Objective 1: Determine the genetic basis of the host ranges and climatic tolerances of pest herbivorous insects and parasitoids of these pests with a focus on using molecular genetic methods to elucidate factors responsible for the evolution of host specificity, to predict responses to climate change, and to develop methods for management of pest impacts. [NP304, C1, PS1A; C3, PS3A, 3B and 3C] Sub-objective 1.A – Determine genetic basis of differences in host specificity of aphid parasitoids. Sub-objective 1.B – Measure genetic variation in responses of parasitoids and their aphid hosts to different temperature regimes. Sub-objective 1.C – Develop and test mathematical models for impact of climate change on population dynamics and evolution of parasitoids and aphid hosts. Objective 2: Determine interactions between biological control, plant resistance, and aphid virulence in their effects on virulence frequencies. [NP304, C1, PS1A; C3, PS3A and 3B] Sub-objective 2.A – Measure interactions between resistance, virulence, and parasitoids in their effects on virulence frequencies. Sub-objective 2.B – Develop and test mathematical models of interactions between plants, aphids, and parasitoids in their effects on virulence frequencies. Objective 3: Determine molecular phylogenetic relationships, test host specificity, and introduce parasitoids for biological control of target aphids. [NP304, C1, PS1A; C3, PS3A and 3B] Sub-objective 3.A – Determine molecular phylogenetic relationships of parasitoids. Sub-objective 3.B – Test host specificity of parasitoids of pest aphids. Sub-objective 3.C – Introduce parasitoids against pest aphids. Sub-objective 3.D – Measure the impact of the introduced parasitoids on distribution and abundance target and non-target aphid species.
Hypotheses about genetic architecture of host specificity have two extremes: (a) few genes with large effects that interact additively; (b) many genes of small effects that interact epistatically. We will take two approaches to testing these hypotheses. In one approach, we will use hybridization of A. atriplicis and A. certus in the laboratory to map QTL affecting parasitism of D. noxia, a host for A. atriplicis and a non-host for A. certus. We will cross A. atriplicis into A. certus with multi-parent advanced generation intercrosses and inbreed the intercrosses to make recombinant inbred lines (RILs). We will genotype SNPs in the RILs, make a linkage map, measure parasitism of D. noxia by these lines, and map QTL affecting parasitism. In the other approach, we will map QTL involved in differences between lines of A. rhamni reared for >140 generations on R. padi versus control lines reared on A. glycines. We will cross and backcross these lines, make a linkage map based on SNPs, measure parasitism of R. padi by backcross females, and map QTL affecting parasitism. We will test whether genes that diverge in sequence and/or expression between RIL and backcross females are associated with QTL found above. We will do this by genotyping RIL and backcross females for alleles in divergent genes and integrating these genes onto the QTL maps. To find genes that diverge in sequence or expression between A. atriplicis and A. certus and between selection and control A. rhamni, we have sequenced and assembled their genomes and transcriptomes. We will test whether divergent genes associated with QTL are expressed in sensilla on antennae, ovipositor, or mouth partsby hybridizing probes for the genes in parasitoids and imaging whole-mounts microscopically. We will whether divergence in gene sequences or expression levels correlate with differences in host specificity, host acceptance, and host suitability among Aphelinus species. We will measure genetic variation in temperature responses of parasitoids and their hosts that could allow adaptation to new temperature regimes using isofemale lines. We will develop and test mathematical models for the impact of climate change on population dynamics and evolution of parasitoids and hosts. To test this whether parasitoids slow the increase in frequency of virulent aphid genotypes that can overcome host plant resistance, we will do within-generation to get parameter estimates and multi-generation experiments to test the effects of parasitoids on viruluence freqencies. We will also develop and test mathematical models of interactions between plants, aphids, and parasitoids in their effects on virulence frequencies. We will continue to determine molecular phylogenetic relationships of parasitoids in the genus Aphelinus. We will continue to test the host specificity of parasitoids of pest aphids, and we will introduce parasitoids with narrow host range and measure their impact on the distribution and abundance target and non-target aphid species.
