Bactericera cockerelli vitellogenin-6 like, a vitellogenin without a direct reproductive function?

Vitellogenin-like proteins are members of the large lipid transfer proteins, a family of proteins involved in reproduction, lipid circulation and immune defences. In this study, we identified a new Bactericera cockerelli vitellogenin-like (Vg-like) transcript, and named it BcVg6-like based on its similarity to Acyrthosiphon pisum Vg6. In silico analyses predicted different conserved domains in BcVg6-like compared with the conventional Ba. cockerelli vitellogenin, BcVg1-like, previously described by our research group. Phylogenetic analyses determined that BcVg6-like clustered with Vg-like-B proteins and not the conventional vitellogenins involved in vitellogenesis. Also, the expression analyses showed differences in BcVg6-like transcript expression between 7-day-old males and 3- and 7-day-old females. BcVg6-like was not upregulated after exogenous application of juvenile hormone III, but its relative expression increased significantly in alimentary canals of adult females exposed to tomato plants infected by the bacterial plant pathogen 'Candidatus Liberibacter solanacearum'. Our results suggest that in Ba. cockerelli, both vitellogenin genes may have different functions: BcVg1-like is a conventional vitellogenin that conserved its ancestral function as an egg yolk precursor whereas BcVg6-like might have acquired a function in lipid and/or other molecule transport, and could potentially play a role in immune defence.

Selection of reference genes for expression analysis in the potato psyllid, Bactericera cockerelli.

The selection of reference genes is a crucial step for quantitative real-time PCR analyses and increasingly the use of more than one reference gene for accurate and reliable normalization is being recommended. In this study, a set of six genes was selected and their stability was assessed in different life stages and female organs of Bactericera cockerelli harbouring or not the bacterial pathogen 'Candidatus Liberibacter solanacearum' (Lso) haplotype B. The stability of each gene was determined using the BestKeeper, NormFinder and GeNorm programs. These analyses identified elongation factor-1a, ribosomal protein subunit L5 and ribosomal protein subunit 18 as the most stable genes to analyse gene expression during the insect life stages irrespective of Lso presence; Lso haplotype B only affected their respective ranking. By contrast, a common set of normalizers could not be found amongst the different female organs tested (bacteriomes, alimentary canals and reproductive organs).

Settling and Ovipositional Behavior of Bactericera cockerelli (Hemiptera: Triozidae) on Solanaceous Hosts Under Field and Laboratory Conditions.

Potato psyllid, Bactericera cockerelli (Sulc), is a seasonal insect pest in the Lower Rio Grande Valley of Texas, where it transmits the bacterial pathogen "Candidatus Liberibacter solanacearum" that causes zebra chip disease of potato. Studies were conducted to evaluate host preference of B. cockerelli adults for different plant species, and plant size and density. Settling and oviposition behavior of B. cockerelli was studied on its wild and cultivated solanaceous hosts, including potato, tomato, pepper, eggplant, and silverleaf nightshade, under both field and laboratory conditions. Naturally occurring B. cockerelli were used to evaluate host preference under open field conditions throughout the growing season. Settling and oviposition preference studies in the laboratory were conducted as cage-release experiments using pairs of plants, and observations were recorded over a 72-h period. Results of field trials indicated that naturally occurring B. cockerelli preferred potato and tomato equally for settling and oviposition, but settled on pepper, eggplant, and silverleaf nightshade only in the absence of potato and tomato. Under laboratory conditions, B. cockerelli adults preferred larger host plants, regardless of the species tested. Results also showed that movement of B. cockerelli was minimal after initial landing and settling behavior was influenced by host plant density. Lone plants attracted the most psyllids and can be used as sentinel plants to monitor B. cockerelli activity. Information from both field and laboratory studies demonstrated that not only host plant species determined host selection behavior of B. cockerelli adults, but also plant size and density.

Genetics coupled to quantitative intact proteomics links heritable aphid and endosymbiont protein expression to circulative polerovirus transmission.

Yellow dwarf viruses in the family Luteoviridae, which are the causal agents of yellow dwarf disease in cereal crops, are each transmitted most efficiently by different species of aphids in a circulative manner that requires the virus to interact with a multitude of aphid proteins. Aphid proteins differentially expressed in F2 Schizaphis graminum genotypes segregating for the ability to transmit Cereal yellow dwarf virus-RPV (CYDV-RPV) were identified using two-dimensional difference gel electrophoresis (DIGE) coupled to either matrix-assisted laser desorption ionization-tandem mass spectrometry or online nanoscale liquid chromatography coupled to electrospray tandem mass spectrometry. A total of 50 protein spots, containing aphid proteins and proteins from the aphid's obligate and maternally inherited bacterial endosymbiont, Buchnera, were identified as differentially expressed between transmission-competent and refractive aphids. Surprisingly, in virus transmission-competent F2 genotypes, the isoelectric points of the Buchnera proteins did not match those in the maternal Buchnera proteome as expected, but instead they aligned with the Buchnera proteome of the transmission-competent paternal parent. Among the aphid proteins identified, many were involved in energy metabolism, membrane trafficking, lipid signaling, and the cytoskeleton. At least eight aphid proteins were expressed as heritable, isoelectric point isoform pairs, one derived from each parental lineage. In the F2 genotypes, the expression of aphid protein isoforms derived from the competent parental lineage aligned with the virus transmission phenotype with high precision. Thus, these isoforms are candidate biomarkers for CYDV-RPV transmission in S. graminum. Our combined genetic and DIGE approach also made it possible to predict where several of the proteins may be expressed in refractive aphids with different barriers to transmission. Twelve proteins were predicted to act in the hindgut of the aphid, while six proteins were predicted to be associated with the accessory salivary glands or hemolymph. Knowledge of the proteins that regulate virus transmission and their predicted locations will aid in understanding the biochemical mechanisms regulating circulative virus transmission in aphids, as well as in identifying new targets to block transmission.

