SYSTEMATICS, TAXONOMY, BIODIVERSITY, AND FOOD-WEB INTERACTIONS OF ICHNEUMONIDAE (INSECTA: HYMENOPTERA)

• Sponsoring Institution: National Institute of Food and Agriculture
• Project Status: TERMINATED
• Funding Source: HATCH
• Reporting Frequency: Annual
• Accession No.: 1002019
• Project No.: KY008065
• Project Start Date: Jan 22, 2014
• Project End Date: Sep 30, 2018

Project Director
Sharkey, M.

Recipient Organization
UNIVERSITY OF KENTUCKY
500 S LIMESTONE 109 KINKEAD HALL
LEXINGTON,KY 40526-0001

Performing Department
Entomology

Non Technical Summary
Understanding the Earth's biodiversity is central to many environmental issues facing human society today. If the most recent estimates of approximately 6.1 million arthropod species (the majority of non-bacterial species) are accurate, then over 250 years of modern taxonomy has only described about a fifth of the Earth's biodiversity (Hamilton et al. 2013; Bisby et al. 2010). Knowledge of animals, plants, and microorganisms is fundamental to our understanding, and successful management, of the earth's biosphere and to the maintenance of the health and economy of the nation. All biological research, pure and applied, is dependent on systematics. An understanding of the phylogenetic relationships among biological entities provides the foundation of all predictive classifications. These in turn are tools crucial to all aspects of resource identification, exploitation and preservation. Both parts of the project presented herein will help to address this issue by resulting in new species descriptions of species of potential economic importance (in biological control). Part one will result in published revisions of groups of braconid wasps, of which there are estimated to be 13,000-33,000 undescribed species (Jones et al. 2009; Dolphin & Quicke 2001). Because they attack and kill other arthropods, parasitic wasps are a critical component of our arsenal against insect pests that are of agricultural, silvicultural, domestic, and medical importance. We will collect thousands of specimens with the appropriate traps, sequence key parts of their genomes that will allow us to discover new species, revise the species-groups to which they belong, and through phylogenetics, understand how they are related to other members of their respective groups. Knowledge of evolution is important for predicting what parasitoids may be important for the biological control. The second part of this proposal will greatly aid in the understanding of connections between species. Here, we fog trees with pesticides in order to sample the canopy for arthropods. Focusing on caterpillars, we will use DNA-based methods to determine (1) their identity, (2) what plant species they are feeding on (in addition to the tree itself, rainforest trees house a large number of epiphytes, which will be sequenced), and (3) what wasp and fly species are parasitizing them. Like part one, this study will result in specimens for revisionary work, as collections will be made to compare DNA sequences for which we have no specimens (caterpillar parasites) to those associated with specimens collected with traps. This part of the project will also yield knowledge about how species in little-known ecosystems are connected. Understanding how species are connected provides more evidence for what might happen if certain species go extinct. Furthermore, understanding what species parasitize the caterpillars in this study would be extremely helpful to biological control programs in cases where these caterpillar species may be become pests if accidentally introduced into areas where they have few to no natural enemies (~40% of insect pests in the U.S. are exotic in origin).

Animal Health Component

0%

Research Effort Categories

Basic

90%

Applied

(N/A)

Developmental

10%

Classification

Knowledge Area (KA)	Subject of Investigation (SOI)	Field of Science (FOS)	Percent
135	3110	1130	100%

Knowledge Area
135 - Aquatic and Terrestrial Wildlife;

Subject Of Investigation
3110 - Insects;

Field Of Science
1130 - Entomology and acarology;

Keywords

systematics

taxonomy

biodiversity

food-web connections

ichneumonoidea

braconidae

Goals / Objectives
1. To describe, identify, and produce phylogenetic hypotheses (classifications) for genera and higher taxa of selected parasitic wasps and describe and provide identification keys for constituent taxa. 2. To determine and compare arthropod species diversity, host diversity, and interaction diversity in various habitats.

