Location: Beneficial Insects Introduction Research Unit
2020 Annual Report
Objectives
Objective 1: Determine the physiological, behavioral, ecological, and genetic basis of host ranges of noctuid moths and parasitoids of pest insects, such as soybean aphid, Russian wheat aphid, sugar cane aphid, and spotted-wing Drosophila, with a focus on using molecular genetic methods to elucidate factors responsible for the evolution of host specificity.
Subobjective 1.1 – Determine the genetic basis of host ranges of noctuid moths and of parasitoids of pest insects.
Subobjective 1.2 – Test whether bacterial endosymbionts affect acceptance and suitability of hosts and determine mechanisms of these effects.
Subobjective 1.3 – Test whether the host specificity of Aphelinus species changes with stress or experience.
Objective 2: Determine interactions between biological control and host plant resistance in their effects on survival, reproduction, and population dynamics of pest insects, such as soybean aphid, Russian wheat aphid, sugar cane aphid, and spotted-wing Drosophila, in laboratory and field experiments.
Objective 3: Determine molecular phylogenetic relationships, test host specificity, and introduce parasitoids for biological control of pest insects, such as soybean aphid, Russian wheat aphid, sugar cane aphid, and spotted-wing Drosophila, and determine the impact of the introduced parasitoids on the abundance and distribution of target and non-target species.
Subobjective 3.1 – Determine phylogenetic relationships among parasitoids whose members are candidates for biological control introductions.
Subobjective 3.2 – Measure host specificity of parasitoids that are candidates for biological control introductions.
Subobjective 3.3 – Introduce parasitoids to control pest insects, such as soybean aphid, Russian wheat aphid, sugar cane aphid, and spotted-wing Drosophila, and measure the impact of the introduced parasitoids on the abundance and distribution of target and non-target species.
Approach
We will use analysis of genomes for genes that are divergent in sequence or expression, QTL mapping, co-localization of probes for QTL markers and divergent genes with chromosomal fluorescence in-situ hybridization and allele genotyping, analysis of tissue-specific expression (antenna, ovipositor), and gene knock-out with CRISPR/Cas9 and RNAi technology to identify genes involved in host recognition and acceptance. To test whether defensive bacterial endosymbionts affect acceptance and suitability of hosts of parasitoids and to determine mechanisms underlying these effects, we will assay more species of parasitoids on more species of aphids with and without their defensive endosymbionts. To test whether host ranges of Aphelinus species are ever dynamic, we will test the effects of starvation, age, and experience on parasitism of sub-optimal hosts by parasitoid species with broad host ranges. We will do additional experiments on the interactions between host plant resistance and parasitism by Aphelinus species. Continued development of the molecular phylogeny of Aphelinus species will provide a framework for other results. We will conduct host specificity testing of parasitoids for release against D. noxia, M. sacchari and D. suzukii. We will introduce parasitoid species with narrow host ranges and monitor their impact on target and non-target species.
Progress Report
We studied the genetics of differences in host specificity among Aphelinus species that parasitize aphids. We crossed A. atriplicis, which readily parasitizes the Russian wheat aphid, Diuraphis noxia, with A. certus, which rarely parasitizes this aphid. We measured parasitism of the aphid by backcross progeny, genotyped them, and used these data to map genomic regions and candidate genes affecting parasitism. We identified 8 regions that explained 40% of the variance in parasitism and determined that 13 of 15 candidate genes that diverged between these species mapped to these regions. To test whether these genes were chemosensory, we measured tissue-specific expression and found that all 13 genes were expressed in sensilla on ovipositors or mandibles. In marker-phenotype analysis among backcross progeny from A. rhamni, a soybean aphid parasitoid, selected by three generations of rearing on a non-preferred aphid, Rhopalosiphum padi, versus unselected A. rhamni, 35 genetic markers were associated with increased parasitism. After 144 generations of rearing on R. padi, parasitism of this aphid doubled. To study the evolution of host specificity, we sequenced, assembled, and annotated the genomes/transcriptomes of 20 populations of Aphelinus in 15 species and measured host-specificity for these populations. Genome assembly sizes were 308-477 Mb, and these assemblies captured 92-97 percent of a set of 1658 conserved insect genes. We identified 28-55k genes per species, and among them, 136-192 were chemoreceptors, 115-156 venom proteins, and 103-154 cytochrome-p450s. While the amount of genetic variation was similar between specialists and generalists, some variants were phenotype specific. In generalists only, we found variants in three of the main histone proteins which could affect the regulation of genes involve in host use. In specialists only, we found variants in several chemosensory genes that might affect host specificity. Objective 1.1 (Agreement number 60-8010-7-002).
