Tsutomu Kakuda

Publications (23)48.58 Total impact

• Article: Isolation of Rhodococcus equi from wild boars (Sus scrofa) in Japan.

  Mizuki Sakai, Ryoko Ohno, Chihiro Higuchi, Masato Sudo, Kazuo Suzuki, Hiroshi Sato, Ken Maeda, Yukako Sasaki, Tsutomu Kakuda, Shinji Takai
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  ABSTRACT: Rhodococcus equi was isolated from the submaxillary lymph nodes of wild boars (Sus scrofa) in Wakayama and Hyogo, Japan, with a high prevalence. Rhodococcus equi isolation rates between locations, sexes, or body weights were not different, except in the prevalence of vapB-positive R. equi between locations.
  Journal of wildlife diseases 07/2012; 48(3):815-7. · 1.08 Impact Factor
• Article: Characterization of two putative mechanosensitive channel proteins of Campylobacter jejuni involved in protection against osmotic downshock.

  Tsutomu Kakuda, Yuuki Koide, Aya Sakamoto, Shinji Takai
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  ABSTRACT: Acute hypotonic stress becomes a threat to the survival of bacteria in the environment. Mechanosensitive channels play an essential role in the maintenance of bacterial cell integrity during hypoosmotic shock. A database search suggested that Campylobacter jejuni, a major worldwide cause of bacterial gastroenteritis in humans, possesses two putative mechanosensitive channels, designated Cjj0263 and Cjj1025, in C. jejuni strain 81-176. Osmotic downshock experiments demonstrated that a mutant lacking Cjj0263 showed a severe defect in survival of hypoosmotic shock, while a mutant lacking Cjj1025 exhibited the same survival capacity as the wild type. We further examined the colonization ability of each mutant using the one-day old chick model. Cjj0263 or Cjj1025 mutants were able to colonize chick ceca at the same level as the wild type, but a Cjj0263 Cjj1025 double mutant revealed significantly reduced ability to colonize chick ceca. To examine whether C. jejuni that have grown in the digestive tract of chicks are protected against acute hypotonic stress, bacteria in ceca were directly exposed to water. The wild type was able to survive acute osmotic downshift, but the Cjj0263 mutant suffered a substantial loss of viability when subjected to a rapid osmotic downshock. Immunoblot analysis suggested that both Cjj0263 and Cjj1025 were glycosylated via the N-linked protein glycosylation pathway, but glycan modification of these proteins was unlikely to have a major effect on their function and stability. Our data suggest that Cjj0263, a mechanosensitive channel, has a pivotal role in protection against hypoosmotic stress experienced during environmental transmission.
  Veterinary Microbiology 05/2012; 160(1-2):53-60. · 3.33 Impact Factor
• Article: Participation of CheR and CheB in the chemosensory response of Campylobacter jejuni.

  Doungjit Kanungpean, Tsutomu Kakuda, Shinji Takai
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  ABSTRACT: Campylobacter jejuni is a leading cause of bacterial gastroenteritis in humans and a commensal bacterium of the intestinal tracts of animals, especially poultry. Chemotaxis is an important determinant for chicken colonization of C. jejuni. Adaptation has a crucial role in the gradient-sensing mechanism that underlies chemotaxis. The genome sequence of C. jejuni reveals the presence of genes encoding putative adaptation proteins, CheB and CheR. In-frame deletions of cheB, cheR and cheBR were constructed and the chemosensory behaviour of the resultant mutants was examined on swarm plates. CheB and CheR proteins significantly influence chemotaxis but are not essential for this behaviour to occur. Increased mobility of two methyl-accepting chemotaxis proteins (MCPs), DocC and Tlp1, during SDS-PAGE was detected in the mutants lacking functional CheB in the presence of CheR, presumably resulting from stable methylation of receptors. In vitro studies using tissue culture revealed that deletion of cheR resulted in hyperadherent and hyperinvasive phenotypes, while deletion of cheB resulted in nonadherent, noninvasive phenotypes. Furthermore, the ΔcheBR mutant showed significantly reduced ability to colonize chick caeca. Our data suggest that modification of chemoreceptors by the CheBR system is involved in regulation of chemotaxis in C. jejuni although CheB is apparently not controlled by phosphorylation.
  Microbiology 02/2011; 157(Pt 5):1279-89. · 3.06 Impact Factor
• Article: False positive responses of Campylobacter jejuni when using the chemical-in-plug chemotaxis assay.

