Journal of Microbiology

Journal Article

Genetic changes in plaque-purified varicella vaccine strain Suduvax during in vitro propagation in cell culture: Hye Rim Hwang , Se Hwan Kang , Chan Hee Lee; J. Microbiol. 2021;59(7):702-707. Published online June 1, 2021; DOI: https://doi.org/10.1007/s12275-021-1062-3

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Abstract: Infection by varicella-zoster virus (VZV) can be prevented by using live attenuated vaccines. VZV vaccine strains are known to evolve rapidly in vivo, however, their genetic and biological effects are not known. In this study, the plaque-purified vaccine strain Suduvax (PPS) was used to understand the genetic changes that occur during the process of propagation in in vitro cell culture. Full genome sequences of three different passages (p4, p30, and p60) of PPS were determined and compared for genetic changes. Mutations were found at 59 positions. The number of genetically polymorphic sites (GPS) and the average of minor allele frequency (MAF) at GPSs were not significantly altered after passaging in cell culture up to p60. The number of variant nucleotide positions (VNPs), wherein GPS was found in at least one passage of PPS, was 149. Overall, MAF changed by less than 5% at 52 VNPs, increased by more than 5% at 42 VNPs, and decreased by more than 5% at 55 VNPs in p60, compared with that seen in p4. More complicated patterns of changes in MAF were observed when genetic polymorphism at 149 VNPs was analyzed among the three passages. However, MAF decreased and mixed genotypes became unequivocally fixed to vaccine type in 23 vaccine-specific positions in higher passages of PPS. Plaque-purified Suduvax appeared to adapt to better replication during in vitro cell culture. Further studies with other vaccine strains and in vivo studies will help to understand the evolution of the VZV vaccine.

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

In Vitro Development and Transfer of Resistance to Chlortetracycline in Bacillus subtilis: Menghong Dai , Junjie Lu , Yulian Wang , Zhenli Liu , Zonghui Yuan; J. Microbiol. 2012;50(5):807-812. Published online November 4, 2012; DOI: https://doi.org/10.1007/s12275-012-1454-5

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16 Scopus

Abstract: The present criteria and rules controlling the approval of the use of probiotics are limited to antibiotic resistance patterns and the presence of antibiotic resistance genes in bacteria. There is little information available in the literature regarding the risk of the usage of probiotics in the presence of antibiotic pressure. In this study we investigated the development and transfer of antibiotic resistance in Bacillus subtilis selected in vitro by chlortetracycline in a stepwise manner. Bacillus subtilis was exposed to increasing concentrations of chlortetracyclineto induce in vitro resistance to chlortetracycline, and the minimal inhibitory concentrations were determinedfor the mutants. Resistant B. subtilis were conjugated with Escherichia coli NK5449 and Enterococcus faecalis JH2-2 using the filter mating. Three B. subtilis tetracycline resistant mutants (namely, BS-1, BS-2, and BS-3) were derived in vitro. A tetracycline resistant gene, tet (K), was found in the plasmids of BS-1 and BS-2. Three conjugates (BS-1N, BS-2N, and BS-3N) were obtained when the resistant B. subtilis was conjugated with E. coli NK5449. The conjugation frequencies for the BS-1N, BS-2N, and BS-3N conjugates were 4.57×10-7, 1.4×10-7, and 1.3×10-8, respectively. The tet(K) gene was found only in the plasmids of BS-1N. These results indicate that long-term use of probiotics under antibiotic selection pressure could cause antibiotic resistance, and the resistance gene could be transferred to other bacteria. The risk arising from the use of probiotics under antibiotic pressure should be considered in the criteria and rules for the safety assessment of probiotics.

