Eui-Chong Kim

A Case of Streptococcus gallolyticus subsp. gallolyticus Infective Endocarditis with Colon Cancer: Identification by 16S Ribosomal DNA Sequencing

Although the association between Streptococcus bovis endocarditis and colon carcinoma is well known, very few cases of S. bovis infection associated with underlying malignancies have been reported in Korea. The S. bovis group has been recently reclassified and renamed as Streptococcus gallolyticus and Streptococcus infantarius subspecies under a new nomenclature system. We report a case of infective endocarditis with colon cancer caused by S. gallolyticus subsp. gallolyticus (previously named S. bovis biotype I). A 59-yr-old woman presented with a 1-month history of fever. Initial blood cultures were positive for gram-positive cocci, and echocardiography showed vegetation on mitral and aortic valves. Antibiotic treatment for infective endocarditis was started. The infecting strain was a catalase-negative and bile-esculin-positive alpha-hemolytic Streptococcus susceptible to penicillin and vancomycin. The strain was identified as S. gallolyticus subsp. gallolyticus with the use of the Vitek 2 GPI and API 20 Strep systems (bioMérieux, USA). The 16S rDNA sequences of the blood culture isolates showed 100% homology with those of S. gallolyticus subsp. gallolyticus reported in GenBank. The identification of the infecting organism, and the subsequent communication among clinical microbiologists and physicians about the changed nomenclature, led to the detection of colon cancer. The patient recovered after treatment with antibiotics, valve surgery, and operation for colon cancer. This is the first report of biochemical and genetic identification of S. gallolyticus subsp. gallolyticus causing infective endocarditis associated with underlying colon cancer in a Korean patient.

First case of Mycobacterium longobardum infection.

Mycobacterium longobardum is a slow-growing, nontuberculous mycobacterium that was first characterized from the M. terrae complex in 2012. We report a case of M. longobardum induced chronic osteomyelitis. A 71-yr-old man presented with inflammation in the left elbow and he underwent a surgery under the suspicion of tuberculous osteomyelitis. The pathologic tissue culture grew M. longobardum which was identified by analysis of the 65-kDa heat shock protein and full-length 16S rRNA genes. The patient was cured with the medication of clarithromycin and ethambutol without further complications. To the best of our knowledge, this is the first report of a M. longobardum infection worldwide.

Evaluation of Two Hepatitis C Virus Genotyping Assays Based on the 5′ Untranslated Region (UTR): the Limitations of 5′ UTR-Based Assays and the Need for a Supplementary Sequencing-Based Approach

ABSTRACT We evaluated two genotyping methodologies that characterize the 5′ untranslated region (5′ UTR) of the hepatitis C virus (HCV) genome. The limitations of these genotype assays need to be thoroughly evaluated, and sequencing-based approaches may be needed to complement these methods in clinical settings.

Comparison of the analytical and clinical performances of Abbott RealTime High Risk HPV, Hybrid Capture 2, and DNA Chip assays in gynecology patients

The detection of high-risk (HR) HPV in cervical cancer screening is important for early diagnosis of cervical cancer or pre-cancerous lesions. We evaluated the analytical and clinical performances of 3 HR HPV assays in Gynecology patients. A total of 991 specimens were included in this study: 787 specimens for use with a Hybrid Capture 2 (HC2) and 204 specimens for a HPV DNA microarray (DNA Chip). All specimens were tested using an Abbott RealTime High Risk HPV assay (Real-time HR), PGMY PCR, and sequence analysis. Clinical sensitivities for severe abnormal cytology (severe than high-grade squamous intraepithelial lesion) were 81.8% for Real-time HR, 77.3% for HC2, and 66.7% for DNA Chip, and clinical sensitivities for severe abnormal histology (cervical intraepithelial neoplasia grade 2+) were 91.7% for HC2, 87.5% for Real-time HR, and 73.3% for DNA Chip. As compared to results of the sequence analysis, HC2, Real-time HR, and DNA Chip showed concordance rates of 94.3% (115/122), 90.0% (117/130), and 61.5% (16/26), respectively. The HC2 assay and Real-time HR assay showed comparable results to each other in both clinical and analytical performances, while the DNA Chip assay showed poor clinical and analytical performances. The Real-time HR assay can be a good alternative option for HR HPV testing with advantages of allowing full automation and simultaneous genotyping of HR types 16 and 18.

