Abstract

   Plutella xylostella has become the major lepidopteran pest of Brassica
   owing to its strong ability of resistance development to a wide range
   of insecticides. Destruxin A, a mycotoxin of entomopathogenic fungus,
   Metarhizium anisopliae, has broad-spectrum insecticidal effects.
   However, the interaction mechanism of destruxin A with the immune
   system of P. xylostella at genomic level is still not well understood.
   Here, we identified 129 immunity-related genes, including pattern
   recognition receptors, signal modulators, few members of main immune
   pathways (Toll, Imd, and JAK/STAT), and immune effectors in P.
   xylostella in response to destruxin A at three different time courses
   (2 h, 4 h, and 6 h). It is worthy to mention that the immunity-related
   differentially expressed genes (DEGs) analysis exhibited 30, 78, and 72
   up-regulated and 17, 13, and 6 down-regulated genes in P. xylostella
   after destruxin A injection at 2 h, 4 h, and 6 h, respectively,
   compared to control. Interestingly, our results revealed that the
   expression of antimicrobial peptides that play a vital role in insect
   immune system was up-regulated after the injection of destruxin A. Our
   findings provide a detailed information on immunity-related DEGs and
   reveal the potential of P. xylostella to limit the infection of fungal
   peptide destruxin A by increasing the activity of antimicrobial
   peptides.

Introduction

   The diamondback moth, Plutella xylostella, has become the major
   lepidopteran pest of Brassica worldwide in the past four decades
   costing approximately US$4 billion annually on its management^[44]1.
   The attributes like high reproductive potential, lack of natural
   enemies, and its strong ability of resistance development to a wide
   range of insecticides and growth regulators, are the reasons for its
   continued success against modern pest management approaches^[45]2.
   Until now, P. xylostella has evolved resistance to almost all classes
   of insecticides and Bacillus thuringiensis-based products^[46]2, [47]3.
   At present, there is a need to develop novel biological control
   methods, to reduce harmful effects of insecticides, as alternative
   control strategies^[48]4.

   The entomopathogenic fungi, such as Metarhizium anisopliae and
   Beauveria bassiana, are widely considered as important biological
   control agents^[49]5–[50]7, and M. anisopliae has commercially been
   used for controlling insect pests^[51]8–[52]11. The reason for
   successfully infecting a wide range of insects could be secretion of
   virulence factors by some fungi during pathogenesis. Destruxins, the
   secondary metabolites of fungi, produced by entomopathogenic fungi like
   M. anisopliae and Aschersonia spp. are considered as vital virulence
   factors accelerating the death of insects^[53]12–[54]14.

   Chemically, destruxins have a typical composition containing α-hydroxy
   acid and five amino acids which form cyclic hexadepsipeptides. Until
   now, 39 analogs of destruxins have been extracted from various fungal
   species^[55]15–[56]17. Among them, few destruxins such as Destruxin A,
   Destruxin B, and Destruxin E have exhibited significant insecticidal
   activities against various insect pests^[57]12, [58]18, [59]19.
   Previously, it has been shown that destruxins inhibit V-type ATPase
   hydrolytic activity of Galleria mellonella, prompt oxidative stress in
   Spodoptera litura and affect the Ca^2+ channel in muscle cells of
   Manduca sexta ^[60]20–[61]22. Additionally, destruxins are also
   reported to affect the immune system of insects, such as Drosophila
   melanogaster innate immune response was suppressed by destruxin A
   following the inhibition of antimicrobial peptides^[62]23, however, no
   significant changes in the expression of antimicrobial peptides were
   observed in hemocytes of Bombyx mori in response to destruxin A^[63]24.

   Invertebrates, unlike mammals, don’t have an adaptive immune system,
   but instead, they rely on a sophisticated innate immune system for
   defense against invading microbes. The innate immune system of insects
   is comprised of two main components, cellular and humoral immune
   responses^[64]25. The former relies majorly on the action of hemocytes
   in the phagocytosis of pathogens^[65]26, while the latter refers to the
   process of melanization with phenoloxidases^[66]27 and synthesis of
   immune effector molecules^[67]28.

   To date, with the help of genome-wide analysis, immunity-related genes
   and gene families have been identified in various insect species
   including P. xylostella ^[68]29–[69]32. Prior to the genome sequence of
   P. xylostella, immunity-related genes were identified by using
   expressed sequence tags and cDNA microarray analysis^[70]33, however,
   recently Xia et al.^[71]32 reanalyzed the immunity-related genes of P.
   xylostella in response to bacterial infection, to better understand the
   mechanism of immunity-related genes, at the genomic level. Similarly,
   Etebari et al.^[72]34 identified microRNAs from P. xylostella in
   response to parasitization by Diadegma semiclausum using the genome of
   other lepidopteran species as proxy references. Recently, Etebari et