Emergence of optrA-Mediated Linezolid-Nonsusceptible Enterococcus faecalis in a Tertiary Care Hospital

This study investigated resistance mechanisms and epidemiology of emerging linezolid-nonsusceptible Enterococcus faecalis (LNSEF) in a tertiary care hospital. LNSEF isolated from clinical samples were collected from November 2017 to June 2019. The isolates were investigated for linezolid resistance and the associated molecular mechanisms, including mutations of 23S rRNA domain V and acquisition of the cfr or optrA resistance gene. We used pulsed-field gel electrophoresis (PFGE) and multilocus sequence typing for the molecular typing of the isolates. Among 4,318 E. faecalis isolates, 10 (0.23%) were linezolid-nonsusceptible. All LNSEF isolates were optrA-positive and cfr-negative. Of these isolates, five were sequence type (ST) 476, two ST585, one ST16, one ST16-like, and one ST480. Six LNSEF isolates obtained in the first year clustered to three types in the PFGE analysis: two ST476 isolates of type A, two ST585 isolates of type B, and two ST16 or ST16-like isolates of type C. Seven cases were of community-onset and three were hospital acquired, but total of eight were healthcare-associated including five community-onset. None of the patients had a history of linezolid treatment, and in one patient, we detected linezolid-susceptible E. faecalis one month before LNSEF detection. In conclusion, heterogenous clones of optrA-positive LNSEF emerged in the hospital mainly via community-onset.

A Case of Chronic Strongyloidiasis with Recurrent Hyperinfection

Strongyloides stercoralis is an intestinal nematode that often causes chronic diarrhea and may develop severe complicated form of hyperinfection or disseminated infection in immunocompromised patients. Here, we report a case of recurrent strongyloidiasis presenting with pulmonary and meningeal involvement. A 55-year-old male diagnosed with pancreatic cancer 4 months ago was admitted due to chronic diarrhea, abdominal pain, and weight loss for 2–3 months. He had been treated with albendazole for chronic recurrent strongyloidiasis 13 years ago and again 2 years ago. He developed sepsis of Klebsiella pneumoniae and Escherichia coli on Days 3 and 7, respectively, and then meningitis of E. coli on Day 42. Strongyloidiasis was diagnosed by detection of abundant filariform larvae in sputum specimens on Day 15. He was treated for disseminated strongyloidiasis with albendazole and ivermectin for five weeks until clearance of larvae was confirmed in sputum and stool specimens. Laboratory diagnosis is important to guide appropriate treatment and to prevent chronic and recurrent strongyloidiasis.

Performance of MALDI Biotyper for Species Identification of Carbapenem-Resistant Enterobacteriaceae by Media Types and Incubation Time

BACKGROUND: This study was conducted to evaluate the impact of the media type used for direct identification of colonies on the surveillance culture of carbapenem-resistant Enterobacteriaceae (CRE) by matrix assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS). METHODS: CRE surveillance culture isolates were subjected to species identification using the MALDI Biotyper (Bruker Daltonics, Germany) for 2 months starting in March 2017. Four types of media were evaluated: blood agar (BA), Mueller Hinton agar (MH), MacConkey agar (Mac), and MacConkey agar containing imipenem of 1 µg/mL (IMP-Mac). CRE-like colonies on IMP-Mac and their subculture colonies on the other media were tested after overnight incubation and extended incubation for one additional day. The percent identification and score value were analyzed for each media types and incubation time when the identification was correct at the genus level. RESULTS: A total of 117 isolates were identified as 84 Klebsiella pneumoniae, 12 Escherichia coli, 9 Enterobacter cloacae, 5 Klebsiella oxytoca, 4 Enterobacter aerogenes, and 2 Raoultella ornithinolytica. The successful identification rates (SIR) for BA and MH were 98.3% and 97.4% (P=0.9), respectively, while those for Mac and IMP-Mac were 82.1% (P < 0.001) and 70.9% (P < 0.001), respectively. After extended incubation, SIRs were decreased to 96.6%, 96.6% (P=1.0), 61.5% (P < 0.001), and 58.1% (P < 0.001) on BA, MH, Mac, and IMP-Mac, respectively. The average score values were significantly lower for Mac (2.017±0.22) and IMP-Mac (1.978±0.24) than for BA (2.213±0.16) (P < 0.001). CONCLUSIONS: The low performance of the MALDI Biotyper applied directly to the colonies grown on Mac or IMP-Mac indicates that subculture on BA or MH is preferable before identification by MALDI-TOF MS.

