[Show abstract][Hide abstract] ABSTRACT: A fast and accurate diagnosis is necessary to control and eliminate tuberculosis (TB). In Korea, TB continues to be a serious public health problem. In this study, diagnostic tests on clinical samples from patients suspected to have TB were performed and the sensitivity and specificity of the various techniques were compared. The main objective of the study was to compare various diagnostic tests and evaluate their sensitivity and specificity for detecting tuberculosis.
From January 2013 to December 2013, 170,240 clinical samples from patients suspected to have TB were tested with smear microscopy, acid-fast bacilli culture, and real-time polymerase chain reaction (PCR). The test results were compared and data were analyzed.
A total of 8216 cultures tested positive for TB (positive detection rate, 4.8%). The contamination rate in the culture was 0.6% and the isolation rate of nontuberculous mycobacteria was 1.0%. The sensitivity and specificity of smear microscopy were 56.8% and 99.6%, respectively. The concordance rate between the solid and liquid cultures was 92.8%. Mycobacterium isolates were not detected in 0.4% of the cases in the liquid culture, whereas no Mycobacterium isolates were detected in 6.8% of the cases in the solid culture. The sensitivity and specificity of real-time PCR for the solid culture were 97.2% and 72.4%, respectively, whereas the corresponding data for the liquid culture were 93.5% and 97.2%.
The study results can be used to improve existing TB diagnosis procedure as well as for comparing the effectiveness of the assay tests used for detecting Mycobacterium tuberculosis isolates.
[Show abstract][Hide abstract] ABSTRACT: In Korea, a large portion of tuberculosis (TB) patients are diagnosed and treated in private institutes. Laboratory tests are crucial for TB control. There are many possible problems using laboratory tests in the private sector. In this study, we aimed to investigate the characteristics and trends of utilizing laboratory tests for TB and mycobacterial diseases in the private sector by analyzing the National Health Insurance (NHI) database.
After selecting TB or other mycobacteria-related test items, we searched the number and cost of each item on the website of the Health Insurance Review and Assessment Service using the code of each test from 2007 to 2012.
Our data revealed that the number and cost of tests drastically increased between 2007 and 2012. Culture and molecular tests primarily contributed to the tremendous increases. For each year, concentrated smearing and fluorochrome staining were more commonly used. The number of serologic tests for latent TB infection stagnated, despite the expansion of contact investigation.
The NHI data could be considerably useful for understanding the utilization trends of laboratory tests for TB and mycobacterial diseases in Korea. Our data showed that TB laboratory systems have recently improved. In this study, many issues were noticed. Therefore, solutions to these issues are required and the continued monitoring of NHI data regarding laboratory diagnosis.
[Show abstract][Hide abstract] ABSTRACT: Background
The tuberculin skin test (TST) frequently yields false positive results among BCG-vaccinated persons thereby limiting its diagnostic value particularly in settings with high BCG vaccination rate. We determined the agreement between IGRA and TST using 2 cutoff values and identified possible relationships between the results of these tests and the development of active tuberculosis.
Adolescents aged 11–19 years in close contact with smear-positive tuberculosis cases and with normal chest radiographs were recruited from middle and high schools in South Korea. The TST was conducted by trained nurses, and blood was drawn for the QuantiFERON-TB Gold In-Tube (QFT-GIT). Participants were followed up for 2 years to check for incidence tuberculosis.
A total of 2,982 subjects were included in the study, the average age was 15.1 years (SD 1.3), 61% had BCG vaccination scars. The agreement of QFT-GIT and the TST was low (κ = 0.38, 95% CI 0.32 to 0.42) using 10 mm cutoff; however, when the 15 mm cutoff was used, the agreement was intermediate (κ = 0.56, 95% CI 0.50 to 0.61). The odds ratio (OR) for the development of active tuberculosis was 7.9 (95% CI 3.46 to 18.06) for QFT-GIT positive patients, 7.96 (95% CI 3.14-20.22) for TST/QFT-GIT+ and the OR 4.62 (95% CI 2.02 to 10.58) and 16.35 (95% CI 7.09 to 37.71) for TST 10 mm and 15 mm cutoff respectively.
The results of this study suggest that the TST cutoff point for patients aged 11–17 years would be 15 mm in other study. The OR of QFT-GIT for the development of active tuberculosis and its intermediate agreement with TST using 15 mm cutoff demonstrates its role as an adjunct diagnostic tool to current clinical practice. Positive responders to both TST and QFT-GIT at the outset may benefit from chemoprophylaxis.
