Abstract
Investigation of the in vitro activity of tigecycline against Burkholderia pseudomallei and Burkholderia thailandensis revealed that the inhibition zone diameters of tigecycline against all isolates were ≥20 mm and that the MIC50 values were 0.5 and 1 μg/ml and the MIC90 values were 2 and 1.5 μg/ml for B. pseudomallei and B. thailandensis, respectively.
Burkholderia pseudomallei, a gram-negative bacterium, causes in humans and animals a disease called melioidosis (22). The bacterium is a soil organism found mainly in Southeast Asia and northern Australia. Recently, two distinct biotypes of B. pseudomallei strains have been defined based on their ability to assimilate l-arabinose and their difference in pathogenicity (2, 7, 17, 23). Both biotypes have been found in soil of areas where melioidosis is endemic in Thailand (20, 21). The Ara+ B. pseudomallei is much less virulent than Ara− B. pseudomallei (2, 17). However, Ara+ B. pseudomallei has been reported to cause disease in humans (11). Subsequently, a distinct new species, Burkholderia thailandensis, was proposed for the Ara+ B. pseudomallei strain (3). B. pseudomallei is usually resistant to many antibiotics. Antibiotics currently recommended for therapy of melioidosis are ceftazidime, imipenem, meropenem, amoxicillin/clavulanate, cefoperazone/sulbactam, trimethoprim-sulfamethoxazole, doxycycline, and chloramphenicol (22). The development of resistance of B. pseudomallei to these antibiotics was recognized (6, 8, 19, 24), and hence a search for new agents effective against B. pseudomallei is needed.
Tigecycline is a glycylcycline antibiotic that shows promising activity against a wide range of organisms (25). Tigecycline is active against gram-positive cocci, including methicillin-resistant staphylococci, penicillin-resistant Streptococcus pneumoniae, and vancomycin-resistant enterococci. Tigecycline is also active against many gram-negative bacilli, including those resistant to multiple antibiotics as well as anaerobes. However, the activity of tigecycline against B. pseudomallei has not been reported. The present study was undertaken to explore the activity of tigecycline against B. pseudomallei and B. thailandensis.
One hundred twenty-six strains of B. pseudomallei and B. thailandensis were selected from our collection. One hundred two strains of B. pseudomallei were isolated from different infected patients, and 24 strains of B. thailandensis were isolated from 23 soil samples collected from different sites and from one infected patient. All Burkholderia species were identified with the API 20NE (bioMerieux, France). B. pseudomallei and B. thailandensis were differentiated by the arabinose assimilation test (20). In vitro susceptibilities were determined by Kirby-Bauer disk diffusion, Etest, and MicroScan. Paper disks containing tigecycline at 15 μg per disk (Becton Dickinson), Etest strips (AB Biodisk), and gram-negative MicroScan MIC panels (Dade Behring Inc.) were provided by Wyeth Research. Susceptibility testing was done by direct colony suspension according to guidelines suggested by CLSI (4). Quality control was performed by testing the susceptibility of Escherichia coli ATCC 25922 as recommended by Wyeth Research.
The distribution of inhibition zone diameters of tigecycline against B. pseudomallei and B. thailandensis is shown in Table 1. All strains had an inhibition zone diameter of ≥20 mm. The MIC50 and MIC90 values of tigecycline as determined by Etest are shown in Table 2. The MIC50 values were 0.5 and 1 μg/ml for B. pseudomallei and B. thailandensis, respectively. The MIC90 values were 2 and 1.5 μg/ml for B. pseudomallei and B. thailandensis, respectively. There was a significant correlation between inhibition zone diameters and MICs from Etest (P < 0.001; r = −0.68). The mean inhibition zone diameters of the strains with MIC of 3, 2, 1.5, 1. 0.75, and 0.5 μg/ml were 20, 21.8, 22.6, 23.5. 24.4, and 26.7 mm, respectively. The correlation of MICs determined by Etest and MicroScan was satisfactory, as shown in Table 3. Thirty-two strains (50%) had identical MICs determined by Etest and MicroScan, whereas another 50% had a difference in MICs of 0.25 to 0.5 μg/ml.
TABLE 1.
Distribution of tigecycline susceptibility as determined by disk diffusion for B. pseudomallei and B. thailandensis
| Organism (no. of isolates) | No. (%) of isolates for which the inhibition zone diameter (mm) was:
|
|||||||||
|---|---|---|---|---|---|---|---|---|---|---|
| 20 | 21 | 22 | 23 | 24 | 25 | 26 | 27 | 28 | 30 | |
| B. pseudomallei (102) | 12 (11.8) | 16 (15.7) | 26 (25.5) | 21 (20.6) | 2 (2) | 24 (23.5) | 1 (1) | |||
| B. thailandensis (24) | 1 (4.2) | 3 (12.5) | 5 (20.8) | 13 (54.2) | 1 (4.2) | 1 (4.2) | ||||
TABLE 2.
