Skip to main content

METARHIZIUM ANISOPLIAE AS A MODEL FOR STUDYING BIOINSECTICIDAL HOST PATHOGEN INTERACTIONS

  • Conference paper
Novel Biotechnologies for Biocontrol Agent Enhancement and Management

Part of the book series: NATO Security through Science Series ((NASTA))

Abstract

Molecular biology methods have elucidated pathogenic processes in several biocontrol agents including one of the most commonly applied entomopathogenic fungi, Metarhizium anisopliae. In this article I will describe how a combination of EST and microarray approaches, gene disruption strategies, manipulation of gene expression and use of marker genes has: (1) identified and characterized genes involved in infection; (2) manipulated the genes of the pathogen to improve biocontrol performance; (3) allowed expression of a neurotoxin from the scorpion Androctonus australis; (4) allowed assessments of environmental risks posed by these modifications and (5) identified differences in genic constituents and gene expression that account for differences between strains.

This is a preview of subscription content, log in via an institution to check access.

Access this chapter

Subscribe and save

Springer+ Basic
$34.99 /Month
  • Get 10 units per month
  • Download Article/Chapter or eBook
  • 1 Unit = 1 Article or 1 Chapter
  • Cancel anytime
Subscribe now

Buy Now

Chapter
USD 29.95
Price excludes VAT (USA)
  • Available as PDF
  • Read on any device
  • Instant download
  • Own it forever
eBook
USD 259.00
Price excludes VAT (USA)
  • Available as PDF
  • Read on any device
  • Instant download
  • Own it forever
Softcover Book
USD 329.99
Price excludes VAT (USA)
  • Compact, lightweight edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info
Hardcover Book
USD 329.99
Price excludes VAT (USA)
  • Durable hardcover edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info

Tax calculation will be finalised at checkout

Purchases are for personal use only

Institutional subscriptions

Preview

Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.

Similar content being viewed by others

References

  1. T. M. Butt, C. Jackson, and N. Magan, Introduction-fungal biological control agents: Progress, problems and potential, inFungal Biocontrol Agents: Progress, Problems and Potential, edited by T. M. Butt, C. Jackson, and N. Morgan (CAB International, Wallingford, UK, 2001), pp. 1–8.

    Google Scholar 

  2. R. P. Bateman, Controlled droplet application of mycoinsecticides: An environmentally friendly way to control locusts. Antenna 16, 6–13 (1992).

    Google Scholar 

  3. R. J. Milner, Selection and characterization of strains of Metarhizium anisopliae for control of soil insects in Australia, in Biological Control of Locusts and Grasshoppers, edited by C. J. Lomer and C. Prior (CAB International, Wallingford, UK, 1992), pp. 200–207.

    Google Scholar 

  4. C. Prior, Discovery and characterization of fungal pathogens for locust and grasshopper control, in Biological Control of Locusts and Grasshoppers, edited by C. J. Lomer and C. Prior (CAB International, Wallingford, UK, 1992), pp. 159–180.

    Google Scholar 

  5. A. Hajek, S. P. Wraight, and J. D. Vandenberg, Control of arthropods using pathogenic fungi, in Bio-exploitation of Filamentous Fungi, edited by S. P. Pointing and K. D. Hyde (Fungal Diversity Press, Hong Kong, 2001).

    Google Scholar 

  6. R. L. Harrison and B. C. Bonning, Genetic engineering of biocontrol agents for insects, in Biological and Biotechnological Control of Insect Pests, edited by J. E. Rechcigl and N. A. Rechcigl (Lewis Publishers, Boca RAton, FL, 1998), pp. 243–280.

    Google Scholar 

  7. S. P. Wraight, M. A. Jackson, and S. L. deKock, Production, stabilization and formulation of fungal bocontrol agents, inFungal biocontrol agents: Progress, problems and potential, edited by T. M. Butt, C. Jackson, and N. Morgan (CAB International, Wallingford, UK, 2001), pp. 253–287.

    Google Scholar 

  8. M. G. Klein, P. S. Grewal, and T. A. Jackson, Lawn, turf and grassland pests, in Field Manual of Techniques in Invertebrate Pathology, edited by L. A. Lacey and H. K. Kaya (Kluwer Academic Publishers, Dordrecht, 2000), pp. 681–706.