[Show abstract][Hide abstract] ABSTRACT: Mycobacterium abscessus complex causes the most frequent rapidly growing mycobacterium infection of nontuberculous mycobacterium (NTM), causing pulmonary infections in Korean populations. M. abscessus, M. massiliense, and M. bolletii are members of this complex, and the discrimination of these strains is clinically important because of their different antibiotic susceptibilities. In this study, we analyzed the antibiotic susceptibility and genetic characteristics of 23M. abscessus, 22M. massiliense, and 2M. bolletii strains from Korean patients. The M. massiliense isolates obtained from the pulmonary infections were further classified into three genotypes (I, II-1, and II-2) based on the hsp65 sequence analysis, previously described. A total of 47M. abscessus complex strains from Korean patients were grouped into three major clusters by Ase I digested pulsed-field gel electrophoresis (PFGE) which reflected the taxonomic status of the species and the hsp65 genotype. Multi-locus sequence typing (MLST) patterns also reflected their taxonomic status. In conclusion, our molecular typing methods showed that the strains of M. massiliense hsp65 genotype II-1 were clearly separated from strains of M. massiliense hsp65 genotype I or II-2 as well as strains of M. abscessus or M. bolletii; thus, our data suggest that M. massiliense hsp65 genotype II-1 might represent an additional subspecies of M. massiliense.
No preview · Article · Jun 2014 · Journal of Microbiological Methods
[Show abstract][Hide abstract] ABSTRACT: Certain Escherichia coli (E. coli) strains have the ability to cause diarrheal disease. Five types of diarrheagenic E. coli have been identified, including EHEC, ETEC, EPEC, EAEC, and EIEC. To detect these five diarrheagenic types rapidly, we developed a one-step multiplex PCR (MP-PCR) assay using nine primer pairs to amplify nine virulence genes specific to the different virotypes, with each group being represented, i.e., stx1 and stx2 for EHEC, lt, sth, and stp for ETEC, eaeA and bfpA for EPEC, aggR for EAEC, and ipaH for EIEC. The PCR primers were constructed using MultAlin (http://bioinfo.genotoul.fr/multalin/multalin.html). The sensitivity and specificity of the constructed multiplex PCR primers were measured using DNA isolated from diarrheagenic E. coli strains representing each group. The limits of detection (LODs) were as follows: 5 × 10(1) cfu/ml for EHEC, 5 × 10(3) cfu/ml for ETEC expressing lt and sth, 5 × 10(4) cfu/ml for ETEC expressing stp, 5 × 10(2) cfu/ml for EPEC, 5 × 10(4) cfu/ml for EAEC, and 5 × 10(2) cfu/ml for EIEC. To confirm specificity, C. jejuni, C. perfringens, S. Typhimurium, V. parahaemolyticus, L. monocytogenes, Y. enterocolitica, B. cereus, and S. aureus were used as negative controls, and no amplification was obtained for these. Moreover, this kit was validated using 100 fecal samples from patients with diarrhea and 150 diarrheagenic E. coli strains isolated in Korea.
Full-text · Article · Mar 2014 · Journal of Microbiology and Biotechnology
[Show abstract][Hide abstract] ABSTRACT: Seventy-four Shiga toxin-producing Escherichia coli (STEC) isolates belonging to the serotype O91:H21 were isolated from 1,643 asymptomatic human carriers in a STEC outbreak at Gwangju in Korea. Although the isolates did not cause any symptoms, all of them produced Shiga toxins 1 (Stx1) and 2 (Stx2). In order to determine why these strains cause no symptoms, we explored the differences in virulence potential between the asymptomatic STEC O91:H21 isolates and symptomatic STEC O91:H21 strains (ATCC 51435 and ATCC 51434). The asymptomatic STEC O91:H21 isolates showed strongly reduced cytopathic effects compared with the symptomatic strains when intact bacterial cells were used as an inoculant. Moreover, we found a reduced adherence phenotype when testing asymptomatic strains on HeLa cells. Real-time quantitative PCR results suggest that transcriptional repression of the genes encoding type-1 fimbriae occurs in the asymptomatic isolates but not in the symptomatic strains.
Full-text · Article · May 2013 · Journal of Microbiology and Biotechnology
[Show abstract][Hide abstract] ABSTRACT: Enteroaggregative Escherichia coli (EAEC) was recently reported as a major diarrheagenic pathogen in infant and adult travelers, both in developing and developed countries. EAEC strains are known to be highly resistant to antibiotics including quinolones. Therefore in this study we have determined the various mechanisms of quinolone resistance in EAEC strains isolated in Korea.
