Source: THE UNIVERSITY OF TEXAS AT AUSTIN submitted to
UNDERSTANDING THE COEVOLUTIONARY DYNAMICS OF OBLIGATE INSECT SYMBIOSIS USING PEA APHIDS
Sponsoring Institution
National Institute of Food and Agriculture
Project Status
TERMINATED
Funding Source
Reporting Frequency
Annual
Accession No.
1008486
Grant No.
2016-67012-24676
Project No.
TEXW-2015-03702
Proposal No.
2015-03702
Multistate No.
(N/A)
Program Code
A7201
Project Start Date
Jan 15, 2016
Project End Date
Jan 14, 2018
Grant Year
2016
Project Director
Chong, R. A.
Recipient Organization
THE UNIVERSITY OF TEXAS AT AUSTIN
101 EAST 27TH STREET STE 4308
AUSTIN,TX 78712-1500
Performing Department
Integrative Biology
Non Technical Summary
Symbiotic relationships between bacteria and animals play a key role in generating and maintaining biodiversity in all ecosystems. Many plant species are significantly impacted by sap feeding insects, which are able to subsist on a nutritionally unbalanced diet of plant sap with the aid of symbiotic bacterial partners that live within specialized cells of their insect hosts. After millions of years of co-existence, some symbiotic relationships are now obligate where neither can exist without its partner. Though these relationships have persisted over time, little is known about what mechanisms allow for the coexistence between insects and bacteria. This research will investigate how insect hosts functionally regulate bacterial symbiont populations and will address to what extent these insects can tolerate and coexist with bacterial symbionts. This project will use experiments performed on laboratory populations of pea aphids, which harbor obligate symbiotic bacteria. In addition to experiments, this project will use new genomic and molecular technologies to determine to how pea aphids maintain their symbiotic bacteria. This project will provide key insights into the interactions of obligate symbionts with their insect hosts, which will extend current understanding of how symbiotic bacteria affect the ecological tolerance and geographic range of their hosts. Aphids are considered one of the most significant pestiferous and invasive insect species in North America. Thus, all experiments will be performed on a North American pest species. These results will provide new insight into how these pest species interact with microorganisms, and will provide critical information on the role of environmental change in the persistence of local pests and the potential emergence of novel pest species.
Animal Health Component
0%
Research Effort Categories
Basic
100%
Applied
(N/A)
Developmental
(N/A)
Classification

Knowledge Area (KA)Subject of Investigation (SOI)Field of Science (FOS)Percent
21131101080100%
Knowledge Area
211 - Insects, Mites, and Other Arthropods Affecting Plants;

Subject Of Investigation
3110 - Insects;

