Plant-mediated RNA interference (RNAi) has been successfully used as a tool to study gene function in aphids. The persistence and transgenerational effects of plant-mediated RNAi in the green peach ...aphid (GPA) Myzus persicae were investigated, with a focus on three genes with different functions in the aphid. Rack1 is a key component of various cellular processes inside aphids, while candidate effector genes MpC002 and MpPIntO2 (Mp2) modulate aphid–plant interactions. The gene sequences and functions did not affect RNAi-mediated down-regulation and persistence levels in the aphids. Maximal reduction of gene expression was ~70% and this was achieved at between 4 d and 8 d of exposure of the aphids to double-stranded RNA (dsRNA)-producing transgenic Arabidopsis thaliana. Moreover, gene expression levels returned to wild-type levels within ~6 d after removal of the aphids from the transgenic plants, indicating that a continuous supply of dsRNA is required to maintain the RNAi effect. Target genes were also down-regulated in nymphs born from mothers exposed to dsRNA-producing transgenic plants, and the RNAi effect lasted twice as long (12–14 d) in these nymphs. Investigations of the impact of RNAi over three generations of aphids revealed that aphids reared on dsMpC002 transgenic plants experienced a 60% decline in aphid reproduction levels compared with a 40% decline of aphids reared on dsRack1 and dsMpPIntO2 plants. In a field setting, a reduction of the aphid reproduction by 40–60% would dramatically decrease aphid population growth, contributing to a substantial reduction in agricultural losses.
Various animals derive nutrients from symbiotic microorganisms with much-reduced genomes, but it is unknown whether, and how, the supply of these nutrients is regulated. Here, we demonstrate that the ...production of essential amino acids (EAAs) by the bacterium Buchnera aphidicola in the pea aphid Acyrthosiphon pisum is elevated when aphids are reared on diets from which that EAA are omitted, demonstrating that Buchnera scale EAA production to host demand. Quantitative proteomics of bacteriocytes (host cells bearing Buchnera) revealed that these metabolic changes are not accompanied by significant change in Buchnera or host proteins, suggesting that EAA production is regulated post-translationally. Bacteriocytes in aphids reared on diet lacking the EAA methionine had elevated concentrations of both methionine and the precursor cystathionine, indicating that methionine production is promoted by precursor supply and is not subject to feedback inhibition by methionine. Furthermore, methionine production by isolated Buchnera increased with increasing cystathionine concentration. We propose that Buchnera metabolism is poised for EAA production at certain maximal rates, and the realized release rate is determined by precursor supply from the host. The incidence of host regulation of symbiont nutritional function via supply of key nutritional inputs in other symbioses remains to be investigated.
DNA methods are useful to identify ingested prey items from the gut of predators, but reliable detection is hampered by low amounts of degraded DNA. PCR‐based methods can retrieve minute amounts of ...starting material but suffer from amplification biases and cross‐reactions with the predator and related species genomes. Here, we use PCR‐free direct shotgun sequencing of total DNA isolated from the gut of the harlequin ladybird Harmonia axyridis at five time points after feeding on a single pea aphid Acyrthosiphon pisum. Sequence reads were matched to three reference databases: Insecta mitogenomes of 587 species, including H. axyridis sequenced here; A. pisum nuclear genome scaffolds; and scaffolds and complete genomes of 13 potential bacterial symbionts. Immediately after feeding, multicopy mtDNA of A. pisum was detected in tens of reads, while hundreds of matches to nuclear scaffolds were detected. Aphid nuclear DNA and mtDNA decayed at similar rates (0.281 and 0.11 h⁻¹ respectively), and the detectability periods were 32.7 and 23.1 h. Metagenomic sequencing also revealed thousands of reads of the obligate Buchnera aphidicola and facultative Regiella insecticola aphid symbionts, which showed exponential decay rates significantly faster than aphid DNA (0.694 and 0.80 h⁻¹, respectively). However, the facultative aphid symbionts Hamiltonella defensa, Arsenophonus spp. and Serratia symbiotica showed an unexpected temporary increase in population size by 1–2 orders of magnitude in the predator guts before declining. Metagenomics is a powerful tool that can reveal complex relationships and the dynamics of interactions among predators, prey and their symbionts.
Many plant viruses depend on aphids and other phloem‐feeding insects for transmission within and among host plants. Thus, viruses may promote their own transmission by manipulating plant physiology ...to attract aphids and increase aphid reproduction. Consistent with this hypothesis, Myzus persicae (green peach aphids) prefer to settle on Nicotiana benthamiana infected with Turnip mosaic virus (TuMV) and fecundity on virus‐infected N. benthamiana and Arabidopsis thaliana (Arabidopsis) is higher than on uninfected controls. TuMV infection suppresses callose deposition, an important plant defense, and increases the amount of free amino acids, the major source of nitrogen for aphids. To investigate the underlying molecular mechanisms of this phenomenon, 10 TuMV genes were over‐expressed in plants to determine their effects on aphid reproduction. Production of a single TuMV protein, nuclear inclusion a‐protease domain (NIa‐Pro), increased M. persicae reproduction on both N. benthamiana and Arabidopsis. Similar to the effects that are observed during TuMV infection, NIa‐Pro expression alone increased aphid arrestment, suppressed callose deposition and increased the abundance of free amino acids. Together, these results suggest a function for the TuMV NIa‐Pro protein in manipulating the physiology of host plants. By attracting aphid vectors and promoting their reproduction, TuMV may influence plant–aphid interactions to promote its own transmission.