For 26 EAEC strains highly resistant to fluoroquinolone, minimal inhibitory concentrations for fluoroquinolones were determined, mutations in the quinolone target genes were identified by PCR and sequencing, the presence of transferable quinolone resistance mechanism were identified by PCR, and the contribution of the efflux pump was determined by synergy tests using a proton pump inhibitor. The expression levels of efflux pump-related genes were identified by relative quantification using real-time PCR.
Apart from two, all tested isolates had common mutations on GyrA (Ser83Leu and Ser87Gly) and ParC (Ser80Gln). Isolates EACR24 and EACR39 had mutations that have not been reported previously: Ala81Pro in ParC and Arg157Gly in GyrA, respectively. Increased susceptibility of all the tested isolates to ciprofloxacin and norfloxacin in the presence of the pump inhibitor implies that efflux pumps contributed to the resistance against fluoroquinolones. Expression of the efflux pump-related genes, tolC, mdfA, and ydhE, were induced in isolates EACR 07, EACR 29, and EACR 33 in the presence of ciprofloxacin.
These results indicate that quinolone resistance of EAEC strains mainly results from the combination of mutations in the target enzyme and an increased expression of efflux pump-related genes. The mutations Ala81Pro in ParC and Arg157Gly in GyrA have not been reported previously the exact influence of these mutations should be investigated further.
[Show abstract][Hide abstract] ABSTRACT: We aimed at evaluating the virulence of atypical Shigella flexneri II:(3)4,7(8) by DNA microarray and invasion assay.
We used a customized S. flexneri DNA microarray to analyze an atypical S. flexneri II:(3)4,7(8) gene expression profile and compared it with that of the S. flexneri 2b strain.
Approximately one-quarter of the atypical S. flexneri II:(3)4,7(8) strain genes showed significantly altered expression profiles; 344 genes were more than two-fold upregulated, and 442 genes were more than 0.5-fold downregulated. The upregulated genes were divided into the category of 21 clusters of orthologous groups (COGs), and the "not in COGs" category included 170 genes. This category had virulence plasmid genes, including the ipa-mxi-spa genes required for invasion of colorectal epithelium (type III secretion system). Quantitative reverse-transcription polymerase chain reaction results also showed the same pattern in two more atypical S. flexneri II:(3)4,7(8) strains. Atypical S. flexneri II:(3)4,7(8) showed four times increased invasion activity in Caco-2 cells than that of typical strains.
Our results provide the intracellularly regulated genes that may be important for adaptation and growth strategies of this atypical S. flexneri.
[Show abstract][Hide abstract] ABSTRACT: Serotyping has been the gold standard for identifying Salmonella, but it requires large amounts of standard antisera. Multilocus sequence typing (MLST) has been applied to identify Salmonella serovars, but the recombination of 4-7 housekeeping genes and multiple analytic steps diminish its applicability. In the present study, we determined the complete sequences of the RNA polymerase beta subunit gene (rpoB) and 7 housekeeping genes (aroC, dnaN, hemD, hisD, purE, sucA, and thrA) for 76 strains of 33 Salmonella enterica serovars and conducted phylogenetic analyses together with the corresponding gene sequences of 24 reference strains registered in the GenBank database. Based on the phylogenetic analyses, 100 strains from 40 serovars and 91 strains from 37 serovars were classified into 60 rpoB (RST) and 49 multilocus sequence types (ST), respectively. The nucleotide similarities were 98.8-100% and 96.9-100% for the complete rpoB gene and the seven concatenated housekeeping genes, respectively. The strains of 35 and 30 serovars formed serovar-specific branches or clusters in the rpoB and housekeeping gene phylogenetic trees, respectively. Therefore, complete rpoB gene sequencing and phylogenetic analysis may be a useful method for identifying Salmonella serovars that is a simpler, more cost-effective, and less time-consuming alternative or complementary method to MLST and conventional serotyping.
No preview · Article · Dec 2012 · The Journal of Microbiology
[Show abstract][Hide abstract] ABSTRACT: Enterohemorrhagic Escherichia coli (EHEC) causes a disease involving diarrhea, hemorrhagic colitis, and hemolytic-uremic syndrome (HUS). Here we present the
draft genome sequence of NCCP15647, an EHEC isolate from an HUS patient. Its genome exhibits features of EHEC, such as genes
for verotoxins, a type III secretion system, and prophages.
