Identification and Characterization of Cowpea Aphid Salivary Proteins and the Cowpea Resistance Mechanism to the Cowpea Aphid

Cowpea (Vigna unguiculata) is a vital crop for semiarid regions of the world. Cowpea is able to withstand harsh abiotic stresses prevalent in these regions but is susceptible to the cowpea aphid (Aphis craccivora). Unlike most aphids, cowpea aphids are phytotoxic and damage cowpea even at low populations. Aphids feed on plant phloem sap and while feeding deposit saliva which contain proteinaceous effectors to disrupt plant defenses and alter plant physiology. The composition of the cowpea aphid saliva has not been well studied and how cowpea aphids manipulate their hosts remain unresolved. While resistance to cowpea aphids has been identified in an African cowpea line, and the genetic determinants of the resistance mapped to two QTLs, the underlying resistance mechanisms remain unknown. In Chapter One, liquid chromatography tandem mass spectrometry (LC-MS/MS) was used to identify the proteome of the cowpea aphid saliva. One of the proteins identified, diacetyl/L-xylulose reductase (DCXR), was functionally characterized using Agrobacterium-mediated transient expression and in vitro biochemical analyses. DCXR is a member of short-chain dehydrogenases/reductases invovled in both carbohydrate and dicarbonyl metabolism. I showed that cowpea aphid infestation transiently induces the cytotoxic dicarbonyl, methylglyoxal, in cowpea. Recombinant cowpea aphid DCXR (AcDCXR) was able to detoxify methylglyoxal in vitro and to oxidize the carbohydrate xylitol to xylulose and expression of AcDCXR in pea (Pisum sativum) improved aphid fecundity. Using resistant and susceptible cowpeas, and various aphid infestation approaches and behavioral assays, I determined the nature of the resistance in Chapters Two and Three. In Chapter Two, the susceptible interaction is elaborated and the resistance mechanism revealed to be localized in the phloem and to involve both antibiosis and antixenosis. In Chapter Three, aphid dispersal assays indicate aphids prefer susceptible cowpea to resistant. In addition, using timecourse infestation of resistant and susceptible cowpeas and RNASeq, I determined the genes regulated in both susceptible and resistant responses. The transcriptome analyses identified major differences in susceptible and resistant cowpea including involvement of multiple plant hormones and defense related genes. A subset of these genes are candidates for further exploration as the source of resistance