[show abstract][hide abstract] ABSTRACT: We report a rare case of lung disease caused by Mycobacterium lentiflavum in a previously healthy woman. A 54-year-old woman was referred to our hospital due to chronic cough and sputum. A computed tomography scan of the chest revealed bilateral bronchiectasis with bronchiolitis in the right middle lobe and the lingular division of the left upper lobe. Nontuberculous mycobacteria were isolated twice from three expectorated sputum specimens. All isolates were identified as M. lentiflavum by multilocus sequence analysis based on rpoB, hsp65, and 16S rRNA fragments. To the best of our knowledge, this is the first documented case of M. lentiflavum lung disease in an immunocompetent adult in Korea.
Tuberculosis and Respiratory Diseases 04/2013; 74(4):187-90.
[show abstract][hide abstract] ABSTRACT: Mycobacterium chelonae lung disease is very rare. We report a case of lung disease caused by M. chelonae in a previously healthy woman. A 69-year-old woman was referred to our hospital because of hemoptysis. A computed tomography (CT) scan of the chest revealed bronchiolitis associated with bronchiectasis in the lingular division of the left upper lobe. Nontuberculous mycobacteria were isolated three times from sputum specimens. All isolates were identified as M. chelonae by various molecular methods that characterized rpoB and hsp65 gene sequences. Although some new lesions including bronchiolitis in the superior segment of the left lower lobe developed on the chest CT scan 35 months after diagnosis, she has been followed up without antibiotic therapy because of her mild symptoms. To the best of our knowledge, this is the first case of M. chelonae lung disease in Korea in which the etiologic organisms were confirmed using molecular techniques.
Tuberculosis and Respiratory Diseases 04/2013; 74(4):191-4.
[show abstract][hide abstract] ABSTRACT: Background. Lung disease caused by Mycobacterium abscessus and Mycobacterium massiliense are variable in their clinical manifestation and progression. We hypothesized that mycobacterial genotypes represent their pathogenic phenotypes, which would result in particular genotypes being associated with disease progression. Methods. Variable number tandem repeat (VNTR) loci were selected to establish a genotype assay that was capable of differentiating patients with heterogeneous prognoses in the development cohort (48 isolates). The analysis was re-evaluated in the validation cohort (63 isolates). Results. A total of 53 M. abscessus and 58 M. massiliense isolates were assembled into three clusters based on their VNTR genotyping. The patients in cluster A were more likely to have stable disease of the nodular bronchiectatic form; 100% of M. abscessus patients and 96% of M. massiliense patients were followed without antibiotic treatment for more than 24 months after a diagnosis. In contrast, the patients in cluster B were more likely to have progressive disease of the nodular bronchiectatic form; 96% of M. abscessus patients and 81% of M. massiliense patients started antibiotic treatment within 24 months after a diagnosis. All patients in cluster C had fibrocavitary disease and started antibiotic treatment immediately after a diagnosis. The genetic distance of each clinical isolate from the reference strain was associated with the highest likelihood of disease progression and a disease phenotype of the fibrocavitary form (P<0.001). Conclusions. Mycobacterial genotyping of M. abscessus and M. massiliense may provide valuable information for predicting disease phenotype and progression.