    Google Scholar 

  9. D. W. Roberts and R. J. St. Leger, Metarhizium spp. Cosmopolitan insect-pathogenic fungi: Mycological aspects, Adv. Appl. Microbiol. 54, 1–70 (2004).

    Article  PubMed  CAS  Google Scholar 

  10. M. A. Benjamin, E. Zhioua, and R. S. Ostfeld, Laboratory and field evaluation of the entomopathogenic fungus Metarhizium anisopliae (Deuteromycetes) for controlling questing adult Ixodes scapularis (Acari: Ixodidae), J. Med. Entomol. 39, 723–728 (2002).

    PubMed  Google Scholar 

  11. M. Wright, A. K. Raina, and A. R. Lax, A strain of the fungus Metarhizium anisopliae for controlling subterranean termites, J. Econ. Entomol. 98, 1451–1458 (2005).

    Article  PubMed  Google Scholar 

  12. F. M. Freimoser, S. Screen, S. Bagga, G. Hu, and R. J. St. Leger, EST analysis of two subspecies of Metarhizium anisopliae reveals a plethora of secreted proteins with potential activity in insect hosts, Microbiology 149, 239–247 (2003).

    Article  PubMed  CAS  Google Scholar 

  13. F. M. Freimoser, S. Screen, G. Hu, and R. J. St. Leger, EST analysis of genes expressed by the zygomycete Conidiobolus coronatus during optimized secretion of proteins, Microbiology 149, 1893–1900 (2003).

    Article  PubMed  CAS  Google Scholar 

  14. F. M. Freimoser, G. Hu, and R. J. St. Leger, Variation in gene expression patterns as the insect pathogen Metarhizium anisopliae adapts to different host cuticles or nutrient deprivation in vitro, Microbiology 151, 361–371 (2005).

    Article  PubMed  CAS  Google Scholar 

  15. C. Wang, G. Hu, and R. J. St. Leger, Differential gene expression by Metarhizium anisopliae growing in root exudate and host (Manduca sexta) cuticle or hemolymph reveals mechanisms of physiological adaptation, Fungal Genet. Biol. 42, 704–718 (2005).

    Article  PubMed  CAS  Google Scholar 

  16. C. Wang, T. M. Butt, and R. J. St. Leger, Colony sectorization of Metarhizium anisopliae is a sign of ageing, Microbiology 151, 3223–3236 (2005).

    Article  PubMed  CAS  Google Scholar 

  17. C. Wang and R. J. St. Leger, A collagenous protective coat enables Metarhizium anisopliae to evade insect immunity, Proc. Natl. Acad. Sci. 103, 6647–6652 (2006).

    Article  PubMed  CAS  Google Scholar 

  18. R. J. St. Leger, L. Joshi, M. J. Bidochka, and D. W. Roberts, Construction of an improved mycoinsecticide over-expressing a toxic protease, Proc. Natl. Acad. Sci. 93, 6349–6354 (1996).

    Article  Google Scholar 

  19. S. E. Screen, G. Hu, and R. J. St. Leger, Transformants of Metarhizium anisopliae sf. anisopliae overexpressing chitinase from Metarhizium anisopliae sf. acridum show early induction of native chitinase but are not altered in pathogenicity to Manduca sexta, J. Invertebr. Pathol. 78, 260–266 (2001).

    Article  PubMed  CAS  Google Scholar 

  20. A. Leclerque, H. Wan, A. Abschultz, S. Chen, G. V. Mitina, G. Zimmerman, and H. U. Scharer, Agrobacterium-mediated insertional mutagenesis (AIM) of the entomopathogenic fungus Beauveria bassiana, Curr. Genet. 45, 111–119 (2004).

    Article  PubMed  CAS  Google Scholar 

  21. R. J. St. Leger, B. May, L. Allee, D. C. Frank, D. W. Roberts, and R. C. Staples, Genetic differences in allozymes and in formation of infection structures among isolates of the entomopathogenic fungus Metarhizium anisopliae, J. Invertebr. Pathol. 60, 89–101 (1992).

    Article  Google Scholar 

  22. M. J. Bidochka, R. J. St. Leger, and D. W. Roberts, Differentiation of species and strains of entomopathogenic fungi by random amplification of polymorphic DNA (RAPD), Curr. Genet. 25, 107–113 (1994).