Full-text · Article · Jul 2012 · Journal of bacteriology
[Show abstract][Hide abstract] ABSTRACT: Enterohemorrhagic Escherichia coli causes severe food-borne disease in the guts of humans and animals. Here, we report the high-quality draft genome sequence
of E. coli NCCP15658 isolated from a patient in the Republic of Korea. Its genome size was determined to be 5.46 Mb, and its genomic
features, including genes encoding virulence factors, were analyzed.
Full-text · Article · Jul 2012 · Journal of bacteriology
[Show abstract][Hide abstract] ABSTRACT: Shiga toxin-producing Escherichia coli causes bloody diarrhea and hemolytic-uremic syndrome and serious outbreaks worldwide. Here, we report the draft genome sequence
of E. coli NCCP15657 isolated from a patient. The genome has virulence genes, many in the locus of enterocyte effacement (LEE) island,
encoding a metalloprotease, the Shiga toxin, and constituents of type III secretion.
Full-text · Article · Jul 2012 · Journal of bacteriology
[Show abstract][Hide abstract] ABSTRACT: Whole genome-scale transcriptome analysis of enterohemorrhagic Escherichia coli (EHEC) O157:H7 EDL933 was performed to investigate the influence of mucin components on the EHEC gene expression. Here we report that the 732 candidate genes were differentially expressed by the presence of 0.5% porcine stomach mucin, including the 8 flagella-related genes. Quantitative real-time PCR analyses revealed that the transcription expression of the flg genes (encoding the structural components for flagella basal body) was down-regulated by the mucin components. Indeed, bacterial swarming motility was drastically reduced when grown on 0.3% trypton agar plates containing the mucin. These results imply that gastrointestinal (GI) mucin is a possible environmental signal which negatively regulates the flagellation of EHEC O157:H7 in the GI tract.
Full-text · Article · Jun 2012 · Biochemical and Biophysical Research Communications
[Show abstract][Hide abstract] ABSTRACT: In this study, we measured the drug resistance conferred by mdfA mutations in two Shigella flexneri strains. A mutant in mdfA genes was constructed by polymerase chain reaction-based, one-step inactivation of chromosomal genes. The antimicrobial susceptibility of parent and mutant strains to fluoroquinolones was determined by minimal inhibitory concentration (MICs). The △mdfA mutants were somewhat more susceptible to fluoroquinolones than the parent strains. The low level changes in MICs of the △mdfA mutants suggest that mdfA contributed the fluoroquinolone resistance in S flexneri. This finding found that the increased expression level of an MdfA efflux pump mediated fluoroquinolone resistance, but it is not likely a major effecter of higher resistance levels.
[Show abstract][Hide abstract] ABSTRACT: Slaughterhouse workers are in direct contact with cattle nearly every day. The purpose of this study was to survey the presence and distribution of anti-Shiga toxin 1 (Stx1) immunoglobulin G (IgG) in slaughterhouse workers, enabling a study of the serologic response to this toxin while working in an area at high-risk of Stx-producing Escherichia coli (STEC) infection.
One thousand seven hundred and twenty-nine serum samples from healthy slaughterhouse employees were collected and surveyed by indirect enzyme-linked immunosorbent assay (ELISA).
Among the 5 slaughterhouse positions, slaughterers had the highest distribution of anti-Stx1 IgG values by an ELISA. Based on the ELISA values, 25% (433/1729) of the workers had anti-Stx1 IgG. Slaughterers, residual products handlers, inspectors, livestock hygiene controllers, and grading testers had anti-Stx1 IgG-positive rates of 28%, 25%, 20%, 19%, and 17%, respectively. The ELISA values of anti-Stx1 IgG increased with increases in the number of years worked by slaughterers, but not by residual products handlers, inspectors, livestock hygiene controllers, or grading testers.
From these results, slaughterhouse workers are healthy and asymptomatic; slaughterers in particular are at high-risk for STEC exposure.
No preview · Article · Nov 2011 · Scandinavian Journal of Infectious Diseases
[Show abstract][Hide abstract] ABSTRACT: We encountered a patient with hemolytic uremic syndrome (HUS) with persistent isolation of shiga toxin-producing Escherichia coli (STEC) for 3 weeks despite of having no clinical symptoms. STEC has been recognized as an important food-borne pathogen that causes severe diseases such as HUS. We characterized this STEC strain via a polymerase chain reaction, reverse-passive latex agglutination and the slide agglutination method. In this STEC strain, stx2 (shiga toxin), eaeA, tir, iha (adherence genes), espADB (type III secretion genes), and hlyA, ehxA, clyA (hemolysin genes) were present. The O antigen of the strain was non-typable.
