Journal of Microbiology

Review

Trans-acting regulators of ribonuclease activity: Jaejin Lee , Minho Lee , Kangseok Lee; J. Microbiol. 2021;59(4):341-359. Published online March 29, 2021; DOI: https://doi.org/10.1007/s12275-021-0650-6

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Abstract: RNA metabolism needs to be tightly regulated in response to changes in cellular physiology. Ribonucleases (RNases) play an essential role in almost all aspects of RNA metabolism, including processing, degradation, and recycling of RNA molecules. Thus, living systems have evolved to regulate RNase activity at multiple levels, including transcription, post-transcription, post-translation, and cellular localization. In addition, various trans-acting regulators of RNase activity have been discovered in recent years. This review focuses on the physiological roles and underlying mechanisms of trans-acting regulators of RNase activity.

Journal Article

Lentibacillus cibarius sp. nov., isolated from kimchi, a Korean fermented food: Young Joon Oh , Joon Yong Kim , Hee Eun Jo , Hyo Kyeong Park , Seul Ki Lim , Min-Sung Kwon , Hak-Jong Choi; J. Microbiol. 2020;58(5):387-394. Published online April 11, 2020; DOI: https://doi.org/10.1007/s12275-020-9507-7

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Abstract: Two bacterial strains designated NKC220-2T and NKC851-2 were isolated from commercial kimchi from different areas in Korea. The strains were Gram-positive, aerobic, oxidaseand catalase-positive, rod-shaped, spore-forming, non-motile, and halophilic bacteria. Both strains grew without NaCl, unlike type species in the genus Lentibacillus. The optimal pH for growth was 8.0, higher than that of the type species in the genus Lentibacillus, although growth was observed at pH 5.5–9.0. 16S rRNA gene sequence-based phylogenetic analysis indicated that the two strains (99.3–99.9% similarity) are grouped within the genus Lentibacillus and most closely related to Lentibacillus juripiscarius IS40-3T (97.4–97.6% similarity) isolated from fish sauce in Thailand. OrthoANI value between two novel strains and Lentibacillus lipolyticus SSKP1- 9T (79.5–79.6% similarity) was far lower than the species demarcation threshold. Comparative genomic analysis displayed differences between the two strains as well as among other strains belonging to Lentibacillus. Furthermore, each isolate had strain-specific groups of orthologous genes based on pangenome analysis. Genomic G + C contents of strains NKC- 220-2T and NKC851-2 were 41.9 and 42.2 mol%, respectively. The strains contained meso-diaminopimelic acid in their cell walls, and the major menaquinone was menaquinone-7. Phosphatidylglycerol, diphosphatidylglycerol, and an unidentified glycolipid, aminophospholipid, and phospholipid were the major polar lipid components of both strains. The major cellular fatty acids of the strains were anteiso-C15:0 and anteiso- C17:0. Based on phenotypic, genomic, phylogenetic, and chemotaxonomic features, strains NKC220-2T and NKC851-2 represent novel species of the genus Lentibacillus, for which the name Lentibacillus cibarius sp. nov. is proposed. The type strain is NKC220-2T (= KACC 21232T = JCM 33390T).

Review

[MINIREVIEW] Alanine dehydrogenases in mycobacteria: Ji-A Jeong , Jeong-Il Oh; J. Microbiol. 2019;57(2):81-92. Published online January 31, 2019; DOI: https://doi.org/10.1007/s12275-019-8543-7

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11 Citations

Abstract: Since NAD(H)-dependent L-alanine dehydrogenase (EC 1.1.4.1; Ald) was identified as one of the major antigens present in culture filtrates of Mycobacterium tuberculosis, many studies on the enzyme have been conducted. Ald catalyzes the reversible conversion of pyruvate to alanine with concomitant oxidation of NADH to NAD+ and has a homohexameric quaternary structure. Expression of the ald genes was observed to be strongly upregulated in M. tuberculosis and Mycobacterium smegmatis grown in the presence of alanine. Furthermore, expression of the ald genes in some mycobacteria was observed to increase under respiration-inhibitory conditions such as oxygen-limiting and nutrient-starvation conditions. Upregulation of ald expression by alanine or under respiration-inhibitory conditions is mediated by AldR, a member of the Lrp/AsnC family of transcriptional regulators. Mycobacterial Alds were demonstrated to be the enzymes required for utilization of alanine as a nitrogen source and to help mycobacteria survive under respiration-inhibitory conditions by maintaining cellular NADH/NAD+ homeostasis. Several inhibitors of Ald have been developed, and their application in combination with respiration-inhibitory antitubercular drugs such as Q203 and bedaquiline was recently suggested.