  Doungjit Kanungpean, Tsutomu Kakuda, Shinji Takai
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  ABSTRACT: Campylobacter jejuni reportedly exhibits chemotactic behavior towards fucose, several amino acids and organic acids, mucin and bile. The chemotaxis of C. jejuni has mainly been studied using the chemical-in-plug chemotaxis assay. In this study, a nonchemotactic mutant (cheY mutant) and nonmotile mutant (flhA mutant) were constructed and used as negative controls in an assay. Apparent zones of accumulation around test plugs containing several amino acids and organic acids were observed with both of the mutants. Our results suggest that positive responses of C. jejuni in the chemical-in-plug assay are not always indicative of chemotaxis.
  Journal of Veterinary Medical Science 10/2010; 73(3):389-91. · 0.85 Impact Factor
• Article: Isolation and characterisation of Rhodococcus equi from submaxillary lymph nodes of wild boars (Sus scrofa).

  László Makrai, Ayumi Kobayashi, Misa Matsuoka, Yukako Sasaki, Tsutomu Kakuda, Béla Dénes, István Hajtós, István Révész, Katalin Jánosi, László Fodor, János Varga, Shinji Takai
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  ABSTRACT: Rhodococcus equi has been isolated from the submaxillary lymph nodes of domesticated pigs, but little is known about the presence of R. equi in wild boars. The aim of the study was the evaluation of the incidence of R. equi in wild boars and the characterisation of them. Of 482 submaxillary lymph nodes of wild boars shot in 39 settlements throughout Hungary, R. equi was isolated from 60 specimens, and plasmid types of 82 isolates were examined. The isolates were tested for the presence of 15-17-kDa (VapA) and 20-kDa virulence-associated protein antigen (VapB) genes by polymerase chain reaction (PCR). Plasmid DNAs were isolated and analysed by digestion with restriction endonucleases to estimate size and compare their polymorphisms. None of the 82 isolates contained vapA gene but 21 isolates (25.6%) were positive for vapB gene showing 827bp product of the expected size in the PCR amplification. Sixty-one strains (74.4%) did not contain plasmid. The 21 isolates of intermediate virulence contained virulence plasmids that were identified as types 1 (1 isolate), 5 (16 isolates), 21 (1 isolate), and three new distinct plasmid variants (1-1-1 isolate), respectively. On the basis of restriction digestion patterns of plasmid DNAs, we tentatively designated the new variants as types 25-27, respectively. The prevalence of R. equi strains of intermediate virulence among the isolates originated from the submaxillary lymph nodes of wild boars (25.6%) is very similar to those of domestic pigs (26.8%) in Hungary, and plasmid type 5 is the predominating one in both groups. This is the first report of isolation of VapB-positive R. equi from wild boars in the world.
  Veterinary Microbiology 06/2008; 131(3-4):318-23. · 3.33 Impact Factor
• Article: Filamentous-haemagglutinin-like protein genes encoded on a plasmid of Moraxella bovis.

  Tsutomu Kakuda, Nopporn Sarataphan, Tetsuya Tanaka, Shinji Takai
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  ABSTRACT: The complete nucleotide sequence of a plasmid, pMBO-1, from Moraxella bovis strain Epp63 was determined. We identified 30 open reading frames (ORFs) encoded by the 44,215bp molecule. Two large ORFs, flpA and flpB, encoding proteins with similarity to Bordetella pertussis filamentous haemagglutinin (FHA), were identified on the same plasmid. The gene for a specific accessory protein (Fap), which may play a role in the secretion of Flp protein, was also identified. Reverse transcriptase PCR analysis of total RNA isolated from M. bovis Epp63 indicated that the flpA, flpB, and fap genes are all transcribed. Southern blot analysis indicated that the flp and fap genes are present in other clinical isolates of geographically diverse M. bovis.
  Veterinary Microbiology 12/2006; 118(1-2):141-7. · 3.33 Impact Factor
• Article: Rhodococcus equi in the soil environment of horses in Inner Mongolia, China.