Project Methods
I. Phylogenetic analyses and descriptions of various groups of Hymenoptera (Insecta) using morphology and molecular sequence data. In revisionary studies the first step is to obtain specimens and specimen records. This is done by borrowing material from museums, visiting museum collections, collecting specimens and searching the literature. Taxa of concern are then separated and their characteristics are compiled. Using the software DELTA (Partridge et al. 1992), hard-copy keys and descriptions are produced. Using the software INTKEY (Dallwitz and Paine 1990) internet-based interactive keys are produced. Phylogenetic hypotheses (trees) employing the parsimony criterion will be generated using the software packages TNT (Golobof et al. 2008) and discriminate compatibility (Sharkey 1994). The data sets are analyzed under Bayesian Inference with MrBayes (Huelsenbeck & Ronquist 2001) and Maximum Likelihood using Garli (Zwickl 2006) under the appropriate model of evolution as determined via jModelTest (Posada 2008) and conducted for as many generations as necessary to produce robust relationships. Standard methods for DNA extraction, amplification and sequencing are as follows: a. DNA extraction. Appendages are extracted using the Qiagen DNEasy extraction kit following the animal tissue protocol. The specimen from which a leg was extracted is given a unique identification number so all molecular data generated from that specimen can be tracked. b. PCR amplification. PCR reactions (25 μL total) consist of 1X Takara PCR buffer, 0.2 mM of each dNTP, 0.2 mM of each primer, 0.625 U of Takara Ex Taq and 1-2 μL of template DNA. PCR cycling conditions for COI primers (LepF1 & LepR1; Hebert et al, 2004) are 40 cycles of 95°C for 50 s, 40°C for 45 s, and 72°C for 45 s. Cycling conditions for 28S (28SD2hymF & 28SD2hymR; Sharkey & Sloelb 2012) are similar except the annealing temperature is 53°C. CAD is amplified with 2 primer pairs: (I) 54F & 405R follow the protocols published in Moulton & Wiegmann (2004) and (2) CADforI & CADrevImod follow the protocols published in Danforth (2007). c. DNA sequencing: PCR reactions that yield product are purified with the QIAGEN MinElute PCR purification kit according to the manufacturer's guidelines. Both strands are cycle sequenced using labeled dideoxynucleotides (ABI Big~Dye Tenninator mix v. 3.0) and sequenced at a commercial lab. The collection and phylogenetic analysis of DNA data will be used to test morphologically-based species concepts. In our previous studies, the COI gene has been very useful at uncovering species not detected with morphology alone. Once DNA illuminates the presence of these 'cryptic' species, it is usually possible to find morphological characters that are diagnostic of the newly-detected species (e.g., Sharkey & Stoelb 2013, 2012a,b; van Achterberg et al. in press). Two formal courses teaching many of the techniques outlined above are taught at the University of Kentucky every other year: Insect Taxonomy teaches the principles of insect identification with labs twice weekly to gain practical experience in using taxonomic keys. In Phylogenetic Systematics, DNA extraction, PCR, sequencing and phylogenetic analysis methods are covered in detail. Here, students conduct a study that usually results in the revision of a small taxonomic group in which they generate molecular data, analyze it and make decisions regarding species composition. The class project from the spring semester, 2010 course resulted in a publication (Tucker et al. 2011). As with all taxonomic revisions, success is measured in publication of results. PI Sharkey has consistently averaged publishing at least three revisions per year for the last decade, each including interactive (web-based) keys and Delta data matrices and image files freely accessible on the internet (see http://sharkeylab.org/sharkeylab/sharkeyCV.php for a complete list). II. Determine and compare arthropod species diversity, host diversity, and interaction diversity in various habitats. Methods are as follows using ash trees as an example: Step I. Ash trees are fogged with pyrethrum in the first light of day (Erwin 1983) using a Curtis Superhawk XP commercial fogging machine. Below each tree, 5 sheets, each 10m x 10m, are spread and falling arthropods are collected. Each fogging event is given a unique identifier. Plant samples are taken and chloroplast genes rbcL and trnL are obtained. Step 2. Non-target arthropods are placed in 95% ethanol and stored at -20°C. Step 3. Lepidoptera parasitoids and hyperparasitoids are separated, placed in 95% ethanol, and stored at -20°C. Step 4. Caterpillars are sorted to morphospecies, given a unique identifier, imaged on a sheet of graph paper to allow for measurements (for later identification of larval instar), and stored at -20C. Each specimen is housed in a separate vial, each with a collection event label and barcoded specimen label, and databased following protocols developed by the TriTrophic Thematic Collection Network (NSF:ADBC 1115144) and stored at -80°C. Step 5. Refine caterpillar species concepts with COI data. A proleg of 2 or 3 morphologically disparate specimens of each morphospecies is clipped and used to obtain COI. Caterpillars are resorted and re-identified based on these results in combination with morphological inference. Caterpillar COI is blasted on BOLD and GenBank to obtain genus and species-level identifications. Where one or both of these is not available, specimens are identified to the lowest possible taxon and given unique species identifiers. Step 6. Confirm the host food of each species. Caterpillars are pulverized and their gut contents sequenced for rbcl and trnL for three samples of each caterpillar species, for each fogging event, and identified via comparison to GenBank and/or the rbcl and trnL libraries built in Step I. Step 7. We have immunological assays (ELISA) for Diptera- and Hymenoptera-specific antibodies allowing us to determine which caterpillar specimens are parasitized (estimated parasitism rate = 20-30%). Compared to PCR, ELISA is inexpensive and hundreds of samples can be tested simultaneously. Step 8. Obtain 16S data for each larval parasitoid. We have designed two 16S primer pairs, one specific for Diptera, and the other for Hymenoptera. Depending on the parasitoid present (determined with ELISA in step 6) the appropriate 16S primers are applied to host extractions. Step 9. Match the 16S data obtained from parasitoid larvae with the 16S library of adult parasitoids collected from foggings and traps. Adult insects are also collected in and near the focal tree using a suite of techniques (Malaise and light traps). These specimens are treated as follows: Parasitoids of Lepidoptera are separated and placed in fresh 95% ethanol, stored at -20°C, and sorted to morphospecies. All other hymenopteran parasitoids are sorted to morphospecies in the Sharkey lab. Several representatives of each morphospecies are sequenced for 16S and COI, and morphospecies hypotheses are reexamined based on the results. This procedure will make food-web connections between forest caterpillars, their food plants and their dipteran and hymenopteran parasites. Plant-specific and parasite-specific primers will allow amplification and sequencing of plant and parasite DNA from caterpillar DNA extractions. Success of this part of the project will be determined by the number of new host and parasite associations discovered, and because general collecting will be done, new species will be discovered, leading to more revisionary work with braconids. The molecular methods that we develop to uncover food-web connections in forest canopies will be useful to those wishing to conduct similar studies in other areas.

