McNamara, Louise (2016) Interactions of entomopathogenic fungi and other control agents: mechanism and field potential against Hylobius abietis larvae. PhD thesis, National University of Ireland Maynooth.
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Abstract
Biological control is the beneficial application of natural enemies such as
pathogens, predators and parasites in managing pests and their damage.
Entomopathogenic fungi (EPF) have a crucial role in natural ecosystems and are
being developed as alternative control agents for insect pests. Both Beauvaria
bassiana and Metarhizium anisopliae have been proven to be effective biological
control agents against a range of pests and are commercially produced. Advantages
of using EPF for biological pest control include their degree of specificity, absence
of effects on mammals, reduced probability of insects developing resistance and they
may persist for long periods in some environments which could provide long term
control effects. Disadvantages include that it takes EPF longer to kill insects than
their chemical counterparts, application needs to be timed for high relative humidity
and low pest numbers, and efficacy varies among different insect species.
If a combination of treatments resulted in a synergistic interaction then the
efficacy of these biopesticides would be increased. A number of laboratory and field
studies have used combinations of entomopathogenic nematodes (EPN) and EPF
against insect pests with resulting interactions ranging from antagonistic to
synergistic. In the case of synergism resulting from combined applications it is
suggested that EPF may make the host more susceptible through suppressing its
immune system. To understand how this putative synergistic interaction between
control agents could occur mechanistically, the effect of EPF supernatant was tested
on the immune response of the forestry pest, Hylobius abietis, to screen for species
with immunomodulating properties. The potential of the commonly used model
organism, the greater wax moth, Galleria mellonella, as a model for the study of the
immune response of H. abietis to pathogens was also explored.
Hylobius abietis, the large pine weevil, is a major pest of reforestation in Europe.
It is estimated that H. abietis costs the forestry industry approximately €140
million/year. Current control measures rely heavily on the synthetic chemical
cypermethrin. However, due to concerns over its environmental impact cypermethrin
is being phased out across Europe. Therefore there is an interest in the use of
entomopathogens as biological control agents in integrated pest management. Thus
one objective of this work was to assess the efficacy of EPF, EPN and EPF-EPN
combinations for H. abietis suppression in the field in order to ultimately determine
if the treatments used exert synergistic control over H. abietis.
The effect of EPF supernatant on the immune response of insects was assessed
through a number of bioassays that investigated the effect of EPF on haemocyte
densities and yeast proliferation in the insect haemocoel, as well as testing whether
pre-treatment with EPF increases larval susceptibility to subsequent pathogens. The
effect of EPF supernatant on the humoral immune response was investigated by
subjecting larval haemolymph to label free quantitative (LFQ) proteomic analysis.
To enable proteomic investigations into the effects of EPF on the immune response
of H. abietis and to compensate for the lack of genomic information for H. abietis; a
de novo transcriptome study of H. abietis larvae was performed with Beijing
Genomics Institute (BGI, Hong Kong).
Bioassays indicated that M. anisopliae, B. bassiana and Beauveria caledonica,
demonstrated immunomodulating effects on H. abietis larvae, while M. anisopliae
and B. caledonica modulated the immune response of G. mellonella making the
insect more susceptible to subsequent pathogens. LFQ analysis on larval
haemolymph showed that in response to EPF supernatant both insect species
displayed altered abundance of proteins involved in antimicrobial defence, the
prophenoloxidase cascade, detoxification and detection and sensing. These patterns
of alteration may be integral to the modulation of the host immune response by EPF.
A major difference observed between the proteomic profiles of H. abietis and G.
mellonella haemolymph was that H. abietis injected with B. caledonica supernatant
had a major alteration in metabolic proteins involved in cellulose cleavage, reflective
of its wood based diet. It was concluded G. mellonella may have an application as a
model for looking for secondary metabolites or natural products that display
immunomodulating properties so that EPF isolates could be screened for production
of these products. However G. mellonella are not a substitute for the target pest for
proteomic analysis. Moreover laboratory bioassays with either G. mellonella or H.
abietis are not predictive of whether synergy will occur in the field as there are many
more factors involved than just the ability of EPF to modulate the immune response.
To investigate the ability of EPF and EPN to suppress H. abietis populations in
the field, three field studies were carried out over three consecutive years.
Treatments were applied to tree stumps harbouring H. abietis developmental stages.
The efficacy of EPF and EPN was investigated alone and in combination through
emergence trapping and destructive sampling. Three EPF strains were utilised in
these field studies, commercial strains of M. anisopliae and B. bassiana and a strain
of B. caledonica native to Ireland. Two EPN species were utilised in these field
studies, Steinernema carpocapsae and Heterorhabditis downesi, the latter is native to
Ireland. In this work EPN were found to offer superior control over H. abietis in the
field than EPF, with all treatments that caused significant reduction in adult
emergence being EPN alone or EPN in combination with EPF. Ultimately EPF are
not suited to control of H. abietis using this strategy. Synergy between EPF-EPN was
not achieved in any of the three field studies with all combinations tested giving
additive results.
The final aim of this work was to investigate if the little-studied native fungus B.
caledonica produces immunomodulating compounds active against H. abietis.
Identification, large scale production and structural determination of an abundant
secreted natural product of B. caledonica was carried out using High Performance
Liquid Chromatography (HPLC) and Nuclear magnetic resonance (NMR)
spectroscopy, and the metabolite of interest was found to be oosporein. Subsequently
assessment of the potential insecticidal, anti-feedant and immunomodulating effects
of oosporein was carried out. Oosporein was identified as an abundant metabolite in
B. caledonica supernatant that displayed immunomodulating properties in H. abietis
larvae as well as being a feeding stimulant to H. abietis adults.
The work presented throughout this thesis offers the first report of the effect of
EPF on the cellular and humoral immune system of H. abietis. It presents a critical
assessment of G. mellonella as a model organism for immune studies of H. abietis. It
highlights that EPN are a superior biocontrol agent for the control of H. abietis in the
context of the current approach used. However the native B. caledonica isolate tested
throughout this thesis presents an interesting avenue for future research into the
utilisation of EPF as biocontrol agents in a wider sense than researched in this work.
Item Type: | Thesis (PhD) |
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Keywords: | Interactions; entomopathogenic fungi; control agents; mechanism; field potential; Hylobius abietis larvae; |
Academic Unit: | Faculty of Science and Engineering > Biology |
Item ID: | 10420 |
Depositing User: | IR eTheses |
Date Deposited: | 10 Jan 2019 15:48 |
URI: | https://mural.maynoothuniversity.ie/id/eprint/10420 |
Use Licence: | This item is available under a Creative Commons Attribution Non Commercial Share Alike Licence (CC BY-NC-SA). Details of this licence are available here |
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