Publications by authors named "Tomoya Maeda"

49 Publications

Mutational property of newly identified mutagen l-glutamic acid γ-hydrazide in Escherichia coli.

Mutat Res 2021 Jul 21;823:111759. Epub 2021 Jul 21.

RIKEN Center for Biosystems Dynamics Research, 6-2-3 Furuedai, Suita, Osaka, 565-0874, Japan; Department of Physics, The University of Tokyo, 7-3-1 Hongo, Tokyo, 113-0033, Japan; Universal Biology Institute, The University of Tokyo, 7-3-1 Hongo, Tokyo, 113-0033, Japan.

We previously found that an l-glutamine analog l-glutamic acid γ-hydrazide has high mutagenic activity through the high-throughput laboratory evolution of Escherichia coli. In this study, mutagenicity and mutational property of l-glutamic acid γ-hydrazide were examined by the Ames test and mutation accumulation experiments using E. coli. The Ames test revealed that l-glutamic acid γ-hydrazide showed higher mutagenic activity without metabolic activation than known mutagens 2-aminoanthracene, and cobalt(II) acetate tetrahydrate. This result indicates that l-glutamic acid γ-hydrazide does not require metabolic activation for mutagenic activity in E. coli. Mutation accumulation experiments and whole-genome sequencing analysis revealed the number and spectrum of the accumulated mutations with or without l-glutamic acid γ-hydrazide. In the presence of l-glutamic acid γ-hydrazide, MDS42 strain accumulated 392.3 ± 116.2 point mutations during 30 passages corresponding to 777 generations, while MDS42 strain accumulated 1.5 ± 2.5 point mutations without l-glutamic acid γ-hydrazide during 50 passages corresponding to 1341 generations. The mutational spectrum of l-glutamic acid γ-hydrazide was G/C to A/T transition (82.2 ± 4.3 %) and A/T to G/C transition (17.4 ± 4.3 %). These results indicated that l-glutamic acid γ-hydrazide has a strong mutagenic activity.
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http://dx.doi.org/10.1016/j.mrfmmm.2021.111759DOI Listing
July 2021

Laboratory evolution of Mycobacterium on agar plates for analysis of resistance acquisition and drug sensitivity profiles.

Sci Rep 2021 Jul 23;11(1):15136. Epub 2021 Jul 23.

RIKEN Center for Biosystems Dynamics Research, 6-2-3 Furuedai, Suita, Osaka, 565-0874, Japan.

Drug-resistant tuberculosis (TB) is a growing public health problem. There is an urgent need for information regarding cross-resistance and collateral sensitivity relationships among drugs and the genetic determinants of anti-TB drug resistance for developing strategies to suppress the emergence of drug-resistant pathogens. To identify mutations that confer resistance to anti-TB drugs in Mycobacterium species, we performed the laboratory evolution of nonpathogenic Mycobacterium smegmatis, which is closely related to Mycobacterium tuberculosis, against ten anti-TB drugs. Next, we performed whole-genome sequencing and quantified the resistance profiles of each drug-resistant strain against 24 drugs. We identified the genes with novel meropenem (MP) and linezolid (LZD) resistance-conferring mutation, which also have orthologs, in M. tuberculosis H37Rv. Among the 240 possible drug combinations, we identified 24 pairs that confer cross-resistance and 18 pairs that confer collateral sensitivity. The acquisition of bedaquiline or linezolid resistance resulted in collateral sensitivity to several drugs, while the acquisition of MP resistance led to multidrug resistance. The MP-evolved strains showed cross-resistance to rifampicin and clarithromycin owing to the acquisition of a mutation in the intergenic region of the Rv2864c ortholog, which encodes a penicillin-binding protein, at an early stage. These results provide a new insight to tackle drug-resistant TB.
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http://dx.doi.org/10.1038/s41598-021-94645-zDOI Listing
July 2021

Morphological change of coiled bacterium Spirosoma linguale with acquisition of β-lactam resistance.

Sci Rep 2021 Jun 24;11(1):13278. Epub 2021 Jun 24.

RIKEN Center for Biosystems Dynamics Research, 6-2-3 Furuedai, Suita, Osaka, 565-0874, Japan.

Spirosoma linguale is a gram-negative, coiled bacterium belonging to the family Cytophagaceae. Its coiled morphology is unique in contrast to closely related bacteria belonging to the genus Spirosoma, which have a short, rod-shaped morphology. The mechanisms that generate unique cell morphology are still enigmatic. In this study, using the Spirosoma linguale ATCC33905 strain, we isolated β-lactam (cefoperazone and amoxicillin)-resistant clones. These clones showed two different cell morphological changes: relatively loosely curved cells or small, horseshoe-shaped cells. Whole-genome resequencing analysis revealed the genetic determinants of β-lactam resistance and changes in cell morphology. The loose-curved clones commonly had mutations in Slin_5958 genes encoding glutamyl-tRNA amidotransferase B subunit, whereas the small, horseshoe-shaped clones commonly had mutations in either Slin_5165 or Slin_5509 encoding pyruvate dehydrogenase (PDH) components. Two clones, CFP1ESL11 and CFL5ESL4, which carried only one mutation in Slin_5958, showed almost perfectly straight, rod-shaped cells in the presence of amoxicillin. This result suggests that penicillin-binding proteins targeted by amoxicillin play an important role in the formation of a coiled morphology in this bacterium. In contrast, supplementation with acetate did not rescue the growth defect and abnormal cell size of the CFP5ESL9 strain, which carried only one mutation in Slin_5509. These results suggest that PDH is involved in cell-size maintenance in this bacterium.
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http://dx.doi.org/10.1038/s41598-021-92787-8DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8225782PMC
June 2021

Relevance of NADH Dehydrogenase and Alternative Two-Enzyme Systems for Growth of With Glucose, Lactate, and Acetate.

