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Involvement of Alternative Oxidase in the Regulation of Sensitivity of Sclerotinia sclerotiorum to the Fungicides Azoxystrobin and Procymidone
Ting Xu , Ya-Ting Wang , Wu-Sheng Liang , Fei Yao , Yong-Hong Li , Dian-Rong Li , Hao Wang , Zheng-Yi Wang
J. Microbiol. 2013;51(3):352-358.   Published online April 26, 2013
DOI: https://doi.org/10.1007/s12275-013-2534-x
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AbstractAbstract
Sclerotinia sclerotiorum is a filamentous fungal pathogen that can infect many economically important crops and vegetables. Alternative oxidase is the terminal oxidase of the alternative respiratory pathway in fungal mitochondria. The function of alternative oxidase was investigated in the regulation of sensitivity of S. sclerotiorum to two commercial fungicides, azoxystrobin and procymidone which have different fungitoxic mechanisms. Two isolates of S. sclerotiorum were sensitive to both fungicides. Application of salicylhydroxamic acid, a specific inhibitor of alternative oxidase, significantly increased the values of effective concentration causing 50% mycelial growth inhibition (EC50) of azoxystrobin to both S. sclerotiorum isolates, whereas notably decreased the EC50 values of procymidone. In mycelial respiration assay azoxystrobin displayed immediate inhibitory effect on cytochrome pathway capacity, but had no immediate effect on alternative pathway capacity. In contrast, procymidone showed no immediate impact on capacities of both cytochrome and alternative pathways in the mycelia. However, alternative oxidase encoding gene (aox) transcript and protein levels, alternative respiration pathway capacity of the mycelia were obviously increased by pre-treatment for 24 h with both azoxystrobin and procymidone. These results indicate that alternative oxidase was involved in the regulation of sensitivity of S. sclerotiorum to the fungicides azoxystrobin and procymidone, and that both fungicides could affect aox gene expression and the alternative respiration pathway capacity development in mycelia of this fungal pathogen.
Evaluation of the Sensitivity and Specificity of Primer Pairs and the Efficiency of RNA Extraction Procedures to Improve Noroviral Detection from Oysters by Nested Reverse Transcription-Polymerase Chain Reaction
Cheonghoon Lee , Sooryun Cheong , Hee-Jung Lee , Miye Kwon , Ilnam Kang , Eun-Gyoung Oh , Hong-Sik Yu , Soon-Bum Shin , Sang-Jong Kim
J. Microbiol. 2010;48(5):586-593.   Published online November 3, 2010
DOI: https://doi.org/10.1007/s12275-010-0047-4
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  • 6 Scopus
AbstractAbstract
Noroviruses (NoV) are the key cause of acute epidemic gastroenteritis, and oysters harvested from NoVpolluted sea areas are considered as the significant vectors of viral transmission. To improve NoV detection from oyster using nested reverse transcription-polymerase chain reaction (RT-PCR), we evaluated the sensitivity and specificity of previously published primer pairs and the efficiency of different RNA extraction procedures. Among the primer pairs used for RT-PCR, the sensitivity of GIF1/GIR1-GIF2/GIR1 and GIIF1/GIIR1-GIIF2/GIIR1 was higher than that of other primer pairs used in nested RT-PCR for the detection of NoV genogroup I (NoV GI) and NoV GII from both NoV-positive stool suspension and NoVseeded oyster concentrates, respectively; the resulting products showed neither unspecific bands in the positive samples nor false-positive bands in the negative controls. The extraction of NoV RNA from oyster samples using a QIAamp? Viral RNA Mini kit with a QIAshredderTM Homogenizer pretreatment afforded more efficient recovery (mean recovery for NoV GI and GII, 6.4%) and the procedure was less time consuming (<30 min) than most other RNA extraction procedures. The results of RNA extraction procedure and primer pairs evaluated by nested RT-PCR assay in this study can be useful for monitoring NoV contamination in oysters, which is an indicator of possible public health risks.