  Shinji Takai, Ding Zhuang, Xiao-Wei Huo, Hiroo Madarame, Ming-Hua Gao, Zhong-Tian Tan, Shuang-Cheng Gao, Li-Jun Yan, Chun-Mei Guo, Xue-Feng Zhou, Fumiko Hatori, Yukako Sasaki, Tsutomu Kakuda, Shiro Tsubaki
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  ABSTRACT: Little is known about the distribution of Rhodococcus equi in the soil environment of native horses in China. One hundred and eight soil samples were collected from native-horse farms in the Hulun Beier grasslands of eastern Mongolia, the Xilin Goler grasslands of southern Mongolia, and Tongliao City in Inner Mongolia, China. The isolation rates of R. equi from soil samples from the Hulun Beier and Xilin Goler grasslands ranged from 25.9% to 30.0%. In contrast, isolation rates from soil samples from Tongliao City were as high as 82.3% and the mean number of R. equi in soil samples from Tongliao City was 10 times more than those of samples from the grasslands. The 488 isolates were examined using PCR for the presence of genes that encode virulence-associated 15-17 kDa antigen protein (VapA) and the 20 kDa antigen protein (VapB). All isolates were negative for virulence-associated proteins. Plasmid profiles of these avirulent isolates showed that cryptic plasmids of various sizes were present with an incidence of 13.3% to 21.5%. The results of the present study contrast with those of our recent study (J. Vet. Med. Sci. 67:611-613, 2005), in which we reported that R. equi was absent from Mongolian horses in Ulaanbaatar, Mongolia. It is suggested that the difference between the results of these two studies is due to the mobile pasturing system in Mongolia and nonmobile pasturing system in Inner Mongolia.
  Journal of Veterinary Medical Science 08/2006; 68(7):739-42. · 0.85 Impact Factor
• Source

  Available from: affrc.go.jp

  Article: Molecular typing of VapA-positive Rhodococcus equi isolates from Jeju native horses, Korea.

  Won-Geun Son, Du-Sik Lee, Noriko Yamatoda, Fumiko Hatori, Nanako Shimizu, Tsutomu Kakuda, Yukako Sasaki, Shiro Tsubaki, Shinji Takai
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  ABSTRACT: We recently demonstrated the presence of virulence-associated protein antigen (VapA)-positive Rhodococcus equi in Jeju Island, Korea. These bacteria contained one of two distinct plasmid types, a 90-kb type II plasmid, which has been found in isolates from the native Kiso horses of Japan, and a new variant, a 90-kb type V plasmid. However, the genotypic characters of the VapA-positive R. equi from Jeju native horses and their environments are poorly understood. Ninety-eight isolates from soil samples and 89 isolates from fecal samples were collected from five farms that breed or have bred Jeju native horses, and were tested for the presence of VapA by immunoblotting and PCR. Of the 98 soil isolates and 89 fecal isolates, seven and 13 were VapA-positive R. equi, respectively. In 2003, two Jeju foals died suddenly and were brought to the Faculty of Veterinary Medicine, Cheju National University, for postmortem examination. Pure cultures of R. equi were isolated from the lung lesions of both foals. Of the 16 clinical isolates, 14 were VapA-positive R. equi. Of the 34 VapA-positive clinical and environmental isolates, 16 contained the 90-kb type II plasmid and 18 contained a 90-kb type V plasmid. Pulsed-field gel electrophoresis (PFGE) patterns of the VapA-positive isolates from Jeju horses and Kiso horses, containing the 90-kb type II plasmid, were compared and formed two distinct groups. Furthermore, 18 virulent isolates containing the 90-kb type V plasmid formed two distinct PFGE groups (of 16 and two isolates). These results demonstrate that two virulence plasmid types are widespread in R. equi in Jeju native horses. However, there is little diversity in the PFGE patterns of virulent isolates, suggesting the clonal spread of virulent R. equi. The PFGE pattern of the virulent R. equi isolates from Jeju native horses in Korea is not identical to those of Kiso native horses in Japan.
  Journal of Veterinary Medical Science 04/2006; 68(3):249-53. · 0.85 Impact Factor
• Article: Variation in the N-terminal region of an M-like protein of Streptococcus equi and evaluation of its potential as a tool in epidemiologic studies.