Field Of Science
1080 - Genetics;
Goals / Objectives
The major goal of this project is to understand how obligate insect symbioses are functionally regulated and what role symbioses play in the adaptation and diversification of insect hosts. The environmental and dietary flexibility of many pestiferous insect species are regulated by their obligate bacterial symbionts, which provision hosts with essential nutrients that are lacking from their specialized diets of plant phloem sap. These symbionts have highly reduced genomes that lack many regulatory genes and pathways; thus, hosts play a critical role in integrating and regulating symbiont function and symbiont numbers. These coordinated efforts are further complicated when hosts encounter variable environments that impact symbiont and host function and can ultimately influence the ability of symbiotic lineages to adapt. This raises important questions about how evolutionary forces interact to determine the evolutionary trajectory of symbiotic lineages. Using the obligate symbiosis of pea aphids as a model, the following supporting objectives will be undertaken: (1) determine the role of host and symbiont genotypes and their interactions in regulating symbiosis, (2) test whether rapid and degenerative evolution of symbiont genomes drives adaptation and diversification in host populations, and (3) determine how interactions between host and symbiont genotypes impact symbioses under different environmental and dietary regimes.In order to determine the role of host and symbiont genotypes and their interactions in regulating symbiosis, novel host-symbiont genotype combinations will be generated from several divergent, naturally occurring pea aphid clones using a new method that allows for experimentally replacement of native symbionts with divergent symbionts from donor aphid clones. Using high-throughput methods to sequence complete symbiont genomes and RNA sequence data (transcriptome) for host genes will allow for characterization of the novel genotypic combinations we generate in our crosses and transfers. Evaluating resulting symbiotic phenotypes and genotypes in a comparative framework will provide key insights into the role of host genotypes in regulating symbiosis. Additionally, the generation and characterization of novel pea aphid - Buchnera genetic combinations will allow for the evaluation of potential hybridization of divergent populations in nature that could give rise to lineages that are more or less fit than parental lineages.A combination of high-throughput genotyping and experimental manipulations will be used to test whether rapid and degenerative evolution of symbiont genomes drives adaptation and diversification in host populations, resulting in genetic incompatibilities within or between pea aphid populations. Specifically, genetic incompatibilities and associations between host and symbiont genotypes in the regulation of symbiosis will be tested for by sequencing progeny of reciprocal sexual crosses for which parental lines have divergent Buchnera genotypes. Clonal lineages that show evidence of pre- or post-zygotic isolation will be further tested using the symbiont replacement method, providing direct experimental tests of potential genetic incompatibilities. These results will provide insights into whether potential hybridization of divergent groups can produce new lineages with lower or higher fitness than parental lineages, or with novel properties enabling them to use new host plants, or invade new habitats.To determine how interactions between host and symbiont genotypes impact the regulation of symbiosis under changing environments and dietary regimes, novel combinations of pea aphid and Buchnera genotypes will be generated and their performance will be evaluated under different environmental conditions and on different diets. These results will provide insights into how interactions within and between host genotypes and symbiont genotypes determine a lineage's ability to adapt to different environmental conditions.
Project Methods
Methods for this project: This project uses the pestiferous pea aphid (Acyrthosiphon pisum) and its obligate symbiont (Buchnera aphidicola) as a model to system in which to investigate the processes mediating symbiotic interactions within and between species across changing environments. This project exploits a combination of genomic tools and experimental manipulations to test hypotheses on the coevolutionary dynamics of these intimate symbioses. Specifically, the impacts of variation in host and symbiont genotypes on symbiosis will be examined using a new technique that enables the transfer of Buchnera genotypes among aphid clonal matrilines and by performing sexual crosses between host populations. Using standard husbandry methods, sexual crosses between different clones will be generated to look at how novel combinations of host loci interacting with identical symbiont genotypes (siblings from the same maternal clone) impact host ability to regulate symbiosis. A new experimental method developed by the Moran lab will allows for the replacement of obligate symbiont genotypes from divergent lineages without altering host genotype. In addition to of experimental manipulations, this project will use high-throughput genome sequencing to test for the effects of host and symbiont genotypes and their responses to varying environments. Using standard genomic techniques, symbiont genomes will be sequenced using Illumina MiSeq V3 with paired end inserts and assembled using a bioinformatics pipeline and de novo assembly tools. Similarly, host genes will be sequenced as transcriptomes using Illumina HiSeq 2500 paired-end (2x125) and assembled using transcriptome based assembly software.Efforts: In an effort to expand knowledge in the field of science and to deliver this knowledge to people, this project will involve hands-on learning experiences in the laboratory, participation in workshops and seminar series, and outreach through scientific training of underrepresented minority groups. Specifically, the PD will gain experiences involving experimental design, molecular laboratory techniques, statistical analyses, and bioinformatics skills; the PD will mentor and train multiple undergraduate researchers in biology using these skills.Evaluation: Evaluation of progress towards completion of the project goals and objectives will be based on the completion of experimental crosses and transfections, data collection and analysis, and manuscript preparation. Specifically, each project objective includes the completion of three phases to assure that the project is on schedule and on track for successful conclusion. Specific education goals will be met through training and practice with new methodologies in experimental manipulations and bioinformatics that are associated with the proposed objectives. During the first six months, project milestones will include the generation of novel genetic combinations through sexual crosses and host-symbiont transfers. Accomplishment of these goals will provide me with training in experimental design and manipulations in an insect symbiont system. Subsequent project milestones will include the collection, assembly, and annotation of sequence data for host and symbiont genotypes (for Objectives 1 and 2), which will provide training in analytical methods and computational tools. Results from Objective 1 and 2 will be prepared for publication by Spring 2017, with involved discussion and input from Dr. Moran. Experimental tests between genotypes for Objective 3 will be initiated Spring 2016 with data collection and analysis completed by Spring 2017; these aims will include project milestones of successful experimental manipulation of host diet availability and thermal regime and the collection and analysis of genomic and transcriptome sequence data. Outcomes from Objective 3 will be written up and submitted by Summer 2017, which will provide another opportunity to synthesize and communicate scientific outcomes.