Modulation of Secreted Virulence Factor Genes by Subinhibitory Concentrations of Antibiotics in Pseudomonas aeruginosa: Lixin Shen , Ying Shi , Dan Zhang , Jinhua Wei , Michael G. Surette , Kangmin Duan; J. Microbiol. 2008;46(4):441-447. Published online August 31, 2008; DOI: https://doi.org/10.1007/s12275-008-0054-x

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46 Scopus

Abstract: Recent studies have shown that subinhibitory antibiotics play important roles in regulating bacterial genes including virulence factor genes. In this study, the expression of 13 secreted virulence related gene clusters of Pseudomonas aeruginosa, an important opportunistic pathogen, was examined in the presence of subinhibitory concentrations of 4 antibiotics: vancomycin, tetracycline, ampicilin and azithromycin. Activation of gene expression was observed with phzA1, rhlAB, phzA2, lasB, exoY, and exoS. Subinhibitory concentrations of vancomycin resulted in more than 10-fold increase of rhlAB and phzA2 transcription. Both rhamnolipid production and pyocyanin production were significantly elevated, correlating phenotypes with the increased transcription. P. aeruginosa swarming and swimming motility also increased. Similar results were observed with subinhibitory tetracycline, azithromycin and ampicillin. These results indicate that the antibiotics at low concentrations can up-regulate virulence factors and therefore influence bacterial pathogenesis.

Development of a Virus Concentration Method and its Application for the Detection of Noroviruses in Drinking Water in China: Junyi Liu , Qingping Wu , Xiaoxia Kou; J. Microbiol. 2007;45(1):48-52.; DOI: https://doi.org/2492 [pii]

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Abstract: A new procedure for the concentration of nonoviruses from water samples has been developed. This procedure (calcium flocculation-citrate dissolution method) uses the following steps: virus flocculation formed by treatment with 1 M CaCl2 and 1 M Na2HPO4, virus release by sodium citrate dissolution (0.3 M Na citrate, pH 3.5), and virus re-concentration by ultrafiltration. When reverse transcription (RT)-PCR was performed after the procedure, the overall detection sensitivity for seeded noroviruses in a one liter drinking water sample was as low as 1 RT-PCR unit, which is equal to a 10-6 dilution of the virus sample. This approach showed at least a 5-fold-higher sensitivity than the current method with its three steps of adsorption-elution-concentration. The newly developed procedure was used to test different brands of bottled drinking water from China for putative contamination with noroviruses. A total of 144 samples were analyzed; all of the samples were negative for norovirus specific nucleic acids.

Journal Article

Regulation of Branched-Chain, and Sulfur-Containing Amino Acid Metabolism by Glutathione during Ultradian Metabolic Oscillation of Saccharomyces cerevisiae: Ho-Yong Sohn , Eun-Joo Kum , Gi-Seok Kwon , Ingnyol Jin , Hiroshi Kuriyama; J. Microbiol. 2005;43(4):375-380.; DOI: https://doi.org/2250 [pii]

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Abstract: Autonomous ultradian metabolic oscillation (T~=50 min) was detected in an aerobic chemostat culture of Saccharomyces cerevisiae. A pulse injection of GSH (a reduced form of glutathione) into the culture induced a perturbation in metabolic oscillation, with respiratory inhibition caused by H_2S burst production. As the production of H_2S in the culture was controlled by different amino acids, we attempted to characterize the effects of GSH on amino acid metabolism, particularly with regard to branched chain and sulfur-containing amino acids. During stable metabolic oscillation, concentrations of intracellular glutamate, aspartate, threonine, valine, leucine, isoleucine, and cysteine were observed to oscillate with the same periods of dissolved O_2 oscillation, although the oscillation amplitudes and maximal phases were shown to differ. The methionine concentration was stably maintained at 0.05 mM. When GSH (100 uM) was injected into the culture, cellular levels of branched chain amino acids increased dramatically with continuous H_2S production, whereas the cysteine and methionine concentrations were noticeably reduced. These results indicate that GSH-dependent perturbation occurs as the result of the promotion of branched chain amino acid synthesis and an attenuation of cysteine and methionine synthesis, both of which activate the generation of H_2S. In a low sulfate medium containing 2.5 mM sulfate, the GSH injections did not result in perturbations of dissolved O_2, NAD(P)H redox oscillations without burst H_2S production. This suggests that GSH-dependent perturbation is intimately linked with the metabolism of branched-chain amino acids and H_2S generation, rather than with direct GSH-GSSG redox control.