Progress 01/22/14 to 09/30/18

Outputs
Target Audience: Hymenoptera systematists/phylogeneticists who could use the DNA sequences generated by my work in other studies, or who are interested in using the relationships uncovered in my work to infer evolutionary and biological processes and patterns. Scientists interested in the use of parasitic wasps for biological control. Scientists interested in investigating host-parasitoid food webs. Scientists who need to identify wasps in the genera I have been revising. Land managers interested in monitoring and maintaining biodiversity and understanding trophic interactions between plants, herbivores and their parasites occurring on the lands they manage. Changes/Problems: Nothing Reported What opportunities for training and professional development has the project provided?In the reporting period, we graduated one Master's student who is now working for the UK department of Entomology in various capacities. Hatch funds directly contributed to her research. We finished hosting a visiting scholar from China who directly benefited from the Hatch funds our lab received. The visiting scholar (Junli Yao) received hundreds of parasitic wasp specimens from Dr. Sharkey that he had collected over the years (partially using Hatch funds). Because they were stored properly for DNA work (kept below -20°C in ethanol), she was able to sequence their DNA and publish three papers in 2018 - without the DNA sequences, this work would have been much more difficult. She presented the results of this work at a pest control symposium in China (see Products section of this report). How have the results been disseminated to communities of interest?We have published the results of our research in 7 publications in the past year. We submitted 236 DNA sequences to the Barcode of Life Database that are now publically available (referenced in said papers). Our Hymenoptera database is continually updated and available to the public at http://symbiota4.acis.ufl.edu/scan/portal/index.php - many hours were spent this year updating the information in this database - many of specimens were added, including digital images of some. A total of 3 presentations were given by our lab members at national and regional scientific meetings. Most of our papers are available for download (depending on journal permissions) from our website: http://sharkeylab.org/sharkeylab/sharkeyCV.html. What do you plan to do during the next reporting period to accomplish the goals? Nothing Reported

