Genetic engineering of fungal biocontrol agents to achieve greater efficacy against insect pests

Molecular biology methods have elucidated pathogenic processes in several fungal biocontrol agents including two of the most commonly applied entomopathogenic fungi, Metarhizium anisopliae and Beauveria bassiana. In this review, we describe how a combination of molecular techniques has: (1) identified and characterized genes involved in infection; (2) manipulated the genes of the pathogen to improve biocontrol performance; and (3) allowed expression of a neurotoxin from the scorpion Androctonus australis. The complete sequencing of four exemplar species of entomopathogenic fungi including B. bassiana and M. anisopliae will be completed in 2010. Coverage of these genomes will help determine the identity, origin, and evolution of traits needed for diverse lifestyles and host switching. Such knowledge combined with the precision and malleability of molecular techniques will allow design of multiple pathogens with different strategies to be used for different ecosystems and avoid the possibility of the host developing resistance.

[1]  D. Frank,et al.  Molecular cloning and regulatory analysis of the cuticle-degrading-protease structural gene from the entomopathogenic fungus Metarhizium anisopliae. , 1992, European journal of biochemistry.

[2]  N. Rizzo,et al.  Biochemical characterization and ultrastructural localization of two extracellular trypsins produced by Metarhizium anisopliae in infected insect cuticles , 1996, Applied and environmental microbiology.

[3]  R. S. St. Leger,et al.  Construction of an improved mycoinsecticide overexpressing a toxic protease. , 1996, Proceedings of the National Academy of Sciences of the United States of America.

[4]  N. Rizzo,et al.  Characterization and Ultrastructural Localization of Chitinases from Metarhizium anisopliae, M. flavoviride, and Beauveria bassiana during Fungal Invasion of Host (Manduca sexta) Cuticle , 1996, Applied and environmental microbiology.

[5]  A. Hajek.  Pathology and Epizootiology of Entomophaga maimaiga Infections in Forest Lepidoptera , 1999, Microbiology and Molecular Biology Reviews.

[6]  R. S. St. Leger,et al.  Cloning, Expression, and Substrate Specificity of a Fungal Chymotrypsin , 2000, The Journal of Biological Chemistry.

[7]  M. Elazar,et al.  AaIT: from neurotoxin to insecticide. , 2000, Biochimie.

[8]  N. Magan,et al.  Prospects for strain improvement of fungal pathogens of insects and weeds. , 2001 .

[9]  J. Gressel.  Potential failsafe mechanisms against the spread and introgression of transgenic hypervirulent biocontrol fungi. , 2001, Trends in biotechnology.

[10]  D. L. Johnson,et al.  Biological control of locusts and grasshoppers. , 2001, Annual review of entomology.

[11]  R. S. St. Leger,et al.  Field Studies Using a Recombinant Mycoinsecticide (Metarhizium anisopliae) Reveal that It Is Rhizosphere Competent , 2002, Applied and Environmental Microbiology.

[12]  C. Dumas,et al.  Effects of the peptide mycotoxin destruxin E on insect haemocytes and on dynamics and efficiency of the multicellular immune reaction. , 2002, Journal of invertebrate pathology.

[13]  P. Punt,et al.  Improving the Pathogenicity of a Nematode-Trapping Fungus by Genetic Engineering of a Subtilisin with Nematotoxic Activity , 2002, Applied and Environmental Microbiology.

[14]  R. Lim,et al.  Risks to the aquatic ecosystem from the application of Metarhizium anisopliae for locust control in Australia. , 2002, Pest management science.

[15]  J. Pell,et al.  Entomopathogenic fungi as biological control agents , 2003, Applied Microbiology and Biotechnology.

[16]  J. Baumgärtner,et al.  A field trial of the entomogenous fungus Metarhizium anisopliae for control of onion thrips, Thrips tabaci , 2003 .

[17]  R. S. St. Leger,et al.  Reconstructing the diversification of subtilisins in the pathogenic fungus Metarhizium anisopliae. , 2004, Gene.

[18]  B. Góngora,et al.  Transformación de Beauveria bassiana cepa Bb9112 con los genes de la proteína verde fluorescente y la proteasa pr1A de Metarhizium anisopliae , 2004 .

[19]  E. Quesada-Moraga,et al.  Bassiacridin, a protein toxic for locusts secreted by the entomopathogenic fungus Beauveria bassiana. , 2004, Mycological research.

[20]  N. Keyhani,et al.  Adhesion of the Entomopathogenic Fungus Beauveria (Cordyceps) bassiana to Substrata , 2005, Applied and Environmental Microbiology.

[21]  D. Sim,et al.  Fungal Pathogen Reduces Potential for Malaria Transmission , 2005, Science.

[22]  Chengshu Wang,et al.  Differential gene expression by Metarhizium anisopliae growing in root exudate and host (Manduca sexta) cuticle or hemolymph reveals mechanisms of physiological adaptation. , 2005, Fungal genetics and biology : FG & B.

[23]  D. Boucias,et al.  EST analysis of cDNA libraries from the entomopathogenic fungus Beauveria (Cordyceps) bassiana. II. Fungal cells sporulating on chitin and producing oosporein. , 2006, Microbiology.

[24]  B. Hammock,et al.  Genetically modified baculoviruses: a historical overview and future outlook. , 2006, Advances in virus research.

[25]  E. Quesada-Moraga,et al.  Insecticidal and antifeedant activities of proteins secreted by entomopathogenic fungi against Spodoptera littoralis (Lep., Noctuidae) , 2006 .

