研究者詳細 - 佐々木　崇

論文 - 佐々木　崇

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Origin and molecular evolution of the determinant of methicillin resistance in staphylococci (Antimicrobial Agents and Chemotherapy (2010) 54, 10 (4352?4359))

Sae Tsubakishita, Kyoko Kuwahara-Arai, Takashi Sasaki, Keiichi Hiramatsu

Antimicrobial Agents and Chemotherapy 55 ( 2 ) 946 2011年02月 [査読有り]

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健康な犬におけるメチシリン耐性Staphylococcus pseudintermedius（MRSP）の疫学調査

須藤哲長, 寺井陽子, 三枝早苗, 椿下早絵, 佐々木崇, 平松啓一

獣医臨床皮膚科 ( 日本獣医皮膚科学会 ) 17 ( 2 ) 79 - 83 2011年 [査読有り]

小動物臨床の高度化にともない広域抗菌薬が広く用いられるになり，メチシリン耐性 Staphylococcus pseudintermedius（MRSP）が犬に世界的に蔓延している。小動物皮膚科領域においてもMRSPは臨床上深刻な問題となっている。本研究では，日本国内の健康な犬104頭のMRSPおよびメチシリン耐性S. aureus (MRSA)の保菌調査を行い，二次診療施設（大学付属施設）を受診した病犬102頭の保菌率と比較した。健康犬および二次診療受診犬の鼻腔内MRSP保菌率はそれぞれ4.8%，21.6%（p<0.001）であり，二次診療受診犬で有意に高かった。二次診療受診犬は当該施設受診までに相当の抗菌薬選択圧を受けていることが示唆された。健康犬のMRSP保菌率は海外の報告とほぼ同等な結果であった。一方，MRSA保菌率は，健康犬で0％，二次診療受診犬で1.96％と両者に有意差はなかった。どのような抗菌薬選択圧下でもMRSA保菌率が有意に上昇しないことから，犬はMRSAのレゼルボアにはならないことが示唆された。

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Origin and Molecular Evolution of the Determinant of Methicillin Resistance in Staphylococci

Sae Tsubakishita, Kyoko Kuwahara-Arai, Takashi Sasaki, Keiichi Hiramatsu

ANTIMICROBIAL AGENTS AND CHEMOTHERAPY ( AMER SOC MICROBIOLOGY ) 54 ( 10 ) 4352 - 4359 2010年10月 [査読有り]

Methicillin-resistant Staphylococcus aureus (MRSA) is one of the most important multidrug-resistant pathogens around the world. MRSA is generated when methicillin-susceptible S. aureus (MSSA) exogenously acquires a methicillin resistance gene, mecA, carried by a mobile genetic element, staphylococcal cassette chromosome mec (SCCmec), which is speculated to be transmissible across staphylococcal species. However, the origin/reservoir of the mecA gene has remained unclear. Finding the origin/reservoir of the mecA gene is important for understanding the evolution of MRSA. Moreover, it may contribute to more effective control measures for MRSA. Here we report on one of the animal-related Staphylococcus species, S. fleurettii, as the highly probable origin of the mecA gene. The mecA gene of S. fleurettii was found on the chromosome linked with the essential genes for the growth of staphylococci and was not associated with SCCmec. The mecA locus of the S. fleurettii chromosome has a sequence practically identical to that of the mecA-containing region (similar to 12 kbp long) of SCCmec. Furthermore, by analyzing the corresponding gene loci (over 20 kbp in size) of S. sciuri and S. vitulinus, which evolved from a common ancestor with that of S. fleurettii, the speciation-related mecA gene homologues were identified, indicating that mecA of S. fleurettii descended from its ancestor and was not recently acquired. It is speculated that SCCmec came into form by adopting the S. fleurettii mecA gene and its surrounding chromosomal region. Our finding suggests that SCCmec was generated in Staphylococcus cells living in animals by acquiring the intrinsic mecA region of S. fleurettii, which is a commensal bacterium of animals.