Impacts
What was accomplished under these goals? Our published work resulted in the description of 3 new genera and 91 species new to science and a number of new synonymies and new combinations. We collected new specimens and added them to the holdings at the Hymenoptera Institute for comparative purposes and morphological study. We have added digital information on hundreds of braconid specimens to an ever-growing, web accessible Integrated Digitized Biocollections (IDigBio)-supported database (Symbiota; http://symbiota4.acis.ufl.edu/scan/portal/collections/misc/collprofiles.php?collid=44). The lab's biggest publication in 2018, a revision of the genus Alabagrus in Costa Rica, resulted in the description of 65 new species, all reared from caterpillars in the Area de Conservacion Guanacaste. The findings of this study have wide-ranging implications. First, the 65 new species were all reared from caterpillars collected from plants identified to species as well. So, we now have a much better understanding of which wasp species feed on which caterpillar species. Secondly, this study has the potential for great economic importance if any of these caterpillar species ever get accidentally transported to an area where they become an invasive pest. In this scenario, we would already know which parasitoid wasp species could be used in the biological control of the new caterpillar pest. Furthermore, our previous work, published in 2017, developed DNA primers to detect the presence of parasitoids inside caterpillars. So not only would scientists be able to determine which species of parasitoid would be suitable for biological control, but they would also be able to collect caterpillars and determine if they had been parasitized and even calculate the rate of parasitism with a simple PCR test. Our other published work involved revising or contributing to the knowledge of species of braconid wasps in Thailand and Egypt. Stemming from NSF-supported work that began in 2006, we published four papers on the braconid fauna collected from diverse habitats in Thailand. The paper first-authored by Sharkey resulted in the description of 19 new species and a new genus. We maintained specimens from the Thai material in 95% ethanol at -20°C so it would be viable for DNA work. Dr. Yao, the visiting scholar, was able to separate out a few hundred specimens of her group of interest and obtain DNA sequences from them. This work led to three publications in 2018 with at least 3-4 more papers to be published from this same material. New species were described and new distribution records for Thailand were published. Lastly, we collaborated in a project to revise a subfamily of braconids in Egypt that resulted in the description of a new species. All of these publications included taxonomic keys to identify the species under study, including full color illustrations. Type specimens were designated and sent to the appropriate museums so that other researchers have access to them.

Publications

• Type: Journal Articles Status: Published Year Published: 2018 Citation: Sharkey MJ, Chapman EG. 2018. Revision of Zosteragathis Sharkey of Thailand (Hymenoptera, Braconidae, Agathidinae, Agathidini). Deutsche Entomologische Zeitschrift 65(2): 225-253.
• Type: Journal Articles Status: Published Year Published: 2018 Citation: Sharkey MJ, Chapman EG. 2018. Addendum to "Phylogeny of the Agathidinae (Hymenoptera: Braconidae) with a revised tribal classification and the description of a new genus" (Sharkey and Chapman, 2017, Proc Entomol Soc Wash 119 (Special Issue):823-842). Proceedings of the Entomological Society of Washington 120(3): 580-586.
• Type: Journal Articles Status: Published Year Published: 2018 Citation: Yao Y, Achterberg C van, Sharkey MJ, Chapman EG, Chen J. 2018. Two species and a genus new for Thailand, with description of a new species of Neurolarthra Fischer (Hymenoptera: Braconidae: Alysiinae). Zootaxa 4438(1): 551-560.
• Type: Journal Articles Status: Published Year Published: 2018 Citation: Yao Y, Achterberg C van, Sharkey MJ, Chen J. 2018. Separatatus Chen & Wu (Hymenoptera: Braconidae: Alysiinae) newly recorded from Thailand, with description of one new species and one new combination. Zootaxa 4433(1): 187-194.
• Type: Journal Articles Status: Published Year Published: 2018 Citation: Edmardash YA, Gadallah NS, Sharkey MJ. 2018. Revision of the subfamily Cardiochilinae Ashmead, 1900 (Hymenoptera: Braconidae) in Egypt, with new records and a new species. Journal of Natural History 52(5-6): 269-297.
• Type: Journal Articles Status: Published Year Published: 2018 Citation: Sharkey MJ, Meierotto S, Chapman EG, Janzen DJ, Hallwachs W, Dapkey T, Solis MA. 2018. Alabagrus Enderlein (Hymenoptera, Braconidae, Agathidinae) species of Costa Rica, with an emphasis on specimens reared from caterpillars in Area de Conservaci�n Guanacaste. Contributions in Science 526: 31-180.
• Type: Conference Papers and Presentations Status: Published Year Published: 2018 Citation: Yao J, Achterberg C van, Sharkey MJ, Chapman EG, Chen J. The taxonomy of Alysiini of Thailand. Oral Presentation. Second symposium on insect resources and pest control in tropical and subtropical regions. 16-19 November 2018. Kunming-Xishuangbanna, China.
• Type: Journal Articles Status: Published Year Published: 2018 Citation: Yao Y, Achterberg C van, Sharkey MJ, Chapman EG, Chen J. 2018. Senwot Wharton, a new genus for the Oriental region with description of two new species from Thailand (Hymenoptera: Braconidae: Alysiinae). Journal of Asia-Pacific Entomology 22(2019): 103-109.

