Hemocyte Apoptosis Induced by Entomopathogenic Bacteria, Xenorhabdus and Photorhabdus, in Bombyx mori

  • Sunghwan Cho (School of Bioresource Sciences, College of Natural Sciences, Andong National University) ;
  • Kim, Yonggyun (School of Bioresource Sciences, College of Natural Sciences, Andong National University)
  • Published : 2004.06.01

Abstract

Entomopathogenic bacteria of Xenorbabdus nematophila, Xenorhabdus sp., and Photorhabdus temperata subsp. temperata are symbionts of entomopathogenic nematodes including Steinernema and Heterorhabditis. These bacteria have potent insecticidal pathogenicity causing hemolymph septicemia. To explain a mechanism of the septicemia, this research raises a hypothesis that the hemolymph septicemia is due to the induction of the programmed cell death (=apoptosis) of the hemocytes by the entomopathogenic bacteria. Injection of the bacteria into the hemocoel of the fifth instar larvae of Bombyx mori led to septicemia (few viable hemocytes) at 12h in all three bacterial treatments. During the pathogenicity, the infected hemocytes exhibited the membrane b1ebbing, nuclear chromatin condensation and DNA fragmentation, which reflected typical morphological changes of cells undergoing apoptosis. The apoptosis began as early as 4h after bacterial injection and increased with post-injection time. Three pathogenic bacteria, however, differed in the degree of apoptosis-inducing effect on B. mori by the effective bacterial dose and time. These results support the hypothesis that hemolymph septicemia caused by the bacteria is due to their apoptosis-inducing effect.