    Article  PubMed  CAS  Google Scholar 

  23. G. H. Sung, J. W. Spatafora, R. Zare, K. T. Hodge, and W. Gams, A revision of Verticillium sect. Prostrata, II: Phylogenetic analysis of SSU and LSU nuclear rDNA sequences from anamorphs and teleomorphs of the Clavicipitaceae, Nova Hedwigia 72, 311–328 (2001).

    Google Scholar 

  24. F. Driver, R. J. Milner, and J. W. H. Trueman, A taxonomic revision of Metarhizium based on a phylogenetic analysis of rDNA sequence data, Mycol. Res. 104, 134–150 (2000).

    Article  CAS  Google Scholar 

  25. J. Fargues, Adhesion of the fungal spore to the insect cuticle in relation to pathogenicity, in Infection Processes of Fungi, Conference Report, edited by D. W. Roberts and J. R. Aist (Rockefeller Foundation, 1984), pp. 90–110.

    Google Scholar 

  26. R. J. St. Leger, M. J. Bidochka, and D. W. Roberts, Germination triggers of Metarhizium anisopliae, Microbiology 140, 1651–1660 (1994).

    Google Scholar 

  27. B. Amiri-Besheli, B. Khambay, S. Cameron, M. L. Deadman, and T. M. Butt, Inter- and intra-specific variation in destruxin production by insect pathogenic Metarhizium spp., and its significance to pathogenesis, Mycol. Res. 104, 447–452 (2000).

    Article  CAS  Google Scholar 

  28. T. M. Butt and M. Brownbridge, Increasing the efficacy of entomogenous fungi, in Enhancing Biocontrol Agents and Handling Risks, edited by M. Vurro, J. Gressel, T. Butts, G. Harman, A. Pilgeram, R. J. St. Leger, and D. Nuss (IOS Press, Amsterdam, The Netherlands, 2001), pp. 52–63.

    Google Scholar 

  29. G. D. Inglis, M. S. Goettel, T. M. Butt, and H. Strasser, Use of hyphomycetous fungi for managing insect pests, in Fungi as Biocontrol Agents, edited by T. M. Butt, C. Jackson, and N. Magan (CAB International, Wallingford, UK, 2001), pp. 23–69.

    Google Scholar 

  30. J. Drummond, J. B. Heale, and A. T. Gillespie, Germination and effect of reduced humidity on expression of pathogenicity in Verticillium lecani, Ann. Appl. Biol. 111, 193–201 (1987).

    Article  Google Scholar 

  31. M. J. Bidochka, A. M. Kamp, T. M. Lavender, J. Dekoning, and J. N. A. De Croos, Habitat association in two genetic groups of the insect-pathogenic fungus Metarhizium anisopliae: Uncovering cryptic species? Appl. Environ. Microbiol. 67, 1335–1342 (2001).

    Article  PubMed  CAS  Google Scholar 

  32. D. E. N. Rangel, G. U. L. Braga, A. J. Anderson, and D. W. Robert, Influence of growth environment on tolerance to UV-B radiation, germination speed, and morphology of Metarhizium anisopliae var. acridum conidia, J. Invetebrate Pathol. 90, 55–58 (2005).

    Article  Google Scholar 

  33. G. U. L. Braga, D. E. N. Rangel, S. D. Flint, A. J. Anderson, and D. W. Roberts, Conidial pigmentation is important to tolerance against solar-simulated radiation in the entomopathogenic fungus Metarhizium anisopliae, Photochem. Photobiol. 82, 418–422 (2006).

    Article  PubMed  CAS  Google Scholar 

  34. H. De Grotte, O. Douro-Kpindou, Z. Ouambama, C. Gbongboui, D. Müller, S. Attignon, and Chris Lomer, Assessing the feasibility of biological control of locusts and grasshoppers in West Africa: Incorporating the farmers’ perspective, Agricu. Human Values 18, 413–428 (2001).

    Article  Google Scholar 

  35. Z. Long and D. M. Hunter, Laboratory and field trials of Green Guard® (Metarhizium anisopliase var. acridium) (Deuteromycotina: Hyphomycetes) against the oriental migratory locust (Locusta migratoria manilensis) (Orthoptera: Acrididae), China 14, 27–30 (2005).

