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IMMUNE RESPONSE TO ENTOMOPATHOGENIC

FUNGUS Metarhizium anisopliae INFECTION IN


Guimares, M.Q.,
de
Paula,
A.
R.,
Samuels
,
R.I.,
Gomes,
S.A.,
Fonseca,
R.N.,
Sorgine,
M.H.,
VionetteRhodnius prolixus
(Hemiptera:
Reduviidae)
Amaral, R.J., Moraes, J. and Mury, F.B.
4

LIBHM and LICM, NUPEM, UFRJ, Maca, RJ. LEF, UENF, Campos dos Goytacazes, RJ. 4IBqM, UFRJ, Rio de Janeiro, RJ.
FAT BODY
The Rhodnius prolixus is a hematophagous insect and vector of Trypanosoma cruzi, a
protozoan parasite that causes Chagas disease. Insects are exposed to a wide range of
microorganisms and have interconnected powerful immune reactions. Innate immunity
is the first line of defense being divided into humoral response that is related to
antimicrobial peptides (AMP), lectins and the prophenoloxidase cascade and the
cellular

response

which

includes

phagocytosis,

hemocytes

aggregation

and

encapsulation of pathogens. The Metarhizium anisopliae is an entomopathogenic


fungus used as biological control agents and start the infection process mainly by
TOLL Pathway

IMD Pathway

JAK-STATPathway

penetration through the insect cuticle.

Thomas & Read, 2007


Nature Biotechnology (25;
12)

Silverman & Maniatis 2001

Souza-Neto et al.,

Genes & Development. 15: 2321-2342.

2009
PNAS 20;106(42)

To monitor the effect of entomopathogenic fungus Metarhizium anisopliae infection


on the mortality of Rhodnius prolixus;
To investigated the effect of entomopathogenic fungus Metarhizium anisopliae
infection on the modulation immune response and to linking with embryogenesis
Figure 2 Relative expression of the Dorsal and Cactus (Toll pathway), Relish (IMD pathway), Eigher (TNF
ortholog), STAT and SOCS (Jack-STAT pathway) and Defensin and Lysozymes (LysA, LysB) (AMPs) genes
from fat body of female R. prolixus. Control (Ctl) and infected (I) with M. anisopliae. Female insects were blood fed or
fasting in absence or presence of Metarhizium anisopliae. The expression of the genes were evaluated with the expression of the
housekeeping gene EF-1 (Majerowicz et al., 2011). The samples were assayed with the Applied Biosystems StepOne platform.

process in Rhodnius.

MIDGUT
1107 conidia mL1
were sprayed directly
onto Rhodnius prolixus.
qRT-PCR

Potter Tower

Midgut + fat
body

TTTTT
AAAAA

cDNA

TotalRNA

RT

RNA extraction, cDNA synthesis and Polymerase Chain


Reaction (PCR)

A.

B.

Figure 3 Relative expression of the Dorsal and Cactus (Toll pathway), Relish (IMD pathway), Eigher (TNF
ortholog), STAT and SOCS (Jack-STAT pathway) and Defensin and Lysozymes (LysA, LysB) (AMPs) genes
from midgut of female R. prolixus. Control (Ctl) and infected (I) with M. anisopliae. Female insects were blood fed or
fasting in absence or presence of Metarhizium anisopliae. The expression of the genes were evaluated with the
expression of the housekeeping gene EF-1 (Majerowicz et al., 2011). The samples were assayed with the Applied
Biosystems StepOne platform.

C.

D.

Figure 1 Female insects were blood fed in absence or presence of Metarhizium anisopliae. 24 and 72h
after feeding, individual parameters were determined for each group A) Number of egg per female, as
determined at 20 days after meal; B) Hatching rate, as determined at 25 days after oviposition, C) general
aspect of egg control and D) general aspect of egg injected with Metarhizium anisopliae. Results are expressed
as mean (S.E.M.) of 2 independent experiments. *p < 0.05 (compared to control Rhodnius blood feeding in
absence of Metarhizium anisopliae) (non-paired Students t-test). Each group contained 6 females.

Majerowicz D. et al., (2011) Looking for reference genes for real-time quantitative PCR experiments in Rhodnius
prolixus (Hemiptera: Reduviidae). Insect Mol Biol. 20(6):713-22.
Muller H. et al., (2014) Considerations for the process development of insect-derived antimicrobial peptide
production. Biotechnol Prog. 31(1):1-11.
Silverman N. and Maniatis T. (2001). NF-B signaling pathways in mammalian and insect innate immunity. Genes &
Development. 15: 2321-2342.
Souza-Neto JA et al., (2009). An evolutionary conserved function of the JAK-STAT pathway in anti-dengue defense. Proc
Natl Acad Sci U S A. 20;106(42):17841-6.
Thomas MB and Read AF. (2007). Fungal bioinsecticide with a sting. Nat Biotechnol. 25(12):1367-8.