Progress 01/15/16 to 01/14/18

Outputs
Target Audience:The target audience of this work includes members of the scientific and agricultural community, undergraduate and graduate students, and the general public. During this project P.D. (Dr. Chong) has successfully presented this research at scientific conferences in the fields of Evolution, Symbioses, and Molecular Evolution. In addition to an oral presentation at the Evolution conference in Austin, TX in 2016, the P.D. has presented posters at the Gordon Research Conference: Animal-microbe symbioses in West Dover, VT (June 2017) and at the annual Society for Molecular Biology and Evolution Conference in Austin, TX (July 2017), where she also organized a symposium on symbiosis and interactions. She will also be participating in another conference where she will present her current research. Dr. Chong has presented two additional seminars on this work hosted by the Biology Department at The University of Akron (March 2017) and the Department of Biology at the University of Hawaii at Manoa (May 2017). The PD has also communicated this research through a recent publication of an article in a scientific journal (Chong and Moran, The ISME Journal 2018) that is currently available to the academic and research community. The P.D. has mentored three undergraduate students from the University of Texas at Austin including one student from a small rural town (C. Cole) and two non-traditional minority students (D. Tran and E. Umanzor). Undergraduate students actively participated in research and gained valuable experiences and developed skills important for professional and research careers including experiences with molecular techniques (e.g. DNA extraction, PCR, gel electrophoresis, DNA sequencing, and microscopy), insect dissection, data collection and analysis, and animal care. The P.D. has also worked to engage the general public by participating in science education and outreach events sponsored by the University of Texas at Austin, including "Explore UT" (March 2017), which brings together local GK-12 students and other members of the community to learn about educational and research opportunities at the University of Texas. Changes/Problems: Nothing Reported What opportunities for training and professional development has the project provided?This project has provided several training and professional development opportunities for the P.D. (Dr. Chong). Training opportunities include developing skills in scientific and experimental approaches including insect husbandry and maintaining experimental lab cultures, developing qPCR assays and data collection and analysis using these methods, training in microscopy and microinjection techniques, experience generating high-throughput metagenomic sequence data and performing genomics analyses. The P.D. also gained experiences in preparing manuscripts to summarize and communicate the research conducted and successfully published her work in peer-reviewed scientific journals. Dr. Chong also presented her work at three different conferences and three seminars at multiple universities during the project period and was also able to organize a symposium on "Symbiosis and interactions" at the annual Society of Molecular Biology and Evolution (SMBE) conference held in Austin Texas in 2017. She is also currently co-organizing another symposium on "Genomics and evolution of symbiotic interactions" for the 2018 SMBE conference being held in Yokohama, Japan, where she will also present her current research. By participating in these conferences, Dr. Chong had the opportunity to interact with prominent scientists in her field and was able to establish new collaborations. Specifically, the P.D. has collaborated with Dr. Gordon Bennett (currently at the University of California at Merced) on a project studying the role of differential gene expression in obligate bacterial symbionts of the leafhopper pest, Macrosteles quadrilineatus (Cicadellidae), which was recently published (Bennett and Chong, 2017, G3: Genes, Genomes, Genetics). As a broad extension of this work, Dr. Chong and Dr. Moran have also co-authored an extensive annotated bibliography on Endosymbiosis (Chong and Moran, Oxford Bibliographies, under revision). Additionally, the P.D. mentored three undergraduate researchers during this project and provided students with valuable training in insect rearing and experimentation and molecular techniques including DNA extraction, PCR and sequencing, qPCR, and analyzing genomic data. With the support of this project, Dr. Chong has successfully obtained a new position as an Assistant Professor at the University of Hawaii at Manoa in the Department of Biology. In her new position, using the research, communication, and mentoring skills developed, she will continue to pursue projects building on the genomics and evolution of insect-microbe symbioses in her new lab. How have the results been disseminated to communities of interest?Project results have been presented at multiple conferences and seminars encompassing a diverse audience of scientists. Specifically, this research has been presented to evolutionary biologists (annual Evolution conference in Austin, TX; June 2016), molecular biologists (annual Society for Molecular Biology and Evolution (SMBE) in Austin, TX; July 2017), and symbiosis researchers (Gordon Research Conference: Animal-Microbe Symbioses: Host microbe associations at the foundation of the biosphere in West Dover, VT; June 2017). The more current results from this project will be presented at the annual SMBE conference, which will be held in Yokohama, Japan this summer (July 2018). Results have also been presented to broad biology departments at University of Texas at Austin (October 2016), University of Akron (March 2017), and University of Hawaii at Manoa (May 2017). Additionally, results have been published in peer-reviewed scientific journals (Chong and Moran 2016, Proc. Natl. Acad. Sci.; Chong and Moran 2018, ISME Journal). Some of this work is still ongoing and will be presented at scientific conferences and submitted for publication upon completion. What do you plan to do during the next reporting period to accomplish the goals? Nothing Reported

