Entomopathogenic Fungi as Potent Agents of Biological Control

Text-only Preview


International Journal of Engineering and Technical Research (IJETR)
ISSN: 2321-0869, Volume-2, Issue-3, March 2014
Entomopathogenic Fungi as Potent Agents of
Biological Control
Saba Hasan

constraints include the slow efficacy in comparison to the
Abstract-- The various risk factors associated with the use of
chemicides and on-field exposure to various biotic and abiotic
chemical insecticides such as development of resistance,
stresses. EPFs have significantly higher host specificity in
associated resurgence in insects, accumulation of pesticidal
comparison to the conventional biocontrol agents like
residues in food chain, environmental pollution, health risks and
bacteria, protozoa, nematodes, predatory insects and viruses.
high costs have led to development of alternative strategies of
They are unique when compared to other microbes causing
pest management, thus necessitating interest on the search for
biological control agents that can control destructive pests of

diseases in insects because they cause infection by growing
crops. Entomogenous fungi are potentially the most versatile
through the insect cuticle and not required to be ingested, thus
biological control agents due to their wide host range. These
showing great potential for control of even sucking insect
fungi comprise a diverse group of over 100 genera with
pests. An attractive feature of these fungi is that infectivity is
approximately 750 species, reported from different insects.
by contact and the action is through penetration (Nadeau et
Entomopathogenic fungi vary considerably in their mode of
al.,1996). Many of these offer a great potential in pest
action and virulence. The present review gives a brief picture of
management. The most prominent fungal pathogens are
environmental factors affecting pathogenicity as well as mode of
Metarhizium spp., Beauveria spp., Nomuraea rileyi,
virulence of entomopathogenic fungi with special mention to
Verticillium lecanii and Hirsutella spp. living in diverse
their virulence factors involved in pathogenesis

habitats including fresh water, soil surfaces and aerospaces
Index Terms-- Entomopathogenic, Virulence, Biological
(Hajek and Ledger, 1994). They belong to Zygomycotina,
control, Pathogenicity
Ascomycotina, Basidiomycotina and Deuteromycotina.

Among 85 genera of entomopathogenic fungi only six species
are commercially available for field application. However,
I. INTRODUCTION
comparatively few have been investigated as potential
The increased use of conventional chemical pesticides over
mycoinsecticides. Fungal pathogens particularly B. bassiana,
the years has contributed significantly to an increase in food
I. fumosorosea and M. anisopliae are being evaluated against
production, but on the other hand, has resulted in unfavorable
numerous agricultural and urban insect pests. Several species
effects like resistance, pest resurgence, environmental
belonging to order IIsoptera (Hussain et al., 2010a ; Hussain
pollution and risks to human health on the environment and
et al., 2011), Lepidoptera (Hussain et al., 2009), Coleoptera
non-target organisms. Henceforth, the necessity for
(Ansari et al., 2006), Hemiptera (Leite et al., 2005) and
sustainable crop production through eco-friendly pest
Diptera (St. Leger et al., 1987) are susceptible to various
management technique is being largely felt in recent times.
fungal infections. This has led to a number of attempts to use
The use of bio-control agents is considered as a suitable
entomopathogenic fungi for pest control with varying degrees
alternative to the use of chemical pesticides (Dhaliwal and
of success.
Koul, 2007). These biological control agents like bacteria,

viruses, protozoa, nematodes and most fungi exert
considerable control of target populations.
II. ORIGIN
Among micro-organisms, entomopathogenic fungi constitute
The origin of the entomopathogenic lifestyle may have arisen
the largest single group of insect pathogens. Entomogenous
several times from a common saprophytic ancestor inhabiting
fungi are potentially the most versatile biological control
soil and leaf litter (Spatafora and Blackwell, 1993). The
agents. They belong to the orders Entomophthorales and
biggest emission into different host groups occurred within
Hypocreales
(formerly

called
Hyphomycetes).
Clavicipitaceae (Ascomycotina), and involved multiple
Entomopathogenic fungi from other taxonomic groups are
inter-kingdom jumps between animals (e.g. insects), fungi
also known. Until now, over 700 species of fungi are known
and plants (Nikoh and Fukatsu, 2000; Artjariyasripong et al.,
to infest insects (Wraight et al, 2007).
2001; Spatafora et al., 2007). Hyphomycete species exist as

separate asexual (anamorph) and sexual (teleomorph) forms.
The advantages of EPFs over chemical pesticides are their
Entire of the known genera of entomopathogenic
significantly higher host specificity, the reduction of hazards
Hyphomycetes,
now
proven
teleomorphs
in
the
and the inability of the insects to develop resistance as the
Clavicipitales, and life cycle stages for Hyphomycetes may
EPFs simultaneously use several modes of actions and as a
have become simplified in agricultural situations because of a
"living-pesticide" is subjected to adaptation too. The
superabundance of insect hosts (Evans 2003).


