Molecular Identification and Amphotericin B Susceptibility of Clinical Isolates of Aspergillus from 11 Hospitals in Korea
Min Seok Heo1, Jong Hee Shin1, Min Ji Choi1, Yeon-Joon Park2, Mi-Na Kim3, Sun Hoe Koo4, Won Gil Lee5, Soo Hyun Kim1, Myung-Geun Shin1, Soon-PalSuh1, Dong-WookRyang1
Department of Laboratory Medicine1,ChonnamNationalUniversityMedicalSchool,Gwangju;Department of Laboratory Medicine2, The Catholic University of Korea College of Medicine, Seoul; Department of Laboratory Medicine3, University of Ulsan College of Medicine and Asan Medical Center, Seoul; Department of Laboratory Medicine4, College of Medicine, Chungnam National University, Daejeon; Department of Laboratory Medicine5, Kyungpook National University School of Medicine, Daegu, Korea
Background: The antifungal agent amphotericin B (AmB) is among the gold standards for treating a wide range of fungal infections. However, the in vitro susceptibility to AmB of clinical strains of Aspergillus isolated in Korea has not been fully surveyed. We performed a multicenter study to determine the species distribution and antifungal susceptibilities to AmB of clinical Aspergillus isolates.
Methods: A total of 136 Aspergillus isolates obtained from 11 university hospitals during a 3-month period in 2013 were tested. Species complex identification was performed by sequencing the ITS regions or D1/D2 domains, and species identification was performed through partial sequencing of beta-tubulin. Minimal inhibitory concentrations (MICs) of AmB were determined with the CLSI and European Committee on Antimicrobial Susceptibility Testing (EUCAST) broth microdilution methods, Etest using Mueller-Hinton agar supplemented with glucose and methylene blue (Etest-MH), and Etest using RPMI agar supplemented with glucose (Etest-RPG).
Results: Molecular methods identified 6 Aspergillus species complexes: A.fumigatus (42.6%), A. niger (23.5%), A. flavus (17.6%), A. terreus (11.0%), A. versicolor (4.4%), and A. ustus (0.7%). Cryptic species (identifiable with beta-tubulin sequencing only) accounted for 11.0% of the isolates, the most frequent isolates being A. tubingensis (10), A. lentulus (2), A.tamarii (2), and A. calidoustus (1). The mean AmB MIC50(μg/mL) values determined for the complexes with CLSI were 1, 0.25, 2, 2, and 2 for A. fumigatus, A niger, A flavus, A. terreus, and A. versicolor, respectively. The percentages of resistance (MIC ≥ 4 μg/mL) determined with CLSI, EUCAST, Etest-MH, and Etest-RPG were 5.9%, 11.8%, 36.0%, and 26.5%, respectively, across all isolates. Individual species resistances were, respectively, as follows: A. fumigates complex (3.4%, 6.9%, 13.8%, 10.3%), A. niger complex (0%, 0%, 25%, 0%), A. flavus complex (8.3%, 16.7%, 66.7%, 70.8%), A. terreus complex (26.7%, 53.3%, 93.3%, 86.7%), and A. versicolor complex (0%, 0%, 50%, 0%). Of cryptic species, A. lentulus isolates had AmB MICs of ≥4g/mL, whereas A. tamarii isolates were susceptible (MIC ≤ 1 μg/mL) according to all methods.
Conclusions: Non-fumigatus Aspergillus species constitute ~60% of clinical Aspergillus isolates, and resistance to AmB is not uncommon among these isolates in Korea. Species identification and AMB susceptibility testing are required for Aspergillus isolates of clinical relevance.
Key words: Aspergillus, Amphotericin, MIC, CLSI, EUCAST, Etest
Weakening of B Antigen with Hemolytic Anemia as a Diagnostic Clue in Myelodysplastic Syndrome Belonging to Blood Type B: A Case Report
Min Seok Heo, Duck Cho, Myung-Geun Shin, Jong-Hee Shin, Soon-Pal Suh, Dong-Wook Ryang
Department of Laboratory Medicine, Chonnam National University Medical School and Chonnam National University Hwasun Hospital, Hwasun, Korea
Many patients with MDS are asymptomatic at diagnosis. However, others mostly present with symptoms resulting from a blood cytopenia; it is rarely manifested with unusual findings such as hemolytic anemia, weaking of ABO antigen and so on. A 44-year-old woman was admitted to Chonnam National University Hospital with features of persistent anemia without any signs of bleeding. Laboratory tests revealed an elevated reticulocyte count (approximately 11.01%) and total bilirubin level (approximately 1.3 mg/dL). Hemolytic anemia was initially suspected. Her hemoglobin level decreased to 8.2 g/dL, and ABO typing as a pre-transfusion test was performed. The patient's red blood cells demonstrated Bweak with no reactivity with anti-A sera and trace (+/−) agglutination with anti-B sera in her cell type. A strong anti-A 1 antibody (4+) was present and no detectable antibody was in the reverse type. Exons 6 and 7 of the ABO gene were sequenced, and the genotyping of the proposita was B101/O01. The preliminary finding of bone marrow biopsy was hemolytic anemia, but it was revised to MDS, RAEB-1 (refractory anemia with excessive blasts) on re-review and cytogenetic findings with trisomy 8. Based on this case, we suggest that weakening of ABO antigen with hemolytic anemia is a diagnostic clue of a myeloid malignancy such as AML and MDS.
Key words: ABO blood group, MDS, Diagnostic clue, Myeloid malignancy