Contents lists available at ScienceDirect Journal of Thermal Biology journal homepage: www.elsevier.com/locate/jtherbio Effect of thermal stress on the immune responses of Chilo suppressalis walker (Lepidoptera: Crambidae) to Beauveria bassiana Leila Shamakhi a , Arash Zibaee a,* , Azadeh Karimi-Malati a , Hassan Hoda b a Department of Plant Protection, Faculty of Agricultural Sciences, University of Guilan, Rasht, 416351314, Iran b Iranian Research Institute of Plant Protection, Agricultural Research, Education and Extension Organization (AREEO), Tehran, Iran ARTICLE INFO Keywords: Temperature Immunity Chilo suppressalis Gene expression Entomopathogen ABSTRACT Temperature is one of the important environmental elements affecting ecological fitness of insects through alterations in physiological systems. In the current study, a comparison was made on the cellular and humoral immune responses of the Chilo suppressalis larvae exposed to thermal stress (34 °C) and optimal rearing tem- perature (24 °C). Although total hemocyte count increased in the injected larvae by Beauveria bassiana, elevation of hemocyte numbers was significantly different in the larvae exposed to 34 °C for a short-time period compared to long-term exposure and control. A similar trend was observed in plasmatocyte and granulocyte counts as well as phenoloxidase activity. Gene expression of some antimicrobial peptides, including attacin1, attacin2, ce- cropin1, cecropin2, defensin, gallerimycin, lysozyme and prophenoloxidase-activating proteinase-3, was com- pared in the larvae exposed to thermal regimes and injection challenges. In all cases, expression of the target genes was relatively higher in the larvae injected by B. bassiana and short-term exposure at 34 °C. The present results confirmed that C. suppressalis could modulate the immune system in response to different thermal stress conditions mainly over a short period. 1. Introduction Insects, similar to other organisms, during most of their lifetime, simultaneously encounter a vast array of stressful stimuli that may threaten their survival (Williams et al., 2015). Pathogens (such as fungi, viruses, bacteria, and protozoan/metazoan parasite) and fluctuation in environmental factors like temperature are among the most important stressors simultaneously or consecutively experienced by insects, which may have disruptive effects on their biological and physiological fit- ness; therefore, insects have evolved a set of behavioral, physical and immunological barriers to deal with the effects of these threats (Kaunisto et al., 2016; Wojda, 2017). Behavioral defence (e.g. biting, acquiring genetic resistance through selection of suitable mates, com- mitting suicide to favor kin survival, or developing a fever response) are the first line of defence in insects minimizing or eliminating negative effects of parasites and pathogens (Greeney et al., 2012). The cuticle and epidermis are the second lines of defense forming an efficient protective barrier over external surface extending into the trachea, foregut, and hindgut (Gillespie and Kanost, 1997). Nevertheless, many pathogens and parasites are able to breach these barriers; therefore, insects must also employ their innate immune systems as the final line of their defense. The innate immune system is comprised of cellular and humoral responses, both of them are mediated through various sig- naling pathways (Lavine and Strand, 2002). Humoral defenses contain antimicrobial molecules involved in melanin formation, coagulation and toxicity against pathogens, while cellular responses refer to he- mocyte mediated processes such as phagocytosis, encapsulation, and nodulation (Strand, 2008). Environmental conditions like ambient temperature profoundly affect the performance of the insect immune system (Le Moullac and Haffner, 2000; Adamo, 2004; Mydlarz et al., 2006; De Block and Stoks, 2008). Since insects are poikilotherm organisms, and their immune system relies on temperature-dependent cellular and enzymatic activities, fluctuations in temperature will directly affect immune activity simply based on thermodynamics (Murdock et al., 2012). Early studies have demonstrated that warmer temperatures increase various immune re- sponses (e.g. Ouedraogo et al., 2002; Ouedraogo et al., 2003; Zibaee et al., 2009; Catalán et al., 2012a, 2012b; Wojda and Taszłow, 2013a, 2013b). In fact, the thermoregulatory set-point known as behavioral fever makes a key contribution to immune resistance of both vertebrate and invertebrate toward infection (Kluger, 1979). In particular, the immune system appears to be tightly – albeit complexly – linked to stress response in insects, with both immuno-suppressive and immuno- enhancing effects (Adamo, 2014, 2016). For example, since stress and https://doi.org/10.1016/j.jtherbio.2019.07.006 Received 11 December 2018; Received in revised form 20 May 2019; Accepted 1 July 2019 * Corresponding author. E-mail addresses: arash.zibaee@guilan.ac.ir, arash.zibaee@gmx.com (A. Zibaee). Journal of Thermal Biology 84 (2019) 136–145 Available online 02 July 2019 0306-4565/ © 2019 Elsevier Ltd. All rights reserved. T