Development of a Latex Agglutination Test for Norovirus Detection
Heetae Lee , YoungBin Park , Misoon Kim , Youngmee Jee , Doo-sung Cheon , Hae Sook Jeong , GwangPyo Ko
J. Microbiol. 2010;48(4):419-425.   Published online August 20, 2010
DOI: https://doi.org/10.1007/s12275-010-0071-4
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  • 9 Scopus
AbstractAbstract
Norovirus (NoV) is the leading cause of acute gastroenteritis worldwide. Currently, reverse transcription polymerase chain reaction (RT-PCR) is used commonly to detect NoVs in both clinical and environmental samples. However, RT-PCR requires expensive equipment and cannot be performed on site. In this study, a latex agglutination test (LAT) using antibody-labeled latex beads for detecting NoVs was developed. Two kinds of polyclonal antibodies, one generated from synthetic peptides and the other from E. coli-expressed NoV capsid proteins, were used to develop the LAT. Each of these polyclonal antibodies was immobilized on the surface of latex beads and tested for the ability to detect NoVs. Under optimized conditions, our LAT detected GII.4 NoV at concentrations as low as 3.3×105 RT-PCR units/ml in stool samples. The detection limit for the LAT was approximately 1.7×103 RT-PCR units. Forty-eight stool samples were tested for NoVs using this LAT. In comparison with an RT-PCR assay, the sensitivity and specificity of the LAT were 35% and 100%, respectively. With further optimization, this LAT used with appropriate antibodies could be applied for convenient detection of NoVs in clinical diagnosis and food monitoring.
Alteration of chromosomal structure within β-Tubulin and flagellar calmodulin genes during differentiation of Naegleria gruberi Amebae into Flagellates
Bok, Jin Woong , Lee, Joo Hun
J. Microbiol. 1995;33(3):222-227.
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AbstractAbstract
We have examined DNase I sensitivity of β-tubulin and flagellar calmodulin genes which are transiently and coordinately activated differentiation of Naegleria gruberi amebae into flagellates. The DNase I sensitivity of β-tubulin and flagellar calmodulin genes changed in parallel with the changes in transcriptional activity of the respective genes during differentiation. The two genes were resistant to DNase I inamebae stage when transcription of the two genes was inactive. Forty minutes after initiation of differentiation, when the two genes were most actively being transcribed, the two genes showed the highest sensitsivity to DNase I. One hundred and twenty minutes after initiation, the differentiation was completed and transcriptional activity of the two genes decreased to a low level. At this stage, the two genes were resistant to DNase I treatment like the ones at the ameba stage. This change in the DNase I sensitivity of the two genes was not observed when transcription of the two genes was blocked by adding cycloheximide at the beginning of differentiation.
Reorganization of chromatin conformation from an active to an inactive state after cessation of transcription
Lee , Myeong Sok
J. Microbiol. 1996;34(1):54-60.
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AbstractAbstract
Taking advantage of the heat inducible HSP82 gene in yeast, chromatin structure after transcription cessation was investigated. Alteration of chromating conformation within the HSP82 gene transcription unit into an active state has been shown to correlate with its transcriptional induction. It was thus of interest to examine whether the active chromatin state within the HSP82 mRNA analysis, the gene ceased its transcription within a few hours of cultivation at a normal condition after heat induction. In this condition, an active chromatin conformation in the HSP82 gene body was changed into an inactive state which was revealed by DNase I resistance and by typical nucleosomal cutting periodicity in the corresponding chromatin. These results thus ruled out the possibility of a long-term maintenance of the DNase I sensitive chromatin after transcription cessation. DNA replication may be a critical event for the chromatin reprogramming.
A conditional lethal mutation of a nucleoporin gene, NUP49 in saccharomyces cerevisiae
Lee, Youn Soo , Song, Young Ja , Hwang, Mi Kyung , Lee, Woo Bok , Kim, Jin Mi
J. Microbiol. 1997;35(3):234-238.
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AbstractAbstract
Conditional lethal mutation nup49-1 of a nuclear pore complex component gene was constructed in Saccharomyces cerevisiae. This mutation deleted one third of the essential NUP49 gene at the carboxy-terminal, but retained 13 repeats of the highly conserved GLFG domain. The nup49-1 mutant strain was viable with a slow-growth phenotype, indicating that the C-terminal is dispensable at normal growth temperature. This strain exhibited both temperature-sensitivity at 37℃ and cold-sensitivity at 16℃. Temperature shift experiments revealed that the arrest phenotype at 37℃ was random in the cell division cycle. The nup49-1 mutation was tested to be recessive and is expected to be useful for the functional analysis of nuclear pore complex proteins as well as for studies of nuclear transport systems.

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