Utility of a Direct 16S rDNA PCR and Sequencing for Etiological Diagnosis of Infective Endocarditis

BACKGROUND: Cases of infective endocarditis (IE) require prompt etiological diagnosis for effective treatment. Molecular methods can aid in rapid and reliable diagnosis of culture-negative IE cases. We evaluated the utility of 16S rDNA PCR and sequencing in determining the causative agents of IE in valve tissues, especially when specimens were obtained after initiation of antimicrobial therapy. METHODS: We performed 16S rDNA PCR and sequencing in heart valve specimens and medical records review of 80 patients who underwent protocol-based cardiac surgery from 2013 to 2015. One patient did not meet the criteria for IE. Sixty-five (81.3%) and 14 pa-tients (17.5%) were diagnosed as having definite IE and possible IE, respectively. Blood and heart valve biopsy tissue were examined by using routine microbiological methods. RESULTS: Blood cultures in our hospital were IE-positive for 33 patients (41.8%), whereas 49 patients (62.0%) showed positive blood cultures when initial blood cultures performed at the referring hospital were included. Eighteen (22.8%) and 40 patients (50.6%) were IE-positive in valve tissue cultures and 16S rDNA PCR, respectively. Bacteria in the Streptococcus mitis group (n=26) were the most common etiological agents of IE. Eight (10.1%) culture-negative specimens tested positive by 16S rDNA PCR. In five of eight PCR-positive and culture-negative cases, fastidious or anaerobic organisms were the cause of IE. CONCLUSIONS: Direct 16S rDNA PCR and sequencing can be used as a supplementary method to conventional blood and biopsy culture testing, especially in culture-negative IE cases that are negative for IE by culture.

Practical Aspects of Cytomegalovirus DNA Quantitative PCR / 대한임상미생물학회지

Human cytomegalovirus (CMV) is a clinically important pathogen that causes significant morbidity and mortality in immunocompromised patients and is typically monitored using real-time polymerase chain reaction (real-time PCR). International standards and certified reference materials were recently developed by the WHO, providing the opportunity to standardize viral load reporting. Clinical microbiologists who conduct quantitative CMV DNA testing should be aware of technical issues that can affect the analytical and clinical performance of the method used. These include specimen type, limits of detection and quantification, linear range, reproducibility, and wide variability in viral load values among different assays. Specimen types for testing include whole blood, plasma, serum, and peripheral blood leukocytes. The tests that use whole blood and peripheral blood leukocytes have higher sensitivities. Adsorption chromatography methods are widely used for nucleic acid extraction, and efficiencies can differ between manual and automatic processes. The most common method for quantitative CMV DNA testing is real-time PCR. All CMV testing methods require quality control at the pre-analytic, analytic, and post-analytic stages. In transplant patients, specific quantitative results and monitoring of any changes at follow-up are important. Five to seven days is an adequate follow-up interval in this regard, and drug-resistant CMV should be suspected if there is no response to therapy. One specimen type should be chosen for follow-up quantitative CMV DNA testing, optimized according to WHO standards. Further studies are needed to better standardize CMV testing approaches and to determine the appropriate clinical cut-off level.

Annual Report on the External Quality Assessment Scheme for Clinical Microbiology in Korea (2015)