Research Support, Non-U.S. Gov'ts

Performance of a real-time PCR assay for the rapid identification of Mycobacterium species: Hye-young Wang , Hyunjung Kim , Sunghyun Kim , Do-kyoon Kim , Sang-Nae Cho , Hyeyoung Lee; J. Microbiol. 2015;53(1):38-46. Published online January 4, 2015; DOI: https://doi.org/10.1007/s12275-015-4495-8

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18 Citations

Abstract: Mycobacteria cause a variety of illnesses that differ in severity and public health implications. The differentiation of Mycobacterium tuberculosis (MTB) from nontuberculous mycobacteria (NTM) is of primary importance for infection control and choice of antimicrobial therapy. The diagnosis of diseases caused by NTM is difficult because NTM species are prevalent in the environment and because they have fastidious properties. In the present study, we evaluated 279 clinical isolates grown in liquid culture provided by The Catholic University of Korea, St. Vincent’s Hospital using real-time PCR based on mycobacterial rpoB gene sequences. The positive rate of real-time PCR assay accurately discriminated 100% (195/195) and 100% (84/84) between MTB and NTM species. Comparison of isolates identified using the MolecuTech REBA Myco-ID? and Real Myco-ID? were completely concordant except for two samples. Two cases that were identified as mixed infection (M. intracellulare-M. massiliense and M. avium-M. massiliense co-infection) by PCRREBA assay were only detected using M. abscessus-specific probes by Real Myco-ID?. Among a total of 84 cases, the most frequently identified NTM species were M. intracellulare (n=38, 45.2%), M. avium (n=18, 23.7%), M. massiliense (n=10, 13.2%), M. fortuitum (n=5, 6%), M. abscessus (n=3, 3.9%), M. gordonae (n=3, 3.9%), M. kansasii (n=2, 2.4%), M. mucogenicum (n=2, 2.4%), and M. chelonae (n= 1, 1.2%). Real Myco-ID? is an efficient tool for the rapid detection of NTM species as well as MTB and sensitive and specific and comparable to conventional methods.

Evaluation of the Cell Growth of Mycobacteria Using Mycobacterium smegmatis mc2 155 as a Representative Species: Jorge A. Gonzalez-y-Merchand , Ruben Zaragoza-Contreras , Rosalina Guadarrama-Medina , Addy C. Helguera-Repetto , Sandra Rivera-Gutierrez , Jorge F. Cerna-Cortes , Leopoldo Santos-Argumedo , Robert A. Cox; J. Microbiol. 2012;50(3):419-425. Published online June 30, 2012; DOI: https://doi.org/10.1007/s12275-012-1556-0

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Abstract: The study of the in vitro cell growth of mycobacteria still remains a fastidious, difficult, and time-consuming procedure. In addition, assessing mycobacterial growth in the laboratory is often complicated by cell aggregation and slow growth-rate. We now report that the use of a stainless steel spring in the culture led to an absence of large cell clumps, to a decrease of dead cells in the exponential phase and to growth of a more homogeneous population of large cells. We also report that flow cytometry is a rapid, simple and reliable approach to monitor mycobacterial cell growth and viability. Here, we monitored Mycobacterium smegmatis cellular growth by optical density, dry cell mass, and colony forming units; in addition, viability, cell size and granularity profiles were analyzed by flow cytometry, and cell morphology by electron microscopy. Cultures monitored by flow cytometry may lead to a better understanding of the physiology of mycobacteria. Moreover, this methodology may aid in characterizing the cell growth of other fastidious species of microorganisms.

Evaluation of Three Molecular Methods of Repetitive Element Loci for Differentiation of Mycobacterium avium subsp. paratuberculosis (MAP): Amr El-Sayed , Abdulwahed Ahmed Hassan , Saleh Natour , Amir Abdulmawjood , Michael Bulte , Wilfried Wolter , Michael Zschock; J. Microbiol. 2009;47(3):253-259. Published online June 26, 2009; DOI: https://doi.org/10.1007/s12275-008-0257-1

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15 Citations

Abstract: The aim of the present study is to evaluate the efficiency of three methods to determine the molecular diversity of 34 Mycobacterium avium subsp. paratuberculosis (MAP) strains isolated from 17 cattle herds. The applied methods included the analysis of sequence polymorphism of the mononucleotide (G1 and G2) and trinucleotide sequences (GGT) of the Short Sequence Repeats (SSR) and the determination of size polymorphism of 9 different Mycobacterial Interspersed Repetitive Units (MIRU) and 6 Variable Number Tandem Repeats (VNTR). Sequence analysis of SSR of 34 isolates showed 4, 6, and 2 alleles of G1, G2, and GGT repeats, respectively. The amplification of the investigated 9 MIRU units revealed only two discriminatory genotyping systems (MIRU2 and MIRU3). Out of 6 VNTR PCR differentiation methods, only one method could be recommended for genotyping purposes. The profile 7g-12g-4ggt-II-b-2 of the combination systems G1-G2-GGT-MIRU2-MIRU3-VNTR1658 dominates among the examined isolates and was
detected in 14.7% of the isolates. The use of certain repetitive loci of SSR, MIRU, and VNTR techniques in this study showed greater potential than others for the characterization of MAP isolates. The recommended loci can be used for the epidemiological tracing of MAP field strains and to determine the relationships
between isolates in different herds.