MICs of tigecycline for B. pseudomallei and B. thailandensis as determined by Etest
| Organism (no. of isolates) | MIC (μg/ml)
|
||
|---|---|---|---|
| Range | For 50% of isolates | For 90% of isolates | |
| B. pseudomallei (102) | 0.5-3 | 0.5 | 2 |
| B. thailandensis (24) | 0.5-1.5 | 1 | 1.5 |
TABLE 3.
Correlation between MICs determined by Etest and MicroScan
| MIC (μg/ml) determined by MicroScan | No. of strains with the following MIC (μg/ml) determined by Etest:
|
||||
|---|---|---|---|---|---|
| 0.50 | 0.75 | 1.00 | 1.50 | 2.00 | |
| 0.50 | 1 | 1 | 1 | ||
| 0.75 | |||||
| 1.00 | 1 | 2 | 22 | 24 | |
| 1.50 | |||||
| 2.00 | 3 | 9 | |||
The MICs of tigecycline for B. pseudomallei and B. thailandensis observed in our study were higher than those for S. pneumoniae, Staphylococcus aureus, Enterococcus spp., and non-ESBL-producing Enterobacteriaceae (1). However, they were comparable to the MICs of tigecycline for Acinetobacter spp., Enterobacter aerogenes, and ESBL-producing Klebsiella pneumoniae (1). The breakpoints for inhibition zone diameter and MIC of tigecycline against B. pseudomallei and B. thailandensis are not available. The breakpoint of doxycycline against B. pseudomallei, adapted from data compiled by the National Committee for Clinical Laboratory Standards for similar organisms to be used for susceptibility testing, was 4 μg/ml (15). The U.S. FDA-approved breakpoints of tigecycline against Enterobacteriaceae to be used by local laboratories were an inhibition zone diameter of ≥19 mm and a MIC of ≤2 μg/ml (4). With the aforementioned breakpoints used to determine susceptibility of Burkholderia spp. to tigecycline, all isolates of B. pseudomallei and B. thailandensis were susceptible to tigecycline according to the inhibition zone diameter criteria and 98% of B. pseudomallei isolates and all B. thailandensis isolates were susceptible to tigecycline according to the MIC criteria. The MICs determined by Etest were significantly correlated with the inhibition zone diameters and the MICs determined by MicroScan. Therefore, disk diffusion and Etest methods are reliable for determination of susceptibility of tigecycline against B. pseudomallei and B. thailandensis.
Pharmacokinetic and pharmacodynamic studies of tigecycline in healthy subjects after a 100-mg loading dose given intravenously followed by 50 mg every 12 h have been reported (5, 12-14, 18). The mean maximum concentration (Cmax), the mean time to maximum concentration, the mean minimum concentration (Cmin), the mean area under the curve, and the mean half-life of tigecycline in serum were 0.72 μg/ml, 0.52 h, 0.1 μg/ml, 1.73 μg · h/ml, and 15 h, respectively. These profiles were favorable for many organisms, such as S. pneumoniae, Chlamydia pneumoniae, Moraxella catarrhalis, Mycoplasma pneumoniae, and Haemophilus influenzae, since the MIC90 values of tigecycline for such organisms were very low. The mean Cmax of tigecycline in serum (0.72 μg/ml) after the conventional dose of tigecycline (100-mg loading dose followed by 50 mg every 12 h) was above the MICs of tigecycline for only 6% of B. pseudomallei and B. thailandensis isolates. However, it was found that tigecycline had a large volume of distribution (7 to 10 liters/kg), indicating extensive distribution into the tissues (13). In addition, the mean Cmax, the mean time to maximum concentration, the mean Cmin, the mean area under the curve, and the mean half-life of tigecycline in the alveolar cells were 15.2 μg/ml, 2 h, 6.4 μg/ml, 134 μg · h/ml, and 23.7 h, respectively (5). These observations imply that tigecycline accumulates in the cells and could be effective for infections caused by intracellular organisms. The mean Cmax and the mean Cmin of tigecycline in the alveolar cells were much higher than the MICs for all isolates of B. pseudomallei and B. thailandensis. Therefore, tigecycline should be a suitable antibiotic for therapy of melioidosis, since B. pseudomallei is an intracellular bacterium (9, 10, 16). This hypothesis needs further investigation by conducting clinical trials on therapy of melioidosis with tigecycline.
Acknowledgments
We thank Wyeth Research for providing tigecycline susceptibility disks, Etest strips, and gram-negative MicroScan tests.
We thank the Thailand Research Fund for supporting the study.
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