Progress 10/01/16 to 09/30/17

Outputs
Target Audience:• Hymenoptera systematists/phylogeneticists who could use the DNA sequences generated by my work in other studies, or who are interested in using the relationships uncovered in my work to infer evolutionary and biological processes and patterns. • Scientists interested in the use of parasitic wasps for biological control. • Scientists interested in investigating host-parasitoid food webs - we published new primers that are very useful for this purpose. • Scientists who need to identify wasps in the genera I have been revising. • Land managers interested in monitoring and maintaining biodiversity and understanding trophic interactions between plants, herbivores and their parasites occurring on the lands they manage. Changes/Problems: Nothing Reported What opportunities for training and professional development has the project provided?In the reporting period, we graduated one Master's student who went on as a Ph. D. student in Dr. Michael Stout's lab at Louisiana State University. Our other Master's student matriculated and is now working on her Ph. D. in our lab. Hatch funds directly contributed to their research. We are currently hosting a visiting scholar from China who will indirectly benefit from the Hatch funds our lab receives. Morphological studies of braconid wasps are made possible by the many thousands of specimens in the Hymenoptera Institute Collection available for study by our graduate students. Furthermore, the connections that PI Sharkey has to museums around the world facilitates the borrowing of specimens by graduate students in our lab which are critical for their studies. One of our Master's students collaborated with Dr. Dan Janzen whose ongoing project is discovering host plant-caterpillar-parasitoid relationships in Costa Rica - he received hundreds of specimens from Dr. Janzen and studied the evolution of host associations in the braconid genus Lytopylus and described over 30 new species (Kang et al. 2017; see publication list). Our students presented their research at the annual meeting of the Entomological Society of America (ESA) in Denver, Colorado in November of 2017 - this research is improved by the available Hatch funds. Additionally, our Ph. D. student won 2nd place in the student competition at the North-Central branch meeting of ESA Indianapolis, Indiana in June, 2017. How have the results been disseminated to communities of interest?We have published the results of our research in 5 publications in the past year. We submitted 380 DNA sequences to GenBank that are now publically available (referenced in said papers). Our Hymenoptera database is continually updated and available to the public at http://symbiota4.acis.ufl.edu/scan/portal/index.php - many hours were spent this year updating the information in this database - hundreds of specimens were added, including digital images of some. A total of 5 presentations were given by our lab members at national and regional scientific meetings. Most of our papers are available for download (depending on journal permissions) from our website: http://sharkeylab.org/sharkeylab/sharkeyCV.html. What do you plan to do during the next reporting period to accomplish the goals?We have numerous ongoing studies in Hymenoptera systematics, including (1) a key to the genera of Agathidinae (Braconidae) which is necessary due to the plethora of new synonymies and combinations we have published and (2) a revision of the genus Crassomicrodus, with multiple new species described. We are continuing to develop our methods for molecular determination of biodiversity. Other ongoing projects include (1) using high-throughput sequencing to determine biodiversity from bulk insect samples taken from a site in Kentucky and (2) comparing the diversity of parasitoid wasps between cites in central Kentucky and French Guiana, and (3) revisionary work on the Alysiinae (Braconidae) of Thailand. Hatch funds continue to play an integral role towards obtaining data.