  Toru Anzai, Yasushi Kuwamoto, Ryuichi Wada, Sigeo Sugita, Tsutomu Kakuda, Shinji Takai, Tohru Higuchi, John F Timoney
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  ABSTRACT: To develop a method for typing Streptococcus equi on the basis of the DNA sequence of the genes that produce an M-like protein and to compare isolates among the United States, Japan, and other countries. S equi strains CF32, Hidaka/95/2, and NCTC9682 as well as 82 other isolates from the United States, Japan, and other countries obtained during 1975 to 2001. DNA sequences of the structural genes ( SeM and SzPSe) that produce M-like proteins were determined for 3 representative strains to find a variable region. Variability in this region of SeM was then determined for the other isolates. Amino acid sequences were deduced and analyzed phylogenetically by use of the neighbor-joining method. Sequence diversity was detected in the N-terminal region of SeM but not in SzPSe of the 3 representative strains. Base substitutions in the variable region of SeM varied in a nonsynonymous manner, resulting in variation in the amino acid sequence. Eighty-five isolates were categorized as 32 types of SeM on the basis of differences in the deduced amino acid sequences. This study documented a region in the N-terminal portion of SeM that varies in a nonsynonymous manner. This information should be useful in molecular epidemiologic studies of S equi.
  American Journal of Veterinary Research 01/2006; 66(12):2167-71. · 1.27 Impact Factor
• Article: The absence of Rhodococcus equi in Mongolian horses.

  Shinji Takai, Sugar Sengee, Hiroo Madarame, Fumiko Hatori, Kaori Yasuoka, Erdene Ochirel, Yukako Sasaki, Tsutomu Kakuda, Shiro Tsubaki, Namkhai Bandi, Ruuragch Sodnomdarjaa
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  ABSTRACT: In native Mongolian horses, the incidence and distribution of Rhodococcus equi are poorly understood. One hundred and fourteen equine fecal samples and 71 soil samples were collected from the camp sites of 26 nomadic families located in three areas less than 100 km from Ulaanbaatar, Mongolia. Five fecal samples were also collected from foals of Przewalski's Horses introduced into the Hustai National Park, Mongolia. No R. equi was isolated from the Mongolian horses or the soil samples. However, three colonies of R. equi were isolated from two fecal samples collected from foals of Przewalski's Horses. These isolates were avirulent, with neither 15- to 17-kDa antigens (VapA) nor a 20-kDa antigen (VapB) genes being detected. We concluded that native Mongolian horses and their environment appear free from contamination with R. equi.
  Journal of Veterinary Medical Science 07/2005; 67(6):611-3. · 0.85 Impact Factor
• Source

  Available from: ncbi.nlm.nih.gov

  Article: Characterization of virulence plasmids and serotyping of rhodococcus equi isolates from submaxillary lymph nodes of pigs in Hungary.

  László Makrai, Saki Takayama, Béla Dénes, István Hajtós, Yukako Sasaki, Tsutomu Kakuda, Shiro Tsubaki, Andrea Major, László Fodor, János Varga, Shinji Takai
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  ABSTRACT: The plasmid types and serotypes of 164 Rhodococcus equi strains obtained from submaxillary lymph nodes of swine from different piggeries in 28 villages and towns located throughout the country were examined. The strains were tested by PCR for the presence of 15- to 17-kDa virulence-associated protein antigen (VapA) and 20-kDa virulence-associated protein antigen (VapB) genes. Plasmid DNAs were isolated and analyzed by digestion with restriction endonucleases to estimate size and compare their polymorphism characteristics. None of the 164 isolates contained the vapA gene, and 44 (26.8%) isolates were positive for the vapB gene, showing a product of the expected 827-bp size in the PCR amplification. The 44 isolates of intermediate virulence contained virulence plasmids that were identified as types 1 (3 isolates), 4 (1 isolate), 5 (36 isolates), 6 (1 isolate), and 7 (2 isolates) and as a new variant (1 isolate). On the basis of restriction digestion patterns of plasmid DNAs, we tentatively designated the variant as type 17. Use of the serotyping method of Prescott showed that 110 (67.1%) out of the 164 isolates were typeable and that serotype 2 predominated (83 isolates [50.6%]), followed by serotype 1 (26 strains [15.9%]). Only one isolate belonged to serotype 3. A total of 54 (32.9%) isolates were untypeable in Prescott's system. The prevalence of R. equi strains of intermediate virulence among the isolates that came from the submaxillary lymph nodes of swine in Hungary was lower than that seen with isolates obtained elsewhere.
  Journal of Clinical Microbiology 04/2005; 43(3):1246-50. · 4.15 Impact Factor
• Article: Molecular epidemiology of virulent Rhodococcus equi from foals in Brazil: virulence plasmids of 85-kb type I, 87-kb type I, and a new variant, 87-kb type III.