Impacts
What was accomplished under these goals? Intimate interactions between bacteria and animals play a critical role in generating and maintaining diversity in all ecosystems. By studying the dynamics of these symbiotic interactions, we gain a deeper understanding of how interactions between different organisms can promote biodiversity as well as how these systems can be directly impacted by changes in biological and environmental factors. Many insects, including aphids, harbor bacterial symbionts that allow them to feed on plant sap by provisioning hosts with essential nutrients lacking in the host diet. The extent to which hosts are able to regulate their symbionts, and the ability of symbionts to provision nutrients, will directly influence the range of hosts plants. Understanding how hosts regulate microbial partners can reveal key factors in maintaining mutually obligate symbiosis, which can allow for better management of pestiferous insects feeding on crop plants. We use a combination of experimental methods, molecular approaches, and high-throughput sequencing to elucidate the relationship between microbial symbionts and their aphid hosts. Ultimately, we show that the hosts play a significant role in regulating their symbionts and that symbiont abundance significantly varies across different lineages, which influence host fitness. Additionally, genetic diversity in microbial symbionts reflect differences in ability to provisions essential nutrients, which further reinforces the need for hosts to regulate symbionts. This variation potentially can lead to shifts in preferred plant hosts and influence the ability of different lineages to be more significant pests. (I) To determine the role of host and symbiont genotypes and their interactions in regulating symbiosis, we generated a large set of genetically distinct clonal lines of pea aphids (n=39) with known relatedness by performing sexual crosses between four divergent parental lines. We founds differences in egg hatching/survivorship of progeny based on parental crosses, suggesting genetic incompatibilities play an important role in limiting the genetic combinations that result in successful progeny. For parental and surviving progeny clones, we estimated symbiont abundance using quantitative PCR (new symbiont assay developed) and measured multiple metrics of host fitness including, development time, weight at maturity, time to reproduction, and reproductive rate. We find variation in symboint abundance and differences in host fitness based on host genotype and find a significant negative correlation between symbiont abundance and host fitness. Specifically, in a controlled lab setting, we show that host genotypes with more symbionts are less fit compared to lineages with fewer symbionts that have higher reproductive rates. Interesting, these results suggest hosts that maintain lower symbiont levels proliferate more quickly and may represent increased potential to become pest species. Through this objective, the PD has gained valuable experiences by designing experiments, performing insect husbandry, conducting new molecular techniques, and preparing a manuscript, which resulted in a publication in a scientific journal (Chong and Moran. PNAS. 2016). Additionally, the progeny clones from the sexual crosses (novel genetic resources) and the molecular resources (qPCR assays) developed will serve as valuable tools for the research community and for future studies. (II) To test whether degenerative evolution of symbiont genomes drives adaptation and diversification, we generated novel genetic clones (above) and conducted symbiont transfers between several lineages. While we were successful in replacing symbionts in hosts that share genetically similar symbionts, repeated attempts to transfer more divergent symbionts failed to produce successful transfers, resulting in either complete mortality or a significant recovery of the nearly depleted native symbiont populations rather than acquisition of genetically divergent symbionts. Our experimental results suggest that the relationships between host and symbiont genotypes are tightly correlated and that hosts may not be able to survive with genetically divergent symbionts, which may be a result of genetic incompatibilities with specific symbionts and that host are unable to maintain and regulate symbionts that are divergent from their native symbionts. Through this objective, the PD has gained valuable experiences with microscopy, microinjection techniques, and genetic screening of experimental lines. We also generated genomic sequences from several aphid hosts and their symbionts and are characterizing the patterns of genome degradation in symbionts across these lineages. Specifically, we used next-generation sequencing (Illumina HIseq 4000) to generate large metagenomic datasets for multiple strains (n=25) of Buchnera and their aphid host. Following sequence assembly, symbiont sequences were identified and extracted, and complete genomes were annotated. The PD is currently conducting comparative genomic analyses and initial results suggest that the degree of genome degradation significantly varies between lineages and that these patterns of mutation and gene loss determine the degree of functionality of symbionts, which would significantly impact hosts ability to feed on different host plants. Deep sequencing of more divergent aphid hosts revealed that one host species lack the ancient symbiont Buchnera, which is usually found in all aphids. Based on genomic analyses, molecular techniques, and microscopy, we identified a novel Bacteroidetes symbiont, which has replaced the ancient symbiont. Compared to related free-living bacteria, the genome of the Bacteroidetes symbiont is more degenerate (reduced genome size and gene loss), though it retains nearly all genes necessary to synthesize essential amino acids. These results suggest the new symbiont has the genomic features of an obligate symbiont and serves as a functional replacement of Buchnera, which was likely unable to provision nutrients and was eventually eliminated. The PD gained additional experience collecting and analyzing large genomic datasets, using new microscopy techniques, and submitting manuscripts for publication (Chong and Moran 2018, ISME Journal). (III) To further investigate how interactions between host and symbiont genotypes impact symbioses, we are examining the role of symbiont regulation under changing environment and dietary regimes. Preliminary data on the impacts of changing environments (increasing temperatures) show different effects on host fitness across lineages that differ in symbiont abundance, where clones with higher symbiont abundance seem to be more robust under exposure to higher temperatures compared to lineages with lower symbiont abundance. This work in ongoing and through her new position at the University of Hawaii at Manoa, Dr. Chong will continue her work on investigating the role of genetic interaction in regulating symbiosis. We will complete experiments to examine how symbiont regulation influences host fitness under different diet treatments. Collectively, these results will help elucidate the role of genetic interactions in driving the regulation of symbiosis and the potential of pestiferous insects to expand their range of plant hosts and colonize new habitats. Additionally, these results will shed light on the impacts of changing environment on the biodiversity and microbial interactions in non-pest insects that are a part of unique ecosystems.