[show abstract][hide abstract] ABSTRACT: RATIONALE: Macrolides, such as clarithromycin (CLR) and azithromycin (AZM), are frequently the only oral antibiotics that are active against Mycobacterium abscessus and Mycobacterium massiliense infections. Neither CLR nor AZM have previously been demonstrated to provide superior treatment efficacy for these infections. OBJECTIVES: We compared the treatment efficacies of CLR and AZM and determined the correlation between efficacy and induced erm(41) expression in experimental models of M. abscessus and M. massiliense infections. MEASUREMENTS AND MAIN RESULTS: In all tested M. abscessus isolates, a high level of inducible CLR resistance developed (minimal inhibitory concentration [MIC] on day 3 vs. day 14; P<0.001). Whereas the AZM MIC increased on day 14 (P<0.01 vs. day 3), the level was significantly lower than the CLR MIC on day 14 (P<0.001). However, the MICs of CLR and AZM for the M. massiliense isolates did not change. Compared to CLR, AZM presented greater antibiotic activity against M. abscessus in vitro, ex vivo, and in vivo (P<0.05), whereas both macrolides were comparably effective against M. massiliense. In M. abscessus infection, the level of erm(41) expression was higher following exposure to CLR than following exposure to AZM (P<0.001). Experiments using an erm(41)-knockout M. abscessus mutant and an M. massiliense transformant expressing M. abscessus erm(41) confirmed that erm(41) was responsible for inducible CLR resistance. CONCLUSIONS: CLR induces greater erm(41) expression and thus higher macrolide resistance than AZM in M. abscessus infection. AZM may be more effective against M. abscessus, whereas both macrolides appear to be equally effective against M. massiliense.
American Journal of Respiratory and Critical Care Medicine 08/2012; · 11.04 Impact Factor
[show abstract][hide abstract] ABSTRACT: Infections caused by Mycobacterium abscessus and Mycobacterium massiliense are on the rise among humans. Although macrolides, including clarithromycin (CLR) and azithromycin (AZM), are key antibiotics for the treatment of M. abscessus and M. massiliense infections, treatment regimens for these infections are still largely undefined. In this study, we evaluated the in vitro, ex vivo, and in vivo activities of moxifloxacin (MXF) in combination with macrolides against clinically isolated M. abscessus and M. massiliense strains. Overall, CLR, AZM, and MXF alone showed activity against both species in vitro, ex vivo, and in vivo. When MXF was combined with a macrolide against M. abscessus isolates, antagonism was observed in 65.4% (17/26) of the strains with CLR and 46.2% (12/26) of the strains with AZM in vitro as well as in 66.7% (10/15) of the strains with CLR and 40.0% (6/15) of the strains with AZM in macrophages as determined by the fractional inhibitory concentration index. In contrast, either indifferent or synergistic effects of the MXF-macrolide combinations were observed against only M. massiliense strains. Moreover, a murine infection model showed similar results. Antagonism between the MXF and macrolide combinations was observed in five out of seven M. abscessus strains, while indifferent and synergistic effects for these combinations were observed for three of the six M. massiliense strains tested, respectively. In conclusion, the activity of MXF in combination with a macrolide differed for M. abscessus and M. massiliense infections and the addition of MXF to macrolide therapy had no benefit for the treatment of M. abscessus infections.
Antimicrobial Agents and Chemotherapy 05/2012; 56(7):3549-55. · 4.57 Impact Factor
[show abstract][hide abstract] ABSTRACT: Mycobacterium abscessus subsp. bolletii is an increasing cause of human pulmonary disease and infections of the skin and soft tissues. Consistent reports of human infections indicate that M. bolletii is a highly pathogenic, emerging species of rapidly growing mycobacteria (RGM). Here we report the first whole-genome sequence of M. abscessus subsp. bolletii BD(T).
Journal of bacteriology 05/2012; 194(10):2756-7. · 3.94 Impact Factor
[show abstract][hide abstract] ABSTRACT: A disseminated infection caused by Mycobacterium avium subspecies avium (MAA) was diagnosed in a 57-yr-old male Asian elephant (Elephas maximus) housed at the Seoul Zoo, Gyeonggi, Republic of Korea. An apparent granulomatous inflammation with central caseous necrosis was evident in the lung sections. To confirm mycobacterial infection, polymerase chain reaction-restriction enzyme polymorphism analysis (PCR-RFLP) of the rpoB and hsp65 genes was performed from multiple organs and cultured bacteria. The PCR-RFLP revealed a M. avium subspecies. MAA was identified by multiplex PCR for detection of IS901 and IS1311. Thus, it is believed that MAA caused the disseminated infection in this case. Although the source of infection was not determined, the elephant may have become infected through contamination of soil and feed by free-living birds infected with MAA. This is the first reported case of disseminated infection due to MAA in a captive elephant in the Republic of Korea.
