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Kazuhiro Iiyama Last modified date:2019.06.05

Associate Professor / Agricultural Bioresource Sciences
Department of Bioresource Sciences
Faculty of Agriculture


Graduate School
Undergraduate School


E-Mail
Homepage
http://www.agr.kyushu-u.ac.jp/lab/plantpp/
A research outline, research achievements, a research theme, etc. .
Phone
092-802-4563
Academic Degree
PhD
Country of degree conferring institution (Overseas)
No
Field of Specialization
Plant pathology, Biological control, Insect pathology, Bacteriology
Total Priod of education and research career in the foreign country
00years00months
Outline Activities
Evolution in virulence, and genetic diversity analyses are carried out by using genome information of plant pathogenic bacteria. Isolation and identification of bacteria that have biological control activity against Ralstonia solanacearum, an plant pathogenic bacteria are conducted. Furthermore, the effects of microorganisms being virulence to pests on plant pathogens are investigated.
Research
Research Interests
  • Studies on the evolution in pathogenesis
    keyword : pathogenicity, evolution, genome comparison
    2017.01~2018.05.
  • Studies on the dual biological control against plant pathogen and insect
    keyword : plant pahogen, entomopahogenic organism, virulence, pathogenicity
    2017.01~2018.05.
  • Studies on the pathogenicity of plant pathogenic bacteria
    keyword : plant pahogenic bacteria, virulence, pathogenicity
    2017.01~2018.05.
  • Study on pathogenicity of Hafnia sp. to insects
    keyword : Hafnia sp. pathogenicity silkworm
    2015.04.
  • Genome analysis of Paenibacillus popilliae and evolution in pathogenicity
    keyword : Genome analysis, Paenibacillus popilliae, evolution, pathogenicity
    2012.04.
  • Pathogenicity of Pseudomonas aeruginosa in insect hosts
    keyword : Pseudomonas aeruginosa, Bombyx mori, virulence
    2006.04.
  • Study on phospholipases produced by Pseudomonas aeruginosa
    keyword : Pseudomonas aeruginosa, Bombyx mori, phospholipase C, plcB, plcH, plcN
    2006.04~2011.01.
  • Study on superoxide dismutases produced by Pseudomonas aeruginosa
    keyword : Pseudomonas aeruginosa, Bombyx mori, virulence,superoxide dismutase,sodM, sodB
    2004.04~2012.05Pseudomonas aeruginosa is an opportunistic pathogen of insects and possesses a manganese- and an iron- superoxide dismutases (SODs). To ascertain the role of these SODs in virulence to the silkworm, Bombyx mori, we have generated three mutants carrying an interrupted sodM (encoding manganese- SOD, Mn-SOD) or sodB (encoding iron- SOD, Fe-SOD) and a sodM sodB double mutation. Total SOD activities of sodM, sodB and sodM sodB mutants in Luria broth and tryptic soy broth were ca. 100%, 0% and 0% of the activity of a wild-type strain, respectively. Whereas the relative activities in low phosphate-succinate broth were 0%, 96% and 0%, respectively. This result indicates that the two SODs of P. aeruginosa are highly regulated by the environmental conditions. To investigate their virulence to the silkworm, the fourth instar larvae were inoculated with various doses of the wild-type and three mutants. The virulence remarkably differed among these strains. The sodM mutant was indistinguishable from the wild-type in aspect of virulence at all inoculation levels (103-106 cells / larva). Whereas the virulence of sodB mutant reduced, when relatively low cells number (103-104) was inoculated. Moreover, the double mutant could not cause the any fatal damage at 105 cells or lower. These results suggest that the SODs are required for full virulence of P. aeruginosa to the silkworm and Fe-SOD is more important than Mn-SOD in the infection process..
  • Study on the toxic protein produced by the bacteria isolated from insect
    keyword : entomopathogenic bacteria, bacterial isolates from ant lion, virulence, virulence factor
    2003.04.
