Shuttle Vectors Derived from pN315 for Study of Essential Genes in <i>Staphylococcus aureus</i>

Matsuo, Miki,Kurokawa, Kenji,Lee, Bok-Luel,Sekimizu, Kazuhisa Pharmaceutical Society of Japan 2010 BIOLOGICAL & PHARMACEUTICAL BULLETIN Vol.33 No.2

<P>Using the <I>par</I> to <I>rep</I> region of the 24653 bp plasmid pN315, which is present in <I>Staphylococcus aureus</I> strain N315, we constructed three vectors that can be shuttled between <I>Escherichia coli</I> and <I>S. aureus</I> and maintained stably in <I>S. aureus</I>. Due to plasmid incompatibility, the resident plasmid in <I>S. aureus</I> cells can be replaced <I>via</I> transformation with an entering plasmid, which carries a different drug resistance gene. To evaluate the applicability of this plasmid-based approach for identifying genes essential for <I>S. aureus</I> cell growth, the chromosomal <I>mraY</I> gene, which is involved in peptidoglycan biosynthesis, was deleted in cells harboring a resident plasmid with an intact <I>mraY</I> gene. The resultant disruptant was then transformed with an empty vector. Cells with a chromosomal <I>mraY</I> deletion but lacking the plasmid supplying <I>mraY</I> could not be recovered, suggesting that <I>mraY</I> is indispensable for staphylococcal cell growth or viability. In contrast, other two genes were shown to be dispensable by this system. Thus, the pN315-based plasmids appear to be useful for studying genes essential for <I>S. aureus</I> cell growth.</P>

Development of Technology for Determining Rice Amylose Content using Near-infrared Spectroscopy

( Miki Matsuo ),( Shuso Kawamura ),( Mizuki Kato ),( Edenio Olivares Diaz ),( Shigenobu Koseki ) 한국농업기계학회 2018 한국농업기계학회 학술발표논문집 Vol.23 No.1

Rice is staple food for people in the large part of the world, especially in Asian countries. The major chemical constituent contents of rice are moisture, protein and starch (amylose and amylopectin). Those chemical constituents associate with eating quality of rice. Near-infrared (NIR) spectroscopy is one of the non-destructive methods for determining grain chemical contents. Moisture and protein contents can be measured with high accuracy using NIR spectrometer at rice grain elevator in Japan. However, the accuracy to measure amylose content is not sufficient. Thus, there is a strong need for the highly accurate measurement of rice amylose content using non-destructive method. The overall objective of this study was to develop non-destructive techniques to determine rice amylose content for practical use at rice grain elevator. Amylose content measurement was performed using an auto-analyzer for reference (chemical) analysis. Spectra data of rice were obtained using an NIR spectrometer with a wavelength range of 850 to 1048 nm. Calibration model to determine amylose content was developed using non-waxy Japonica-type rice samples (calibration set, n=974, including 14 cultivars) grown in Japan from 2008 to 2016. Partial least squares (PLS) regression analysis was used to develop calibration model. The accuracy of the model was validated using other rice samples (validation set, n=95, including 10 cultivars) grown in 2017. There are two cultivar groups of non-waxy Japonica-type rice produced in Japan. One is ordinary amylose content rice cultivar and the other is low amylose content rice cultivar. The accuracy of determination of rice amylose content was improved by increasing production years of calibration sample set. When one calibration model to determine amylose content was developed using all cultivars together, validation statistics were shown: coefficient of determination (r²) was 0.72, bias was -0.04%, standard error of prediction (SEP) was 0.92%, and ratio of SEP to standard deviation of reference data (RPD) was 1.90. To improve the accuracy of determination of rice amylose content, two calibration models were developed using ordinary amylose content rice cultivars or low amylose content rice cultivars, respectively. As a result, the accuracy of two calibration models was better than that of one calibration model. Validation statistics of the two calibration models were shown: coefficient of determination (r²) was 0.93, bias was 0.01%, standard error of prediction (SEP) was 0.64%, and ratio SEP to standard deviation of reference data (RPD) was 2.76. The production year of validation set (2017) was different from those of calibration set (2008 to 2016). This is the same condition as practical use of this non-destructive method at rice grain elevator. The result obtained in this study indicated that the two calibration models enables non-destructive determination of rice amylose content at rice grain elevator.

Evaluation of Target Specificity of Antibacterial Agents Using Staphylococcus aureus ddlA Mutants and d-Cycloserine in a Silkworm Infection Model

Kurokawa, Kenji,Hamamoto, Hiroshi,Matsuo, Miki,Nishida, Satoshi,Yamane, Noriko,Lee, Bok Luel,Murakami, Kazuhisa,Maki, Hideki,Sekimizu, Kazuhisa American Society for Microbiology 2009 Antimicrobial Agents and Chemotherapy Vol.53 No.9

<B>ABSTRACT</B><P>The availability of a silkworm larva infection model to evaluate the therapeutic effectiveness of antibiotics was examined. The 50% effective doses (ED50) of d-cycloserine against the <I>Staphylococcus aureus ddlA</I> mutant-mediated killing of larvae were remarkably lower than those against the parental strain-mediated killing of larvae. Changes in MICs and ED50 of other antibiotics were negligible, suggesting that these alterations are d-cycloserine selective. Therefore, this model is useful for selecting desired compounds based on their therapeutic effectiveness during antibiotic development.</P>