[26]  Yuehua Xiao,et al.  Increased Insect Virulence in Beauveria bassiana Strains Overexpressing an Engineered Chitinase , 2006, Applied and Environmental Microbiology.

[27]  Chengshu Wang,et al.  A collagenous protective coat enables Metarhizium anisopliae to evade insect immune responses. , 2006, Proceedings of the National Academy of Sciences of the United States of America.

[28]  Li Liu,et al.  EST analysis of cDNA libraries from the entomopathogenic fungus Beauveria (Cordyceps) bassiana. I. Evidence for stage-specific gene expression in aerial conidia, in vitro blastospores and submerged conidia. , 2006, Microbiology.

[29]  Chengshu Wang,et al.  A scorpion neurotoxin increases the potency of a fungal insecticide , 2007, Nature Biotechnology.

[30]  N. Keyhani,et al.  Phage display cDNA cloning and expression analysis of hydrophobins from the entomopathogenic fungus Beauveria (Cordyceps) bassiana. , 2007, Microbiology.

[31]  B. Hammock,et al.  Delivery methods for peptide and protein toxins in insect control. , 2007, Toxicon : official journal of the International Society on Toxinology.

[32]  A. Gatehouse,et al.  BIOTECHNOLOGY IN CROP PROTECTION: TOWARDS SUSTAINABLE INSECT CONTROL , 2007 .

[33]  J. Gressel,et al.  Novel biotechnologies for biocontrol agent enhancement and management , 2007 .

[34]  M. Goettel,et al.  Biological Control: A Global Perspective , 2007 .

[35]  Chengshu Wang,et al.  The Metarhizium anisopliae Perilipin Homolog MPL1 Regulates Lipid Metabolism, Appressorial Turgor Pressure, and Virulence* , 2007, Journal of Biological Chemistry.

[36]  Chengshu Wang,et al.  The MAD1 Adhesin of Metarhizium anisopliae Links Adhesion with Blastospore Production and Virulence to Insects, and the MAD2 Adhesin Enables Attachment to Plants , 2007, Eukaryotic Cell.

[37]  R. Leger.  METARHIZIUM ANISOPLIAE AS A MODEL FOR STUDYING BIOINSECTICIDAL HOST PATHOGEN INTERACTIONS , 2007 .

[38]  Chengshu Wang,et al.  MOS1 Osmosensor of Metarhizium anisopliae Is Required for Adaptation to Insect Host Hemolymph , 2007, Eukaryotic Cell.

[39]  M. Goettel,et al.  Green MuscleTM, a fungal biopesticide for control of grasshoppers and locusts in Africa. , 2007 .

[40]  Xingyong Yang,et al.  Directed evolution for increased chitinase activity , 2007, Applied Microbiology and Biotechnology.

[41]  A. Read,et al.  Fungal bioinsecticide with a sting , 2007, Nature Biotechnology.

[42]  Stephen P. Wraight,et al.  Mycoinsecticides and Mycoacaricides: A comprehensive list with worldwide coverage and international classification of formulation types ☆ , 2007 .

[43]  A. Read,et al.  Can fungal biopesticides control malaria? , 2007, Nature Reviews Microbiology.

[44]  N. Keyhani,et al.  Surface characteristics of the entomopathogenic fungus Beauveria (Cordyceps) bassiana. , 2007, Microbiology.

[45]  R. S. St. Leger,et al.  Fungal Peptide Destruxin A Plays a Specific Role in Suppressing the Innate Immune Response in Drosophila melanogaster* , 2007, Journal of Biological Chemistry.

[46]  Chengshu Wang,et al.  Increased pathogenicity against coffee berry borer, Hypothenemus hampei (Coleoptera: Curculionidae) by Metarhizium anisopliae expressing the scorpion toxin (AaIT) gene. , 2008, Journal of invertebrate pathology.

[47]  Y. Pei,et al.  Characterization of a Highly Active Promoter, PBbgpd, in Beauveria bassiana , 2008, Current Microbiology.

[48]  Chengshu Wang,et al.  Insecticidal evaluation of Beauveria bassiana engineered to express a scorpion neurotoxin and a cuticle degrading protease , 2008, Applied Microbiology and Biotechnology.

[49]  R. Leger.  Studies on adaptations of Metarhizium anisopliae to life in the soil , 2008 .

[50]  I. Glazer,et al.  Insect pathogens: molecular approaches and techniques. , 2009 .

[51]  N. Keyhani,et al.  Lectin mapping reveals stage-specific display of surface carbohydrates in in vitro and haemolymph-derived cells of the entomopathogenic fungus Beauveria bassiana. , 2009, Microbiology.

[52]  N. Keyhani,et al.  Uptake of the fluorescent probe FM4-64 by hyphae and haemolymph-derived in vivo hyphal bodies of the entomopathogenic fungus Beauveria bassiana. , 2009, Microbiology.

[53]  Chengshu Wang,et al.  Entomopathonic fungi and the genomics era. , 2009 .

[54]  Chengshu Wang,et al.  A phosphoketolase Mpk1 of bacterial origin is adaptively required for full virulence in the insect-pathogenic fungus Metarhizium anisopliae. , 2009, Environmental microbiology.

[55]  R. S. St. Leger,et al.  Protein kinase A regulates production of virulence determinants by the entomopathogenic fungus, Metarhizium anisopliae. , 2009, Fungal genetics and biology : FG & B.