  • Study on a two component, GacS-GacA in an oppotunistic bacteria Pseudomonas aeruginosa
    keyword : entomopathogenic bacteria, two component system
    2001.04~2013.01To investigate the pathogenicity of Pseudomonas aeruginosa in insects, a gacA mutant of P. aeruginosa PA01 was constructed by site-directed mutagenesis. The mutant was designated as C1. C1 was less virulent to Bombyx mori than the parent strain. To complement the gacA gene, P. aeruginosa C1 was transformed with the broad host range plasmid pJB3Km1 carrying a 3.9-kbp gacA fragment. The expression of the gacA mRNA in C1 (pgacA) was detected. In addition, the complemented mutant restored the level and timing of pyocyanin production, indicating that functional GacA is produced in the complemented strain. However, no significant difference was observed between C1 and C1 (pgacA) with respect to the killing of B. mori larvae..
Academic Activities
Papers
1. Mai Morishita, Hiroaki Mon, Jae Man Lee, Takahiro Kusakabe, Maximiano Corrêa Cassal, Chisa Yasunaga-Aoki and Kazuhiro Iiyama, Toxin complex is a major virulence determinant of Enterobacter sp. 532 against the silkworm, Bombyx mori, J. Insect Biotechnol. Sericol. , 88, 1, 21-25, 2019.02.
2. Mai Morishita, Akitsu Masuda, Hiroaki Mon, Jae Man Lee, Takahiro Kusakabe, Kosuke Tashiro, Chisa Yasunaga-Aoki, Kazuhiro Iiyama, Identification of insecticidal toxin produced by Enterobacter sp. strain 532 isolated from diseased Bombyx mori silkworms, FEMS Microbiology Letters, 366, 2, fny295, 2019.01, Although Enterobacter sp. 532 shows pathogenicity in Bombyx mori, the insecticidal mechanisms are unclear. Here, we identified and characterised an insecticidal protein from Enterobacter. The insecticidal protein was purified from the strain and inoculated into B. mori larvae. Intracellular proteins were prepared, purified and separated by preparative native polyacrylamide gel electrophoresis (PAGE); one protein band had insecticidal activity. Sodium dodecyl sulfate-PAGE showed the presence of several bands, indicating that the insecticidal protein formed a complex. Peptide mass fingerprinting of a prominent 255.3-kDa band revealed 64 peptides that matched one protein with 33.0% sequence coverage. This protein was a homologue of the A component of the toxin complex (Tc), and the VRP1 domain was conserved; thus, the gene was named itcA (insecticidal toxin complex A). In the itcA downstream region, B and C component gene homologues were found, and these genes were located on an 86.2-kb contig sequence. Two repA genes and 27 genes related to conjugation transfer of plasmids were located on the contig, suggesting that the contig originated from a mobilisable plasmid. Therefore, these findings suggested that the strain may have acquired the Tc genes by horizontal transfer. This is the first description of Tc produced by the genus Enterobacter..
3. Kenji Kikuhara, Kazuhiro Iiyama, Masaru Matsumoto, Naruto Furuya, First report on DMI fungicide resistance in Gymnosporangium asiaticum, the causal agent of Japanese pear rust, in Japan, JOURNAL OF GENERAL PLANT PATHOLOGY, 10.1007/s10327-018-0820-5, 85, 1, 49-56, 2019.01, Japanese pear rust, caused by Gymnosporangium asiaticum Miyabe ex Yamada, is one of the most important fungal plant diseases affecting Japanese pear cultivation. Although sterol demethylation inhibitor (DMI) fungicides have controlled this disease effectively, outbreaks have been frequent in some areas of Japan since 2008. We, therefore, evaluated fungal populations from Fukuoka Prefecture for DMI fungicide sensitivity in vivo. Aeciospores on diseased pear leaves were collected in 2013 from four orchards in Fukuoka Prefecture and one control orchard in Ibaraki Prefecture. Juniperus chinensis var. kaizuka Hort. test plants were sprayed with difenoconazole and fenarimol at recommended rates, and the plants were inoculated with aeciospores. Difenoconazole and fenarimol effectively controlled the population of rust fungus from Ibaraki Prefecture. In contrast, fenarimol and difenoconazole were found to be ineffective against fungal populations from Fukuoka Prefecture. Subsequently, telia were collected from diseased juniper twigs in five areas in Fukuoka Prefecture and one control area in Ibaraki Prefecture in 2013 and 2014. Similarly, test plants of Japanese pear ‘Kousui’ were treated with difenoconazole or fenarimol and inoculated with basidiospores taken from those telia. Both difenoconazole and fenarimol were largely ineffective against the populations from Fukuoka Prefecture. In addition, field trials conducted in an experimental orchard in 2014 and 2015 showed that difenoconazole and fenarimol were not effective in controlling this disease. These results indicate the occurrence of Japanese pear rust resistant to DMI fungicides in Fukuoka Prefecture..