Liver cirrhosis caused by Exophiala dermatitidis.

We report a case of liver cirrhosis caused by Exophiala dermatitidis in a previously healthy child. The infecting organism was initially mistaken as capsule-deficient Cryptococcus neoformans.

Comparison of Modified Multiple-locus Variable-number Tandem-repeat Fingerprinting with Pulsed-field Gel Electrophoresis for Typing Clinical Isolates of Staphylococcus aureus

Background Multiple-locus variable-number tandem-repeat fingerprinting (MLVF) is based on multiplex PCR, utilizing variable number tandem repeat. Our goal was to compare the performance of MLVF in distinguishing clinical Staphylococcus aureus isolates with that of pulsed-field gel electrophoresis (PFGE), which has traditionally been the gold standard. Methods Sixty-three clinically significant S. aureus isolates were tested using both PFGE and MLVF. Multiplex PCR for MLVF was performed using PCR primers for clfA, clfB, sdrCDE, sspA, and spa. PFGE was performed with genomic DNA fragments generated by SmaI endonuclease digestion. Banding patterns of MLVF or PFGE were analyzed using InfoQuestFP software. Results The hands-on time of our modified method was about 3 h, on average, for each of 18 isolates. PFGE (80% cutoff) or MLVF (75% cutoff) separated all of the 63 isolates into 13 and 12 types, respectively. Three types generated by PFGE were identical to those generated by MLVF. PFGE and MLVF yielded similar Simpsons diversity indices, indicating similar discriminatory power. The overall concordance between PFGE and MLVF was low, as represented by adjusted Rand indices (0.266-0.278). PFGE predicted MLVF type better than MLVF predicted PFGE type, as reflected by Wallace coefficients (PFGE cutoff 80% vs. MLVF cutoff 75%, 0.389 vs. 0.233). Analysis of the relationship between a pair of isolates showed 91.0% concordance between the PFGE (80% cutoff) and MLVF (75% cutoff). Conclusions Our simple, low-cost, modified MLVF protocol can effectively discriminate between S. aureus clinical isolates. MLVF can replace PFGE for the hospital infection control of S. aureus.

The First Korean Case of Moraxella osloensis Bacteremia in a Patient with Acute Myeloid Leukemia

The genus Moraxella comprises approximately 20 species of aerobic, non-motile, oxidase-positive, and gram-negative coccobacilli. Besides M. catarrhalis, a well-known pathogen, M. atlantae, M. canis, M. lacunata, M. lincolnii, M. nonliquefaciens, and M. osloensis are also occasionally isolated from clinical samples [1]. M. osloensis is a commensal microorganism in the human respiratory tract, but has also been reported as a rare causative pathogen in human infections [1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12]. Here, we report a case of M. osloensis bacteremia in a patient with a hematologic malignancy. To our knowledge, this is the first report of M. osloensis bacteremia in Korea. n nA 66-yr-old man with acute myeloid leukemia (AML) was admitted to our hospital for consolidation chemotherapy. Two months prior to this hospitalization, the patient was diagnosed with AML, not otherwise specified, according to the 2008 WHO classification system [13], and he subsequently received cytarabine and idarubicin chemotherapy. After 1 month, follow-up bone marrow examination was performed, and he achieved complete remission. n nOn admission, the patient presented with symptoms of an upper respiratory infection including coughing and chills. He had a temperature of 37.2℃, a blood pressure of 117/57 mmHg, a pulse of 106/min, and a respiration rate of 20 breaths/min. The results of the laboratory investigation were as follows: Hb, 11.7 g/dL; leukocyte count, 5.7×109/L; platelet count, 177×109/L; C-reactive protein level, 6.48 mg/dL; blood urea nitrogen (BUN)/creatinine, 15/1.2 mg/dL; and total protein/albumin, 6.8/3.6 g/dL. A total of four blood culture sets were collected from two separate peripheral veins and central venous catheter. There were no abnormalities in the chest radiographic images, but paranasal sinus radiographic imaging indicated a suspected maxillary sinusitis. Therefore, empirical antibiotic therapy with intravenous ampicillin-sulbactam was administered. n nBacterial growth was detected in an aerobic culture bottle that contained the blood from the central venous catheter after 1 day of incubation, and in the other 3 aerobic culture bottles after 2 days of incubation (Fig. 1A). The broths from positive culture bottles were inoculated onto blood agar plates (BAPs) and MacConkey agar plates (MACs) that were subsequently incubated for 24 hr at 35℃ in a 5% CO2 atmosphere. Whereas no colony was observed on the MACs, grey-white colored, non-hemolytic colonies grew on the BAPs (Fig. 1B) and gram-negative coccobacilli were observed from gram stain smear preparations (Fig. 1C). The isolates were oxidase-positive and indole-negative. n n n nFig. 1 n nColonial and microscopic morphology of Moraxella osloensis. (A) Gram-negative coccobacilli from positive aerobic blood culture smear preparations (Gram stain, ×1,000). (B) Grey-white colored colonies on blood agar plate

