Peptidomics Approaches for the Identification of Bioactive Molecules from Diaphorina citri.

Huanglongbing (HLB), a deadly citrus disease, is primarily associated with Candidatus Liberibacter asiaticus (CLas) and spread by the hemipteran insect Diaphorina citri. Control strategies to combat HLB are urgently needed. In this work, we developed and compared workflows for the extraction of the D. citri peptidome, a dynamic set of polypeptides produced by proteolysis and other cellular processes. High-resolution mass spectrometry revealed bias among methods reflecting the physiochemical properties of the peptides: while TCA/acetone-based methods resulted in enrichment of C-terminally amidated peptides, a modification characteristic of bioactive peptides, larger peptides were overrepresented in the aqueous phase of chloroform/methanol extracts, possibly indicative of reduced co-analytical degradation during sample preparation. Parallel reaction monitoring (PRM) was used to validate the structure and upregulation of peptides derived from hemocyanin, a D. citri immune system protein, in insects reared on healthy and CLas-infected trees. Mining of the data sets also revealed 122 candidate neuropeptides, including PK/PBAN family neuropeptides and kinins, biostable analogs of which have known insecticidal properties. Taken together, this information yields new, in-depth insights into peptidomics methodology. Additionally, the putative neuropeptides identified may lead to psyllid mortality if applied to or expressed in citrus, consequently blocking the spread of HLB disease in citrus groves.

Arrestment behavior in the polyphagous tropical root weevil Diaprepes abbreviatus.

A semiochemical-based attractant for the tropical root weevil, Diaprepes abbreviatus L., has been sought since the introduction of this polyphagous pest to Florida in the 1960s. A six-unit wind tunnel apparatus was constructed to allow multiple runs to be conducted simultaneously to test the response of D. abbreviatus adults to volatile and contact semiochemicals. Arrestment was measured as the number of weevils stopping and remaining on a target when presented with upwind target choices. No response (movement upwind) was observed when weevils were presented with a control (no odor). Some movement upwind but no arrestment occurred when weevils were presented with young citrus leaves (flush). Citrus flush that had been fed on by D. abbreviatus adults was highly attractive and induced arrestment of both male and female adults. The effect was duplicated by placing organdy-enclosed feces (frass) on undamaged flush. The arrestment effect was eliminated, however, when fed-on flush was enclosed in a cage so that weevils could not contact the leaves. In these experiments, weevils continued to move upwind. The results of these tests lead us to conclude that host and mate location by D. abbreviatus is mediated by volatile and contact semiochemicals.