Keywords

References

  1. Albina, J.E., S. Cui, R.B. Mateo and J.S. Reichner. 1993. Nitric oxide mediated apoptosis in murine peritoneal macrophages. J. Immunol. 150: 5080-5085
  2. Arends, M.J. and A.H. Wyllie. 1991. Apoptosis: mechanisms and roles in pathology, Int. Rev. Exp. Path. 32: 223-354
  3. Boemare, N. and R.J. Akhurst. 1988. Biochemical and physiological characterization of colony form variants in Xenorhabdus and Photorhabdus spp. (Enterobacteriaceae). J. Gen. Microbiol. 134: 751-761
  4. Brillard, J., C. Ribeiro, N. Boemare, M. Breh$\acute{e}$lin and A. Givaudan. 2001. Two distinct hemolytic activities in Xenorhabdus nematophila are active against immunocompetent insect cells. Appl. Environ. Microbiol. 67:2515-2525 https://doi.org/10.1128/AEM.67.6.2515-2525.2001
  5. Buttke, T.M. and P.A. Sandstrom. 1994. Oxidative stress asa mediator of apoptosis. Immunol. Today 15: 7-10 https://doi.org/10.1016/0167-5699(94)90018-3
  6. Chen, G., Y. Zhang, J. Li, G.B. Dunphy, Z.K. Punja and J.M. Webster. 1996. Chitinase activity of Xenorhabdus and Photorhabdus species, bacterial associates of entomopathogenic nematodes. J. Invertebr. Pathol. 68: 101-108 https://doi.org/10.1006/jipa.1996.0066
  7. Chen, Y., and A. Zychlinsky. 1994. Apoptosis induced by bacterial pathogens. Microb. Pathog. 17: 203-212 https://doi.org/10.1006/mpat.1994.1066
  8. Cohen, G.M. 1997. Caspases: the executioners of apoptosis. Biochem. J. 326: 1-16
  9. Dunphy, G.B. and J.M. Webster. 1988. Lipopolysaccharides of Xenorhabdus nematophilus (Enterobacteriaceae) and their haemocyte toxicity in non-immune Galleria mellonella (Insecta: Lepidoptera) larvae. J. Gen. Microbiol. 134: 1017-1028
  10. Dunphy, G.B. and J.M. Webster. 1991. Antihemocytic surface components of Xenorhabdus nematophilus var. dutki and their modification by serum of nonimmune larvae of Galleria mellonella. J. Invertebr. Pathol. 58: 40-51 https://doi.org/10.1016/0022-2011(91)90160-R
  11. Forst, S., D. Barbara, N. Boemare and E. Stackebrandt. 1997. Xenorhabdus and Photorhabdus spp.: bugs that kill bugs. Annu. Rev. Microbiol. 51: 47-72 https://doi.org/10.1146/annurev.micro.51.1.47
  12. Givaudan, A. and A. Lanois. 2000. flhDC, the flagellar master operon of Xenorhabdus nematophilus: requirement for motility, lipolysis, extracellular hemolysis, and full virulence in insects. J. Bacteriol. 182: 107-115 https://doi.org/10.1128/JB.182.1.107-115.2000
  13. Helmreich, E.J. 2001. Regulation of cell death, pp. 234-249, in The biochemistry of cell signaling. 328pp. Oxford University Press, Oxford, UK
  14. Hwang, S.Y., S. Paik, S.H. Park, H.S. Kirn, I.S. Lee, S.P. Kim, W.K. Baek, M.H. Suh, T.K. Kwon, J.W. Park, J.B. Park, J.J. ee and S.I. Suh. 2003. N-phenethyl-2-phenylacetamide isolated from Xenorhabdus nematophilus induced apoptosis through caspases activation and calpainmediated Bax cleavage in U937 cells, Int. J. Oncol. 22:151-157
  15. Ji, D. and Y. Kim. 2004. An entomopathogenic bacterium, Xenorhabdus nematophila, inhibits the expression of an antibacterial peptide, cecropin, of the beet armyworm, Spodoptera exigua. J. Insect Physiol. 50: (In press)
  16. Kang, S. 2003. Identification of an entomopathogenic nematode, Heterorhabditis megidis, and its symbiotic bacterium, Photorhabdus temperata subsp. temperata. Master Thesis. Andong National University, Andong, Korea
  17. Kumar, S. and P.A. Colussi. 1999. Prodomains-adaptors-oligomerization: the pursuit of caspase activation in apoptosis. Trends Biochem. Sci. 24: 1-4 https://doi.org/10.1016/S0968-0004(98)01332-2
  18. Mutunga, M., P.M. Preston and K.J. Sumption. 1998. Nitric oxide is produced by Cowdria ruminantium-infected bovine pulmonary endothelial cells in vitro and is stimulated by gamma interferon. Infect. Immun. 66: 2115-2122
  19. Paik, S., Y.H. Park, S.I. Suh, H.S. Kim, I.S. Lee, M.K. Park, C.S. Lee and S.H. Park. 2001. Unusual cytotoxic phenethylamides from Xenorhabdus nematophilus. Bull. Korean Chem. Soc. 22: 372-374
  20. Park, Y. and Y. Kim. 2000. Eicosanoids rescue Spodoptera exigua infected with Xenorhabdus nematophilus, the symbiotic bacteria to the entomopathogenic nematode, Steinernema carpocapsae. J. Insect Physiol. 46:1469-1476 https://doi.org/10.1016/S0022-1910(00)00071-8
  21. Park, Y. and Y. Kim. 2003. Xenorhabdus nematophila inhibits p-bromophenacy1 bromide (BPB)-sensitive $PLA_2$ of Spodoptera exigua. Arch. Insect Biochem. Physiol. 54:134-142 https://doi.org/10.1002/arch.10108
  22. Park, Y. Y. Lee and Y. Kim. 1999. Identification and characterization of a symbiotic bacterium associated with Steinernema carpocapsae in Korea. J. Asia-Pacific Entomol. 2: 105-111 https://doi.org/10.1016/S1226-8615(08)60038-2
  23. Park, Y., Y. Kim, H. Tunaz and D.W. Stanley. 2004. An entomopathogenic bacterium, Xenorhabdus nematophita, inhibits hemocytic phsopholipase $A_2$ ($PLA_2$) in tobacco hornworms, Manduca sexta. J. Invertebr. Pathol. (In press)
  24. Ribeiro, C., M. Vignes and M. Brehelin. 2003. Xenorhabdusnematophila (Enterobacteriaceae) secretes a cation-selective calcium-independent porin which causes vacuolation of the rough endoplasmic reticulum and cell lysis. J. Biol. Chem. 278: 3030-3039 https://doi.org/10.1074/jbc.M210353200
  25. SAS Institute. 1989. SAS/STAT user's guide, release 6.03 ed. SAS Institute, Cary, North Carolina
  26. Silverman, D.J. and L.A. Santucci. 1988. Potential for free radical-induced lipid peroxidation as a cause of endothelial cell injury in Rocky Mountain spotted fever. Infect Immun. 56: 3110-3115
  27. Susin, S.A., H.K. Lorenzo, N. Zamzami, I. Marzo, B.E. SnowG.M. Brothers, J. Mangion, E. Jacotot, P. Constantini, M. Loeffler, N. Larochette, D.R. Goodlett, R. Aebersold, D.P. Siderovski, J.M. Penninger and G. Kroemer. 1999. Molecular characterization of mitochondrial apoptosisinducing factor. Nature 397: 441-446 https://doi.org/10.1038/17135
  28. Wyllie, A.H., J.F.R. Kerr, A.R. Currie. 1980. Cell death: the significance of apoptosis. Int. Rev. Cytol. 68: 251-306 https://doi.org/10.1016/S0074-7696(08)62312-8
  29. Zychlinsky, A. and P. Sansonetti. 1997. Apoptosis in bacterial pathogenesis. J. Clin. Invest. 17: 203-212