Manuscript received March 20, 2014.
Dr Saba Hasan, Amity Institute of Biotechnology, Amity University,
Lucknow (U.P.), India, Mobile No. : +91-8756858224
234 www.erpublication.org

Entomopathogenic Fungi as Potent Agents of Biological Control

III. ENVIRONMENTAL INFLUENCE ON NATURAL
Pathogenesis is the process of chain of events in the disease
PATHOGENICITY
development in a host upon infection. Unlike bacteria and

viruses, fungal pathogenesis in insects occurs via a series of
Environmental factors which influence the virulence of
systematically integrated events progressing upon spore
entomopathogens must be considered for the successful
attachment to germination, penetration, growth and
development of the fungus as a biocontrol agent. A wide
proliferation within the body of the host, interaction with
range of factors such as water, ions, fatty acids, nutrients on
insect defense mechanism and finally re-emergence on the
the cuticle surface and the physiological state of the host,
cadavers (Nadeau et al., 1996; Thomas et al., 1996).
influence spore germination and behavior (Hassan et al.,
The cuticle is the first barrier to infection by fungi. Hence,
1989). Environmental conditions, especially temperature and
rapid and direct penetration of the cuticle is important for
humidity, have long been recognized to play a significant role
virulence (Pekrul and Grula, 1979). The insect procutide is
in the incidence of epizootics of insect pathogens (Benz
primarily chitin fibrils embedded in a protein matrix and
1987). For example, Daoust and Pereira (1986) demonstrated
penetration involves both mechanical and enzymatic
that temperature and humidity affect both survival and
components (Charnley and St. Leger, 1989). Penetration is a
germination rates in B. bassiana.
stage of infection where specificity may be determined since,
Of all the ecofactors that influence epizootics of a
many pathogens are virulent after being injected into the
mycopathogen, high humidity is most critical for sporulation,
haemolymph of an otherwise nonsusceptible host. In
germination and invasion of the host (>90% RH) (Getzin,
terrestrial environment, fungal conidial germination proceeds
1961). Pathogenesis occurs at much lower ambient values
with the formation of germtube (Boucias & Pendland, 1991)
(Ramoska, 1982) probably because of high humidity in the
or appressorium (Zacharuk, 1970a),which forms a thin
microclimate at the insect cuticle. However, the external
penetration peg that breaches the insect cuticle via mechanical
sporulation never occurs on the killed insect, if the relative
(turgor pressure) or enzymatic means (proteases) (Zacharuk,
humidity is too low. Milner et al. (2002) studied the effect of
1970b). In M. anisopliae, appressorium formation,
relative humidities (RH) from 90 to 100 percent on
hydrophobins, and the expression of cuticle-degrading
germination of a termite-active isolate of M. anisopliae
proteases are triggered by low nutrient levels (St. Leger et al,
(isolate F125 and FI610) using a liquid germinating medium.
1992), demonstrating that the fungus senses environmental
Germination was found to be delayed at water activities
conditions or host cues at the initiation of infection.
equivalent to 99, 98 and 96 per cent RH and was completely
The production of cuticle-degrading enzymes, chitinases,
inhibited at 94, 92 and 90 percent. Although much higher
lipases and proteases, has been
humidity is required for conidial germination, B. bassiana
recognized since long as an important determinant of the
infections have been shown to occur at ambient humidities as
infection process in various fungi, facilitating penetration as
low as 50%, probably due to micro environmental conditions
well as providing nourishment for further development
on the insect cuticle (Ramoska 1984, Marcandier and
(Hussain et al, 2010b). Among the proteases found in
Khachatourians 1987b).
entomopathogenic fungi, the spore bound Pr1 has been well