The First Case of Eggerthella lenta Bacteremia in Korea

Eggerthella lenta, previously Eubacterium lentum, is a non-motile, non-spore-forming anaerobic gram-positive bacillus. In 1999, the designation Eggerthella gen. nov. (named in honor of Arnold Eggerth, who published the first description) was proposed as a substitute for Eubacterium lentum on the basis of 16S rRNA gene analysis and G+C content comparisons, and Eggerthella lenta was only species in that genus [1, 2]. To date, 2 species, Eggerthella lenta and Eggerthella sinensis, have been assigned to the genus Eggerthella. Eggerthella hongkongensis, which was previously included in the genus Eggerthella, has been classified under a new genus and renamed Paraeggerthella hongkongensis since 2009, based on chemotaxonomic data [3]. E. lenta is a causative pathogen in appendicitis, bacteremia, cutaneous abscess, genitourinary tract infection, liver abscess, peritonitis, spondylodiscitis, and wound infection, but reports of E. lenta bacteremia are rare [4-13]. Here we report a case of E. lenta bacteremia in a rectal cancer patient. To the best of our knowledge, this is the first report of E. lenta bacteremia in Korea. n nA 53-yr-old man was admitted to our hospital with fever and chills following the replacement of a double-J stent in the outpatient clinic on August 16, 2013. The patient was previously diagnosed with rectal cancer and underwent laparoscopic ultralow anterior resection with ileostomy in 2011. He underwent Hartmanns operation for recurrence in 2012 and received chemotherapy from August 2012 to April 2013. Following chemotherapy, left kidney hydronephrosis, recurrent mass in the left pelvic wall, and peritoneal seeding was observed in the follow-up abdominal computed tomography, and he underwent left double-J stent insertion for obstructive acute kidney injury in April 2013. n nOn admission, the patient had a temperature of 39.5℃, blood pressure of 157/93 mmHg, pulse of 98/min, and respiratory rate of 20 breaths/min. Laboratory investigation showed a Hb of 11.4 g/dL, leukocyte count of 2.2×109/L, platelet count of 255×109/L, C-reactive protein (CRP) level of 9.55 mg/dL, blood urea nitrogen (BUN)/creatinine of 20/1.3 mg/dL, and total protein/albumin level of 7.1/3.5 g/dL. A urine sample and two sets of blood samples were collected, and the patient received empirical antibiotic therapy. After three days of incubation, gram-positive bacilli grew in an anaerobic culture bottle, but no microorganisms were detected in the urine culture. The positive culture broth was inoculated onto a Brucella agar plate and anaerobically cultured for 48 hr. Small and translucent colonies grew on the Brucella agar plate, and gram-positive coccobacilli were observed by using Gram staining (Fig. 1). The organism was identified as E. lenta by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) (Bruker Daltonik GmbH, Bremen, Germany), with a score of 2.223. To confirm the identity of the isolate, 16S rRNA gene sequencing was conducted using the MicroSeq 500 system (Applied Biosystems, Foster City, CA, USA). PCR and sequencing kits were designed with universal primers to cover all bacteria. Sequences were analyzed by using an ABI PRISM 3730 Series DNA Analyzer (Applied Biosystems). In the first 500 bp of the 16S rRNA gene sequence, the isolate showed a 100% identity match with GenBank sequence {type:entrez-nucleotide,attrs:{text:NR_074377.1,term_id:444303954}}NR_074377.1 (Eggerthella lenta) and 96.8% identity match with GenBank sequence {type:entrez-nucleotide,attrs:{text:AY321958.1,term_id:32481963}}AY321958.1 (Eggerthella sinensis). The patient was diagnosed as having E. lenta bacteremia. n n n nFig. 1 n n(A) Small and translucent colonies of Eggerthella lenta on a Brucella agar plate. (B) Gram-positive coccobacilli from smear preparations from the colonies on Brucella agar plate (×1,000). n n