  Márcio Garcia Ribeiro, Izumi Seki, Kaori Yasuoka, Tsutomu Kakuda, Yukako Sasaki, Shiro Tsubaki, Shinji Takai
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  ABSTRACT: We investigated the prevalence of virulent Rhodococcus equi in clinical isolates from 41 foals (19 sporadic and seven endemic cases) in Brazil between 1991 and 2003. Of the 41 virulent isolates, six contained an 85-kb type I plasmid, 33 contained an 87-kb type I plasmid, both of which have been found in isolates from the Americas, and the remaining two contained a new variant, which did not display the EcoRI, EcoT22I and BamHI digestion patterns of the 11 representative plasmids already reported (85-kb types I-IV; 87-kb types I and II; 90-kb types I-V). We tentatively designated the new variant as the '87-kb type III' plasmid, because its BamHI digestion pattern is similar to that of the 87-kb type I plasmid. This is the first report of the molecular epidemiology surveillance of virulent R. equi in clinical isolates from Brazilian foals.
  Comparative Immunology Microbiology and Infectious Diseases 02/2005; 28(1):53-61. · 2.34 Impact Factor
• Article: Isolation of Rhodococcus equi from the feces of indigenous animals and soil from the Lower Zambezi National Park and Lochinvar National Park, Zambia.

  Shinji Takai, Michelo Syakalima, Jun Yasuda, Yukako Sasaki, Hisako Tsutsumi, Emiko Miyagawa, Kaya Wada, Tsutomu Kakuda, Shiro Tsubaki, Chihiro Sugimoto
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  ABSTRACT: Rhodococcus equi is an important pathogen in foals; however, its incidence in African indigenous animals is poorly understood. Fecal samples (92 from nine indigenous species) and 43 soil samples were collected from two Zambian National Parks. The presence of R. equi was investigated and 533 isolates were tested for the presence of 15- to 17-kDa antigens (VapA) and a 20-kDa antigen (VapB) by immunoblotting and PCR. R. equi was isolated (10(2)-10(4) colony forming units/g) from 75% of fecal and 74% of soil samples. Neither antigen was detected; however, about 20% of the isolates contained cryptic plasmids of various sizes. There was no evidence of virulent R. equi, but the avirulent form was widespread in the animals and the soil.
  Journal of Veterinary Medical Science 07/2004; 66(6):743-6. · 0.85 Impact Factor
• Article: Molecular epidemiology of Rhodococcus equi of intermediate virulence isolated from patients with and without acquired immune deficiency syndrome in Chiang Mai, Thailand.

  Shinji Takai, Prasit Tharavichitkul, Piyawan Takarn, Banyoung Khantawa, Mami Tamura, Azusa Tsukamoto, Saki Takayama, Noriko Yamatoda, Ayumi Kimura, Yukako Sasaki, Tsutomu Kakuda, Shiro Tsubaki, Niwat Maneekarn, Thira Sirisanthana, Teruo Kirikae
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  ABSTRACT: We investigated the prevalence of virulent Rhodococcus equi in clinical isolates from 69 sporadic cases (60 men, 8 women, and 1 patient of unknown sex) in Chiang Mai, Thailand, between 1993 and 2001. Fifty were human immunodeficiency virus (HIV) positive, 3 were HIV negative, and HIV status was unknown for 16. Fifty-two (75%) of 69 isolates were strains of intermediate virulence that contained the virulence-associated 20-kDa antigen, and 17 isolates (25%) were avirulent. No virulent strains with the virulence-associated 15-17-kDa antigens were identified. R. equi was isolated from HIV-positive patients' houses and those of their neighbors: avirulent strains were widespread, but only 1 strain of intermediate virulence was isolated. R. equi strains of intermediate virulence were isolated from 4 (0.8%) of 500 submaxillary lymph nodes from apparently healthy pigs in Chiang Mai. The routes of R. equi acquisition should be investigated from the viewpoint of zoonosis and public health.
  The Journal of Infectious Diseases 01/2004; 188(11):1717-23. · 6.41 Impact Factor
• Article: Rhodococcus equi virulence plasmids recovered from horses and their environment in Jeju, Korea: 90-kb type II and a new variant, 90-kb type V.

