Transcriptome Analysis of Psacothea hilaris: De Novo Assembly and Antimicrobial Peptide Prediction

SIMPLE SUMMARY: Multidrug resistance for classical antibiotics is the major problem faced in clinical infections. Insects have antimicrobial peptides (AMPs) which are a part of the immune repertoire to protect it from microbial pathogens. AMPs are short chain protein molecules that function as antibacterial, antifungal, antiparasitic and antiviral factors. Insect AMPs are present in their hemolymph or produced in response to entry of pathogens and kill them to protect the host. The objective of this study is to identify novel AMPs from Psacothea hilaris, the yellow spotted long horned beetle. The beetle was immunized with bacteria and fungi and RNA was isolated. The RNA was processed and screened by in silico strategies to identify novel AMPs. We obtained one potential candidate which was tested to be effective against harmful bacteria and fungi. This will be useful in treating multidrug resistant microbes alone or in combination with antibiotics with future validations. ABSTRACT: Antimicrobial peptides (AMPs) are the frontline innate defense system evolutionarily preserved in insects to combat invading pathogens. These AMPs could serve as an alternative to classical antibiotics to overcome the burden of treating multidrug resistant bacteria. Psacotheasin, a knottin type AMP was isolated from Psacothea hilaris and shown to exhibit antimicrobial activity, especially against fungi through apoptosis mediated cell death. In this study, we aimed to identify novel probable AMPs from Psacothea hilaris, the yellow spotted longicorn beetle. The beetle was immunized with the two bacterial strains (E. coli and S. aureus), and the yeast strain C. albicans. After immunization, total RNA was isolated and sequenced in Illumina platform. Then, beetle transcriptome was de novo assembled and searched for putative AMPs with the known physiochemical features of the AMPs. A selection of AMP candidates were synthesized and tested for antimicrobial activity. Four peptides showed stronger activity against E. coli than the control AMP, melittin while one peptide showed similar activity against S. aureus. Moreover, four peptides and two peptides showed antifungal activity stronger than and similar to melittin, respectively. Collectively one peptide showed both antibacterial and antifungal activity superior to melittin; thus, it provides a potent antimicrobial peptide. All the peptides showed no hemolysis in all the tested concentrations. These results suggest that in silico mining of insects’ transcriptome could be a promising tool to obtain and optimize novel AMPs for human needs.