Front Bioeng Biotechnol 2020 20;8:621213. Epub 2021 Jan 20.

IBG-1: Biotechnology, Institute of Bio- and Geosciences, Forschungszentrum Jülich, Jülich, Germany.

The oxidation of NADH with the concomitant reduction of a quinone is a crucial step in the metabolism of respiring cells. In this study, we analyzed the relevance of three different NADH oxidation systems in the actinobacterial model organism by characterizing defined mutants lacking the non-proton-pumping NADH dehydrogenase Ndh (Δ) and/or one of the alternative NADH-oxidizing enzymes, L-lactate dehydrogenase LdhA (Δ) and malate dehydrogenase Mdh (Δ). Together with the menaquinone-dependent L-lactate dehydrogenase LldD and malate:quinone oxidoreductase Mqo, the LdhA-LldD and Mdh-Mqo couples can functionally replace Ndh activity. In glucose minimal medium the Δ mutant, but not the Δ and Δ strains, showed reduced growth and a lowered NAD/NADH ratio, in line with Ndh being the major enzyme for NADH oxidation. Growth of the double mutants ΔΔ and ΔΔ, but not of strain ΔΔ, in glucose medium was stronger impaired than that of the Δ mutant, supporting an active role of the alternative Mdh-Mqo and LdhA-LldD systems in NADH oxidation and menaquinone reduction. In L-lactate minimal medium the Δ mutant grew better than the wild type, probably due to a higher activity of the menaquinone-dependent L-lactate dehydrogenase LldD. The ΔΔ mutant failed to grow in L-lactate medium and acetate medium. Growth with L-lactate could be restored by additional deletion of , suggesting that repression by the transcriptional regulator SugR prevented growth on L-lactate medium. Attempts to construct a ΔΔΔ triple mutant were not successful, suggesting that Ndh, Mdh and LdhA cannot be replaced by other NADH-oxidizing enzymes in .
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http://dx.doi.org/10.3389/fbioe.2020.621213DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7874156PMC
January 2021

Prognostic impact of peripheral blood mRNA expression levels in response to azacytidine in MDS: A single-centre analysis.

Leuk Res Rep 2021 8;15:100231. Epub 2020 Dec 8.

Department of Haemato-Oncology, Saitama International Medical Centre, Saitama Medical University, Hidaka, Saitama, Japan.

To determine the impact of peripheral blood (PB) () mRNA levels in patients with primary myelodysplastic syndromes (MDS), we analysed the relationships between several clinical variables at the time of diagnosis and the haematological response of patients treated with azacytidine. We observed overall responses in 20 (63%) patients; there were no significant differences in clinical variables, including bone marrow blast counts, IPSS scores and IPSS-R risk scores, between responders and non-responders. The responders' PB mRNA levels were significantly lower than those of non-responders ( = 0.03). PB mRNA expression could be a marker for predicting the response to azacytidine in patients with MDS.
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http://dx.doi.org/10.1016/j.lrr.2020.100231DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7744716PMC
December 2020

High-throughput laboratory evolution reveals evolutionary constraints in Escherichia coli.

Nat Commun 2020 11 24;11(1):5970. Epub 2020 Nov 24.

RIKEN Center for Biosystems Dynamics Research, 6-2-3 Furuedai, Suita, Osaka, 565-0874, Japan.

Understanding the constraints that shape the evolution of antibiotic resistance is critical for predicting and controlling drug resistance. Despite its importance, however, a systematic investigation of evolutionary constraints is lacking. Here, we perform a high-throughput laboratory evolution of Escherichia coli under the addition of 95 antibacterial chemicals and quantified the transcriptome, resistance, and genomic profiles for the evolved strains. Utilizing machine learning techniques, we analyze the phenotype-genotype data and identified low dimensional phenotypic states among the evolved strains. Further analysis reveals the underlying biological processes responsible for these distinct states, leading to the identification of trade-off relationships associated with drug resistance. We also report a decelerated evolution of β-lactam resistance, a phenomenon experienced by certain strains under various stresses resulting in higher acquired resistance to β-lactams compared to strains directly selected by β-lactams. These findings bridge the genotypic, gene expression, and drug resistance gap, while contributing to a better understanding of evolutionary constraints for antibiotic resistance.
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http://dx.doi.org/10.1038/s41467-020-19713-wDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7686311PMC
November 2020

[Therapy-related acute promyelocytic leukemia developing during chemotherapy for thymic carcinoma].

Rinsho Ketsueki 2020 ;61(8):874-878

Clinical Laboratory, Saitama Medical University International Medical Center.

A 74-year-old man was admitted to hospital due to suspected acute leukemia. He had a history of thymic carcinoma, which had been treated with carboplatin in combination with either paclitaxel or amrubicin. However, the tumor remained unresponsive to these treatments. Administration of tegafur/gimeracil/oteracil (TS-1) was initiated, which resulted in tumor size reduction and a partial response. However, leukopenia persisted after the last TS-1 treatment, and four years after the initial treatment, increased blast cell counts were found in a blood film . Bone marrow analysis showed blasts with Auer rods, faggot cells, and dysplastic promyelocytes. Flow cytometry was positive for CD13, CD33, CD34, CD117, and myeloperoxidase, but negative for HLA-DR. PML-RARA fluorescence in situ hybridization was positive. Cytogenetic analysis revealed 47,XY,t (15;17) (q22;q21),+21. Thus, therapy-related acute promyelocytic leukemia (tAPL) was diagnosed. The patient achieved and maintained complete remission for more than 20 months by a de novo APL-treatment regimen including all-trans retinoic acid, arsenic trioxide and tamibarotene. Moreover, the thymic carcinoma has remained stable. Although secondary malignancies of thymic carcinoma have been previously reported, therapy-related leukemia, especially tAPL, is very rare.
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http://dx.doi.org/10.11406/rinketsu.61.874DOI Listing
October 2020

Assessment of dysplasia in bone marrow smear with convolutional neural network.