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

Isolation, Characterization, and Investigation of Surface and Hemolytic Activities of a Lipopeptide Biosurfactant Produced by Bacillus subtilis ATCC 6633: Gholamreza Dehghan-Noudeh , Mohammadreza Housaindokht , Bibi Sedigeh Fazly Bazzaz; J. Microbiol. 2005;43(3):272-276.; DOI: https://doi.org/2213 [pii]

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Abstract: Bacillus subtilis ATCC 6633 was grown in BHIB medium supplemented with Mn^2^+ for 96 h at 37^oC in a shaker incubator. After removing the microbial biomass, a lipopeptide biosurfactant was extracted from the supernatant. Its structure was established by chemical and spectroscopy methods. The structure was confirmed by physical properties, such as Hydrophile-Lipophile Balance (HLB), surface activity and erythrocyte hemolytic capacity. The critical micelle concentration (cmc) and erythrocyte hemolytic capacity of the biosurfactant were compared to those of surfactants such as SDS, BC (benzalkonium chloride), TTAB (tetradecyltrimethylammonium bromide) and HTAB (hexadecyltrimethylammonium bromide). The maximum hemolytic effect for all surfactants mentioned was observed at concentrations above cmc. The maximum hemolytic effect of synthetic surfactants was more than that of the biosurfactant produced by B. subtilis ATCC 6633. Therefore, biosurfactant would be considered a suitable surface-active agent due to low toxicity to the membrane.

Effects of Sulfate Concentration on the Anaerobic Dechlorination of Polychlorinated Biphenyls in Estuarine Sediments: Young-Cheol Cho , Kyoung-Hee Oh; J. Microbiol. 2005;43(2):166-171.; DOI: https://doi.org/2167 [pii]

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Abstract: In order to determine the effects of sulfate concentration on the anaerobic dechlorination of polychlorinated biphenyls, sediments spiked with Aroclor 1242 were made into slurries using media which had various sulfate concentrations ranging from 3 to 23 mM. The time course of dechlorination clearly demonstrated that dechlorination was inhibited at high concentration of sulfate due to less dechlorination of meta-substituted congeners. When the dechlorination patterns were analyzed by the calculation of Euclidean distance, the dechlorination pathway in the 3 mM sulfate samples was found to be different from that observed in the 13 mM samples, although the extent of dechlorination in these two samples was similar. It is possible that the dechlorination in the high sulfate concentration samples is inhibited by the suppression of growth of methanogen, which have been shown to be meta-dechlorinating microorganisms.

Biosynthesis of polyhydroxybutyrate and poly(3-hydroxybutyrate-co-3-hydroxyvalerate) by bacillus thuringiensis R-510: Park, Sang Kyu , Lee, Kang Tae , Kim, Young Baek , Rhee, Young Ha; J. Microbiol. 1997;35(2):127-133.

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Abstract: Biosynthesis of polyhydroxybutyrate and copolymer consisting of 3-hydroxybutyrate and 3-hydroxyvalerate [poly(3HB-co-3HV)] by Bacillus thuringiensis R-510 grown with glucose or with mixtures of glucose and propionate was investigated. n-Alkanoic acids other than propionate were not precursors of 3HV units. The fraction of 3HV unit in the copolymer increased from 0 to 84 mol% of 3HV. Polymer yield decreased as the fraction of propionate was increased but the molecular weight distribution was not affected by the composition of carbon substrate. The minimum melting temperature (around 65℃) of poly (3HB-co-3HV) copolymers was observed for the polymer bearing approximately 35 mol% of 3HV. Polyhydroxyalkanoates production by this organism was not dependent on nutritional limitation, but remarkably influenced by dissolved oxygen concentration in the culture medium. Low level of dissolved oxygen concentration prevented spore formation in the cells and stimulated the synthesis of polyhydroxyalkanoate. The composition of poly (3HB-co-3HV) produced by B. thuringiensis R-510 lyhydroxyalkanoate. The composition of poly(3HB-co-3HV) produced by B. thuringiensis R-510 varied according to the growth time. However, there was no evidence that polymers isolated from cells were mixtures of immiscible polymers.

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