With the University of Delaware, Delaware Biotechnology Institute, North Shore University Health System, and the University of Chicago, we studied the effects of missing data and confounding genetic markers from different genomic regions in genotyping-by-sequencing (GBS), while developing a new GBS platform. Phased GBS in maize revealed spurious genotypes from variants from different parts of the genome undetectable in the single-nucleotide genetic variation. Objective 1.1 (60-8010-7-002).
With North Carolina State University and the American Museum of Natural History, we mapped genomic regions involved in host use by herbivores in Chloridea, a genus of noctuid moths that includes pests. The genetic architectures of intra- and interspecific variation were very similar, involving a large number of interchangeable genomic regions. Objective 1.1
With Oregon State University, we studied adaptation in cinnabar moth, which was introduced to control tansy ragwort, a toxic rangeland weed. We sequenced, assembled, and annotated genomes of insects from mountain versus valley habitats that harbor different host species. With 36 variants per kilobase between habitats, 72-79 percent of genes were divergent. Highly divergent genes (=30 percent), comprised 0.1 percent of all genes, and these divergent genes had 3-5 fold fewer homologs than all genes. Development time varies between valley and mountain populations and within valley populations. Analysis of gene expression in larvae that differed in development time revealed 20 genes that differed in expression. Objective 1.1 (58-8010-6-006)
Cowpea aphid and pea aphid may be infected by the symbiotic bacteria Hamiltonella defensa and its bacteriophage, APSE (Acyrthosiphon pisum secondary endosymbiont), that defend these aphids against some parasitoid species. Parasitism of infected aphids by two Aphelinus species did not differ from that of uninfected aphids. While A. atriplicis showed no difference in fitness components between infected and uninfected aphids, A. glycinis produced more adult progeny and larger female progeny on infected than on uninfected aphids. With the University of Georgia, the University of Kentucky, and the University of Minnesota, and the French Institut Nationale de la Recherche Agronomic (INRA), we found support for the hypothesis that the diversity of bacteria in aphids is lower among introduced aphid populations. Laboratory cultures of aphids can lose these bacteria so they should be monitored for symbionts to ensure that laboratory tests produce field-relevant results. Objective 1.2 (58-8010-6-010)
With the University of Illinois, Iowa State University, and Ohio State University, we studied the interaction between plant resistance in soybean, virulence in the soybean aphid, Aphis glycines, and parasitism by Aphelinus species. In an experiment with two parasitoid species, both species parasitized virulent and avirulent aphids on resistant and susceptible soybean. Although parasitism by A. glycinis did not vary with virulence or resistance, A. certus parasitized virulent aphids more than avirulent aphids, perhaps because the former were more abundant. Such density dependent mortality may slow the spread of virulence. With Texas A&M University, Kansas State University, and Pennsylvania State University, we studied the interaction between plant resistance in sorghum and parasitism of the sugarcane aphid, Melanaphis sacchari. Aphelinus nigritus parasitized aphids on resistant and susceptible sorghum. With Colorado State University, we studied the interaction between plant resistance in wheat, virulence in D. noxia, and parasitism by A. hordei. Parasitism by A. hordei did not vary with aphid virulence or wheat resistance and was high in all treatments. Objective 2