4. Shahadat Hossain, Abu Ashraf Khan, Md Rahman, Kazuhiro Iiyama, Naruto Furuya, First Report of Soft Rot Disease of Papaya Caused by Klebsiella variicola in Bangladesh, J. Fac. Agr., Kyushu Univ., 63, 2, 201-205, 2018.12.
5. Htet Wai Wai Kyaw, Kenichi Tsuchiya, Masaru Matsumoto, Kazuhiro Iiyama, Seint San Aye, Myo Zaw, Daisuke Kurose, Mitsuo Horita, Naruto Furuya, Genetic diversity of Ralstonia solanacearum strains causing bacterial wilt of solanaceous crops in Myanmar, J Gen Plant Pathol, 59, 221-233, 2017.07, In 2013 and 2014, an extensive survey of bacterial wilt in Myanmar was performed, and 70 strains of Ralstonia solanacearum (Rs) were collected from wilting plants of tomato, potato, chili and eggplant. Myanmar Rs strains were characterized by traditional and molecular methods. Polymerase chain reaction (PCR) test using Rs-specific primer set amplified one specific band (281-bp) from template DNA of all strains. Pathogenicity tests on the four solanaceous plants differentiated the strains into six pathogenic groups. Biovar determination tests showed that biovar 3 strains predominated (63%) among all Rs strains. Biovar 4 strains (7%) were obtained from both tomato and chili strains, whereas biovar 2 (30%) strains were isolated only from potato. Multiplex-PCR analysis indicated that tomato, eggplant and chili strains belonged to phylotype I, whereas potato strains comprised phylotype I and phylotype II. Strains in phylotype I, which was suggested to have originated from Asia, were the most prevalent in all surveyed areas. Phylogenetic analysis based on the endoglucanase (egl) gene sequences revealed that Myanmar strains partitioned into two major clusters that corresponded to phylotype I and II. Strains in phylotype I were further divided into seven subclusters, each corresponding to a distinct sequevar (15, 17, 46, 47, 48, unknown 1 or unknown 2). All strains in phylotype II belonged to sequevar 1. This is the first comprehensive report of the presence of diverse Rs strains in Myanmar..
6. Kazuhiro Iiyama, Mai Morishita, Jae Man Lee, Hiroaki Mon, Takahiro Kusakabe, Kosuke Tashiro, Taiki Akasaka, Chisa Yasunaga-Aoki and Kazuhisa Miyamoto, A reconsideration of the taxonomic position of two bacterial strains isolated from flacherie-diseased silkworms in 1965, Journal of Insect Biotechnology and Sericology, 86, 35-41, 2017.04, Recent advances in bacterial characterization methodologies have made taxonomic categorization significant- ly more accurate. Here, we re-evaluated the position of bacterial strains (532 and 652) belonging to the genus Hafnia, isolated from flacherie-diseased silkworms in 1965. Phylogenetic analysis based on the 16S rRNA gene sequences of these strains suggests that they belong to genus Enterobacter. Using multilocus sequence analy- sis (MLSA), these strains were further classified to MLSA group A, which is a “core” group of Enterobacter con- taining E. cloacae (the type species of the genus). Although these strains were closely related to E. mori, E. tabaci, and E. asburiae, they also had other MLSA characteristics that distinguished them from these neighbor- ing bacterial species. These data were supported by further biochemical analysis. Thus, it appears that the 532 and 652 strains isolated almost half a century ago belong to genus Enterobacter, and their unique characteristics strongly suggest that they are a novel bacterial species..