n nEmpirical treatment with intravenous cefotaxime and amikacin was started before the culture results were obtained. During the four day treatment course, the patients fever subsided and he was discharged with a prescription for oral antibiotics. There were no microorganisms in the follow-up urine sample or the two sets of blood cultures. n nE. lenta is a non-motile, non-spore-forming anaerobic gram-positive bacillus. It does not produce acids from glucose, does not produce indole or liquefy gelatin, and produces little or no gas [14]. E. lenta colonies on blood agar are 0.25 to 0.75 mm in diameter, slightly raised, smooth, and gray [14]. E. lenta is a common gastrointestinal commensal [2, 14], but has been identified as the causative pathogen in various conditions, including appendicitis, skin abscesses, and spondylodiscitis [4-6]. There have been infrequent reports of E. lenta bacteremia, which is characterized by a high proportion of polymicrobial infection (about 40%) and a high mortality rate (20-30%) [7-13]. In 1994, Jang et al. [15] reported E. lenta (E. lentum) isolate from expressed prostatic secretion sample from a chronic prostatitis patient, which was the only case reported in Korea. Patients with gastrointestinal tract disease, malignancy, hepatobiliary disease, bed sores, diabetes mellitus, and stroke are prone to E. lenta bacteremia [7-13]. n nBiochemical identification of E. lenta by using the API system (bioMerieux, Marcy lEtoile, France) and the VITEK 2 system (bioMerieux) was reported to be reliable, but many reported cases were confirmed by 16S rRNA sequence analysis [9, 11, 16]. MALDI-TOF MS, recently introduced in the clinical microbiologic laboratory, has been used successfully for the identification of E. lenta, with 100% success at the genus level and about 80% success at the species level [17-19]. MALDI-TOF MS is fast and useful in the identification of clinically important anaerobic bacteria such as E. lenta, although it is limited by the number of databases available for anaerobic bacteria [17-19]. n nE. lenta is susceptible to ampicillin-sulbactam, metronidazole, carbapenems, tigecycline, and daptomycin, but the two strains depending on colony morphology exhibit variable susceptibility to cephalosporins. Translucent-colony coccobacilli are susceptible to cephalosporins, while speckled-colony pleomorphic bacilli are resistant to them [12, 20]. Antibiotics, such as ampicillin-sulbactam, metronidazole, and carbapenems, to which E. lenta is known to be susceptible, have been conventionally used for the treatment of E. lenta bacteremia and are appropriate choices for patient management. However, bacteremia induced by translucent-colony-forming E. lenta as observed in the current case, could be treated effectively by using empirical antibiotics containing third-generation cephalosporins. n nIn the present case, we confirmed the identity of the blood isolate as E. lenta by MALDI-TOF MS and 16S rRNA sequence analysis. Although E. lenta bacteremia is associated with significant mortality and morbidity [7-13], this patient made a quick recovery, possibly because of prompt treatment at an early stage. The isolated E. lenta grew only in an anaerobic blood culture bottle, and the patient had high temperature, elevated heart rate, elevated CRP levels, and decreased leukocyte count and gastrointestinal tract malignancy as a predisposing factor for E. lenta bacteremia. On the basis of these observations and laboratory results, we concluded that the patient had E. lenta bacteremia. The occurrence of gram-positive bacillus in blood should not be ignored and warrants further laboratory investigation to avoid complications. n nTo the best of our knowledge, this is the first report of E. lenta bacteremia in Korea. MALDI-TOF MS is a fast, efficient method for identification of clinically anaerobic bacteria, such as E. lenta.