Sci Rep 2020 09 7;10(1):14734. Epub 2020 Sep 7.

Department of Hemato-Oncology, International Medical Center, Saitama Medical University, Saitama, Japan.

In this study, we developed the world's first artificial intelligence (AI) system that assesses the dysplasia of blood cells on bone marrow smears and presents the result of AI prediction for one of the most representative dysplasia-decreased granules (DG). We photographed field images from the bone marrow smears from patients with myelodysplastic syndrome (MDS) or non-MDS diseases and cropped each cell using an originally developed cell detector. Two morphologists labelled each cell. The degree of dysplasia was evaluated on a four-point scale: 0-3 (e.g., neutrophil with severely decreased granules were labelled DG3). We then constructed the classifier from the dataset of labelled images. The detector and classifier were based on a deep neural network pre-trained with natural images. We obtained 1797 labelled images, and the morphologists determined 134 DGs (DG1: 46, DG2: 77, DG3: 11). Subsequently, we performed a five-fold cross-validation to evaluate the performance of the classifier. For DG1-3 labelled by morphologists, the sensitivity, specificity, positive predictive value (PPV), negative predictive value (NPV), and accuracy were 91.0%, 97.7%, 76.3%, 99.3%, and 97.2%, respectively. When DG1 was excluded in the process, the sensitivity, specificity, PPV, NPV, and accuracy were 85.2%, 98.9%, 80.6%, and 99.2% and 98.2%, respectively.
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http://dx.doi.org/10.1038/s41598-020-71752-xDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7477564PMC
September 2020

Allogeneic hematopoietic stem cell transplantation at the first remission for younger adults with FLT3-internal tandem duplication AML: The JALSG AML209-FLT3-SCT study.

Cancer Sci 2020 Jul 29;111(7):2472-2481. Epub 2020 May 29.

Department of Hematology and Oncology, Nagoya University Graduate School of Medicine, Nagoya, Japan.

In this phase II multicenter study (JALSG AML209-FLT3-SCT), we aimed to prospectively elucidate the role of allogeneic hematopoietic stem cell transplantation (allo-HSCT) at first complete remission (CR1) for FLT3-internal tandem duplication (ITD)-positive AML. Newly diagnosed de novo AML patients with FLT3-ITD were enrolled at the achievement of CR1 and received allo-HSCT as soon as possible after the first consolidation therapy. Mutations of 57 genes in AML cells at diagnosis were also analyzed. Among 48 eligible patients with a median age of 38.5 (17-49) years, 36 (75%) received allo-HSCT at a median of 108 days after CR1. The median follow-up was 1726 days. The primary end-point, 3-year disease-free survival (DFS) based on an intent to treat analysis, was 43.8% (95% confidence interval [CI], 30%-57%), suggesting the efficacy of this treatment because the lower limit of the 95% CI exceeded the threshold response rate of 20%. The 3-year overall survival, post-transplant DFS, and non-relapse mortality rates were 54.2% (95% CI, 39%-67%), 58.3% (95% CI, 41%-72%), and 25.0% (95% CI, 12%-40%), respectively. The median ITD allelic ratio (AR) was 0.344 (0.006-4.099). Neither FLT3-ITD AR nor cooccurring genetic alterations was associated with a poor DFS. This prospective study indicated the efficacy and safety of allo-HSCT for FLT3-ITD AML patients in CR1. This study was registered at: www.umin.ac.jp/ctr/ as #UMIN000003433.
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http://dx.doi.org/10.1111/cas.14448DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7484840PMC
July 2020

Indolent T-cell lymphoproliferative disorder of the stomach successfully treated by radiotherapy.

J Clin Exp Hematop 2020 ;60(1):7-10

Department of Hematology, Comprehensive Cancer Center, International Medical Center, Saitama Medical University, Hidaka, Saitama, Japan.

Successful treatment of indolent T-cell lymphoproliferative disorder of the gastrointestinal tract (ITLPDGI) by chemotherapy is rare and watchful waiting is often performed for asymptomatic patients. We report a case of ITLPDGI successfully treated by involved field radiotherapy (IFRT). The patient presented with slow ITLPDGI localised to the stomach with mild symptoms. IFRT (30 Gy/20f) was administered, after which endoscopy revealed resolution of lesions and blood vessel appearance, and absence of proliferating abnormal lymphocytes was confirmed by biopsy. The patient remains lymphoma-free 1 year post-treatment. Although long-term follow-up and additional cases are essential for the evaluation of IFRT as a treatment option for localised ITLPDGL, complete remission after relatively low-dose IFRT is promising, particularly as this has been rarely achieved by chemotherapy.
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http://dx.doi.org/10.3960/jslrt.19022DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7187675PMC
January 2021

Peripheral T-cell lymphoma with gastrointestinal involvement and indolent T-lymphoproliferative disorders of the gastrointestinal tract.

Leuk Res 2020 04 29;91:106336. Epub 2020 Feb 29.

Departments of Haematology, International Medical Centre, Saitama Medical University, Hidaka, Saitama, Japan.