Impacts
What was accomplished under these goals? Our published work resulted in the description of 10 new genera and 127 species new to science and a long list of new synonymies and new combinations. We collected new specimens and added them to the holdings at the Hymenoptera Institute for comparative purposes and morphological study. This year, we returned thousands of borrowed specimens to many museums that we identified to the lowest possible taxon - by providing identification services, we benefit by capturing specimen information and the museums benefit by having their specimens identified by a world-recognized expert. We have added digital information on many hundreds of braconid specimens to an ever-growing, web accessible Integrated Digitized Biocollections (IDigBio)-supported database (Symbiota; http://symbiota4.acis.ufl.edu/scan/portal/collections/misc/collprofiles.php?collid=44). We have developed a better understanding of the relationships among agathidine braconid wasps and their host associations. To this end, we published the most comprehensive molecular phylogeny of the subfamily Agathidinae (Braconidae) to date and revised the tribal classification. This knowledge could aid in biological control efforts in cases where one or more hosts become pests, as species-specific parasitoids have been among the most successful biological control agents. To further this knowledge, we published primers that will amplify parasitoid wasps (from the superfamilies Ichneumonoidea and Chalcidoidea) or parasitoid flies (from the family Tachinidae) from within lepidopteran hosts. These sequences can be used to identify parasitoids at least to genus which will aid researchers in identifying parasitoid larvae within their hosts, a process formerly only accomplished by rearing, which can be very difficult in some lepidopteran taxa. Our papers have led to a better understanding of the identification and phylogenetic relationships of the parasitoid species contained within, which is essential information when developing biological control programs. A better understanding of the biodiversity in selected study areas is becoming increasingly important as we try to understand how to maintain and manage our natural resources. We continue to develop molecular techniques to identify arthropods in bulk field samples. These techniques will allow land managers to assess the arthropod diversity in natural areas at a very low cost (under $10k), whereas studies involving hundreds of taxonomists and years of study can cost millions of dollars. Consequently, comprehensive biodiversity studies are currently thought to be highly impractical, and therefore almost never done. Our studies aim to make such studies not only feasible, but an essential tool for evaluating the value of natural areas and the ecosystem services provided within. Finally, PI Sharkey prepared and submitted two NSF grant proposals; one to study the parasitoid-host trophic interactions of the Lepidoptera of eastern North America and their ichneumonoid and tachinid parasites, and the other to develop new methods to rapidly describe and publish biodiversity. Both were submitted in January 2017.

Publications

• Type: Journal Articles Status: Published Year Published: 2017 Citation: Sharkey MJ, Chapman EG. 2017. Ten new genera of Agathidini (Hymenoptera: Braconidae: Agathidinae) from Southeast Asia. Zookeys 660: 107-150.
• Type: Journal Articles Status: Published Year Published: 2017 Citation: Kang I, Chapman EG, Janzen DJ, Hallwachs W, Solis MA, Sharkey MJ. Revision of the species of Lytopylus from Area de Conservaci�n Guanacaste, northwestern Costa Rica (Hymenoptera, Braconidae, Agathidinae). ZooKeys 721:93-158.
• Type: Journal Articles Status: Accepted Year Published: 2017 Citation: Sharkey MJ, Chapman EG. 2017. Phylogeny of the Agathidinae (Hymenoptera: Braconidae) with a revised tribal classification and the description of a new genus. Proceedings of the Entomological Society of Washington (in press).
• Type: Journal Articles Status: Accepted Year Published: 2017 Citation: Sharkey MJ, Meierotto S, Chapman EG, Janzen DJ, Hallwachs W, Dapkey T, Solis MA. 2017. Alabagrus Enderlein (Hymenoptera, Braconidae, Agathidinae) species of Costa Rica, with an emphasis on specimens reared from caterpillars in Area de Conservaci�n Guanacaste. Contributions in Science (in press).
• Type: Journal Articles Status: Published Year Published: 2017 Citation: Pook V, Athey KJ, Chapman EG, Clutts-Stoelb S, Sharkey MJ. 2017. New PCR primers enhance investigation of host-parasitoid food-webs. Entomologia Experimentalis et Applicata 162 (3): 309-314.