Possession of the macrophage-induced gene by isolates of the Mycobacterium avium complex is not associated with significant clinical disease.

The Mycobacterium avium complex (MAC) is the most frequently isolated species among non-tuberculous mycobacteria (NTM) clinical isolates. Physicians pay attention to the differential diagnosis of the disease caused by MAC from tuberculosis because of their similar clinical presentations. Expression of the macrophage-induced gene (mig) is one of the virulence phenotypes in MAC, but it has not been determined whether the presence of the mig gene itself has any relationship with clinical disease or whether it is merely a marker for MAC. To uncover the significance of the mig gene among MAC clinical isolates, positive cultures from respiratory specimens from patients in a tertiary referral centre were identified by sequencing the 16S rRNA gene. The mig gene was also evaluated using PCR and sequence analysis. The medical records from the patients were reviewed retrospectively. The diagnostic criteria from the American Thoracic Association were adopted for the diagnosis of NTM lung disease. A total of 45 MAC clinical isolates were identified over a period of 1 year. Following 16S rRNA sequencing, all of the 23 M. avium isolates were categorized as sequevar I. Among the 22 Mycobacterium intracellulare isolates, 18 strains were identified as M. intracellulare sequevar I and the remaining four consisted of one each of sequevars II, III, IV and V. The proportion of cases that fitted the diagnostic criteria of NTM lung disease was 26.7 % (12/45). The mig PCR results were 100 % positive for the MAC isolates studied, irrespective of their species, sequevar or disease-causing properties. However, following bootstrap analysis of the mig sequences, we observed definite grouping between M. avium and M. intracellulare. Thus the mig gene is a species-specific marker with distinct sequence diversity between the two species M. avium and M. intracellulare, but there is poor correlation between disease-causing properties and specific mig sequences.

Low-Colony Counts of Nontuberculous Mycobacteria: Clinical Significance Analysis

Background: Diagnosis of nontuberculous mycobacterium (NTM) is challenging, and clinical, radiological and microbiological criteria should be met. Traditionally, culture results on solid media have been reported semi-quantitatively, but no study exists regarding the clinical significance of low-colony count culture reports. The authors of the present study analyzed the clinical significance of low-colony count specimens of NTM with a greater than three-year follow-up period. Methods: A total of 341 clinical isolates were evaluated among the isolates at Seoul National University Hospital and Seoul National University Borame Hospital from October 2005 to September 2006. Colony count less than 50 was considered a low-colony count specimen. Identifications of NTM from all the isolates were performed using a DNA chip (PCR reverse hybridization, LG Life Science, Korea). Clinical significance was analyzed by reviewing the medical records of patients with greater than three years of follow-up data after NTM isolation from respiratory samples. Results: NTM lung disease was observed in 27.0% of the patients with low-colony count specimens among 167 patients with respiratory samples, and 70.4% of the patients were treated. The low-colony count patients had less NTM lung disease, longer incubation period, and less acid fast bacilli-positivity than patients with a colony count greater than 50. Conclusion: The prevalence of NTM lung disease with a low-colony count specimen was greater than 25%. In a clinical setting, NTM lung disease should not be excluded only on the basis of a low-colony count. (Korean J Clin Microbiol 2012;15:9-13)