  Shinji Takai, Won-Geun Son, Du-Sik Lee, Hiroo Madarame, Izumi Seki, Noriko Yamatoda, Ayumi Kimura, Tsutomu Kakuda, Yukako Sasaki, Shiro Tsubaki, Yoon-Kyu Lim
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  ABSTRACT: Rhodococcus equi was isolated from fecal and soil samples from four native Jeju horse farms and six Thoroughbred farms in Jeju, Korea. The isolates were examined for the presence of virulence-associated 15-17-kDa antigens (VapA) by colony blotting, using the monoclonal antibody 10G5, and for the gene encoding VapA by PCR. R. equi was isolated from all 36 soil samples collected from the 10 farms with between 5.0 x 10(2) and 7.5 x 10(4) colony-forming units (cfu) per gram of soil, and from 37 of 40 fecal samples with between 5.0 x 10(1) and 1.1 x 10 (5) cfu per gram of feces. Virulent R. equi was isolated from seven farms and appeared in 2.0% of isolates (10 of 508). Of the 10 virulent isolates, four contained a 90-kb type II plasmid, which has been found in isolates from the Kiso native horses of Japan, and the other six contained a new variant, which did not display the EcoRI and EcoT22I digestion patterns of the 10 representative plasmids already reported (85-kb types I, II, III, and IV; 87-kb types I and II; 90-kb types I, II, III, and IV). We designated the new variant as the "90-kb type V" plasmid, because its EcoRI digestion pattern is similar to that of the 90-kb type II plasmid. This is the first report of the prevalence of virulent R. equi in Jeju, Korea. The same virulence plasmid type is found in both Korean and Japanese isolates, providing insight into the origin, ancestry, and dispersal of native horses in Korea and Japan.
  Journal of Veterinary Medical Science 01/2004; 65(12):1313-7. · 0.85 Impact Factor
• Article: Virulence of Rhodococcus equi isolated from cats and dogs.

  Shinji Takai, Ronald J Martens, Alan Julian, Márcio Garcia Ribeiro, Marconi Rodrigues de Farias, Yukako Sasaki, Kazuho Inuzuka, Tsutomu Kakuda, Shiro Tsubaki, John F Prescott
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  ABSTRACT: Nine cat isolates and nine dog isolates of Rhodococcus equi from clinical material were investigated for the presence of the virulence-associated antigens (VapA and VapB) and virulence plasmids. Five of the cat isolates and one dog isolate were VapA positive and contained an 85-kb type I or an 87-kb type I plasmid. The remaining 12 isolates were avirulent R. equi strains and contained no virulence plasmids.
  Journal of Clinical Microbiology 10/2003; 41(9):4468-70. · 4.15 Impact Factor
• Article: Molecular cloning and characterization of a 79-kDa iron-repressible outer-membrane protein of Moraxella bovis.

  Tsutomu Kakuda, Daiki Oishi, Shiro Tsubaki, Shinji Takai
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  ABSTRACT: Moraxella bovis expresses an iron-repressible 79-kDa outer-membrane protein, IrpA. DNA and N-terminal amino acid sequence analysis indicate that IrpA is closely related to FrpB of Neisseria meningitidis, FetA of Neisseria gonorrhoeae and CopB of Moraxella catarrhalis. The results of manganese mutagenesis and a gel-shift assay suggested that the transcription of irpA is negatively regulated by the ferric uptake regulator. The insertion of an antibiotic resistance cassette into the irpA gene affected the strain's ability to utilize bovine transferrin and lactoferrin. IrpA was detected in geographically diverse clinical isolates, and the antigenicity of IrpA was conserved in all the isolates tested. Therefore, IrpA may have potential as a candidate vaccine.
  FEMS Microbiology Letters 09/2003; 225(2):279-84. · 2.04 Impact Factor
• Source

  Available from: swu.ac.th

  Article: Survey of benign Theileria parasites of cattle and buffaloes in Thailand using allele-specific polymerase chain reaction of major piroplasm surface protein gene.