Journal of Zoo and Wildlife Medicine 12/2011; 42(4):743-6. · 0.43 Impact Factor
[show abstract][hide abstract] ABSTRACT: Tuberculosis remains a severe public health problem worldwide. Presently, genotyping is used for conducting epidemiologic and clinical studies on tuberculosis cases. We evaluated the efficacy of the repetitive sequence-based PCR (rep-PCR)-based DiversiLab™ system (bioMérieux, France) over the IS6110-restriction fragment length polymorphism analysis for detecting Mycobacterium tuberculosis. In all, 89 clinical M. tuberculosis isolates collected nationwide from Korea were used. The DiversiLab system allocated the 89 isolates to 8 groups with 1 unique isolate when a similarity level of 95% was applied. Seventy-six isolates of the Beijing family and 13 isolates of non-Beijing family strains were irregularly distributed regardless of rep-PCR groups. The DiversiLab system generated a rapid, sensitive, and standardized result. It can be used to conduct molecular epidemiologic studies to identify clinical M. tuberculosis isolates in Korea.
The Korean Journal of Laboratory Medicine 10/2011; 31(4):282-4. · 0.72 Impact Factor
[show abstract][hide abstract] ABSTRACT: We report the first case of Segniliparus rotundus pneumonia in an adult with non-cystic fibrosis bronchiectasis. All isolates were identified as S. rotundus by 16S rRNA gene sequencing and rpoB PCR-restriction analysis. Antibiotic therapy with clarithromycin and ciprofloxacin for 2 months improved the patient's condition and achieved successful sputum conversion.
Journal of clinical microbiology 07/2011; 49(9):3403-5. · 4.16 Impact Factor
[show abstract][hide abstract] ABSTRACT: Mycobacterial speciation by polymerase chain reaction (PCR)-restriction fragment length polymorphism analysis (PRA) of the rpoB gene was evaluated for identification of Mycobacterium avium subsp. paratuberculosis (MAP) and other Mycobacterium avium complex (MAC) members to the species or subspecies level by comparison with conventional methods including hsp65 sequencing, high-performance liquid chromatography, and PCR for accepted species- or subspecies-specific genomic targets. A total of 185 type and clinical mycobacterial strains from humans, animals, and environments were tested. A 360-bp PCR product was subsequently digested with MspI, HaeIII, and SmaI restriction enzymes. The PRA using SmaI restriction showed a unique digestion pattern for MAP distinguishing it from other MAC members and other Mycobacterium spp. Moreover, HaeIII and MspI restriction of the rpoB gene enabled MAC-species and -subspecies discrimination. The rpoB-PRA using SmaI or MspI and HaeIII restriction of the rpoB gene is a simple, convenient, and reliable confirmatory assay for simultaneous identification of MAP and other MAC members.
[show abstract][hide abstract] ABSTRACT: Mycobacteria encounter many different cells during infection within their hosts. Although alveolar epithelial cells play an essential role in host defense as the first cells to be challenged upon contact with mycobacteria, they may contribute to the acquisition of mycobacterial virulence by increasing the expression of virulence or adaptation factors prior to being ingested by macrophages on the side of pathogens. From this aspect, the enhanced virulence of nonpathogenic Mycobacterium smegmatis (MSM) passed through human alveolar A549 epithelial cells (A-MSM) was compared to the direct infection of MSM (D-MSM) in THP-1 macrophages and mouse models. The intracellular growth rate and cytotoxicity of A-MSM were significantly increased in THP-1 macrophages. In addition, compared to D-MSM, A-MSM induced relatively greater interleukin (IL)-1β, IL-6, IL-8, IL-12, TNF-α, MIP-1α, and MCP-1 in THP-1 macrophages. As a next step, a more persistent A-MSM infection was observed in a murine infection model with the development of granulomatous inflammation. Finally, 58 genes induced specifically in A-MSM were partially identified by differential expression using a customized amplification library. These gene expressions were simultaneously maintained in THP-1 infection but no changes were observed in D-MSM. Bioinformatic analysis revealed that these genes are involved mainly in bacterial metabolism including energy production and conversion, carbohydrate, amino acid, and lipid transport, and metabolisms. Conclusively, alveolar epithelial cells promoted the conversion of MSM to the virulent phenotype prior to encountering macrophages by activating the genes required for intracellular survival and presenting its pathogenicity.