7. Kazuhiro Iiyama, Eigo Takahashi, Jae Man Lee, Hiroaki Mon, Mai Morishita, Takahiro Kusakabe, Chisa Yasunaga-Aoki, Alkaline protease contributes to pyocyanin production in Pseudomonas aeruginosa., FEMS Microbiology Letters, doi.org/10.1093/femsle/fnx051, 364, fnx051, 2017.04.
8. Eigo Takahashi, Jae Man Lee, Hiroaki Mon, Yuuka Chieda, Chisa Yasunaga-Aoki, Takahiro Kusakabe, Kazuhiro Iiyama, Effect of antibiotics on extracellular protein level in Pseudomonas aeruginosa., Plasmid, 10.1016/j.plasmid.2016.03.001, 84-85, 44-50, 2016.03.
9. Erika Taira, Hiroaki Mon, Jae Man Lee, Takahiro Kusakabe, Chisa Yasunaga-Aoki, Kazuhiro Iiyama, Virulence of lipopolysaccharide-deficient mutants of Serratia liquefaciens toward the silkworm, Bombyx mori., Journal of Insect Biotechnology and Sericology, 10.11416/jibs.85.1_007, 85, 7-14, 2016.04.
10. Oumi Nishi, Kazuhiro Iiyama, Chisa Yasunaga-Aoki, Susumu Shimizu, Phylogenetic status and pathogenicity of Metarhizium majus isolated from a fruit beetle larva in Japan, Mycological Progress, 10.1007/s11557-015-1082-7, 14, 8, 58-58, 2015.08.
11. Kazuhiro Iiyama, Hiroaki Mon, Kazuki Mori, Takumi Mitsudome, Jae Man Lee, Takahiro Kusakabe, Kousuke Tashiro, Shin-Ichiro Asano, Chisa Yasunaga-Aoki, Characterization of KfrA proteins encoded by a plasmid of Paenibacillus popilliae ATCC 14706T., Meta Gene, 10.1016/j.mgene.2015.03.001, 4, 29-44, 2015.06.
12. Erika Taira, Kazuhiro Iiyama, Hiroaki Mon, Kazuki Mori, Taiki Akasaka, Kousuke Tashiro, Chisa Yasunaga-Aoki, Jae Man Lee, Takahiro Kusakabe, Draft genome sequence of entomopathogenic Serratia liquefaciens strain FK01., Genome Announcements, 2, 3, e00609-14-e00609-14, 2014.06.
13. Kazuhiro Iiyama, Masahiro Otao, Kazuki Mori, Hiroaki Mon, Jae Man Lee, Takahiro Kusakabe, Kousuke Tashiro, Shin-Ichiro Asano, Chisa Yasunaga-Aoki, Phylogenetic relationship of Paenibacillus species based on putative replication origin regions and analysis of an yheCD-like sequence found in this region., Bioscience, Biotechnology, and Biochemistry, 10.1080/09168451.2014.905188, 78, 891-897, 2014.05.
14. Oumi Nishi, Kazuhiro Iiyama, Chisa Yasunaga-Aoki, Susumu Shimizu, Comparison of the germination rates of Metarhizium spp. conidia from Japan at high and low temperatures, Letters in Applied Microbiology, 10.1111/lam.12150, 57, 6, 554-560, 2013.12.
15. Kazuhiro Iiyama, Kazuki Mori, Hiroaki Mon, Yuuka Chieda, Jae Man Lee, Takahiro Kusakabe, Kosuke Tashiro, Shin-ichiro Asano, Chisa Yasunaga-Aoki, Susumu Shimizu, Draft Genome Sequence of Paenibacillus popilliae ATCC 14706T, Journal of Insect Biotechnology and Sericology, 82, 2, 45-48, 2013.06.
16. Kazuhiro Iiyama, Oumi Nishi, Hiroaki Mon, JAE MAN LEE, takahiro kusakabe, Asano Shin-ichiro, Chisa Yasunaga-Aoki, Susumu Shimizu, Phylogenetic analysis of Paenibacillus popilliae and its related taxa based on housekeeping genes., J. Insect Biotechnol. Sericol. , 10.11416/jibs.82.1_001, 82, 1-11, 2013.02.