  Nopporn Sarataphan, Tsutomu Kakuda, Kosum Chansiri, Misao Onuma
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  ABSTRACT: During a year from 1999 to 2000, a total of 247 blood samples were collected from 214 cattle and 33 water buffaloes in 16 distinct geographical locations of Thailand and analyzed by allele-specific PCR amplification of major piroplasm surface protein (MPSP) genes of benign Theileria parasites. Four allelic MPSP gene types were determined namely C-type, I-type, B-type and Thai-type, which were originally designated from Japanese Theileria orientalis (Chitose, Ikeda), Australian T. buffeli (Warwick) and Thai T. sp. (Kamphaeng Saen), respectively. Only two allelic MPSP gene types were successively amplified from 204 (82.6%) blood samples. Among positive cases, 138 (67.6%) and 17 (8.3%) samples contained either Thai-type or C-type parasites, respectively, while 49 (24%) samples contained both types. However, nucleotide sequences of MPSP genes of Thai T. sp. amplified by C-type specific primers revealed higher (96.3%) similarity to Indonesian T. sp. rather than (87.8% similarity) to Japanese T. orientalis (Chitose) designated as C-type.
  Journal of Veterinary Medical Science 02/2003; 65(1):133-5. · 0.85 Impact Factor
• Article: Cloning and characterization of the fur gene from Moraxella bovis.

  Tsutomu Kakuda, Daiki Oishi, Shiro Tsubaki, Shinji Takai
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  ABSTRACT: A homologue of the ferric uptake regulator gene (fur) was isolated from Moraxella bovis by degenerate polymerase chain reaction and cloning. Fur protein of M. bovis exhibited 72.1% amino acid identity with Acinetobacter calcoaceticus Fur. Western blot analysis showed a decrease of Fur expression in response to sufficient-iron conditions compared with deficient-iron conditions. An electrophoretic mobility-shift assay indicated that Fur protein binds to DNA fragments containing a putative Fur-box derived from the upstream region of the M. bovis fur gene. Fur of M. bovis may regulate the expression of iron transport systems in response to iron limitation in the environment.
  Microbiology and Immunology 02/2003; 47(6):411-7. · 1.30 Impact Factor
• Article: Molecular epidemiology of VapA-positive Rhodococcus equi in thoroughbred horses in Kagoshima, Japan.

  Teruhiko Yuyama, Shigemoto Yusa, Kazunori Yoshizumi, Shigeki Yamano, Soichiro Murata, Tomoaki Hirose, Rika Osanai, Yoko Onishi, Satoshi Osato, Chie Sasaki, Yukako Sasaki, Tsutomu Kakuda, Shiro Tsubaki, Shinji Takai
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  ABSTRACT: The prevalence of virulent R. equi having 15- to 17-kDa antigens (VapA) in fecal isolates from 13 thoroughbred foals and their dams on 5 farms in Kagoshima, Japan, and the plasmid profiles of VapA-positive isolates by restriction fragment digestion patterns were investigated to compare the genotypic variation among virulence plasmids of R. equi isolates from Japan. In total, 218 (24.6%) of 886 isolates from the feces of the 13 foals and 13 (12.5%) of 104 isolates from the feces of their dams demonstrated VapA-positive R. equi. Plasmid DNA preparations of 231 virulent isolates from foals and dams were analyzed by restriction enzyme digestion with endonucleases EcoRI, EcoT22I and HindIII and were divided into 3 types: 172 isolates contained a 90-kb type I plasmid, 57 contained a 90-kb type III plasmid and 2 contained a 90-kb type IV plasmid. This study demonstrates a geographic character in the distribution of virulence plasmids found in VapA-positive isolates from thoroughbred foals in Kagoshima.
  Journal of Veterinary Medical Science 09/2002; 64(8):715-8. · 0.85 Impact Factor