Medical Microbiology and Immunology 02/2011; 200(3):177-91. · 3.55 Impact Factor
[show abstract][hide abstract] ABSTRACT: Single-nucleotide polymorphism (SNP) analysis is a powerful strategy for large-scale molecular population studies examining phylogenetic relationships among bacterial strains. Mycobacterial interspersed repetitive units-variable number of tandem repeats (MIRU-VNTR) can be easily digitized to share data among laboratories. This study applied SNP and MIRU-VNTR analyses for molecular strain typing of Mycobacterium tuberculosis isolates collected throughout Korea.
We studied 102 clinical M. tuberculosis isolates, including 6 paired strains, collected from 11 university hospitals in Korea in 2008 and 2009. SNPs were detected using hairpin primer assays, and then, MIRU-VNTR analysis was performed.
Thirty-five SNPs contained polymorphisms that helped differentiate the 96 tested isolates. The isolates were classified into 15 clusters. The Beijing family strains were distributed within closely related clusters in the SNP dendrogram. For MIRU-VNTR analysis, the 96 isolates were divided into 12 groups. The discriminatory index in 8 of these groups (MIRU-10, -23, -26, and -31; ETR-A, -B, -C, and -F) was high (Hunter-Gaston diversity index > 0.6). Unlike the SNP method, MIRU-VNTR analysis did not identify any notable localizations of Beijing or non-Beijing family isolates in specific clusters.
SNP and MIRU-VNTR analyses are surrogate molecular strain-typing methods for M. tuberculosis in Korea where Beijing family isolates are predominant.
The Korean Journal of Laboratory Medicine 01/2011; 31(1):37-43. · 0.72 Impact Factor
[show abstract][hide abstract] ABSTRACT: The Beijing family of Mycobacterium tuberculosis has been emerging in the world. However, there are few nationwide data of genotypic distribution in Korea. This study aimed to identify the genotypic diversity of clinical isolates of M. tuberculosis and to demonstrate the population of Beijing family in Korea. We collected 96 clinical M. tuberculosis isolates from 11 university hospitals nationwide in Korea from 2008 to 2009. We observed 24 clusters in IS6110-RFLP analysis and 19 patterns in spoligotyping. Seventy-five isolates were confirmed to be Beijing family. Two isolates of the K strain and 12 isolates of the K family strain were also found. We found that drug resistance phenotypes were more strongly associated with Beijing family than non-Beijing family (P=0.003). This study gives an overview of the distribution of genotypes of M. tuberculosis in Korea. These findings indicate that we have to pay more attention to control of M. tuberculosis strains associated with the Beijing family.
Journal of Korean medical science 12/2010; 25(12):1716-21. · 0.84 Impact Factor
[show abstract][hide abstract] ABSTRACT: A novel duplex PCR method based on variable-number tandem-repeat targets to discriminate among Mycobacterium abscessus complex isolates was developed and evaluated in 85 clinical isolates. The assay accuracy was confirmed by a multiple-target sequence analysis. The duplex PCR assay is a one-step, reliable, and accurate assay for discriminating M. abscessus species.