characterized and its role in cuticle invasion has been
The optimum temperature for the development of the fungus
established (Hussain et al, 2010b; St. Leger, 1994). Ultra
is not necessarily the same for development of the disease.
structural studies of M. anisopliae penetration sites on
However, temperature has a profound influence on the host
Manduca sexta larvae have shown high levels of Pr1
insect and hence it must be taken into consideration, since
coincident with hydrolysis of cuticular proteins (St. Leger et
very short periods between moults resulting from a high
al, 1989). Furthermore, it has also been reported that
temperature may reduce, for example, the duration of the
successive in vivo passage enhanced the capacity of the
instar to an extent that penetration of the fungus through the
fungus to cause infection (Daoust et al, 1982; Hussain et al,
integument is impeded. The rapidity of mycelial development
2010b), which ultimately increased the activity of spore
and therefore, the rapidity of the evolution of infection
bound Pr1 (Shah et al, 2007). On the death of the insect host,
depends on temperature. Generally, optimum values fall
the fungus appears from the dead host and sporulation occurs
between 20C and 30C (for example, 23C for Beauveria
on the outside of the cadaver. Sporulation can occur internally
brongiartii, 24C for Entomophthora obscura, 25C for
when ambient humidity precludes external sporulation.
Beauveria bassiana and Nomuraea rileyi and 27C-28C for

Metarhizium anisopliae) with limits between 5 and 35C.
Temperatures lower than the optima distinctly retard the
V. ENTOMOPATHOGENIC FUNGI - A NEW
development of mycosis without necessarily affecting the
PARADIGM
total mortality (Ferron, 1978). If temperature affects

germination percentage in conidia, then it also affects the
Despite the publication of approximately 7,000 papers on
concentration of infective units in the field. For example,
topics related to entomopathogenic fungi since 1983 (Vega et
Inglis et al. (1997), found that the incidence of B. bassiana
al, 2009), there is still limited success in solving agricultural
infections decreased when acridids actively increased their
problems with entomopathogenic fungi. Few proposed
body temperature by habitat selection(sunning).
research areas mentioned below should lead to a new

paradigm for entomopathogenic fungi that should refocus our
efforts and hopefully lead to exciting new findings that will
IV. MODE OF ENTRY AND VIRULENCE OF
bring success to the field. Various unexpected roles have been
ENTOMOPATHOGENIC FUNGI
reported for fungal entomopathogens, including their

presence as fungal endophytes, plant disease antagonists,
235 www.erpublication.org


International Journal of Engineering and Technical Research (IJETR)
ISSN: 2321-0869, Volume-2, Issue-3, March 2014
rhizosphere colonizers and plant growth promoting fungi
REFERENCES
(Vega et al., 2009).


[1] G. S. Dhaliwal, and O. Koul, "Biopesticide and Pest Management:

Conventional and Biotechnological Approaches", Kalyani
Publishers, New Delhi, 2007, pp. 455.
A. Standardized Mass Production technology for fungal
[2] S.P. Wraight, G.D. Inglis, and M.S. Goettel, "Fungi, In: Field
biocontrol agents
manual of techniques in invertebrate pathology", L.A. Lacey &
H.K. Kaya, (Eds.), 2007, 223-248, 2nd edition, Springer,