    Google Scholar 

  36. E. J. Scholte, K. Ng’habi, J. Kihonda, W. Takken, K. Paaijmans, S. Abdulla, G. F. Killeen and B. G. J. Knols, An entomopathogenic fungus for control of adult African malaria mosquitoes, Science 308, 1641–1642 (2005).

    Article  PubMed  CAS  Google Scholar 

  37. S. Blanford, B. H. K. Chan, N. Jenkins, D. Sim, R. J. Turner, A. F. Read, and M. B. Thomas, Fungal pathogen reduces potential for malaria transmission, Science 308, 1638–1641 (2005).

    Article  PubMed  CAS  Google Scholar 

  38. A. Flores, Saving bees: Fungus found to attack Varroa mites, Agric. Res. Mag. 52, 18 (2004).

    Google Scholar 

  39. M. S. Goettel, and G. D. Inglis, Fungi: Hyphomycetes, in Manual of Techniques in Insect Pathology, edited by L. Lacey (Kluwer Academic Publishers, 1997), pp. 213–249.

    Google Scholar 

  40. M. J. Bidochka, Monitoring the fate of biocontrol fungi, inFungal Biocontrol Agents: Progress, Problems and Potential, edited by T. M. Butt, C. Jackson, and N. Morgan (CAB International, Wallingford, UK, 2001), pp. 193–218.

    Google Scholar 

  41. K. P. Barley, The configuration of the root system in relation to nutrient uptake, Adv. Agron. 22, 159–201 (1970).

    Article  Google Scholar 

  42. T. F. Duda and S. R. Palumbi, Molecular genetics of ecological diversification: Duplication and rapid evolution of toxin genes of the venomous gastropod Conus, Proc. Natl. Acad. Sci. 96, 6820–6823 (1999).

    Article  PubMed  CAS  Google Scholar 

  43. G. E. Harman and B. G. G. Dozelli, Enhancing crop performance and pest resistance with genes from biocontrol agents, in Enhancing Biocontrol Agents and Handling Risks, edited by M. Vurro, J. Gressel, T. Butts, G. Harman, A. Pilgeram, R. St. Leger, and D. Nuss (IOS Press, Amsterdam, The Netherlands, 2001), pp. 114–125.

    Google Scholar 

  44. J. M. Whipps, Microbial interactions and biocontrol in the rhizosphere, J. Exp. Bot. 52, 487–511 (2001).

    PubMed  CAS  Google Scholar 

  45. G. Hu and R. J. St. Leger, Field studies of a recombinant mycoinsecticide (Metarhizium anisopliae) reveal that it is rhizosphere competent. Appl. Environ. Microbiol. 68, 6383–6387 (2002).

    Article  PubMed  CAS  Google Scholar 

  46. R. J. St. Leger and S. Screen, Prospects for strain improvement of fungal pathogens of insects and weeds, in Fungal Biocontrol Agents: Progress, Problems and Potential, edited by T. M. Butt, C. Jackson, and N. Morgan (CAB International, Wallingford, UK, 2001), pp. 219–238.

    Google Scholar 

  47. R. J. St. Leger, M. J. Bidochka, and D. W. Roberts, Co-transformation of Metarhizium anisopliae by electroporation or the gene gun to produce stable GUS transformants, FEMS Microbiol. Lett. 131, 289–294 (1995).

    Article  Google Scholar 

  48. B. R. Kerry, Rhizosphere interactions and the exploitation of microbial agents for the biological control of plant-parasitic nematodes, Annu. Rev. Phytopathol. 38, 423–441 (2000).

    Article  PubMed  CAS  Google Scholar 

  49. R. Baker, Diversity in biological control, Crop Protection 10, 85–94 (1991).

    Article  Google Scholar 

  50. D. W. Roberts and A. E. Hajek, Entompathogenic fungi as bioinsecticides, in Frontiers in Industrial Mycology, edited by G. F. Leatham (Chapman & Hall, New York, 1992), pp. 144–159.

    Google Scholar 

  51. J. M. Barea, M. J. Pozo, R. Azcon, and C. Azcon-Aguilar, Microbial co-operation in the rhizosphere, J. Exp. Bot. 56, 1761–1778 (2005).