Pleiotropic Roles of Polyglycerolphosphate Synthase of Lipoteichoic Acid in Growth of Staphylococcus aureus Cells

Oku, Yusuke,Kurokawa, Kenji,Matsuo, Miki,Yamada, Sakuo,Lee, Bok-Luel,Sekimizu, Kazuhisa American Society for Microbiology 2009 Journal of Bacteriology Vol.191 No.1

<B>ABSTRACT</B><P>Lipoteichoic acid (LTA) is one of two anionic polymers on the surface of the gram-positive bacterium <I>Staphylococcus aureus</I>. LTA is critical for the bacterium-host cell interaction and has recently been shown to be required for cell growth and division. To determine additional biological roles of LTA, we found it necessary to identify permissive conditions for the growth of an LTA-deficient mutant. We found that an LTA-deficient <I>S. aureus</I> Δ<I>ltaS</I> mutant could grow at 30°C but not at 37°C. Even at the permissive temperature, Δ<I>ltaS</I> mutant cells had aberrant cell division and separation, decreased autolysis, and reduced levels of peptidoglycan hydrolases. Upshift of Δ<I>ltaS</I> mutant cells to a nonpermissive temperature caused an inability to exclude Sytox green dye. A high-osmolarity growth medium remarkably rescued the colony-forming ability of the Δ<I>ltaS</I> mutant at 37°C, indicating that LTA synthesis is required for growth under low-osmolarity conditions. In addition, the Δ<I>ltaS</I> mutation was found to be synthetically lethal with the Δ<I>tagO</I> mutation, which disrupts the synthesis of the other anionic polymer, wall teichoic acid (WTA), at 30°C, suggesting that LTA and WTA compensate for one another in an essential function.</P>

Cigarette Smoking and Pancreatic Cancer Risk: A Revisit with an Assessment of the Nicotine Dependence Phenotype

Nakao, Makoto,Hosono, Satoyo,Ito, Hidemi,Oze, Isao,Watanabe, Miki,Mizuno, Nobumasa,Yatabe, Yasushi,Yamao, Kenji,Niimi, Akio,Tajima, Kazuo,Tanaka, Hideo,Matsuo, Keitaro Asian Pacific Journal of Cancer Prevention 2013 Asian Pacific journal of cancer prevention Vol.14 No.7

Background: Cigarette smoking is a well-established risk factor of pancreatic cancer (PC). Although an association between nicotine dependence phenotype, namely time to first cigarette (TTFC) after waking, and the risk of several smoking-related cancers has been reported, an association between TTFC and PC risk has not been reported. We assessed the impact of smoking behavior, particularly TTFC, on PC risk in a Japanese population. Materials and Methods: We conducted a case-control study using 341 PC and 1,705 non-cancer patients who visited Aichi Cancer Center in Nagoya, Japan. Exposure to risk factors, including smoking behavior, was assessed from the results of a self-administered questionnaire. The impact of smoking on PC risk was assessed with multivariate logistic regression analysis adjusted for potential confounders to estimate odds ratios (ORs) and 95% confidence intervals (CIs). Results: Cigarettes per day (CPD) and/or smoking duration were significantly associated with PC risk, consistent with previous studies. For TTFC and PC risk, we found only a suggestive association: compared with a TTFC of more than 60 minutes, ORs were 1.15 (95%CI, 0.65-2.04) for a TTFC of 30-60 minutes and 1.35 (95%CI, 0.85-2.15) for that of 0-30 minutes (p trend=0.139). After adjustment for CPD or smoking duration, no association was observed between TTFC and PC. Conclusions: In this study, we found no statistically significant association between TTFC and PC risk. Further studies concerning TTFC and PC risk are warranted.

Genomic Basis for Methicillin Resistance in Staphylococcus aureus

Keiichi Hiramatsu,Teruyo Ito,Sae Tsubakishita,Takashi Sasaki,Fumihiko Takeuchi,Yuh Morimoto,Yuki Katayama,Miki Matsuo,Kyoko Kuwahara-Arai,Tomomi Hishinuma,Tadashi Baba 대한감염학회 2013 Infection and Chemotherapy Vol.45 No.2

Since the discovery of the first strain in 1961 in England, MRSA, the most notorious multidrug-resistant hospital pathogen, has spread all over the world. MRSA repeatedly turned down the challenges by number of chemotherapeutics, the fruits of modern organic chemistry. Now, we are in short of effective therapeutic agents against MRSA prevailing among immuno-compromised patients in the hospital. On top of this, we recently became aware of the rise of diverse clones of MRSA, some of which have increased pathogenic potential compared to the classical hospital-associated MRSA, and the others from veterinary sources. They increased rapidly in the community, and started menacing otherwise healthy individuals by causing unexpected acute infection. This review is intended to provide a whole picture of MRSA based on its genetic makeup as a versatile pathogen and our tenacious colonizer.