Publications

  • Type: Journal Articles Status: Published Year Published: 2018 Citation: Chong, R.A. and Moran, N.A. (2018) Evolutionary loss and replacement of an obligate endosymbiont in aphids. The ISME Journal. 11:715726
  • Type: Conference Papers and Presentations Status: Other Year Published: 2017 Citation: A case of evolutionary loss and replacement of an obligate endosymbiont in aphids. Poster presentation at Society of Molecular Evolution Conference 2017 in Austin, TX.
  • Type: Conference Papers and Presentations Status: Other Year Published: 2017 Citation: Evolutionary loss and replacement of an obligate endosymbiont in aphids. Poster presentation at Gordon Research Conference: Animal-microbe Symbiosis 2017 in West Dover, VT.
  • Type: Journal Articles Status: Under Review Year Published: 2018 Citation: Moran, N.A. and Chong, R.A. (2018) Endosymbiosis. Oxford Bibliographies. [in revision]
  • Type: Journal Articles Status: Published Year Published: 2016 Citation: Chong, R. A. and Moran, N. A. (2016) Intraspecific genetic variation in hosts affects regulation of obligate heritable symbionts. Proceedings of the National Academy of Sciences USA 113(46): 13114-13119
  • Type: Conference Papers and Presentations Status: Other Year Published: 2016 Citation: Heritability of obligate symbiont titer and link to aphid host fitness. Oral presentation at Evolution Conference 2016 in Austin, TX.


