Ureaplasma urealyticum

Tuberculosis (TB) is an old disease caused by Mycobacterium tuberculosis. Although it has been known for humans for thousands of years, the treatment of this disease still requires a lengthy therapy with multiple antibiotics. Also, the emergence of multidrug-resistant strains made it more difficult to treat TB, calling for a novel treatment approach. In Photodynamic therapy (PDT), a photosensitizer, such as methylene blue (MB), is irradiated by a laser, generating reactive oxygen species and killing microorganisms. Here, using M. smegmatis as a model mycobacterium, we examined the utility of PDT in TB treatment. The photosensitizer MB alone showed weak antimicrobial activity; however, when irradiated by a laser, it efficiently killed M. smegmatis (> 97% killing with 30 mg/ml MB and 54 J/cm2 irradiation). Surprisingly, PDT showed more efficient killing activity toward drug-resistant strains of M. smegmatis than the drug-sensitive wild-type strain. In PDT, when the irradiation step alone (Intermittent PDT) or the entire PDT process was repeated (Repeated PDT), the bactericidal activity was significantly enhanced. Since PDT can be applied locally in a short period of time and kills mycobacterium irrespective of its antibiotic resistance status, we conclude that PDT can be a viable option for TB treatment.

Tuberculosis (TB) patients are normally treated with a combination of antibiotics. However, with improper or incomplete treatment of antibiotics, the disease may progress to multidrug-resistant TB (MDR-TB). The treatment of MDR-TB is very costly and inefficient. Therefore, there is a great demand of new therapeutic approaches for MDR-TB such as photodynamic therapy. In this study, we tried to optimize the conditions for photodynamic inactivation of TB using methylene blue as a photosensitizer. Different combinations of methylene blue concentrations and light doses were tested for their photodynamic effects to A549 cells or Mycobacterium smegmatis (M. smegmatis). We also tested the effect of photodynamic therapy on ciprofloxacin-resistant M. smegmatis. Methylene blue treatment alone did not affect the survival rates of A549 cells or bacteria up to 5 µg/ml. When the A549 and M. smegmatis cells treated with methylene blue were irradiated with laser light (wavelength, 630 nm), photodynamic inactivation of cells was increased in methylene blue concentration- and light dose-dependent manners. Interestingly, the ciprofloxacin-resistant M. smegmatis exhibited higher level of susceptibility to methylene blue-mediated photodynamic inactivation. This study suggests that photodynamic therapy at 3.6 J/cm2 in the presence of 5 µg/ml methylene blue may be an appropriate range for therapy due to the high bactericidal activity against high level of ciprofloxacin-resistant M. smegmatis and the low damaging effect to mammalian cells. This study demonstrates that photodynamic therapy could be a potential alternative for MDR-TB treatment.
Anti-Bacterial Agents ; Bacteria ; Ciprofloxacin ; Humans ; Methylene Blue ; Mycobacterium smegmatis* ; Mycobacterium* ; Photochemotherapy* ; Survival Rate ; Tuberculosis