The 2017 WHO classification includes a new provisional entity of indolent T-lymphoproliferative disorders of the gastrointestinal tract (ITLPD-GIT). We investigated GI involvement of peripheral T-cell lymphoma (PTCL). Eighty-two patients were diagnosed with PTCL during 2007-2017. Eleven patients (13 %) had histologically-confirmed GI tract involvement {3 monomorphic epitheliotropic intestinal lymphoma (MEITL), 3 extranodal NK-/T-cell lymphoma nasal type (ENKL), 2 PTCL, not otherwise specified, 1 adult T-cell leukemia-lymphoma, 2 ITLPD-GIT}. Three patients each had lesions in the small intestine and multiple lesions, two each in the stomach and colon, and one in the duodenum. Six of the 11 patients remained alive. No perforation/stenosis was observed after chemo-radiotherapy, although one patient with ENKL developed gastric bleeding during chemotherapy. One patient with ITLPD-GIT (CD4/CD8/Ki67) with a colonic lesion showing diffuse edema and multiple aphtha by endoscope and diarrhea, initially diagnosed with MEITL, had active but stable disease after various chemotherapies for 1 year and no therapy for the next 5 years. Another patient with ITLPD-GIT (CD4/CD8/Ki67) with a localized gastric lesion and slight epigastralgia was in remission for 1 year after radiation. In conclusion, about 10 % of PTCLs were complicated by GI tract lesions and most had a poor prognosis. ITLPD-GIT should be considered as a differential diagnosis based on histology and clinical course. Local complications after chemo/radiotherapy in PTCL with GI involvement were not frequent.
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http://dx.doi.org/10.1016/j.leukres.2020.106336DOI Listing
April 2020

Suppression of antibiotic resistance evolution by single-gene deletion.

Sci Rep 2020 03 6;10(1):4178. Epub 2020 Mar 6.

Center for Biosystems Dynamics Research, RIKEN, 6-2-3 Furuedai, Suita, Osaka, 565-0874, Japan.

Antibiotic treatment generally results in the selection of resistant bacterial strains, and the dynamics of resistance evolution is dependent on complex interactions between cellular components. To better characterize the mechanisms of antibiotic resistance and evaluate its dependence on gene regulatory networks, we performed systematic laboratory evolution of Escherichia coli strains with single-gene deletions of 173 transcription factors under three different antibiotics. This resulted in the identification of several genes whose deletion significantly suppressed resistance evolution, including arcA and gutM. Analysis of double-gene deletion strains suggested that the suppression of resistance evolution caused by arcA and gutM deletion was not caused by epistatic interactions with mutations known to confer drug resistance. These results provide a methodological basis for combinatorial drug treatments that may help to suppress the emergence of resistant pathogens by inhibiting resistance evolution.
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http://dx.doi.org/10.1038/s41598-020-60663-6DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7060189PMC
March 2020

Bloodstream infection by multidrug-resistant Streptococcus oralis in a leukemic patient with febrile neutropenia after hematopoietic stem cell transplantation.

Transpl Infect Dis 2020 Apr 20;22(2):e13246. Epub 2020 Jan 20.

Department of Laboratory Medicine, Saitama Medical University International Medical Center, Saitama, Japan.

We reported the case of a patient with leukemia who developed febrile neutropenia after hematopoietic stem cell transplantation. Blood culture results revealed the presence of Streptococcus oralis, while antimicrobial susceptibility testing showed the resistance to penicillin and cephem. Furthermore, isolates were not susceptible to either meropenem or daptomycin but not to vancomycin. S oralis is known to belong to Streptococcus mitis group and be a causative agent of bacteremia in the neutropenic patients, but multidrug resistance of S oralis is rare. Our findings suggest that we might pay attention to the emergence of the microorganisms acquiring multidrug resistance in neutropenic patients.
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http://dx.doi.org/10.1111/tid.13246DOI Listing
April 2020

A case series of adult T-cell leukemia-lymphoma, associated with human T-cell leukemia virus type-1, at a single center in a non-viral-endemic metropolitan area.

J Clin Exp Hematop 2019 ;59(3):108-111

We examined 13 patients with adult T-cell leukemia-lymphoma (ATL) diagnosed between 2007 and 2018 at a single center in a metropolitan area non-endemic for human T-cell leukemia virus type I (HTLV-1). The median age of the patients (eight male, five female) was 65 years (range, 48-83). The time from onset of symptoms to referral to our center was relatively short (median, 2 months; range, 1-9 months). Upon referral, all patients were suspected to have lymphoma, five were examined for soluble IL-2 receptor and two were examined for anti-HTLV-1 antibody. In ten of the 13 (77%), the patient themselves or their relatives were born in Kyushu. The birth places of the remaining three patients were unknown. Three patients (23%) had family histories of lymphoma. They all exhibited aggressive ATL (five acute, eight lymphoma type); however, the disease status was generally stable, with relatively stable performance status and low scores for prognostic indices. After combination chemotherapy, eight (62%) achieved remission. However, long-term remission was achieved in only one patient with localized lymphoma-type ATL and one young patient after allogeneic hematopoietic stem cell transplantation. In conclusion, at a center in a metropolitan and HTLV-1 non-endemic area in Japan, patients with ATL were relatively young and mainly presented with aggressive subtypes. At initial referral to our center, all 13 patients were suspected of having lymphoma but only two of having ATL. For centers in similar areas of Japan, prompt diagnosis and appropriate treatment of ATL patients will become increasingly necessary following the recent migration of HTLV-1 carriers to non-endemic areas.
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http://dx.doi.org/10.3960/jslrt.19001DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6798142PMC
February 2020

Complete Genome Sequences of Three Star-Shaped Bacteria, Stella humosa, Stella vacuolata, and Stella Species ATCC 35155.

Microbiol Resour Announc 2019 Aug 8;8(32). Epub 2019 Aug 8.

Center for Biosystems Dynamics Research (BDR), RIKEN, Suita, Osaka, Japan

species are unique star-shaped alphaproteobacteria found in various environments. We report the complete genome sequences of three strains, ATCC 43930, ATCC 43931, and species ATCC 35155. These are the first complete genome sequences of members of the genus .
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http://dx.doi.org/10.1128/MRA.00719-19DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6687932PMC
August 2019

High-throughput identification of the sensitivities of an Escherichia coli ΔrecA mutant strain to various chemical compounds.