Full-text · Article · Nov 2011 · Yonsei medical journal
[Show abstract][Hide abstract] ABSTRACT: This study aimed to investigate the prevalence of antibiotic resistance in fecal Escherichia coli isolates from healthy persons and patients with diarrhea.
E. coli isolates (n = 428) were obtained from fecal samples of apparently healthy volunteers and hospitalized patients with diarrhea. Susceptibility patterns of isolates to 16 antimicrobial agents were determined by agar disc diffusion.
Most E. coli isolates exhibited less than 10% resistance against imipenem, cefotetan, aztreonam, cefepime, cefoxitin, amikacin and netilamicin, although greater than 65% were resistant to ampicillin and tetracycline. No significant difference in resistance rates for all tested antibiotics was found between isolates from the healthy-and diarrheal-patient groups, including for multi-drug resistance (p = 0.22). The highest number of resistant antibiotics was 12 antibiotics. No significant differences in antibiotic resistance were found among the sex and age strata for isolates from healthy individuals. However, antibiotic resistance rates to cefoxitin, cefotaxime, amikacin, and netilamicin were significantly higher in the isolates of men than those of women (p < 0.05) in isolates from patients with diarrhea. Furthermore, isolates from patients with diarrhea older than 40-years of age showed higher resistance to cefepime and aztreonam (p < 0.05).
High resistance to the antibiotics most frequently prescribed for diarrhea was found in isolates from patients with diarrhea and apparently healthy individuals without any significant difference.
[Show abstract][Hide abstract] ABSTRACT: This study was conducted to investigate the phenotypic and genotypic characteristics of Korean isolates of Cronobacter spp. (Enterobacter sakazakii). A total of 43 Cronobacter spp., including 5 clinical isolates, 34 food isolates, 2 environmental isolates, and 2 reference strains (C. sakazakii ATCC 29004 and C. muytjensii ATCC51329) were used in this study. Korean isolates of Cronobacter spp. were divided into 11 biogroups according to their biochemical profiles and 3 genomic groups based on the analysis of their 16S rRNA gene sequences. Biogroups 1 and 2 contained the majority of isolates (n=26), most of which were contained in 16S rRNA cluster 1 (n=34). Korean isolates of Cronobacter spp. showed diverse biochemical profiles. Biogroup 1 contained C. sakazakii GIHE (Gyeonggido Research Institute of Health and Environment) 1 and 2, which were isolated from babies that exhibited symptoms of Cronobacter spp. infection such as gastroenteritis, sepsis, and meningitis. Our finding revealed that Biogroup 1, C. sakazakii, is more prevalent and may be a more pathogenic biogroup than other biogroups, but the pathogenic biogroup was not represented clearly among the 11 biogroups tested in this study. Thus, all biogroups of Cronobacter spp. were recognized as pathogenic bacteria, and the absence of Cronobacter spp. in infant foods should be constantly regulated to prevent food poisoning and infection caused by Cronobacter spp.
No preview · Article · May 2011 · Journal of Microbiology and Biotechnology
[Show abstract][Hide abstract] ABSTRACT: Dairy farmers perform various types of work and are in direct contact with dairy cattle nearly every day. The purpose of this study was to assess the prevalence of Shiga toxin-encoding genes (stx) among dairy farmers and to evaluate the relationship between stx and risk factors.
A questionnaire developed in-house was sent to dairy farmers in Gyeonggi Province, Korea by registered mail. Researchers obtained stool samples and identified or administered the questionnaires by interview. The stool samples were examined for stx genes by polymerase chain reaction.
Twenty (3.4%) of 589 stool samples from dairy farmers were stx-positive. The distribution of stx-positive stool samples revealed an increase in Shiga toxin-producing Escherichia coli infection with age, duration of work, and herd size. There was no association between stx-positive stool samples and type of work. For artificial insemination, taking a shower after work was significant, and the proportion of stx-positive dairy farmers increased as taking a shower after work decreased.
Hygiene-related education to include taking a shower after sessions of artificial insemination should be considered. However, the stx-positive dairy farmers were small in number and the results should be interpreted with caution.
No preview · Article · Apr 2011 · Scandinavian Journal of Infectious Diseases