Journal of clinical microbiology 12/2010; 49(3):1107-9. · 4.16 Impact Factor
[show abstract][hide abstract] ABSTRACT: We evaluated high-resolution melting (HRM) curve analysis as a tool for detecting rifampin (RIF) and isoniazid (INH) resistance in Mycobacterium tuberculosis in an accurate, affordable, and rapid manner. Two hundred seventeen M. tuberculosis clinical isolates of known resistance phenotype were used. Twenty-nine known rpoB mutant DNAs, including rare mutations, were also included. Four pairs of primers were designed: rpoB-F/R (for codons 516 to 539 of rpoB), rpoB-516F/R (for codons 508 to 536 of rpoB), katG-F/R (for the codon 315 region of katG), and inhA-F/R (for the nucleotide substitution of C to T at position -15 of inhA). An HRM curve was generated for each isolate after real-time PCR differentiated the mutant from the wild-type strains. DNA sequencing of the target regions was performed to confirm the results of the HRM curve analysis. All but one of the 73 RIF-resistant (RIF-R) strains and all 124 RIF-susceptible (RIF-S) isolates were correctly identified by HRM curve analysis of rpoB. Twenty-seven of 29 known rpoB mutants were detected. In HRM curve analysis of katG and inhA, 90 INH-R strains that harbored katG or inhA mutations, or both, and all INH-S strains were correctly identified. Ten phenotypically INH-R strains not harboring katG or inhA mutations were not detected. The HRM curve analysis will be a useful method for detection of RIF and INH resistance in M. tuberculosis in a rapid, accurate, simple, and cost-effective manner.
Journal of clinical microbiology 11/2010; 48(11):3893-8. · 4.16 Impact Factor
[show abstract][hide abstract] ABSTRACT: Clonorchis sinensis is still a common parasite in South Korea. An enzyme-linked immunosorbent assay (ELISA) was developed to replace the usual intradermal test, and its performance in an area of endemicity was evaluated. From 2004 to 2006, 182 adults were included. The patient group consisted of 51 adults; 43 patients showed fluke eggs by microscopy, and 8 had their disease diagnosed clinically. The negative-control group included 131 adults: 98 at low risk and 33 at high risk of infestation. Both intradermal testing and ELISA were performed for all patients, and the sensitivity and specificity were evaluated. Statistical analysis for specificity looked at two groups: those at high risk and those at low risk. The sensitivity of the C. sinensis ELISA was 80.4%, while that of the intradermal test was 56.9% (P = 0.019). The specificities of the C. sinensis ELISA in the low-risk and high-risk groups were 93.9% and 33.3%, respectively, while those of the intradermal test were 85.7% and 30.3%, respectively (P = 0.0968, P = 0.9979). The C. sinensis ELISA may be more useful than the intradermal test; however, because of its low specificity, it may not be used independently for the diagnosis of C. sinensis infestation or a prevalence survey in a high-risk group.
Journal of clinical microbiology 07/2010; 48(7):2365-7. · 4.16 Impact Factor
[show abstract][hide abstract] ABSTRACT: It is important to identify and test serologically active antigens in order to devise a mixture of antigens or peptides that is most useful for serodiagnosis. This study evaluated the serodiagnostic potential of CysA2, which has not previously been described as a serological antigen, together with those of PstS1, HspX, antigen 85 complex and CFP-10 proteins.
Serum IgG antibody titres against each antigen and a mixture of the antigens were measured by ELISA, in subjects with pulmonary tuberculosis and in healthy control subjects.
CysA2 showed diagnostic value comparable to that of PstS1 and HspX. Mixtures of these three proteins provided the highest diagnostic sensitivity. CysA2 was useful for identifying patients who did not react to HspX or PstS1, and was most valuable in increasing the sensitivity of testing. Furthermore, CysA2 efficiently overcame the limitation associated with use of PstS1, that is, significantly lower sensitivity for subjects who are negative for acid-fast bacilli.
These findings suggest that CysA2 can be used in combination with HspX and/or PstS1 to increase the accuracy of tuberculosis diagnoses.