Dordrecht, ISBN 978-1-4020-5931-5.
Recently, M. anisopliae was shown to be capable of
[3] M.P., Nadeau, G.B. Dunphy, and J.L., Boisvert, "Development of
producing sclerotia in liquid culture fermentation (Jaronski
Erynia conial (Zygomycetes Entomophthorales) on the cuticle of
and Jackson, 2008). The ability of M. anisopliae to form
the adult black flies Simulium rostratum and Simulium decorum
(Diptera Simuliidae)", Journal of Invertebrate Pathology, 1996,
sclerotia may be important for rhizosphere competence
68: 50-58.
following a pattern seen in phytopathogenic fungi. The use of
[4] A.E. Hajek, and R.J., Ledger, "Interaction between fungal
sclerotial preparations for granular application of biocontrol
pathogens and insect hosts", Annual Review of Entomology,
agents like M. anisopliae in soil and the use of conidia or
1994, 39: 293-322.
[5] J.V., Maddox, "Insect pathogens as biological control agents. In
blastospores in foliar applications for phylloplane insects are
Introduction to Insect Pest Management" (Eds. Metcalf, R.L.,
examples of how the ecology of the fungus-insect interaction
Luckmann, W.H.). John Wiley and Sons Inc., Publication, New
directs the production and use of appropriate infective
York, 1994, pp.199-244.
propagules.
Likewise,
the
use
of
endophytic
[6] J. W., Spatafora, and M. Blackwell, "Molecular systematic of
unitunicate
perithecia
ascomycetes:
the
Clavicipitales-
entomopathogenic fungi for insect control will require an
Hypocreales connection", Mycologia, 1993, 85, 912-922.
understanding of the ecological factors that enhance the
[7] N., Nikoh, and T. Fukatsu, "Interkingdom host jumping
ability of the fungus to become endophytic. Awareness of
underground: phylogenetic analysis of entomoparasitic fungi of
these ecological factors will aid in the development of
the genus Cordyceps". Mol Biol Evol, 2000, 17, 629-638.
[8] S., Artjariyasripong, J. I., Mitchell, N., L., Hywel-Jones, and
production and formulation technologies that deliver
E.B.G. Jones, "Relationship of the genus Cordyceps and related
optimally infective fungal propagules.
genera based on parsimony and spectral analysis of partial 18S

and 28S ribosomal gene sequences", Mycoscience, 2001, 42,

503-517.
[9] J. W., Spatafora, G. H., Sung, J. M., Sung, J. N. L., Hywel, and J.F.
B. Formulation of fungal propagules
White Jr. "Phylogenetic evidence for an animal pathogen origin
of ergot and the grass endophytes", Molecular Ecology, 2007,

16, 1701-1711.
The formulation of propagules of fungal entomopathogenic
[10] H. C. Evans, "Use of Clavicipitalean fungi for the biological
fungi for use in biocontrol has been guided by the need to
control of arthropod pests", In: White JF, et al. (eds)
improve product shelf-life, biocontrol efficacy, and/or the
Clavicipitalean fungi. Marcel Dekker, New York, 2003.
[11] J.

Eilenberg, "Biology of fungi from the order
physical characteristics of the product for application
Entomophthorales, with emphasis on the genera Entomophthora,
(Wraight et al., 2001). Progress in understanding the
Strongwellsea and Eryniopsis", The Royal Veterinary and
mechanisms of pathogenicity is now being made, particularly
Agricultural University, Copenhagen, Denmark, 2002.
in the area of cuticular penetration where the key enzyme is
[12] A.E.M., Hassan, R.J., Dillon and A.K. Charnley, J Invert
Pathol., 1989, 54 :227-279.
probably an endoprotease (St. Leger et al., 1986). To increase
[13] G. Benz, "Environment", In : J. R Fuxa and Y. Tanada [eds.],
the speed of insect kill, it may be possible in the future to
"Epizootiology of insect diseases", Wiley, New York, 1987, pp.
insert toxin genes from Bacillus thuringiensis into fungi.
177-214.
However, improvement in strains achieved by genetic
[14] R.A. Daoust and R.M. Pereira, "Survival of Beauveria bassiana
(Deuteromycetes : Moniliales) conidia on cadavers of cowpea
manipulation could be more useful. Future research should
pests stored outdoors and in the laboratory in Brazil", Environ.
concentrate on the development of formulation and the
Entomol. 1986, 15 : 642-647.
targeting of the pests of economically valuable crops. By
[15] L.W. Getzin, "Spicaria rileyi (Farlow) Charles, an
achieving this aim, agriculture will benefit from prolonged
entomogenous fungus of Trichoplusia ni (Hubner)", J Insect
Pathol.
1961, 3 : 2-10.
pest control, reduced risk of resistance and a high degree of
[16] R.J., Milner, P.R. Samson, and G.K., Bullard, "FI-1045 : A
safety to non-target organisms without disturbing the
profile of a commercially useful isolate of Metarhizium
agro-ecosystem and the quality of environment.
anisopliae var. anisopliae", Biocontrol Science and

Technology, 2002, 12: 43-58.
[17] W. A. Ramoska, "The influence of relative humidity on
Beallveria bas'slana infectivity and replication in the chinch
VI. FUTURE RESEARCH
bug, Blissus leucopterus", J. Invertebr. Pathol., 1984. 43:

389-394.
[18] S Marcandier and G G Khachatourians, "Evolution of relative
Entomopathogenic fungi being component of an integrated
humidity and temperature within a closed chamber used for
approach can provide significant and selective insect control.
entomological studies", Can. Entomol. 1987a, 119 : 893-900.
In the near future, we expect to see synergistic combinations
[19] P. Ferron, "Biological control of insect pests by entomogenous
of microbial control agents with other technologies (in
fungi", Annu. Rev. Entomol. , 1978, 23 : 409-442.
[20] D. G., Inglis, M. S. Goettel, and D. L. Johnson, "Field and
combination with semiochemicals, soft chemical pesticides,
laboratory evaluation of two conidial batches of Beuaveria
other natural enemies, resistant plants, chemigation, remote
bassiana (Balsamo) Vuillemin against grass hoppers", Can.
sensing, etc.) that will enhance the effectiveness and
Entomol., 1997, 129: 171-186.
sustainability of integrated control strategies.
[21] M.P., Nadeau, G.B. Dunphy, and J.L., Boisvert, "Development
of Erynia conica (Zygomycetes : Entomophthorales) on the

cuticle of the adult black flies Simulium rostratum and Simulium
decorum
(Diptera : Simuliidae)", Journal of Invertebrate
Pathology
, 1996, 68: 50-58.
236 www.erpublication.org

Entomopathogenic Fungi as Potent Agents of Biological Control

[22] M.B., Thomas, C. Cbongboui, and C.J., Lomer, "Between
[40] A., Hussain, S. Ahmed, and M. Shahid, "Laboratory and field
season survival of the grasshopper pathogen Metarhizium
evaluation of Metarhizium anisopliae var. anisopliae for
flavoviride in the Sahel", Biocontrol Science Technology, 1996,
controlling subterranean termites", Neotropical Entomology,
6: 569-573.
2011, Vol.40, No.2 pp. 244-250, ISSN 1519-566X
[23] S. Pekrul, and E.A., Grula, "Mode of infection of the corn
[41] A., Hussain, M.Y., Tian, Y.R. He, and S. Ahmed,
earworm (Heliothis zea) by Beauveria bassiana as revealed by
"Entomopathogenic Fungi disturbed the larval growth and
scanning electron microscopy", Journal of Invertebrates
feeding performance of Ocinara varians Walker (Lepidoptera:
Pathology, 1979, 34: 228-247.
Bombycidae) Larvae", Insect Science, 2009, Vol.16, No.6, pp.
[24] A.K. Charnley, and R.J., St. Leger, "The role of cuticle degrading
511-517, ISSN 1672-9609.
enzymes in fungal pathogenesis in insects", In: The Fungal Spore
[42] M.A., Ansari, F.A. Shah, L. Tirry, & M. Moens, "Field trials
and Disease Initiation in Plants and Animals, eds. G.T. Cole and
against Hoplia philanthus (Coleoptera: Scarabaeidae) with a
H.C. Kock, Plenum Press, New York, USA, 1989, pp. 267-286.
combination of an entomopathogenic nematode and the fungus
[25] G.N., El-Sayed, C.M., Ignoffo, T.B., Leathers, and G.N.EL.,
Metarhizium anisopliae CLO 53", Biological Control, 2006,
Sayed, "A semidefined medium for culturing Nomurea rileyi",
Vol.39, No.3, pp. 453-459, ISSN 1049-9644.
Mycopathologia, 1992, 118: 163-165.
[43] L.G., Leite, S.B, Alves, A.B., Filho and D.W. Roberts, "Simple,
[26] D.G. Boucias, and J.C. Pendland, "Attachment of
inexpensive
media
for
mass
production
of
three
mycopathogens to cuticle: the initial event of mycoses in
entomophthoralean fungi", Mycological Research, 2005,
arthropod hosts", In: The fungal spore and disease initiation in
Vol.109, No.3, pp. 326-334, ISSN 0953-7562.
plants and animals. G.T. Cole, H.C. Hoch. (Eds.), Plenum Press,
[44] R.J., St. Leger, R.M. Cooper, and A.K. Charnley, "Production of
New York, 1991, pp. 101-127.
cuticle-degrading enzymes by the entomopathogen Metarhizium
[27] R.Y. Zacharuk, "Fine structure of the fungus Metarhizium
anisopliae during infection of cuticles from Calliphora
anisopliae infecting three species of larval Elateridae
vomitoria and Manduca sexta", Microbiology, 1987, Vol.133,
(Coleoptera): III. Penetration of the host integument", Journal of
No.5, pp. 1371-1382, ISSN 1350-0872.
Invertebrate Pathology, 1970b, Vol.15, 372-396, ISSN