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Multiplex-PCR Method for Species Identification of Coagulase-Positive Staphylococci

Takashi Sasaki, Sae Tsubakishita, Yoshikazu Tanaka, Arihito Sakusabe, Masayuki Ohtsuka, Shintaro Hirotaki, Tetsuji Kawakami, Tsuneo Fukata, Keiichi Hiramatsu

JOURNAL OF CLINICAL MICROBIOLOGY ( AMER SOC MICROBIOLOGY ) 48 ( 3 ) 765 - 769 2010年03月 [査読有り]

In veterinary medicine, coagulase-positive staphylococci (CoPS) other than Staphylococcus aureus have frequently been misidentified as being S. aureus strains, as they have several phenotypic traits in common. There has been no reliable method to distinguish among CoPS species in veterinary clinical laboratories. In the present study, we sequenced the thermonuclease (nuc) genes of staphylococcal species and devised a multiplex-PCR (M-PCR) method for species identification of CoPS by targeting the nuc gene locus. To evaluate sensitivity and specificity, we used this M-PCR method on 374 staphylococcal strains that had been previously identified to the species level by an hsp60 sequencing approach. We could successfully distinguish between S. aureus, S. hyicus, S. schleiferi, S. intermedius, S. pseudintermedius, and S. delphini groups A and B. The present method was both sensitive (99.8%) and specific (100%). Our M-PCR assay will allow the routine species identification of CoPS isolates from various animal species for clinical veterinary diagnosis.

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臨床検査機関でメチシリン耐性黄色ブドウ球菌（MRSA）と同定された犬由来ブドウ球菌株の分類学的再検討

笠井智子, 三枝早苗, 佐々木崇

獣医臨床皮膚科 ( 日本獣医皮膚科学会 ) 16 ( 3 ) 119 - 124 2010年 [査読有り]

人医領域で院内感染菌として重要視されているメチシリン耐性 Staphylococcus aureus（MRSA）が動物から分離される例が近年増加している。一方，犬の常在性ブドウ球菌は S. pseudintermediusであり，メチシリン耐性S. pseudintermedius（MRSP）が問題となっている。 S. pseudintermediusの同定には遺伝子検査が必要なため，臨床検査機関ではその同定に対応できていない。そこで本研究では膿皮症罹患犬のうち過去に臨床検査機関でMRSAの感染が同定された10症例について原因菌を再検討することとした。各症例よりスワブで検体採取し，臨床検査機関2社（A社，B社）への提出と同時に，我々はブドウ球菌を分離しコアグラーゼ陽性ブドウ球菌菌種同定のためのmultiplex-PCR（M-PCR）法を用いた菌種同定を行い，結果を比較検討した。M-PCRの結果，10症例はいずれも S. aureusではなく，8症例はS. pseudintermedius，2症例はS. schleiferiであり，遺伝子検査によって9症例がメチシリン耐性と判定された。一方臨床検査機関2社の検査結果は，A社が9症例をMRSAと同定し，B社は7症例で属レベルの同定（Staphylococcus sp.）にとどまり，全症例でメチシリン耐性判定未実施であった。ブドウ球菌感染症において必須の検査項目であるメチシリン耐性判定の基準は菌種によって異なるため，菌種同定の不備が治療方針に影響を与えることが示唆された。

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Emergence of Quinolone-Resistant Bordetella pertussis in Japan

Masayuki Ohtsuka, Ken Kikuchi, Kenichiro Shimizu, Namiko Takahashi, Yuka Ono, Takashi Sasaki, Keiichi Hiramatsu

ANTIMICROBIAL AGENTS AND CHEMOTHERAPY ( AMER SOC MICROBIOLOGY ) 53 ( 7 ) 3147 - 3149 2009年07月 [査読有り]

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Genetic Diversity of Staphylocoagulase Genes (coa): Insight into the Evolution of Variable Chromosomal Virulence Factors in Staphylococcus aureus

Shinya Watanabe, Teruyo Ito, Takashi Sasaki, Shanshuang Li, Ikuo Uchiyama, Kozue Kishii, Ken Kikuchi, Robert Leo Skov, Keiichi Hiramatsu

PLOS ONE ( PUBLIC LIBRARY SCIENCE ) 4 ( 5 ) e5714 2009年05月 [査読有り]