[show abstract][hide abstract] ABSTRACT: Oligonucleotide chips targeting the bacterial internal transcribed spacer region (ITS) of the 16S-23S rRNA gene, which contains genus- and species-specific regions, were developed and evaluated. Forty-three sequences were designed consisting of 1 universal, 3 Gram stain-specific, 9 genus-specific, and 30 species-specific probes. The specificity of the probes was confirmed using bacterial type strains including 54 of 52 species belonging to 18 genera. The performance of the probes was evaluated using 825 consecutive samples that were positive by blood culture in broth medium. Among the 825 clinical specimens, 708 (85.8%) were identified correctly by the oligonucleotide chip. Most (536 isolates, or 75.7%) were identified as staphylococci, Escherichia coli, or Klebsiella pneumoniae. Thirty-seven isolates (4.5%) did not bind to the corresponding specific probes. Most of these also were staphylococci, E. coli, or K. pneumoniae and accounted for 6.3% of total number of the species. Sixty-two specimens (7.5%) did not bind the genus- or species-specific probes because of lack of corresponding specific probes. Among them, Acinetobacter baumannii was the single most frequent isolate (26/62). The oligonucleotide chip was highly specific and sensitive in detecting the causative agents of bacteremia directly from positive blood cultures.
Journal of clinical microbiology 03/2010; 48(5):1578-83. · 4.16 Impact Factor
[show abstract][hide abstract] ABSTRACT: Antimicrobial susceptibility of Legionella spp. has rarely been studied in Korea. Therefore, we aimed to determine the susceptibility of Legionella spp. to various antibiotics.
We assessed the antimicrobial susceptibility of 66 environmental and clinical Legionella isolates collected between January 2001 and December 2008 from Korea and Japan. The minimum inhibitory concentrations (MICs) of 6 antibiotics, namely, azithromycin, ciprofloxacin, clarithromycin, clindamycin, gatifloxacin, and gemifloxacin were determined by the broth microdilution method using buffered starch yeast extract broth.
The MIC ranges of the 6 antibiotics used against the Legionella isolates were as follows: 0.004-0.062 microg/mL (azithromycin), 0.002-0.5 microg/mL (ciprofloxacin), 0.004-0.5 microg/mL (clarithromycin), 0.12-4 microg/mL (clindamycin), 0.002-0.12 microg/mL (gatifloxacin), and 0.008-1 microg/mL (gemifloxacin).
Legionella spp. isolates from Korea and Japan were most susceptible to gatifloxacin. Azithromycin, clarithromycin, ciprofloxacin, and gemifloxacin were also effective for treating legionellosis.
The Korean Journal of Laboratory Medicine 02/2010; 30(1):28-33. · 0.72 Impact Factor
[show abstract][hide abstract] ABSTRACT: Mycobacterium tuberculosis is one of the most clinically significant infectious agents. Especially during mass outbreaks, accurate identification and monitoring are required. The proportion of Beijing family members is very high among infecting strains, and spoligotyping is not suitable for strain typing. Therefore, we studied the homogeneity of isolates using the mycobacterial interspersed repetitive units-variable number of tandem repeats (MIRU-VNTR) method and identified its utility for carrying out molecular epidemiologic analysis.
Eighty-one clinical M. tuberculosis isolates that had previously been analyzed by spoligotyping were used in this study. We used the 12 standard MIRU loci and further four exact tandem repeat (ETR) loci (ETR-A, -B, -C, and -F). Four strains each of randomly selected Beijing and Beijing-like families were subjected to IS6110- restriction fragment length polymorphism analysis.
All 81 samples showed amplification products of all VNTR loci, and all of them showed differences in at least one locus. The calculation of the Hunter-Gaston diversity index (HGDI) for MIRU-VNTR gave the value of 0.965. Discriminatory index in the six loci (MIRU-10, -16, -26, -31, -39, and ETR-F) were found to be highly discriminated (HGDI >0.6). Beijing and Beijing-like family isolates were discriminated into different MIRU-VNTR types.
MIRU-VNTR analysis by using well-selected loci can be useful in discriminating the clinical M. tuberculosis isolates in areas where the Beijing family is predominant.
The Korean Journal of Laboratory Medicine 08/2009; 29(4):314-9. · 0.72 Impact Factor