Expression and Regulation of Ribulose 1,5-Bisphosphate Carboxylase/Oxygenase Genes in Mycobacterium sp. Strain JC1 DSM 3803: Jae Ho Lee , Dong Oh Park , Sae Woong Park , Eun Ha Hwang , Jeong Il Oh , Young Min Kim; J. Microbiol. 2009;47(3):297-307. Published online June 26, 2009; DOI: https://doi.org/10.1007/s12275-008-0210-3

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10 Citations

Abstract: Ribulose 1,5-bisphosphate carboxylase/oxygenase (RubisCO) is the key enzyme of the Calvin reductive pentose phosphate cycle. Two sets of structural genes (cbbLS-1 and -2) for form I RubisCO have been previously identified in the Mycobacterium sp. strain JC1, which is able to grow on carbon monoxide (CO) or methanol as sole sources of carbon and energy. Northern blot and reverse transcriptase PCR showed that the cbbLS-1 and -2 genes are expressed in cells grown on either carbon monoxide (CO) or methanol, but not in cells grown in nutrient broth. A promoter assay revealed that the cbbLS-2 promoter has a higher activity than the cbbLS-1 promoter in both CO- and methanol-grown cells, and that the activities of both promoters were higher in CO-grown cells than in methanol-grown cells. A gel mobility shift assay and footprinting assays showed that CbbR expressed in Escherichia coli from a cbbR gene, which is located downstream of cbbLS-1 and transcribed in the same orientation as that of the cbbLS genes, specifically bound to the promoter regions of the cbbLS-1 and -2 genes containing inverted repeat sequence. A DNase I footprinting assay revealed that CbbR protected positions -59 to -3 and -119 to -78 of the cbbLS-1 and -2 promoters, respectively. Overexpression of CbbR induced the transcription of RubisCO genes in Mycobacterium sp. strain JC1 grown in nutrient broth. Our results suggest that the CbbR product from a single cbbR gene may positively regulate two cbbLS operons in the Mycobacterium sp. strain JC1 as is the case for Rhodobacter sphaeroides and Cupriavidus necator.

Journal Article

Targeting the rpoB Gene Using Nested PCR-Restriction Fragment Length Polymorphism for Identification of Nontuberculous Mycobacteria in Hospital Tap Water: Ji-Hyun Shin , Hae-Kyung Lee , Eun-Jin Cho , Jae-Yon Yu , Yeon-Ho Kang; J. Microbiol. 2008;46(6):608-614. Published online December 24, 2008; DOI: https://doi.org/10.1007/s12275-008-0102-6

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19 Citations

Abstract: Nontuberculous mycobacteria (NTM) are ubiquitous in the environment and can cause nosocomial infections in immunocompromised patients. Recently the presence of NTM in public drinking water and hospital water distribution systems has been reported. Their ability to form biofilms and their resistance to chlorine both contribute to their survival and colonization in water distribution systems. Here we analyzed thirty-two hospital tap water samples that were collected from different locations in three hospitals so as to evaluate the prevalence of NTM species. The water samples were concentrated by membrane filtration and then eluted with sterilized water following sonication. Two-step direct PCR targeting the rpoB gene, restriction fragment length polymorphism (RFLP) using the MspI restriction enzyme, and sequence analysis were performed for identification of NTM to the species level. The sequences of each PCR product were analyzed using BLASTN. Seven samples (7/32, 21.9%) were positive for NTM as determined by nested-PCR. The PCR-RFLP results indicated five different patterns among the seven positive PCR samples. The waterborn NTM were identified, including M. peregrinum, M. chelonae (2 cases), M. abscessus, M. gordonae (2 cases), and Mycobacterium sp. JLS. The direct two-step PCR-RFLP method targeting the rpoB gene was effective for the detection and the differentiation of NTM species from hospital tap water.

Genomic polymorphism in clinical mycobacterial strains analyzed by pulsed-field gel electrophoresis: Kim, Jeong Ran , Kim, Cheorl Ho; J. Microbiol. 1997;35(3):172-176.

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Abstract: The Mycobacterium tuberculosis clinical isolates in Korea, showing different drug resistances, were analyzed by comparing large restriction fragment (LRF) patterns produced y digestion of genomic DNA with infrequent-cutting endonucleases of SpeI, AsnI and pulsed-field gel electrophoresis (PFGE). SpeI and AsnI allowed with AsnI and SpeI, strains yielded an absolutely identical pattern for Korean type's mycobacteria even though they showed different drug resisstance. However, when three M. tuberculosis strains, showing drug resistance, were digested with XbaI, patterns were different from those of the other M. tuberculosis strians which are susceptible to drugs. This study reveals that the comparison of chromosomal restriction patterns is very useful as an additional aid for the differentiation and identification of M. tuberculosis strains showing drug resistances.

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