17. Mon Hiroaki, JAE MAN LEE, Fukushima Mai, Nagata Yudai, Fujii Mie, Xu Jian, Nishi Oumi, Kazuhiro Iiyama, takahiro kusakabe, Production and characterization of the celery mismatch endonuclease CEL II using baculovirus/silkworm expression system, APPLIED MICROBIOLOGY AND BIOTECHNOLOGY, 10.1007/s00253-012-4583-1, 97, 15, 6813-6822, 2013.08.
18. Arai Hiroyuki, Iiyama Kazuhiro, Role of nitric oxide-detoxifying enzymes in virulence of Pseudomonas aeruginosaagainst the silkworm, Bombyx mori, Biosci. Biotechnol. Biochem, 77, 1, 198-200, 2013.01.
19. Kaibara Fusako, Iiyama Kazuhiro, Chieda Yuuka, Lee Jae Man, kusakabe takahiro, Yasunaga-Aoki Chisa, Shimizu Susumu, Construction of serralysin-like metalloprotease-deficient mutants of Serratia liquefaciens and their virulence in the silkworm, Bombyx mori, J. Insect Biotechnol. Sericol. , 81, 55-61, 2012.12.
20. Chieda, Y., Iiyama, K., Lee, J.M., Kusakabe, T., Yasunaga-Aoki, C. and Shimizu, S., Virulence of an exotoxin A-deficient strain of Pseudomonas aeruginosa toward the silkworm, Bombyx mori., Microbial Pathogenesis, 51, 6, 407-414, 2011.12.
21. Iiyama, K., Lee, J.M., Kusakabe, T., Yasunaga-Aoki, C. and Shimizu, S. , Improvement in pHERD vectors to express recombinant proteins tagged with hexahistidine at either the NH2- or COOH- terminal in Pseudomonas aeruginosa, J. Insect Biotechnol. Sericol., 80, 57-61, 2011.06.
22. Egami, I., Iiyama, K., Zhang, P., Chieda, Y., Ino, N., Hasegawa, K., Lee, J.M., Kusakabe, T., Yasunaga-Aoki, C. and Shimizu, S., Insecticidal bacterium isolated from an ant lion larva from Munakata, Japan., J. Appl. Entomol., vol. 133 No. 2 117-124, 2009.03.
23. Iiyama, K., Chieda, Y., Lee, J.M., Kusakabe, T., Yasunaga-Aoki, C. and Shimizu, S., Virulence of Phospholipase C Mutants of Pseudomonas aeruginosa PAO1 against the Silkworm, Bombyx mori., J. Insect Biotechnol. Sericol., vol. 77, No. 2, 115-120, 2008.06.
24. Chieda, Y., Iiyama, K., Lee, J.M., Kusakabe, T., Yasunaga-Aoki, C. and Shimizu, S., Inactivation of pyocyanin synthesis genes has no effect on the virulence of Pseudomonas aeruginosa PAO1 toward the silkworm, Bombyx mori, FEMS Microbiology Letters, 278 101-107, 2008.01.
25. Chieda, Y., Iiyama, K., Lee, J.M., Kusakabe, T., Yasunaga-Aoki, C. and Shimizu, S., The gacA gene of Pseudomonas aeruginosa PAO1 is not required for full virulence in Bombyx mori., J. Insect Biotechnol. Sericol. , Vol. 76, 89-95, 2007.06.
26. Iiyama K, Chieda Y, Lee JM, Kusakabe T, Yasunaga-Aoki C, Shimizu S., Effect of superoxide dismutase gene inactivation on virulence of Pseudomonas aeruginosa PAO1 toward the silkworm, Bombyx mori, Appl Environ Microbiol., 73(5):1569-75, 2007.03.
27. Chieda, Y., Iiyama, K., Yasunaga-Aoki, C., Lee, J.M., Kusakabe, T. and Shimizu, S., Pathogenicity of gacA mutant of Pseudomonas aeruginosa PA01 in the silkworm, Bombyx mori, FEMS Microbiol. Lett., 10.1016/j.femsle.2005.01.032, 244, 1, 181-186, 244巻 181-186, 2005.03.