Progress 01/15/16 to 01/14/17

Outputs
Target Audience:The target audience of this work includes members of the scientific and agricultural community, undergraduate and graduate students, and the general public. During this project period P.D. (Dr. Chong) has successfully presented this research at a scientific conference (oral presentation at the Evolution conference in Austin, TX) and presented a seminar on this work hosted by the Department of Integrative Biology at the University of Texas at Austin (PopBio seminar at UT Austin). The PD has also communicated this research through publication of an article in a scientific journal (Chong and Moran, Proc. Natl. Acad. Sci. USA. 2016) that is currently available to the academic and research community and will be widely available to the public when the article becomes open access. During the project period P.D. has mentored three undergraduate students from the University of Texas at Austin including one student from a small rural town (C. Cole) and two non-traditional minority students (D. Tran and E. Umanzor). These students actively participated in the research project and gained valuable experiences and developed skills important for professional and research careers. Undergraduate research experiences included molecular techniques (e.g. DNA extraction, PCR, gel electrophoresis, DNA sequencing, cloning, and quantitative PCR), insect dissection and microscopy, data collection and analysis, and animal care. The P.D. has also mentored two underrepresented graduate students from the University of Texas at Austin (Ph.D. student S. Zhao and rotation student N. Xerri). Graduate student training included experimental design, insect husbandry, molecular techniques, and bioinformatic skills. The P.D. has also worked to engage the general public by participating in science education and outreach events sponsored by the University of Texas at Austin (Family Day on October 17, 2015 and Explore UT on March 2016). Changes/Problems: Nothing Reported What opportunities for training and professional development has the project provided?This project has provided numerous training and development opportunities. (1) Animal husbandry and experimental manipulation. The PD has received extensive training in raising and maintaining large numbers of aphid colonies and has learned to perform sexual crosses using aphid clones. Additionally, the PD has performed numerous fitness estimates, which involves measuring development daily, weighing individual aphids using a microbalance, and counting total number of offspring for individual females. (2) Molecular techniques and microscopy. The PD developed additional molecular techniques including quantitative PCR and also learned new skills in dissection, microinjection, and microscopy. (3) Computational skills. The PD has gained skills in statistical analysis and bioinformatics by using high-throughput methods to sequence, assemble, and annotate and analyze complete symbiont genomes. (4) Science communication. The PD participated and presented project research at a scientific conference (oral presentation at the Evolution conference in Austin, TX). (5) The PD has gained experience preparing a manuscript for publication in a scientific journal [Chong and Moran, Proc. Natl. Acad. Sci. USA. 2016] (6) As a USDA postdoctoral fellow, the PD mentored has trained three undergraduate students and two graduate students during this project period including four students from underrepresented groups. The PD also severed as a mentor for the Undergraduate Diversity Program at Evolution 2016. (7) Activities. The PD's proposed symposium topic on symbiosis for SMBE 2017 meeting (Society for Molecular Biology and Evolution annual conference) was selected and the PD is now organizing and selecting speakers for the accepted symposium on symbiosis. How have the results been disseminated to communities of interest?Results from the project have been disseminated through publication, oral presentation, and direct research experiences. The PD has presented project results at a scientific conference (oral presentation at the Evolution conference in Austin, TX on June 17-21, 2016) and presented a seminar on the project at the University of Texas at Austin (PopBio seminar at UT Austin on October 20, 2016). The results from this project have also been presented through publication as a scientific journal article [Chong and Moran, Proc. Natl. Acad. Sci. USA. 2016], which will be available to the public once it is open access in spring 2017. As a USDA postdoctoral fellow, the PD has also disseminated project results by mentoring and training three undergraduate students and two graduate students during this project period. Results from the project have been shared directly with the public at community outreach events hosted by the University of Texas at Austin. What do you plan to do during the next reporting period to accomplish the goals?To accomplish project goals, the PD will perform symbiont transfers, complete genome and transcriptome data collection and analysis, perform environmental stress experiments, and prepare and submit results for publication. To test whether rapid and degenerative evolution of symbiont genomes drives adaptation and diversification in host populations, Buchnera transfers are being performed. New aphid host lines with a replacement of obligate symbiont genotypes have been established, and we are currently working on performing transfers between more divergent host-symbiont lines. In combination with experimental manipulations, high-throughput genotyping will be used to test examine evolution of symbiont genomes. We have collected next-generation sequencing data for a number of different aphid lineages and have started preliminary data analysis while new transfer-genotypes are being generated. Once data collection and analysis is completed project results will be prepared for publication. To determine how interactions between host and symbiont genotypes impact the regulation of symbiosis under changing environments and dietary regimes, novel combinations of pea aphid and Buchnera genotypes have been generated and their fitness performance will be evaluated under different stressed environmental conditions. Experimental manipulations between genotypes are being initiated and testing should be completed by spring 2017. Following experimental manipulations of host environment, genomic and transcriptome sequence data will be collected and analyzed and results will be submitted fall 2017.