Safe, effective biological-control introductions against invasive pests depend on narrowly host-specific natural enemies with the ability to adapt to a changing environment. As part of a project on the genetic architectures of these traits, we assembled and annotated the genomes of two aphid parasitoids, APHELINUS ATRIPLICIS and APHELINUS CERTUS. We made several assemblies using Illumina and PacBio data, which we combined into a meta-assembly. We scaffolded the meta-assembly with markers from a genetic map of hybrids between A. ATRIPLICIS and A. CERTUS. We used the genetic-linkage scaffolded (GLS) assembly of A. ATRIPLICIS to scaffold a de novo assembly of A. CERTUS. The de novo assemblies differed in contiguity, and the meta-assembly was more contiguous than the best de novo assembly. Scaffolding with genetic-linkage data allowed the chromosomal-level assembly of the A. ATRIPLICIS genome and scaffolding a de novo assembly of A. CERTUS with this GLS assembly greatly increased the contiguity of the A. CERTUS assembly. However, completeness of the A. ATRIPLICIS assembly, as measured by percent of the complete, single-copy BUSCO hymenopteran genes, varied little among de novo assemblies and was not increased by meta-assembly or genetic scaffolding. Furthermore, the greater contiguity of the meta-assembly and GLS assembly had little or no effect on the numbers of genes identified, the proportion with homologs, or functional annotations. Increased contiguity of the A. CERTUS assembly provided a modest improvement in assembly completeness, as measured by the percent complete, single-copy BUSCO hymenopteran genes. The total genic sequence increased, and while the number of genes declined, gene length increased, which together suggests greater accuracy of gene models. More contiguous assemblies provide other than gene annotation, such as identifying the genes associated with quantitative trait loci and understanding chromosomal rearrangements associated with speciation. Subobjectives 1.A and B (Agreement 60-8010-7-002). When attempting to create recombinant inbred lines (RIL) from Multi parent Advanced Generation Intercross (MAGIC) populations of crosses between APHELINUS ATRIPLICIS and APHELINUS CERTUS, we lost a significant proportion of lines each generation, making maintaining lines difficult. Some explanations include genetic incompatibility between the two species, inbreeding depression, and random loss (the number of offspring can vary widely between individuals). Several approaches to keeping sufficient lines were attempted, like replicating lines and using multiple parents per line, but none were sustainable, so the creation of RILs was abandoned. Instead, we use the MAGIC hybrid populations of A. ATRIPLICIS x A. CERTUS, which have now been maintained for 34 generations of random mating in 4 population cages of ~200 individuals/generation. These populations are being used for host-use experiments on DIURAPHIS NOXIA (D. NOXIA) to address the original goal of QTL mapping of parasitism of a novel host. We measured the parasitism of D. NOXIA by 980 females from these MAGIC populations after 22 generations of random mating. We are now extracting DNA and making reduced-representation libraries for each female to map QTL involved in the parasitism of D. NOXIA. Subobjective 1.A (Agreement number 60-8010-7-002). We continued selection of APHELINUS RHAMNI for increased parasitism of a non-preferred, low-quality host, RHOPALOSIPHUM PADI, by rearing the parasitoid on this aphid for for 144 generations. Parasitism of R. PADI by A. RHAMNI doubled during this period, and body masses of selected A. RHAMNI were higher on R. PADI than those of unselected A. RHAMNI. Interestingly, A. RHAMNI selected by rearing on R. PADI still parasitized more A. GLYCINES than R. PADI. We crossed and backcrossed the selected and unselected populations and measured the number of R. PADI parasitized by 752 backcross females, which varied from 0 to 44 aphids, with 19 percent of the females parasitizing no aphids. We are extracting DNA and making reduced-representation libraries for each backcross female to map QTL involved in the increased parasitism of R. PADI. Subobjective 1.A (Agreement number 60-8010-7-002). We conducted an experiment on the host-specificity of APHELINUS MACULATUS, an aphid parasitoid collected from PHORODON CANNABIS in New York State. We exposed A. MACULATUS to seven aphid species for one week. We will count the numbers of aphids parasitized and the sex, number, and body masses, of adult parasitoid progeny when COVID-19 pandemic restrictions are lifted. Because A. MACULATUS is in a different sub-genus of APHELINUS than all the other species of APHELINUS, the results should shed light on the evolution of host-specificity of parasitoids in this genus, which has several species important in the biological control of invasive pests, Sub-objective 3.B.
1. Chromosomal-level assembly of the genome of an aphid parasitoid. As part of a project on the genetic architectures of host specificity and climatic adaptation, we assembled and annotated the genomes of two aphid parasitoids, APHELINUS ATRIPLICIS and APHELINUS CERTUS. Scaffolding with genetic-linkage data allowed chromosomal-level assembly of the A. ATRIPLICIS genome, and scaffolding a de novo assembly of A. CERTUS with this genetic-linkage scaffolded assembly greatly increased the contiguity of the A. CERTUS assembly. These more contiguous assemblies will aid in identifying the genes affecting host specificity and climatic adaptation in these aphid parasitoids. The results also extend knowledge of the genetic basis for host specificity that may be applicable to a broad range of biological control agents.
Hopper, K.R., Wittmeyer, K.T., Kuhn, K.L., Lanier, K. 2021. Response to selection for parasitism of a sub-optimal, low-preference host in an aphid parasitoid. Evolutionary Applications. 00:1-13. https://doi.org/10.1111/eva.13254.