The aphid Myzus persicae s.l. (Hemiptera: Aphididae) is an important pest of many crops worldwide with a complex life cycle, intensely controlled by chemical pesticides, and has developed resistance ...to almost all used insecticides. In Greece, the aphid exhibits high genetic variation and adaptability and it is a classic example of evolution in the making. We have been studying M. persicae for over 20 years, on different host plants and varying geographical areas, analyzing its bio-ecology and the ability to develop resistance to insecticides. In this review, we present new and historical data on the effectiveness of insecticides from seven chemical groups used to control the aphid in Greece and the incidence of seven resistance mechanisms, including the new fast-spreading R81T point mutation of the postsynaptic nicotinic acetylcholine receptor. Thousands of samples were tested by biological, biochemical and molecular assays. The aphid populations were found to have developed and maintain resistance at medium to high levels to organophosphates, carbamates, pyrethroids and neonicotinoids for decades. In the latter group, a marked increase is recorded during an ~10-year period. The data analyzed and the extensive bibliography, advocate the difficulty to control the aphid making the design and application of IPM/IRM programs a challenge. We discuss principles and recommendations for the management of resistance, including the use of compounds such as flonicamid, spirotetramat, flupyradifurone and sulfoxaflor. We emphasize that resistance is a dynamic phenomenon, changing in time and space, requiring, therefore, continuous monitoring.
Associations between microbes and animals are ubiquitous and hosts may benefit from harbouring microbial communities through improved resource exploitation or resistance to environmental stress. The ...pea aphid, Acyrthosiphon pisum, is the host of heritable bacterial symbionts, including the obligate endosymbiont Buchnera aphidicola and several facultative symbionts. While obligate symbionts supply aphids with key nutrients, facultative symbionts influence their hosts in many ways such as protection against natural enemies, heat tolerance, color change and reproduction alteration. The pea aphid also encompasses multiple plant-specialized biotypes, each adapted to one or a few legume species. Facultative symbiont communities differ strongly between biotypes, although bacterial involvement in plant specialization is uncertain. Here, we analyse the diversity of bacterial communities associated with nine biotypes of the pea aphid complex using amplicon pyrosequencing of 16S rRNA genes. Combined clustering and phylogenetic analyses of 16S sequences allowed identifying 21 bacterial OTUs (Operational Taxonomic Unit). More than 98% of the sequencing reads were assigned to known pea aphid symbionts. The presence of Wolbachia was confirmed in A. pisum while Erwinia and Pantoea, two gut associates, were detected in multiple samples. The diversity of bacterial communities harboured by pea aphid biotypes was very low, ranging from 3 to 11 OTUs across samples. Bacterial communities differed more between than within biotypes but this difference did not correlate with the genetic divergence between biotypes. Altogether, these results confirm that the aphid microbiota is dominated by a few heritable symbionts and that plant specialization is an important structuring factor of bacterial communities associated with the pea aphid complex. However, since we examined the microbiota of aphid samples kept a few generations in controlled conditions, it may be that bacterial diversity was underestimated due to the possible loss of environmental or transient taxa.
Clothianidin is a second-generation neonicotinoid insecticide, widely used against sap-sucking insect pest including melon aphid, Aphis gossypii Glover (Hemiptera: Aphididae). This pest causes severe ...economic damage to Cucurbitaceae plants worldwide. In this study, we investigated clothianidin resistance development under continuous selection pressure. Moreover, the age-stage, two-sex life table approach was used to evaluate the impact of clothianidin resistance on the fitness of A. gossypii. A clothianidin resistant strain (CT-R) with a 23.17-fold resistance level was developed from a susceptible strain (CT-S) after continuous selection for 24 generations. Life table results showed a significant reduction in the relative fitness (0.847) of CT-R strain compared to the CT-S strain of A. gossypii. The developmental duration, oviposition days, total pre-oviposition period (TPOP), longevity, and fecundity of CT-R strain were found to be significantly lower when compared to CT-S strain. The demographic parameters, including the intrinsic rate of increase (r), finite rate of increase (λ), net reproductive rate (R0), and mean generation time (T) were also significantly decreased in CT-R strain compared to the CT-S strain. Both the reproductive and survival rates were affected by clothianidin resistance in CT-R strain compared with the CT-S strain of A. gossypii. Overall, our results demonstrate that in-depth knowledge about the trade-off at play between resistance degree and fitness cost might be useful to design resistance management strategies against A. gossypii.