J Antibiot (Tokyo) 2019 07 21;72(7):566-573. Epub 2019 Feb 21.

RIKEN Center for Biosystems Dynamics Research, 6-2-3 Furuedai, Suita, Osaka, 565-0874, Japan.

Antibiotic resistance is considered a global threat to public health. Adaptive resistance mutations and the acquisition of resistance genes by horizontal gene transfer are known to be facilitated by the RecA-dependent SOS response during antibiotic treatment, making RecA inhibitors promising agents for the prevention of antibiotic resistance. However, the impact of RecA inactivation on antibiotic sensitivities remains unclear. Therefore, in this study, we performed high-throughput screening to determine the minimum inhibitory concentrations (MICs) of 217 chemicals, including both antibiotics and toxic chemicals of unknown drug action, in the wild-type MDS42 and the ΔrecA mutant strains of Escherichia coli. The ΔrecA mutant showed increased sensitivity to DNA-damaging agents, DNA replication inhibitors, and chromate stress, as well as to other chemicals, such as S-(2-aminoethyl)-L-cysteine, L-histidine, ruthenium red, D-penicillamine, carbonyl cyanide 3-chlorophenylhydrazone (CCCP), cerulenin, and L-cysteine. Microarray analysis showed further that the ΔrecA mutant had lower expressions of glnK, nac, and glnLG, which encode nitrogen assimilation regulators, as well as amtB, which encodes an ammonium transporter, compared with the wild type. These findings suggest that the ΔrecA mutation affects not only the SOS response but also amino acid metabolism.
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http://dx.doi.org/10.1038/s41429-019-0160-5DOI Listing
July 2019

False-positive blood culture results in patients with hematologic malignancies.

J Infect Chemother 2019 May 23;25(5):404-406. Epub 2019 Jan 23.

Department of Laboratory Medicine, Saitama Medical University International Medical Center, Saitama, Japan; Department of Laboratory Medicine, Saitama Medical University, Saitama, Japan.

Blood cultures are the most valuable tool when bacteremia is clinically suspected. Technical advances have led to the development of automated blood culture systems to detect bacterial infections. Usually positive signals in automated blood culture systems result from the proliferation of microorganisms. Cases are classified as false-positive when the automated blood culture system produces a positive signal but no microorganisms are detected on Gram-stained smears and no microorganism growth is observed in blood subcultures. False-positive blood culture results are very rare in patients with hematologic malignancies. Recently, we encountered four patients who had false-positive blood culture results. Two of the patients were diagnosed with acute leukemia, involving hyperleukocytosis and an excess of blasts. The other two patients were diagnosed with acute leukemia and diffuse large B cell lymphoma with leukocytopenia. Although hypercapnia or acidosis, apart from hyperleukocytosis, might also cause false-positive results, our cases clearly did not have these conditions. We should be aware of the possibility that false-positive blood culture results can occur in patients with leukocytopenia, as well as hyperleukocytosis. To understand the mechanisms responsible for the observed false-positive results, additional studies are needed after the accumulation of similar cases.
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http://dx.doi.org/10.1016/j.jiac.2018.12.010DOI Listing
May 2019

Understanding and engineering alcohol-tolerant bacteria using OMICS technology.

World J Microbiol Biotechnol 2018 Oct 19;34(11):157. Epub 2018 Oct 19.

Center for Biosystems Dynamics Research (BDR), RIKEN, 6-2-3 Furuedai, Suita, Osaka, 565-0874, Japan.

Microbes are capable of producing alcohols, making them an important source of alternative energy that can replace fossil fuels. However, these alcohols can be toxic to the microbes themselves, retaring or inhibiting cell growth and decreasing the production yield. One solution is improving the alcohol tolerance of such alcohol-producing organisms. Advances in omics technologies, including transcriptomic, proteomic, metabolomic, and genomic technologies, have helped us understand the complex mechanisms underlying alcohol toxicity, and such advances could assist in devising strategies for engineering alcohol-tolerant strains. This review highlights these advances and discusses strategies for improving alcohol tolerance using omics analyses.
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http://dx.doi.org/10.1007/s11274-018-2542-4DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6208762PMC
October 2018

The progression of severe aplastic anemia to hypoplastic leukemia in a long-term observation after the administration of pegylated rHuMGDF.

Hematol Rep 2018 Sep 5;10(3):7679. Epub 2018 Sep 5.

Department of Hemato-Oncology, Saitama Medical University International Medical Center, Hidaka, Saitama, Japan.

Thrombopoietin (TPO) is a critical regulator of hematopoiesis. We previously reported that a severe aplastic anemia (SAA) who received a short-term administration of pegylated recombinant human megakaryocyte growth and development factor (rHuMGDF). A trilineage hematologic response was induced, however the patient was diagnosed with leukemia after nine years and eight months from administration of rHuMGDF. In recent reports, somatic mutations in myeloid cancer candidate genes were present in one-third of the AA. A mutant clone may be expanded by rHuMGDF in our patient. The long-term safety of patients treated with TPO and eltrombopag remains unknown. Careful observations are warranted hereafter.
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http://dx.doi.org/10.4081/hr.2018.7679DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6151347PMC
September 2018

RNase E/G-dependent degradation of metE mRNA, encoding methionine synthase, in Corynebacterium glutamicum.

J Gen Appl Microbiol 2019 Mar 9;65(1):47-52. Epub 2018 Jul 9.

Department of Life Science and Technology, Tokyo Institute of Technology.