Background: The production of staphylocoagulase (SC) causing the plasma coagulation is one of the important characteristics of Staphylococcus aureus. Although SCs have been classified into 10 serotypes based on the differences in the antigenicity, genetic bases for their diversities and relatedness to chromosome types are poorly understood. Methodology/Principal Findings: We compared the nucleotide sequences of 105 SC genes (coa), 59 of which were determined in this study. D1 regions, which contain prothrombin-activating and -binding domains and are presumed to be the binding site of each type-specific antiserum, were classified into twelve clusters having more than 90% nucleotide identities, resulting to create two novel SC types, XI and XII, in addition to extant 10 types. Nine of the twelve SC types were further subdivided into subtypes based on the differences of the D2 or the central regions. The phylogenetical relations of the D1 regions did not correlate exactly with either one of agr types and multilocus sequence types (STs). In addition, genetic analysis showed that recombination events have occurred in and around coa. So far tested, STs of 126 S. aureus strains correspond to the combination of SC type and agr type except for the cases of CC1 and CC8, which contained two and three different SC types, respectively. Conclusion: The data suggested that the evolution of coa was not monophyletic in the species. Chromosomal recombination had occurred at coa and agr loci, resulting in the carriage of the combinations of allotypically different important virulence determinants in staphylococcal chromosome.

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Smqnr, a new chromosome-carried quinolone resistance gene in Stenotrophomonas maltophilia

Kenichiro Shimizu, Ken Kikuchi, Takashi Sasaki, Namiko Takahashi, Masayuki Ohtsuka, Yuka Ono, Keiichi Hiramatsu

ANTIMICROBIAL AGENTS AND CHEMOTHERAPY ( AMER SOC MICROBIOLOGY ) 52 ( 10 ) 3823 - 3825 2008年10月 [査読有り]

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多剤耐性緑膿菌に対するaztreonamとarbekacinによる併用療法

菊池賢, 星最智, 佐々木崇, 堀賢, 平松啓一

感染症学雑誌 ( (一社)日本感染症学会 ) 82 ( 5 ) 567 - 567 2008年09月

Is Histoplasma capsulatum a native inhabitant of Japan ?

Ken Kikuchi, Takashi Sugita, Koichi Makimura, Kensaku Urata, Takashi Someya, Takashi Sasaki, Katsuhiko Kamei, Masakazu Niimi, Keiichi Hiramatsu, Yoshimasa Uehara

MICROBIOLOGY AND IMMUNOLOGY ( BLACKWELL PUBLISHING ) 52 ( 9 ) 455 - 459 2008年09月 [査読有り]

Histoplasmosis is an infectious disease caused by inhaling spores of the fungal pathogen H. capsulatum and in Japan is considered an imported mycosis. However, some patients in Japan with histoplasmosis have no history of traveling overseas nor of risk of occupational exposure to Histoplasma. To investigate the possibility of native distribution of Histoplasma in Japan, 187 bat guano samples from 67 bat-inhabited caves in 17 prefectures were collected. These were examined for H. capsulatum by culture and Histoplasma-specific PCR in three independent laboratories. No H. capsulatum was detected by either method, therefore H. capsulatum is unlikely to be present in bat guano in Japanese caves.

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Postantibiotic effects and bactericidal activities of Levofloxacin and gatifloxacin at concentrations simulating those of topical ophthalmic administration against fluoroquinolone-resistant, and fluoroquinolone-sensitive methicillin-resistant staphylococcus aureus strains

Saichi Hoshi, Ken Kikuchi, Takashi Sasaki, Chie Sotozono, Shigeru Kinoshita, Keiichi Hiramatsu

ANTIMICROBIAL AGENTS AND CHEMOTHERAPY ( AMER SOC MICROBIOLOGY ) 52 ( 8 ) 2970 - 2973 2008年08月 [査読有り]

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多剤耐性緑膿菌に対するaztreonamとarbekacinによる併用療法

菊池賢, 星最智, 佐々木崇, 堀賢, 平松啓一

感染症学雑誌 ( (一社)日本感染症学会 ) 82 ( 臨増 ) 248 - 248 2008年03月

Reclassification of phenotypically identified Staphylococcus intermedius strains

Takashi Sasaki, Ken Kikuchi, Yoshikazu Tanaka, Namiko Takahashi, Shinichi Kamata, Keiichi Hiramatsu