Differences in bacterial diversity of host-associated populations of Phylloxera notabilis Pergande (Hemiptera: Phylloxeridae) in pecan and water hickory.

Host-associated differentiation (HAD) is the presence of genetically divergent, host-associated populations. It has been suggested that microbial symbionts of insect herbivores may play a role in HAD by allowing their insect hosts to use different plant species. The objective of this study was to document if host-associated populations of Phylloxera notabilis Pergande (Hemiptera: Phylloxeridae) in pecan and water hickory corresponded with differences in the composition of their associated bacteria. To test this hypothesis, we characterized the symbionts present in P. notabilis associated with these two tree species through metagenomic analyses using 454 sequencing. Differences in bacterial diversity were found between P. notabilis populations associated with pecan and water hickory. The bacteria, Pantoea agglomerans and Serratia marcescens, were absent in the P. notabilis water hickory population, whereas both species accounted for more than 69.72% of bacterial abundance in the pecan population.

A genomic analysis of transcytosis in the pea aphid, Acyrthosiphon pisum, a mechanism involved in virus transmission.

Aphids are the primary vectors of plant viruses. Transmission can occur via attachment to the cuticle lining of the insect (non-circulative transmission) or after internalization in the insect cells with or without replication (circulative transmission). In this paper, we have focused on the circulative and non-propagative mode during which virions enter the cell following receptor-mediated endocytosis, are transported across the cell in vesicles and released by exocytosis without replicating. The correct uptake, transport and delivery of the vesicles cargo relies on the participation of proteins from different families which have been identified in the Acyrthosiphon pisum genome. Assemblage of this annotated dataset provides a useful basis to improve our understanding of the molecules and mechanisms involved in virus transmission by A. pisum and other aphid species.

Identification of ion channel genes in the Acyrthosiphon pisum genome.

Aphids are major pests of crops, causing hundreds of millions of dollars worth of damage annually. Ion channel proteins are often the targets of modern insecticides and mutations in ion channel genes can lead to resistance to many leading classes of insecticides. The sequencing of the pea aphid, Acyrthosiphon pisum, genome has now allowed detailed in silico analysis of the aphid ion channels. The study has revealed significant differences in the composition of the ion channel families between the aphid and other insects. For example A. pisum does not appear to contain a homologue of the nACh receptor alpha 5 gene whilst the calcium channel beta subunit has been duplicated. These variations could result in differences in function or sensitivity to insecticides. The genome sequence will allow the study of aphid ion channels to be accelerated, leading to a better understanding of the function of these economically important channels. The potential for identifying novel insecticide targets within the aphid is now a step closer.

Tangible benefits of the aphid Acyrthosiphon pisum genome sequencing for aphid proteomics: Enhancements in protein identification and data validation for homology-based proteomics.

Homology-driven proteomics promises to reveal functional biology in insects with sparse genome sequence information. A proteomics study comparing plant virus transmission competent and refractive genotypes of the aphid Schizaphis graminum isolated numerous candidate proteins involved in virus transmission, but limited genome sequence information hampered their identification. The complete genome of the pea aphid, Acyrthosiphon pisum, released in 2008, enabled us to double the number of protein identifications beyond what was possible using available EST libraries and other insect sequences. This was concomitant with a dramatic increase of the number of MS and MS/MS peptide spectra matching the genome-derived protein sequence. LC-MS/MS proved to be the most robust method of peptide detection. Cross-matching spectral data to multiple EST sequences and error tolerant searching to identify amino acid substitutions enhanced the percent coverage of the Schizaphis graminum proteins. 2-D electrophoresis provided the protein pI and MW which enabled the refinement of the candidate protein selection and provided a measure of protein abundance when coupled to the spectral data. Thus, the homology-based proteomics pipeline for insects should include efforts to maximize the number of peptide matches to the protein to increase certainty in protein identification and relative protein abundance.

Sequence evaluation of four specific cDNA libraries for developmental genomics of sunflower.

Four different cDNA libraries were constructed from sunflower protoplasts growing under embryogenic and non-embryogenic conditions: one standard library from each condition and two subtractive libraries in opposite sense. A total of 22,876 cDNA clones were obtained and 4800 ESTs were sequenced, giving rise to 2479 high quality ESTs representing an unigene set of 1502 sequences. This set was compared with ESTs represented in public databases using the programs BLASTN and BLASTX, and its members were classified according to putative function using the catalog in the Kyoto Encyclopedia of Genes and Genomes (KEGG). Some 33% of sequences failed to align with existing plant ESTs and therefore represent putative novel genes. The libraries show a low level of redundancy and, on average, 50% of the present ESTs have not been previously reported for sunflower. Several potentially interesting genes were identified, based on their homology with genes involved in animal zygotic division or plant embryogenesis. We also identified two ESTs that show significantly different levels of expression under embryogenic and non-embryogenic conditions. The libraries described here represent an original and valuable resource for the discovery of yet unknown genes putatively involved in dicot embryogenesis and improving our knowledge of the mechanisms involved in polarity acquisition by plant embryos.