0022-2011.
[28] R.J., St Leger, D.C., Frank, D.W. Roberts, and R.C., Staples,

"Molecular cloning and regulatory analysis of the
cuticle-degrading
protease
structural
gene
from
the
entomopathogenic fungus Metarhizium anisopliae", European
Journal of
Biochemical, 1992, 204: 991-1001.
[29] A., Hussain, M.Y. He, Tian, Y.R. Bland, J.M. & W.X. Gu,
"Behavioral and electrophysiological responses of C. formosanus
towards entomopathogenic fungal volatiles", Biological Control,
2010a, Vol.55, pp. 166-173, ISSN 1049-9644.
[30] A., Hussain, M.Y. He, Y.R. Tian, and R. Lin, "In vitro and in
vivo culturing impacts on the virulence characteristics of serially
passed entomopathogenic fungi", Journal of Food Agriculture
& Environment
, 2010b, Vol.8, No.3&4, pp. 481-487, ISSN
1459-0255
[31] R.J. St. Leger, "The role of cuticle degrading proteases in fungal
pathogenesis of Insects", Canadian Journal of Botany, 1994,
Vol.73 (Suppl. 1), pp. 1119-1125, ISSN 1480-3305.
[32] R.J., St. Leger, T.M. Butt, R.C. Staples, and D.W. Roberts,
"Synthesis of proteins including a cuticle-degrading protease
during differentiation of the entomopathogenic fungus
Metarhizium anisopliae", Experimental Mycology, 1989,
Vol.13, pp. 253-262, ISSN 0147-5975.
[33] R.A., Daoust, M.G. Ward, and D.W. Roberts, "Effect of
formulation on the virulence of Metarhizium anisopliae conidia
against mosquito larvae", Journal of Invertebrate Pathology,
1982, Vol.40, pp. 228-236, ISSN 0022-2011.
[34] F.A., Shah, N. Allen, C.J. Wright, and T.M. Butt, "Repeated in
vitro subculturing alters spore surface properties and virulence of
Metarhizium anisopliae", FEMS Microbiology Letters, 2007,
Vol.276, pp. 60-66, ISSN 0378-1097.
[35] F. E., Vega, M. S., Goettel, M., Blackwell, D., Chandler, M. A.,
Jackson, S., Keller, M., Koike, N. K., Maniania, A., Monzon,
B.H., Ownley, J. K., Pell, D. E. N., Rangel, and H. E. Roy,
"Fungal entomopathogens: new insights on their ecology" 2009,
http://dx.doi.org/10.1016/j.funeco.2009.05.001.
[36] S.T., Jaronski, M.A., Jackson, " Efficacy of Metarhizium
anisopliae microsclerotial Granules", Biocontrol Science and
Technology
, 2008, 18: 849-863.
[37] S.P., Wraight, M.A. Jackson, S.L.,de Kock, " Production,
stabilization, and formulation of fungal biocontrol agents", In:
Butt TM, Jackson CW, Magan N (Eds.), Fungi as Biocontrol
Agents: Progress, Problems and Potential. CABI Publishing,
Wallingford, United Kingdom, 2001, pp. 253-288.
[38] R. J., St. Leger, R. M., Cooper, and A.K. Charnley, "Cuticle
degrading enzymes of entomopathogenic fungi: Cuticle
degradation in vitro by enzymes from entomopathogens",
Journal of Invertebrate Pathology, 1986, 47,167-177.
[39] A., Hussain, M.Y. He, Y.R. Tian, J.M. Bland, and W.X. Gu,
"Behavioral and electrophysiological responses of C. formosanus
towards entomopathogenic fungal volatiles", Biological Control,
2010a, Vol.55, pp. 166-173, ISSN 1049-9644.

237 www.erpublication.org