Characterization of seminal fluid peptides/proteins of male <i>Helicoverpa armigera </i>and their plausible role in post-copulatory modulation of female reproductive behaviour

In many insect species, female reproductive output is determined by male ejaculate components such as seminal fluid, plasma, and male accessory gland (MAG) peptides. These elements have additional roles in affecting insect receptivity and mating physiology. Studies indicate that the peptides/proteins of the male insects’ seminal fluid are responsible for these behavioural changes in female Helicoverpa armigera insects. In particular, the virgin male’s MAG peptide induces mating receptivity inhibition approximately 12 h after eclosion (emergence), which reaches peak concentration approximately at 24 h. This time coincides with the male’s mating time, and is transferred to female moths during copulation. The aim of the study was to purify and characterize the virgin male’s MAG-duplex peptides/proteins by biochemical and bioinformatics analysis. Analysis yielded a peptide of 5 kDa molecular weight. In insects several conserved proteins belonging to members of multigene families control a wide range of reproduction and related physiological processes. Data mining of the MAG‑duplex peptides utilizing the public protein repository identified six proteins (heat shock proteins, diazepam-binding inhibitor, elongation factor 1 alpha, odorant-binding protein, serpin, and thioredoxin) with diverse functions. The diazepam-binding inhibitor protein, which binds coenzyme A and thiol esters of long fatty acids, was unique to H. armigera. The molecular evolutionary analysis of the proteins independently supports the widely accepted theory that genes in gene families with roles in reproduction have evolved over a generational time scale.