28. Iiyama, K., Chieda, Y., Yasunaga-Aoki, C., Hayasaka, S. and Shimizu, S., Analyses of the ribosomal DNA region in Nosema bombycis NIS 001., J. Eukaryot. Microbiol, 10.1111/j.1550-7408.2004.tb00592.x, 51, 6, 598-604, 51巻 598-604., 2004.01.
Presentations
1. Kyaw, H.W.W., Tsuchiya, K., Matsumoto, M., Kurose, D., Iiyama, K., Horita, M., and Furuya, N., Properties of potato strains of Ralstonia solanacearum in Myanmar, 平成30年度日本植物病理学会, 2018.03.
2. Htet Wai Wai Kyaw, Masaru Matsumoto, Kazuhiro Iiyama, Mitsuo Horita, Daisuke Kurose, Kenichi Tsuchiya, Naruto Furuya, Characterization of Ralstonia solanacearum strains causing bacterial wilt of solanaceous crops in Myanmar, AFELiSA 2017 International Symposium on Agricultural, Food, Environmental and Life Sciences in Asia, 2017 (The 14th International Joint Symposium between Japan and Korea), 2017.11.
3. Kyaw, H.W.W., Matsumoto, M., Kurose, D., Iiyama, K., Horita, M., Tsuchiya, K., Furuya, N., Genetic diversity of Myanmar strains of Ralstonia solanacearum, 平成29年度日本植物病理学会, 2017.04.
4. Erika Taira, Kazuhiro Iiyama, Jae Man Lee, Takahiro Kusakabe, Chisa Yasunaga-Aoki, Lipopolysaccharide contributes to the virulence of Serratia liquefaciens against Bombyx mori, The 4th Asia-Pacific Congress of Sericulture and Insect Biotechnology (APSERI 2015), 2015.04.
5. Shota Fukui, Chisa Yasunaga-Aoki, Takashi Matsuyama, Kazuhiro Iiyama, Study on in vitro excystation of Farinocystis sp. isolated from the West Indian sweet potato weevil, Euscepes postfasciatus, The 4th Asia-Pacific Congress of Sericulture and Insect Biotechnology (APSERI 2015), 2015.04.
6. Shota Fukui, Chisa Yasunaga-Aoki, Takashi Matsuyama, Kazuhiro Iiyama, Study on intestinal factors affecting excystation of Farinocystis sp. in the West Indian sweet potato weevil, Euscepes postfasciatus, International Symposium on Basic and Applied Research on Sericulture and Insect Sciences, Collaboratively Organized by Kyushu Branch of Japanese Society of Sericultural Science, Japan and College of Agriculture and Life Sciences, Kyungpook National Univer, 2015.04.
7. Erika Taira, Kazuhiro Iiyama, Jae Man Lee, Takahiro Kusakabe, Chisa Yasunaga-Aoki, Sensitivity to oxidative stress in lipopolysaccharide mutants of Serratia liquefaciens, International Symposium on Basic and Applied Research on Sericulture and Insect Sciences, Collaboratively Organized by Kyushu Branch of Japanese Society of Sericultural Science, Japan and College of Agriculture and Life Sciences, Kyungpook National Univer, 2015.04.
8. Erika Taira, Kazuhiro Iiyama, Jae Man Lee, Takahiro Kusakabe, Chisa Yasunaga-Aoki, Virulence of Serratia liquefaciens against Bombix mori, International Symposium on Agriculture, Forestry, Environment and Life Sciences in Asia, 2014, 2014.10, Serratia spp. are Gram-negative bacteria of the family Enterobacteriaceae. Serratia spp. are ubiquitous environmental bacteria, and they infect a wide range of species from animals to plants. Unfortunately, Serratia spp. have been reported to kill useful insects, such as the silkworm, Bombyx mori, and the honeybee, Apis mellifera. Therefore, in order to protect these useful insects from diseases caused by Serratia spp., it is important to elucidate their pathogeneses. In our previous work, S. liquefaciens isolated from an antlion larva was highly virulent against the silkworm. Since the virulence of S. liquefaciens remains poorly understood, a comprehensive analysis using transposon-induced mutants was carried out to investigate its virulence against the silkworm in the present study. Firstly, a transposon insertion library of S. liquefaciens FK01 was constructed. Subsequently, 1,200 transconjugants were injected into the hemocoel of silkworm larvae. Then, four transposon mutants showing a reduced virulence were screened. Southern blot analysis confirmed the insertion of transposons at single sites in the transconjugants. Sequences flanking both sides of the transposons indicated that they were inserted into the lipopolysaccharide (LPS) biosynthesis genes in these transconjugants. These results suggest that LPS markedly contributes to the virulence of S. liquefaciens against the silkworm. The role of LPS in the virulence of S. liquefaciens against the silkworm remains to be determined..