Corynebacterium glutamicum is used for the industrial production of various metabolites, including L-glutamic acid and L-lysine. With the aim of understanding the post-transcriptional regulation of amino acid biosynthesis in this bacterium, we investigated the role of RNase E/G in the degradation of mRNAs encoding metabolic enzymes. In this study, we found that the cobalamin-independent methionine synthase MetE was overexpressed in ΔrneG mutant cells grown on various carbon sources. The level of metE mRNA was also approximately 6- to 10-fold higher in the ΔrneG mutant strain than in the wild-type strain. A rifampicin chase experiment showed that the half-life of metE mRNA was approximately 4.2 times longer in the ΔrneG mutant than in the wild-type strain. These results showed that RNase E/G is involved in the degradation of metE mRNA in C. glutamicum.
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http://dx.doi.org/10.2323/jgam.2018.05.001DOI Listing
March 2019

Interobserver concordance of assessments of dysplasia and blast counts for the diagnosis of patients with cytopenia: From the Japanese central review study.

Leuk Res 2018 11 7;74:137-143. Epub 2018 Jun 7.

Department of Hematology and Oncology, Graduate School of Medicine, Kyoto University, Kyoto, Japan.

The diagnosis of myelodysplastic syndromes (MDS) is based on morphology and cytogenetics. However, limited information is currently available on the interobserver concordance of the assessment of dysplastic lineages (<10% or ≥10% in bone marrow (BM)). The revised International Prognostic Scoring System (IPSS-R) described a new threshold (2%) for BM blasts. However, the interobserver concordance of the categories (0-≤2% and >2-<5%) has limited data. The purpose of the present study was to investigate the assessment of dysplastic lineages and IPSS-R reproducibility. Our study was divided into two Steps. In each Step, the microscopic examinations were performed separately by two morphologists. Regarding the category of BM blasts ≤2% and >2-<5%, interobserver agreement was more than 'moderate' in all pairs (kappa test: 0.43-0.90). Regarding dysgranulopoiesis (dysG) and dyserythropoiesis (dysE) in BM, interobserver agreement was more than 'moderate' in all pairs (kappa test, dysG: 0.45-0.96, dysE: 0.45-0.81). Regarding the category of dysmegakaryopoiesis (dysMgk) in BM, interobserver agreement was more than moderate in 4 out of 5 pairs (kappa test: 0.58-1.00), and was fair for one pair (kappa test: 0.37). We consider that high interobserver concordance may be possible for the BM blast cell count (≤2% or >2-<5%) and dysplasia (<10% or ≥10%) of each lineage.
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http://dx.doi.org/10.1016/j.leukres.2018.06.003DOI Listing
November 2018

Final analysis of the JALSG Ph+ALL202 study: tyrosine kinase inhibitor-combined chemotherapy for Ph+ALL.

Ann Hematol 2018 Sep 24;97(9):1535-1545. Epub 2018 Apr 24.

National Hospital Organization Nagoya Medical Center, Nagoya, Japan.

The Japan Adult Leukemia Study Group (JALSG) Ph+ALL202 study reported a high complete remission (CR) rate for Philadelphia chromosome-positive acute lymphoblastic leukemia (Ph+ALL) patients treated with imatinib-combined chemotherapy. However, the long-term treatment efficacy remains uncertain. Here, we report a final analysis of the JALSG Ph+ALL202 study. The outcomes were compared with those of the JALSG ALL93 and ALL97 studies, which were conducted in the pre-imatinib era. Ninety-nine newly diagnosed Ph+ALL patients were enrolled in Ph+ALL202 (median age, 45 years; median follow-up, 4.5 years). CR was achieved in 96/99 (97%) patients. Fifty-nine of these 96 patients (61%) underwent allogeneic hematopoietic stem cell transplantation (allo-HSCT) in their first CR (CR1). The 5-year overall and disease-free survival (DFS) rates were 50 and 43%, respectively, which were significantly higher compared to those in the pre-imatinib era (15 and 19%, respectively). Multivariate analysis revealed that imatinib administration, allo-HSCT in CR1, and a white blood cell count < 30 × 10/L were favorable independent prognostic factors for long-term DFS. Improved odds of receiving allo-HSCT and a lower relapse rate leaded to good long-term outcomes. The 3-year DFS tended to be higher in PCR-negative than that in PCR-positive patients (29 vs. 14%) in the non-HSCT patients, and this tendency was also seen in the allo-HSCT patients (59 vs. 50%). The higher rate of CR upon imatinib use may have contributed to these improvements.
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http://dx.doi.org/10.1007/s00277-018-3323-8DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6097750PMC
September 2018

Glutamine-rich toxic proteins GrtA, GrtB and GrtC together with the antisense RNA AsgR constitute a toxin-antitoxin-like system in Corynebacterium glutamicum.

Mol Microbiol 2018 06 19;108(5):578-594. Epub 2018 Apr 19.

Research Institute of Innovative Technology for the Earth, Kyoto, Japan.

The Corynebacterium glutamicum R grtA (cgR_2936), grtB (cgR_2934) and grtC (cgR_2933) genes were identified as paralogs encoding glutamine-rich toxic proteins. We also identified a new antisense small RNA AsgR (antisense sRNA for grtA) that overlaps the 3' end of the grtA gene. Single over-expressions of grtA, grtB and grtC resulted in complete inhibition of Escherichia coli cell growth. This growth was rescued by co-expression of AsgR. Similar effects were observed in C. glutamicum, although the toxicities of these proteins were moderate. Inhibition of AsgR transcription resulted in increased levels and prolonged half-lives of grtA, grtB and grtC mRNAs. We also found that the expression levels of grtA, grtB and grtC were increased in an RNase III deletion mutant. Primer extension analysis revealed the RNase III cleavage site to be in the 3' untranslated region (3'-UTR) of the grtA mRNA. The expression levels of grtA, grtB and grtC were increased after exposure to several stresses, including heat shock, treatment with penicillin G, lysozyme or H O . The deletions of grtABC and asgR genes resulted in decreased survival rate under several stresses. These results indicate that GrtABC and AsgR constitute a type I toxin-antitoxin-like system in C. glutamicum.
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http://dx.doi.org/10.1111/mmi.13951DOI Listing
June 2018

Toward prediction and control of antibiotic-resistance evolution.