Annual proficiency surveys were conducted in March, June, and September in 2015 by the Clinical Microbiology Subcommittee of the Korean Association of External Quality Assessment Service. The program covers the sections of bacteriology, advanced bacteriology and mycology, mycobacteriology, and parasitology. Each trial was composed of three sets of different combinations of five bacteria and yeasts. These sets were distributed among laboratories for Gram staining, culture, identification, and antimicrobial susceptibility tests. Five slides with fixed sputum smears were provided as part of each trial for acid-fast bacilli detection. The survey material distribution was section-based. Two survey materials were provided in each trial, while five specimens for mycobacterial culture and identification, five specimens for anti-tuberculosis susceptibility testing and two Mycobacterium tuberculosis strains for rapid detection of rifampin and isoniazid resistance were distributed in the March and June trials. Five virtual microscopy files for stool parasite examination were availed by registered participants in the June trial. Out of the 334 enrolled laboratories, 328 (98.2%), 328 (98.2%), and 329 (98.5%) submitted responses in trials I, II, and III, respectively. Identification of bacteria, namely, Escherichia coli, Klebsiella pneumoniae, Enterococcus faecalis, Pseudomonas aeruginosa, Streptococcus pneumoniae, and Vibrio fluvialis by more than 95% of participants was acceptable. Surveillance cultures for vancomycin-resistant enterococci and carbapenem-resistant Enterobacteriaceae were determined accurately by 75.8%–85.3% and 93.1% of the respondents, respectively. Species-level identification of Candida krusei, Candida lusitanae, and Candida guilliermondii was still low at 79.8%, 55.7%, and 42.7%, respectively. Disk diffusion method revealed an unacceptably high false-positive rate of resistance to glycopeptides in E. faecalis and to trimethoprim-sulfamethoxazole in S. pneumoniae. Advanced bacteriology trials revealed unsatisfactory results for species-level identification of moulds. Mycobacterial culture, identification and susceptibility testing, and molecular detection of rifampin and isoniazid resistance were performed exceedingly well by participants. Hymenolepsis diminuta could not be identified by participants, with a correct answer rate of only 46.5% and ‘no parasite seen’ answer rate of only 31.8% for negative specimens. Species-level identification of Candida and moulds was challenging for clinical microbiology laboratories. Disk diffusion method was found to be problematic in testing the susceptibility of microorganisms to glycopeptides and trimethoprim-sulfamethoxazole. Improvement is required in result interpretation of negative specimens in parasitology.

A Case of Pneumonia Caused by Pneumocystis jirovecii Resistant to Trimethoprim/Sulfamethoxazole in the Absence of Previous Drug Exposure

Pneumocystis jirovecii pneumonia is a common opportunistic infection seen in patients with human immunodeficiency virus (HIV) infection. Dihydropteroate synthase (DHPS) is a target of sulfa drugs, and mutations in DHPS gene are associated with failure in treatment and prophylaxis of P. jirovecii infections in HIV-infected patients. Here, we report a case of a patient with P. jirovecii infection, harboring DHPS gene mutations, who had not been previously treated with trimethoprim/sulfamethoxazole (TMP/SMX). A 50-yr-old man was admitted to the hospital with symptoms such as fever, cough, sputum, and sore throat. Chest computed tomography scanning revealed diffuse ground glass opacity in both the lungs, and the patient was diagnosed as having HIV infection with a CD4+ T cell count of 22/µL. Immunohistochemical test results were positive for P. jirovecii. He was treated with TMP/SMX; however, his symptoms and laboratory findings did not improve. The treatment was changed to clindamycin and primaquine, and his symptoms improved after 3 days. Molecular testing of the sample for the detection of DHPS gene mutations and the typing of mitochondrial large subunit rRNA (mtlsurRNA) revealed DHPS gene mutations at codon 55 and 57, respectively, and the case had type 3 mtlsurRNA. This case study illustrates that DHPS mutation test results can be positive even in patients without previous exposure to TMP/SMX.

Concurrent Hematologic and Metastatic Epithelial Malignancies in the Bone Marrow: Report of Three Cases

No abstract available.

Gelatinous transformation of the bone marrow in hepatocellular carcinoma

No abstract available.

A Case of Refractory Thrombocytopenia with 5q Deletion: Myelodysplastic Syndrome Mimicking Idiopathic Thrombocytopenic Purpura

No abstract available.

Isolation of Anti-D after Administration of Intravenous Immune Globulin in a Patient with Immune Thrombocytopenic Purpura / 대한수혈학회지

Intravenous immune globulin (IVIG) is widely used in treatment of hypogammablobulinemia and for immunomodulation. Passive transfer of anti-D activity through administration of IVIG may cause difficulty in serologic assessment of patients. Here we report on a case of passive anti-D from IVIG in a D positive patient. The patient was a 72-year-old Korean woman who was hospitalized for refractory immune thrombocytopenic purpura that is not cured after steroid therapy. IVIG 6,000 mg was administered for treatment of immune thrombocytopenic purpura. After IVIG administration for two days, we identified anti-D in the patient and a positive direct antiglobulin test was demonstrated. The patient's hemoglobin level remained unchanged. After IVIG administration for 10 days, the patient's specimen was negative for anti-D, as would be expected with passively acquired antibody. Antibodies in IVIG may confuse and complicate serologic testing of transfusion candidates. Therefore, passive transfer of anti-D should be considered when anti-D is detected, especially when the patient has received IVIG, as in this case.