9. Oumi Nishi, Kazuhiro Iiyama, Chisa Yasunaga-Aoki, Susumu Shimizu, Comparative virulence analysis of entomopathogenic fungus Metarhizium spp. in different ambient temperature conditions., International Symposium on Agricultural, Food, Environmental and Life Sciences in Asia, 2013, 2013.11.
10. Pseudomonas aeruginosa is a Gram-negative, rod shaped bacterium and human opportunistic pathogen. Since several strains of the P. aeruginosa infect plants, nematodes and insects, some organisms have been used as model hosts of P. aeruginosa infection. To use silkworm, Bombyx mori, as a model host, it is important to compare the bacterial virulence strategies between the different hosts, mammal and B. mori.
We created the mutants of GacS-GacA two component system (gacA), pyocyanin production (phzM, phzS), superoxide dismutase (SOD) production (sodM, sodB, sodM sodB) and nucleotide excision repair (NER) mechanism (uvrC) originated from P. aeruginosa PAO1. Among these genes, gacA, sodB phzM and phzS have been reported to be important in the virulence to mammalian hosts. Gene disruption in each mutant was confirmed by Southern hybridization and change of phenotypic characteristics (pyocyanin production, SOD activity and UV sensitivity). Inoculation test demonstrated that gacA, phzM, phzS mutants had the same level of virulence in comparison with the parent strain, PAO1. Whereas, sodB, sodM sodB and uvrC mutants showed reduced virulence. In addition, the virulence of the mutants was restored by the complementation of the corresponding genes in trans. These results suggested that the importance of these virulence factors depended on the host species, but that of the survival factors were common in P. aeruginosa..
11. It is known that the GacS/GacA two-component system is important for the virulence of Pseudomonas aeruginosa in invertebrate and vertebrate hosts and in plants. In this study, the gacA nonpolar mutant C2 and polar mutant C3 were generated and inoculated into Bombyx mori. The polar mutants showed reduced virulence, whereas no significant difference was observed between the gacA nonpolar mutant and wild type strain with respect to the ability to kill B. mori larvae. This suggests that the reduced virulence was due to a polar effect of the insertions, rather than to the lack of the gacA gene product.
In the genome, the gacA gene is directly followed by a uvrC gene. To clarify the importance of uvrC to virulence, the uvrC mutant strain UC was constructed. UC strains were assessed for the production of virulence factors (pyocyanin, pyoverdine, elastase and total protease) in comparison to UC strains. No difference was observed between PAO1 (parent strain) and UC strains, suggesting that the mutation of uvrC had no effect on the production of these virulence factors.
UvrC is an endonuclease involved in NER, which is one of the excision repair systems. We tested the survival of UC strains exposed to cisplatin or UV. The survival rate of UC (pBBR1MCS2) after cisplatin or UV treatment was clearly lower than that of PAO1 (pBBR1MCS2).
To investigate the influence of uvrC on virulence of hemocoel infection, UC strains were injected into B. mori. UC was shown to be less virulent than PAO1. This result suggests that not gacA rather than uvrC is necessary for the full virulence of P. aeruginosa PAO1 to in B. mori..
Educational
Educational Activities
Research about the newest technical journal announcement paper related to plant pathology, plant pathogenic microbiology.
Study about the characterization, infection / multiplication style, a host and a pathogen interaction, plant pathology.
The instruction of an experiment and advice which are performed with the latest technology.