Curr Opin Biotechnol 2018 12 14;54:45-49. Epub 2018 Feb 14.

Quantitative Biology Center, RIKEN, 6-2-3 Furuedai, Suita, Osaka 565-0874, Japan.

The emergence of antibiotic-resistant bacteria is a serious public concern. To deal with this problem, recent advances in technology and the use of laboratory evolution experiments have provided valuable information on the phenotypic and genotypic changes that occur during the evolution of resistance. These studies have demonstrated the existence of evolutionary constraints on the development of drug-resistance, which suggests predictability in its evolution. In this review, we focus on the possibility to predict and control the evolution of antibiotic resistance, based on quantitative analysis of phenotypic and genotypic changes observed in bacterial laboratory evolution. We emphasize the key challenges in evolutionary biology that will contribute to the development of appropriate treatment strategies for preventing resistance evolution.
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http://dx.doi.org/10.1016/j.copbio.2018.01.026DOI Listing
December 2018

Diagnostic performance of procalcitonin, presepsin, and C-reactive protein in patients with hematological malignancies.

J Clin Lab Anal 2017 Nov 30;31(6). Epub 2017 Jan 30.

Department of Laboratory Medicine, International Medical Center, Saitama Medical University, Saitama, Japan.

Introduction: Infections represent a major complication of hematological malignancies. C-reactive protein (CRP) and procalcitonin (PCT) have been used as diagnostic biomarkers of infections, but do not produce definitive findings. Recently, a new biomarker, presepsin, has been used as a diagnostic tool for detecting infections in the fields of emergency and neonatal medicine. However, the usefulness of presepsin for identifying infections in patients with hematological malignancies, including those who develop febrile neutropenia, remains unclear.

Methods: In this study, we retrospectively analyzed the utility of PCT, presepsin, and CRP as biomarkers of infections during 49 febrile episodes that occurred in 28 patients with hematological malignancies.

Results: The levels of PCT, but not those of CRP or presepsin, were significantly higher in the infection group than in the uninfected group (P<.03), indicating that PCT might be a more sensitive biomarker of infections. No differences in presepsin levels were detected between the patients with and without neutropenia, or between the infected and uninfected patients with neutropenia, indicating that presepsin might have less diagnostic value in patients with neutropenia.

Conclusions: We conclude that PCT might provide additional information and could be used in combination with other biomarkers to detect infections in patients with hematological malignancies.
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http://dx.doi.org/10.1002/jcla.22147DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6817173PMC
November 2017

Phase II study of imatinib-based chemotherapy for newly diagnosed BCR-ABL-positive acute lymphoblastic leukemia.

Am J Hematol 2017 Apr 21;92(4):367-374. Epub 2017 Feb 21.

National Hospital Organization Nagoya Medical Center, Nagoya, Japan.

This study investigated the efficacy of imatinib based therapy with intensified consolidation therapy in patients with Philadelphia chromosome (Ph)-positive acute lymphoblastic leukemia (ALL) to prevent early relapse. We conducted a phase II trial of imatinib-combined chemotherapy for newly diagnosed BCR-ABL-positive ALL in adults. Sixty-eight patients were included in the trial between October 2008 and December 2010. The median age was 49 years, with 28 patients >55 years of age. Sixty-five patients achieved CR (95.6%). The estimated 2-year event-free survival (EFS) and overall survival (OS) were 62.3% and 67.4%, respectively. Allogeneic stem cell transplantation (allo-SCT) at initial CR was performed in 43 patients. Thirty-five of 39 patients <55 years and 8 of 26 patients >55 years underwent allo-SCT at first CR. The 3-year OS in patients <55 years receiving allo-SCT at first CR, patients >55 years receiving allo-SCT at first CR, patients <55 years not receiving allo-SCT at first CR, and patients >55 years not receiving allo-SCT at first CR were 80.4%, 41.1%, 32.5%, and 52.0%, respectively (P = 0.058). The three-year EFS in each group was 76.7%, 53.6%, not reached, and 26.4%, respectively (P = 0.150). A high CR rate was observed with imatinib-based chemotherapy allowing allo-SCT in a high proportion of patients, particularly those <55 years. Moreover, intensified consolidation therapy reduced early relapse rates following induction therapy and resulted in improved OS and EFS rates following allo-SCT. This trial was registered with the UMIN (000001226).
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http://dx.doi.org/10.1002/ajh.24653DOI Listing
April 2017

Polynucleotide Phosphorylase, RNase E/G, and YbeY Are Involved in the Maturation of 4.5S RNA in Corynebacterium glutamicum.

J Bacteriol 2017 03 14;199(5). Epub 2017 Feb 14.

Research Institute of Innovative Technology for the Earth, Kyoto, Japan

has been applied for the industrial production of various metabolites, such as amino acids. To understand the biosynthesis of the membrane protein in this bacterium, we investigated the process of signal recognition particle (SRP) assembly. SRP is found in all three domains of life and plays an important role in the membrane insertion of proteins. SRP RNA is initially transcribed as precursor molecules; however, relatively little is known about its maturation. In , SRP consists of the Ffh protein and 4.5S RNA lacking an Alu domain. In this study, we found that 3'-to-5' exoribonuclease, polynucleotide phosphorylase (PNPase), and two endo-type RNases, RNase E/G and YbeY, are involved in the 3' maturation of 4.5S RNA in The mature form of 4.5S RNA was inefficiently formed in Δ Δ mutant cells, suggesting the existence of an alternative pathway for the 3' maturation of 4.5S RNA. Primer extension analysis also revealed that the 5' mature end of 4.5S RNA corresponds to that of the transcriptional start site. Immunoprecipitated Ffh protein contained immature 4.5S RNA in Δ, Δ, and Δ mutants, suggesting that 4.5S RNA precursors can interact with Ffh. These results imply that the maturation of 4.5S RNA can be performed in the 4.5S RNA-Ffh complex. Overproduction of a membrane protein, such as a transporter, is useful for engineering of strains of , which is a workhorse of amino acid production. To understand membrane protein biogenesis in this bacterium, we investigated the process of signal recognition particle (SRP) assembly. SRP contains the Ffh protein and SRP RNA and plays an important role in the membrane insertion of proteins. Although SRP RNA is highly conserved among the three domains of life, relatively little is known about its maturation. We show that PNPase, RNase E/G, and YbeY are involved in the 3' maturation of the SRP RNA (4.5S RNA) in this bacterium. This indicates that 3' end processing in this organism is different from that in other bacteria, such as .
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http://dx.doi.org/10.1128/JB.00798-16DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5309912PMC
March 2017

