Immune challenges trigger cellular and humoral responses in adults of
Pterostichus melas italicus (Coleoptera, Carabidae)
Q3
Anita Giglio
a, *
, Pietro Brandmayr
a
, Teresa Pasqua
a
, Tommaso Angelone
a
,
Silvia Battistella
b
, Piero G. Giulianini
b
a
Dipartimento di Biologia, Ecologia e Scienze della Terra, Universit a della Calabria, Via P. Bucci, I-87036 Rende, Italy
b
Dipartimento di Scienze della Vita, Universit a di Trieste, Via Giorgieri 5, I-34127 Trieste, Italy Q1
article info
Article history:
Received 17 July 2014
Received in revised form
20 January 2015
Accepted 23 January 2015
Available online xxx
Keywords:
Carabid beetle
Cellular immunity
Microscopy
Nitric oxide synthase
Phenoloxidase
Phagocytosis
abstract
The present study focuses on the ability of Pterostichus melas italicus Dejean to mount cellular and hu-
moral immune responses against invading pathogens. Ultrastructural analyses revealed the presence of
five morphologically distinct types of hemocytes: prohemocytes, plasmatocytes, granulocytes, oenocy-
toids and macrophage-like cells. Differential hemocyte counts showed that plasmatocytes and gran-
ulocytes were the most abundant circulating cell types and plasmatocytes exhibited phagocytic activity
following the latex bead immune challenge. Macrophage-like cells were recruited after the immune
challenge to remove exhausted phagocytizing cells, apoptotic cells and melanotic capsules formed to
immobilize the latex beads. Total hemocyte counts showed a significant reduction of hemocytes after
latex bead treatment. Phenoloxidase (PO) assays revealed an increase of total PO in hemolymph after
immune system activation with lipopolysaccharide (LPS). Moreover, the LPS-stimulated hemocytes
showed increased protein expression of inducible nitric oxide synthase, indicating that the cytotoxic
action of nitric oxide was engaged in this antimicrobial collaborative response. These results provide a
knowledge base for further studies on the sensitivity of the P. melas italicus immune system to the
environmental perturbation in order to evaluate the effect of chemicals on non target species in
agroecosystems.
© 2015 Published by Elsevier Ltd.
1. Introduction
Once pathogens enter the hemocoel of the host, they encounter
a complex system of innate defense mechanism involving cellular
and humoral responses. These responses are based on a sequence of
events including the recognition of invaders and their immobili-
zation (Gillespie et al., 1997; Siva-Jothy et al., 2005; Ottaviani,
2005). This non-self recognition involves a series of membrane
receptors of hemocytes which are essential to recognize pathogen-
associated molecular patterns (PAMPs) and to trigger an immune
response. The cellular immune response involves different hemo-
cyte types which participate in pathogen clearance by phagocy-
tosis, nodule formation, encapsulation and cytotoxic reactions.
Morphological, histochemical and functional characteristics and
monoclonal antibody and genetic markers have been used to
characterize many hemocyte types in insects. The most common
types of hemocytes described in species of diverse orders are pro-
hemocytes, granulocytes, plasmatocytes, spherule cells and oeno-
cytoids (Gillespie et al., 1997; Lavine and Strand, 2002; Giulianini
et al., 2003; Jiravanichpaisal et al., 2006; Giglio et al., 2008). Hu-
moral defenses include the production of antimicrobial peptides
(AMPs), reactive intermediates of oxygen or nitrogen and the
prophenoloxidase enzymatic cascade (proPO) regulating melani-
zation of hemolymph. The inducible isoform of the enzyme nitric
oxide synthase (iNOS) is rapidly synthesized by a wide array of cells
and tissues and it catalyzes nitric oxide synthesis until the substrate
is depleted in response to acute infections in invertebrates (Nappi
and Ottaviani, 2000; Nappi et al., 2000; Nappi and Vass, 2001;
Rivero, 2006). In insect hemocytes, iNOS is synthesized in
response to immune insult and the resulting nitric oxide acts
cytotoxically on many kinds of pathogens (Ratcliffe et al., 1985;
Davies, 2000; Siva-Jothy et al., 2005; Marmaras and Lamp-
ropoulou, 2009). The proPO-activating system comprises a complex
cascade of serine proteases allowing the conversion of proPO to
phenoloxidase (PO) (Marmaras et al., 1996; Gillespie et al., 1997;
Rolff and Siva-Jothy, 2003; Schmid-Hempel, 2005; Siva-Jothy
* Corresponding author. Tel.: þ39 0984492982; fax: þ39 0984492986.
E-mail address: anita.giglio@unical.it (A. Giglio).
Contents lists available at ScienceDirect
Arthropod Structure & Development
journal homepage: www.elsevier.com/locate/asd
http://dx.doi.org/10.1016/j.asd.2015.01.002
1467-8039/© 2015 Published by Elsevier Ltd.
Arthropod Structure & Development xxx (2015) 1e9
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Please cite this article in press as: Giglio, A., et al., Immune challenges trigger cellular and humoral responses in adults of Pterostichus melas
italicus (Coleoptera, Carabidae), Arthropod Structure & Development (2015), http://dx.doi.org/10.1016/j.asd.2015.01.002