1. The pea aphid, Acyrthosiphon pisum, maintains extreme variation in resistance to its most common parasitoid wasp enemy, Aphidius ervi, which is sourced from two known mechanisms: protective ...bacterial symbionts, most commonly Hamiltonella defensa, or endogenously encoded defences. We have recently found that individual aphids may employ each defence individually, occasionally both defences together, or neither. 2. In field populations, Hamiltonella-infected aphids are found at low to moderate frequencies and while less is known about the frequency of resistant genotypes, they show up less often than susceptible genotypes in field collections. To better understand these patterns, we sought to compare the strengths and costs of both types of defence, individually and together, in order to elucidate the selective pressures that maintain multi-modal defence mechanisms or that may favour one over the other. 3. We experimentally infected five aphid genotypes (two lowly and three highly resistant), each with two symbiont strains, Hamiltonella-APSE8 (moderate protection) and Hamiltonella-APSE3 (high protection). This resulted in three sublines per genotype: uninfected, +APSE8 and +APSE3. Each of the 15 total sublines was first subjected to a parasitism assay to determine its resistance phenotype and in a second experiment, a subset was chosen to compare fitness (fecundity and survivorship) in the presence and absence of parasitism 4. In susceptible aphid genotypes, parasitized sublines infected with Hamiltonella generally showed increased protection with direct fitness benefits, but clear infection costs to fitness in the absence of parasitism. In resistant genotypes, Hamiltonella infection rarely conferred additional protection, often further reduced fecundity and survivorship when enemy challenged, and resulted in constitutive fitness costs in the absence of parasitism. We also identified strong aphid genotype×symbiontstrain interactions, such that the best defensive strategy against parasitoids varied for each aphid genotype; one performed best with no protective symbionts, the others with particular strains of Hamiltonella. 5. This surprising variability in outcomes helps explain why Hamiltonella infection frequencies are often intermediate and do not strongly track parasitism frequencies in field populations. We also find that variation in endogenous traits, such as resist-ance, among host genotypes may offer redundancy and generally limit the invasion potential of mutualistic microbes in insects.
•Absence of RNAi response by feeding and injection of dsRNA in pea aphids (Acyrthosiphon pisum).•Degradation of dsRNA by salivary secretions and aphid hemolymph.•Upon dsRNA injection, no effect in ...expression of RNA-related genes dicer-2, argonaute-2, r2d2 and sid-1.•Pea aphids contain an Eri-1-like endocellular nuclease as in nematodes.
Over the past decade, RNA interference (RNAi), the sequence-specific suppression of gene expression, has proven very promising for molecular research in many species, including model insects as Tribolium castaneum and Apis mellifera. It showed its usefulness to analyze gene function and its potential to manage pest populations and reduce disease pathogens. However, in several insects, the efficiency of RNAi is low or very variable at best. One of the factors that could influence RNAi efficiency in insects is degradation of dsRNA after administration to the insect. In this paper, we report on the importance of dsRNA breakdown in the pea aphid (Acyrthosiphon pisum) associated with the absence of an RNAi response upon oral feeding and injection with dsRNA targeting different genes such as the ecdysone hormone receptor and ultraspiracle. In essence, we discovered that both the salivary secretions of aphids and the hemolymph were able to degrade the dsRNA. In parallel, introduction of dsRNA in the aphid body was not able to provoke a response in the expression of the siRNA core machinery genes.
Plant viruses maintain intricate interactions with their vector and non-vector insects and can impact the fitness of insects. However, the details of their molecular and cellular mechanisms have not ...been studied well. We compared the transcriptome-level responses in vector and non-vector aphids (Schizaphis graminum and Rhopalosiphum padi, respectively) after feeding on wheat plants with viral infections (Barley Yellow Dwarf Virus (BYDV) and Wheat dwarf virus (WDV), respectively). We conducted differentially expressed gene (DEG) annotation analyses and observed DEGs related to immune pathway, growth, development, and reproduction. And we conducted cloning and bioinformatic analyses of the key DEG involved in immune.
For all differentially expressed gene analyses, the numbers of DEGs related to immune, growth, development, reproduction and cuticle were higher in vector aphids than in non-vector aphids. STAT5B (signal transducer and activator of transcription 5B), which is involved in the JAK-STAT pathway, was upregulated in R. padi exposed to WDV. The cloning and bioinformatic results indicated that the RpSTAT5B sequence contains a 2082 bp ORF encoding 693 amino acids. The protein molecular weight is 79.1 kD and pI is 8.13. Analysis indicated that RpSTAT5B is a non-transmembrane protein and a non-secreted protein. Homology and evolutionary analysis indicated that RpSTAT5B was closely related to R. maidis.
Unigene expression analysis showed that the total number of differentially expressed genes (DEGs) in the vector aphids was higher than that in the non-vector aphids. Functional enrichment analysis showed that the DEGs related to immunity, growth and reproduction in vector aphids were higher than those in non-vector aphids, and the differentially expressed genes related to immune were up-regulated. This study provides a basis for the evaluation of the response mechanisms of vector/non-vector insects to plant viruses.
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