    Article  PubMed  CAS  Google Scholar 

  52. E. B. Nelson, Microbial dynamics and interactions in the spermosphere, Annu. Rev. Phytopathol. 42, 271–309 (2004).

    Article  PubMed  CAS  Google Scholar 

  53. P. Liang and A. B. Pardee, Analyzing differential gene expression in cancer, Nat. Rev. Cancer 3, 869–876 (2003).

    Article  PubMed  CAS  Google Scholar 

  54. Y. Ben-Shahar, A. Robichon, M. B. Sokolowski, and G. E. Robinson, Influence of gene action across different time scales on behavior, Science 296, 741–744 (2002).

    Article  PubMed  CAS  Google Scholar 

  55. W. Enard, P. Khaitovich, J. Klose, S. Zollner, F. Heissig, P. Giavalisco, K. Nielset-Struwe, E. Muchmore, A. Varki, R. Ravid, G. M. Doxiadis, R. E. Bontrop, and S. Paabo, Intra and inter specific variation in primate gene expression patterns, Science 296(5566), 233–235 (2002).

    Article  Google Scholar 

  56. R. B. Brem, G. Yvert, R. Clinton, and L. Kruglyak, Genetic dissection of transcriptional regulation in budding yeast, Science 296, 752–755 (2002).

    Article  PubMed  CAS  Google Scholar 

  57. T. L. Ferea, D. Botstein, P. O. Brown, and R. F. Rosenzweig, Systematic changes in gene expression patterns following adaptive evolution in yeast, Proc. Natl. Acad. Sci. 96, 9721–9726 (1999).

    Article  PubMed  CAS  Google Scholar 

  58. J. P. Townsend, D. Cavalieri, and D. L. Hartl, Population genetic variation in genome-wide gene expression, Mol. Biol. Evol. 20, 955–963 (2003).

    Article  PubMed  CAS  Google Scholar 

  59. M. V. Olson, When less is more: Gene loss as an engine of evolutionary change, Am. J. Hum. Genet. 64, 18–23 (1999).

    Article  PubMed  CAS  Google Scholar 

  60. G. Hu and R. J. St. Leger, A phylogenomic approach to reconstructing the diversification of serine proteases in fungi, J. Evol. Biol. 17, 1204–1214 (2004).

    Article  PubMed  CAS  Google Scholar 

  61. R. H. Waterston, K. Lindblad-Toh, E. Birney, et al., Initial sequencing and comparative analysis of the mouse genome, Nature 420, 520–562 (2002).

    Article  PubMed  CAS  Google Scholar 

  62. D. G. Boucias and J. C. Pendland, Detection of protease inhibitors in the hemolymph of resistant Anticarsia gemmatalis inhibitory to the entomopathogenic fungus Nomuraea rileyi, Experientia 43, 336–3439 (1997).

    Article  Google Scholar 

  63. S. Bagga, S. E. Screen, and R. J. St. Leger, Reconstructing the diversification of subtilisins in the pathogenic fungus Metarhizium anisopliae. Gene, 324, 159–169 (2004).

    Article  PubMed  CAS  Google Scholar 

  64. C. B. Michielse, P. J. Hooykaas, C. A. van den Hondel, and A. F. Ram, Grobacterium-mediated transformation as a tool for functional genomics in fungi, Curr. Genet. 48, 1–17 (2005).

    Article  PubMed  CAS  Google Scholar 

  65. L. Hoyer, The ALS gene family of Candida albicans, Trends Microbiol. 9, 176–180 (2001).

    Article  PubMed  CAS  Google Scholar 

  66. K. Al-Aidroos and D. W. Roberts, Mutants of Metarhizium anisopliae with increased virulence toward mosquito larvae, Can. J. Genet. Cytol. 20, 211–220 (1978).

    Google Scholar 

  67. A. K. Charnley, Fungal pathogens of insects: cuticle degrading enzymes and toxins, Adv. Bot. Res. 40, 241–321 (2003).

    Article  CAS  Google Scholar 

  68. R. J. St. Leger, Biology and mechanisms of invasion of deuteromycete fungal pathogens, in: Parasites and Pathogens of Insects, edited by N. C. Beckage, S. N. Thompson, and B. A. Federici (Academic Press, New York, 1993), vol. 2, pp. 211–229.

    Google Scholar 

  69. R. J. St. Leger, R. M. Cooper, and A. K. Charnley, The effect of melanization of Manduca sexta cuticle on growth and infection by Metarhizium anisopliae, J. lnvertebr. Pathol. 52, 459–470 (1988).