With the University of California Riverside, Hebei University (China), Indiana University, Zoologisches Forschungsmuseum Alexander Koenig (Germany), University of Rochester, University of Georgia, Texas A&M University, Illinois Natural History Survey, State Museum of Natural History (Germany), and University of Hohenheim (Germany), we made transcriptome-based phylogenetic trees of Chalcidoidea, which is the largest group of parasitic wasps and includes many species used in biological control of pests. We found that the poorly resolved tree backbone could only be marginally improved by adding more genes and species, reducing missing data, and proof-checking for contamination and errors in homology. Concatenation analyses supported egg parasitism as ancestral in chalcidoids, but coalescent analyses gave a different result, unless filtered to include only the most highly supported genes. The results uncovered gene discordances and identified functional bias in genes supporting alternative trees that may indicate ancient adaptive introgression. This research shows that understanding mechanisms resulting in gene-tree discordance is essential to sorting out backbone relationships, especially for groups that have undergone extremely rapid radiation. Objective 3.1 (58-1926-3-002)
With the City University of New York, we studied mixed-strategy extracellular vesicles in the venom of Leptopilina heterotoma, a parasitoid that attacks Drosophila species, some of which are major pests. During this attack, female wasps inject into host larvae venom with vesicles carrying proteins that suppress the immune system of their hosts. We found 246 vesicle proteins in the L. heterotoma proteome, and an enrichment analysis supports the vesicular nature of these structures. Transcripts of more than 90% of these proteins were present in whole-body transcriptomes. Sequencing and assembly of the wasp genome showed that more than 90% of vesicle proteins are coded in the wasp genome. These results explain the stability of vesicles in wasps. Suppression of the immune system of hosts is a major mechanism determining host specificity of parasitic wasps, and understanding its evolution is import for predicting the safety of biological control introductions against invasive pests. Objective 3.3
Aphelinus hordei, a candidate for introduction to control the Russian wheat aphid, had a narrow host range in laboratory experiments. We prepared, submitted, and received approval for a petition to the North American Plant Protection Organization to introduce A. hordei. An environmental assessment has been prepared by APHIS-PPQ, and we expect to receive a field-release permit soon. Objective 3.3
We tested host specific of 36 populations of Aphelinus from Eurasia in at least 17 species that are candidates for introduction against the soybean aphid. Three species (A. coreae, A. glycinis, and A. rhamni) had narrow host ranges. Petitions to the North American Plant Protection Organization (NAPPO) for introductions of A. glycinis and A. rhamni have been approved, and Animal and Plant Health Inspection Service - Plant Protection and Quarantine (APHIS-PPQ) has provided permits for field release. With colleagues at Iowa State University, the University of Minnesota, Ohio State University, and the University of Illinois, we reared and released >600,000 A. glycinis in Iowa and Minnesota during 2013-16. In the summer of 2015, sampling showed a high abundance of aphids parasitized by Aphelinus, peaking at over 50 parasitized aphids per soybean plant. Aphelinus glycinis overwintered successfully in soybean fields and under buckthorn. Objective 3.3 (Agreement numbers 58-8010-5-017, 58-8010-5-018, 58-8010-5-019).
Accomplishments
Review Publications
Zhang, J., Lindsey, A., Peters, R., Heraty, J.M., Hopper, K.R., Werren, J.H., Martinson, E.O., Woolley, J.B., Yoder, M.J., Krogmann, L. 2020. Conflicted Signal in Transcriptomic Markers Leads to a Poorly Resolved Backbone Phylogeny of Chalcidoid Wasps (Hymenoptera: Chalcidoidea). Systematic Entomology. https://doi.org/10.1111/syen.12427.
Wey, B., Heavner, M., Wittmeyer, K.T., Briese, T., Hopper, K.R., Govind, S. 2019. Immune suppressive extracellular vesicle proteins of Leptopilina heterotoma are encoded in the wasp genome. Journal of Hymenoptera Research. 10:1-12. https://doi.org/10.1534/g3.119.400349.
Shirley, X., Woolley, J.B., Hopper, K.R., Isidoro, N., Romani, R. 2019. Evolution of glandular structures on the scapes of males in the genus Aphelinus Dalman, 1820 (Hymenoptera: Aphelinidae). Journal of Hymenoptera Research. 72:27-43. https://doi.org/10.3897/jhr.72.36356.