RNase III mediated cleavage of the coding region of mraZ mRNA is required for efficient cell division in Corynebacterium glutamicum.

Mol Microbiol 2016 Mar 10;99(6):1149-66. Epub 2016 Feb 10.

Research Institute of Innovative Technology for the Earth, Kyoto, Japan.

The Corynebacterium glutamicum R cgR_1959 gene encodes an endoribonuclease of the RNase III family. Deletion mutant of cgR_1959 (Δrnc mutant) showed an elongated cell shape, and presence of several lines on the cell surface, indicating a required of RNase III for maintaining normal cell morphology in C. glutamicum. The level of mraZ mRNA was increased, whereas cgR_1596 mRNA encoding a putative cell wall hydrolase and ftsEX mRNA were decreased in the Δrnc mutant. The half-life of mraZ mRNA was significantly prolonged in the Δrnc and the Δpnp mutant strains. This indicated that the degradation of mraZ mRNA was performed by RNase III and the 3'-to-5' exoribonuclease, PNPase. Northern hybridization and primer extension analysis revealed that the cleavage site for mraZ mRNA by RNase III is in the coding region. Overproduction of MraZ resulted in an elongated cell shape. The expression of ftsEX decreased while that of cgR_1596 unchanged in an MraZ-overexpressing strain. An electrophoretic mobility shift assay and a transcriptional reporter assay indicate that MraZ is a transcriptional repressor of ftsEX in C. glutamicum. These results indicate that RNase III is required for efficient expression of MraZ-dependent ftsEX and MraZ-independent cgR_1596.
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http://dx.doi.org/10.1111/mmi.13295DOI Listing
March 2016

Unrelated bone marrow transplantation or immediate umbilical cord blood transplantation for patients with acute myeloid leukemia in first complete remission.

Eur J Haematol 2016 Sep 20;97(3):278-87. Epub 2016 Jan 20.

Saitama Medical Center, Jichi Medical University, Saitama, Japan.

Background: While unrelated bone marrow transplantation (UBMT) has been widely used as alternative donor transplantation, the use of umbilical cord blood transplantation (UCBT) is increasing recently.

Methods: We conducted a decision analysis to address which transplantation procedure should be prioritized for younger patients with acute myeloid leukemia (AML) harboring high- or intermediate-risk cytogenetics in first complete remission (CR1), when they lack a matched related donor but have immediate access to a suitable umbilical cord blood unit. Main sources for our analysis comprised the data from three phase III trials for a chemotherapy cohort (n = 907) and the registry data for a transplantation cohort (n = 752).

Results: The baseline analysis showed that when the 8/8 match was considered for UBMT, the expected 5-year survival rate was higher for UBMT than for UCBT (58.1% vs. 51.8%). This ranking did not change even when the 7/8 match was considered for UBMT. Sensitivity analysis showed consistent superiority of UBMT over UCBT when the time elapsed between CR1 and UBMT was varied within a plausible range of 3-9 months.

Conclusions: These results suggest that 8/8 or 7/8 UBMT is a better transplantation option than UCBT even after allowing time required for donor coordination.
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http://dx.doi.org/10.1111/ejh.12723DOI Listing
September 2016

Proposal of criteria for dyserythropoiesis in the diagnosis of myelodysplastic syndromes.

Int J Hematol 2016 Feb 25;103(2):227-33. Epub 2015 Nov 25.

Japanese Red Cross Nagasaki Atomic Bomb Hospital, Nagasaki, Japan.

The percentage manifesting dysplasia in bone marrow needed to qualify as significant is ≥10 % in each lineage. However, detailed analyses of this threshold have not been reported. Here, we analyzed dyserythropoiesis (dysE) in 109 myelodysplastic syndromes (MDS) patients with 21 immune thrombocytopenia (ITP)/12 hemolytic anemia (HA) patients as a control. In present study, mild megaloblastic erythroblasts were specifically named 'red cell with abnormal chromatin clumping (RCACC)'. RCACC ≥10 % in erythroblasts was observed in 29 % of ITP patients and 58 % of HA patients. The numbers of MDS patients with RCACC in erythroblasts <10, 10-19 and ≥20 % were 1, 3, and 105, respectively. We analyzed dysE criteria according to the WHO classification (original WHO dysE). Most of our MDS patients (98 %) had original WHO dysE ≥20 %. The ITP patients with original WHO dysE ≥10 % was 48 %, and there were no ITP patients had original WHO dysE ≥20 %. Sixty-seven percent of HA patients had original WHO dysE ≥10 %, and three patients (25 %) had original WHO dysE ≥20 %. Raising the threshold of the original WHO dysE from 10 to 20 or 30 % may provide more suitable criteria. If RCACC is not included in dysE criteria, we think that '10 %' is a suitable threshold for the determination of dyserythropoiesis.
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http://dx.doi.org/10.1007/s12185-015-1916-8DOI Listing
February 2016
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