    Article  Google Scholar 

  70. C. E. Gongora, Transformacion de Beauveria bassiana cepa Bb9112 con les genes de la proteina verde fluorescente y la protease pr1A de M. anisopliae, Rev. Colomiana Entomol. 30, 1–5 (2004).

    Google Scholar 

  71. A. Flores, I. Chet, and A. Herrera Estrella, Improved biocontrol activity of Trichoderma harzianum by overexpression of the proteinase encoding gene, prb1, Curr. Genet. 31, 30–37 (1997).

    Article  PubMed  CAS  Google Scholar 

  72. J. Ahman, T. Johansson, M. Olsson, P. J. Punt, A. M. J. Cees, van den Hondel, and A. Tunlid, Improving the pathogenicity of a nematode-trapping fungus by genetic engineering of a subtilisin with nematotoxic activity, Appl. Environ. Microbiol. 68, 3408–3415 (2002).

    Article  PubMed  CAS  Google Scholar 

  73. A. C. Rath, The use of entomopathogenic fungi for control of termites, Biocont. Sci. Technol. 10, 563–581 (2000).

    Article  Google Scholar 

  74. M. G. Villani, S. R. Krueger, P. C. Schroeder, F. Consolie, N. H. Console, L. M. Preston-Wisey, and D. W. Roberts, Soil application effects of Metarhizium anisopliae on Japanese beetle (Coleoptera: Scarabaeidae) behavior and survival in turfgrass microcosms, Environ. Entomol. 23, 502–513 (1994).

    Google Scholar 

  75. R. L. Samuels, S. E. Reynolds, and A. K. Charnley, Calcium-channel activation of insect muscle by destruxins, insecticidal compounds produced by the entomopathogenic fungus Metarhizium anisopliae. Comp. Biochem. Physiol. C 90, 403–412 (1988).

    Google Scholar 

  76. Vey, A., Matha, V., and Dumas, C. Effects of the peptide mycotoxin destruxin E on insect hemocytes and on dynamics and efficiency of the multicellular immune reaction, J. Invertebr. Pathol. 80, 177–187 (1988).

    Article  CAS  Google Scholar 

  77. S. Pal, R. J. St. Leger, and L. P. Wu, Fungal peptide destruxin A plays a specific role in suppressing the innate immune response in Drosophila melanogaster, J. Biol. Chem. (2007) in press.

    Google Scholar 

  78. E. Zlotkin, Y. Fishman, M. Elazer, AaIT: From neurotoxin to insecticide, Biochemie 82, 869–881 (2000).

    Article  CAS  Google Scholar 

  79. X. Sun, X. Chen, Z. Zhang, H. Wang, F. J. Bianchi, H. Peng, J. M. Vlak, and Z. Hu, Bollworm responses to release of genetically modified Helicoverpa armigera nucleopolyhedroviruses in cotton, J. Invertebre. Pathol. 81, 63–69 (2002).

    Article  Google Scholar 

  80. M. Isaka, P. kittakoop, K. Kirtikra, N. Hywel-Jones, and Y. Thebtaranonth, Bioactive substances from insect pathogenic fungi, Acc. Chem. Res. 38, 813–823 (2005).

    Article  PubMed  CAS  Google Scholar 

  81. C. Wang and R. J. St. Leger, Developmental and transcriptional responses to host and non hostcuticles by the specific locust pathogen Metarhizium anisopliae sf. Acridum, Eukaryotic Cell 4, 937–947 (2005).

    Article  PubMed  CAS  Google Scholar 

Download references

Author information

Authors and Affiliations

Authors

Editor information

Editors and Affiliations

Rights and permissions

Reprints and permissions

Copyright information

© 2007 Springer

About this paper

Cite this paper

Leger, R.J. (2007). METARHIZIUM ANISOPLIAE AS A MODEL FOR STUDYING BIOINSECTICIDAL HOST PATHOGEN INTERACTIONS. In: Vurro, M., Gressel, J. (eds) Novel Biotechnologies for Biocontrol Agent Enhancement and Management. NATO Security through Science Series. Springer, Dordrecht. https://doi.org/10.1007/978-1-4020-5799-1_9

Download citation

Publish with us

Policies and ethics