JOURNAL OF CLINICAL MICROBIOLOGY ( AMER SOC MICROBIOLOGY ) 45 ( 9 ) 2770 - 2778 2007年09月 [査読有り]

To reclassify phenotypically identified Staphylococcus intermedius strains, which might include true S. intermedius strains and novel species such as Staphylococcus pseudintermedius and Staphylococcus delphini, we analyzed molecular phylogenies and phenotypic characteristics of 117 S. intermedius group (SIG) strains tentatively identified as being S. intermedius by the Rapid ID32 Staph assay. From phylogenetic analyses of sodA and hsp60 sequences, the SIG strains were divided into three clusters, which belonged to S. pseudintermedius LMG 22219(T), S. intermedius ATCC 29663 T, and S. delphini LMG 22190(T). All the SIG strains from dogs, cats, and humans were identified as being S. pseudintermedius. The wild pigeon strains, except one, were identified as being S. intermedius, and strains from all domestic pigeons, one wild pigeon, horses, and a mink were identified as being S. delphini. In addition, a phylogenetic analysis of nuc genes revealed that S, delphini strains were divided into two clusters: one was the cluster (S. delphini group A) that belonged to S. delphini LMG 22190T, and the other was the cluster (S. delphini group B) that was more related to S. pseudintermedius LMG 22219T than S. delphini LMG 22190T. The DNA-DNA hybridization results showed that S. delphini group B strains were distinguished from S. delphini group A, S. intermedius, and S. pseudintermedius strains. S. intermedius is distinguishable from S. pseudintermedius or S. delphini by positive arginine dihydrolase and acid production from beta-gentiobiose and D-mannitol. However, phenotypical characteristics to differentiate S. delphini group A, S. delphini group B, and S. pseudintermedius were not found. In conclusion, SIG strains were reclassified into four clusters with three established and one probably novel species.

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Methicillin-resistant Staphylococcus pseudintermedius in a veterinary teaching hospital

Takashi Sasaki, Ken Kikuchi, Yoshikazu Tanaka, Namiko Takahashi, Shinichi Kamata, Keiichi Hiramatsu

JOURNAL OF CLINICAL MICROBIOLOGY ( AMER SOC MICROBIOLOGY ) 45 ( 4 ) 1118 - 1125 2007年04月 [査読有り]

We surveyed methicillin-resistant coagulase-positive staphylococcus (MRCPS) strains from 57 (26 inpatient and 31 outpatient) dogs and 20 veterinary staff in a veterinary teaching hospital. From the staff, three MRCPS strains were isolated, and two were methicillin-resistant Staphylococcus aureus (MRSA). In contrast, 18 MRCPS strains were detected in both inpatient (12 of 26 [46.2%]) and outpatient (6 of 31 [19.4%]) dogs. Among them, only one strain was MRSA. Using direct sequencing of sodA and hsp60 genes, the 18 MRCPS strains other than MRSA from a staff and 17 dogs, were finally identified as Staphylococcus pseudintermedius, a novel species of Staphylococcus from a cat. All of the methicillin-resistant S. pseudintermedius (MRSP) strains were multidrug resistant to erythromycin, clindamycin, trimethoprim-sulfamethoxazole, and levofloxacin. Most of the MRSP strains showed high-level resistance to oxacillin (>= 128 mu g/ml, 15 of 18 [83.3%]), and 10 of 15 (66.7%) high-level oxacillin-resistant MRSP strains carried type III SCCmec. DNA fingerprinting of MRSP strains by pulsed-field gel electrophoresis yielded eight clusters: clone A with four subtypes, clone B with four subtypes, clone C with three subtypes, and five other different single clones. MRSP strains from the staff and some inpatient and outpatient dogs shared three major clones (clones A, B, and Q, but the strains of the other five different clusters were distributed independently among inpatient or outpatient dogs. This genetic diversity suggested that the MRSP strains were not only acquired in this veterinary teaching hospital but also acquired in primary veterinary clinics in the community. To our knowledge, this is the first report of MRSP in dogs and humans in a veterinary institution.

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