Impacts
What was accomplished under these goals? The major goal of this project is to understand how obligate insect symbioses are functionally regulated and what role symbioses play in the adaptation and diversification of insect hosts using the obligate symbiosis of pea aphids as a model system. (1) To determine the role of host and symbiont genotypes and their interactions in regulating symbiosis, we used experimental crosses to identify to impacts of host genotype by estimating symbiont titer levels, measured as the ratio of symbiont genome copy number and host genome copy number, and by conducting fitness experiments to estimate host development time and fecundity. We show that symbiont titer levels significantly differ significantly differ between sibling aphid lines that harbor the same symbiont genotype. We find significant differences in host fitness between aphid clones and show that this variation in fitness is correlated with symbiont titer, where higher titer levels are correlated with lower host fitness. The work has been published in a scientific journal (Chong and Moran, Proc. Natl. Acad. Sci. USA. 2016). The PD has gained experience by designing the experiment, performing insect (and plant) husbandry, conducting experimental crosses with aphids, learning new molecular laboratory techniques including Quantitative PCR, using bioinformatic tools, and preparing a manuscript for publication. (2) Using a combination of sexual crosses and the Buchnera transfer technique, enabling the replacement of obligate symbiont genotypes without altering host genotype, we are testing whether rapid and degenerative evolution of symbiont genomes drives adaptation and diversification in host populations. We have successfully generated novel genetic combinations of symbiont and host genotypes through sexual crosses and are currently conducting host-symbiont transfers. Parental aphid clones have been sequenced using Illumina MiSeqV3 and symbiont genomes have been assembled and annotated. Accomplishment of these goals provides the PD with training in experimental design and genetics manipulations in an insect symbiont system. In addition in insect husbandry, the PD has gained experience in microscopy and microinjection techniques as well as skills in bioinformatics by using high-throughput methods to sequence, assemble, and annotate complete symbiont genomes. (3) This project involves designing and performing experiments, learning new techniques and skills, publishing research, participating in conferences and seminars, and outreach and mentorship through scientific training of underrepresented minority groups. As a USDA postdoctoral fellow, the PD mentored has trained three undergraduate students and two graduate students during this project period including four students from underrepresented groups.

Publications

  • Type: Journal Articles Status: Published Year Published: 2016 Citation: Chong, R. A. and Moran, N. A. (2016) Intraspecific genetic variation in hosts affects regulation of obligate heritable symbionts. Proceedings of the National Academy of Sciences USA 113(46): 13114-13119
  • Type: Conference Papers and Presentations Status: Other Year Published: 2016 Citation: Heritability of obligate symbiont titer and link to aphid host fitness. Oral presentation at Evolution Conference 2016 in Austin, TX.