Progress 10/01/15 to 09/30/16

Outputs
Target Audience:• Hymenoptera systematists/phylogeneticists who could use the DNA sequences generated by my work in other studies, or who are interested in using the relationships uncovered in my work to infer evolutionary and biological processes and patterns. • Scientists interested in the use of parasitic wasps for biological control. • Scientists who need to identify wasps in the genera I have been revising. • Land managers interested in monitoring and maintaining biodiversity and understanding trophic interactions between plants, herbivores and their parasites occurring on the lands they manage. Changes/Problems: Nothing Reported What opportunities for training and professional development has the project provided?In 2016, we graduated one Ph.D. student who went on as a post-doctoral scholar in Dr. Seth Debolt's lab in the Horticulture Department here at the University of Kentucky. We currently have two Master's students in our lab who have made significant progress towards their projects in 2016. Hatch funds have directly contributed to their research. We are currently hosting a visiting scholar from China who will indirectly benefit from the by Hatch funds our lab receives. Morphological studies of braconid wasps are made possible by the many thousands of specimens in the Hymenoptera Institute Collection available for study by our graduate students. Furthermore, the connections that PI Sharkey has to museums around the world facilitates the borrowing of specimens by graduate students in our lab which are critical for their studies. One of our Master's students is collaborating with Dr. Dan Janzen whose ongoing project is discovering host plant-caterpillar-parasitoid relationships in Costa Rica - he has received hundreds of specimens from Dr. Janzen and is studying the evolution of host associations in the braconid genus Lytopylus and describing over 30 new species. Our students presented their research at the joint annual meeting between the Entomological Society of America and the International Congress of Entomology in Orlando, Florida in September of 2016 - this research is improved by the available Hatch funds. One of our Master's students received an NSF fellowship to study in Dr. Meier's lab in the summer of 2016 - she would have been less competitive for this fellowship if it were not for the Hatch funds our lab receives. How have the results been disseminated to communities of interest?We have published the results of our research in 6 publications in the past year. We submitted over 300 DNA sequences to GenBank and are now publically available (referenced in said papers). Our Hymenoptera database is continually updated and available to the public at http://symbiota4.acis.ufl.edu/scan/portal/index.php - many hours were spent this year updating the information in this database - hundreds of specimens were added, including digital images of some. Three presentations were given by our lab members at the International Congress of Entomology. Most of our papers are available for download (depending on journal permissions) from our website: http://sharkeylab.org/sharkeylab/sharkeyCV.html. What do you plan to do during the next reporting period to accomplish the goals?We have numerous ongoing studies in Hymenoptera systematics, including (1) a revision of the genus Alabagrus in Costa Rica in which we will be describing dozens of new species (2) a revision of the genus Crassomicrodus, again with multiple new species described. We are continuing to develop our methods for molecular determination of biodiversity. Other ongoing projects include (1) using high-throughput sequencing to determine biodiversity from bulk insect samples taken from a site in Kentucky and (2) comparing the diversity of parasitoid wasps between cites in central Kentucky and French Guiana. PI Sharkey has prepared and submitted two NSF grant proposals; one to study the parasitoid-host trophic interactions of the Lepidoptera of eastern North America and their ichneumonoid and tachinid parasites, and the other to develop new methods to rapidly describe and publish biodiversity. Both were submitted in January 2017. Hatch funds continue to play an integral role towards obtaining preliminary data to support our grant proposals.

Impacts
What was accomplished under these goals? My published work resulted in the description of 10 genera and 29 species new to science and a long list of new synonymies and new combinations. We have increased in the specimen holdings at the Hymenoptera Institute and other museums for comparative purposes and morphological study. This year, we returned hundreds of borrowed specimens to many museums that we identified to species - by providing identification services, we benefit by capturing specimen information and the museums benefit by having their specimens identified by a world-recognized expert. We have added digital information on many hundreds of braconid specimens to an ever-growing, web accessible Integrated Digitized Biocollections (IDigBio)-supported database (Symbiota; http://symbiota4.acis.ufl.edu/scan/portal/collections/misc/collprofiles.php?collid=44). We have developed a better understanding of the relationships among braconid wasps and their host associations. This knowledge has been published (see publications). This knowledge could aid in biological control efforts in cases where one or more hosts become pests, as species-specific parasitoids have been among the most successful biological control agents. Our papers have led to a better understanding of the identification and phylogenetic relationships of the parasitoid species contained within, which is essential information when developing biological control programs. A better understanding of the biodiversity in selected study areas is becoming increasingly important as we try to understand how to maintain and manage our natural resources. We are developing molecular techniques to identify arthropods in bulk field samples. These techniques will allow land managers to assess the arthropod diversity in natural areas at a very low cost (under $10k), whereas studies involving hundreds of taxonomists and years of study can cost millions of dollars. Consequently, comprehensive biodiversity studies are currently thought to be highly impractical, and therefore almost never done. Our studies aim to make such studies not only feasible, but an essential tool for evaluating the value of natural areas and the ecosystem services provided within. We have developed collaborative relationships with two recognized experts in this field (Matthieu Leray, a post-doc at the Smithsonian Tropical Research Institute in Panama and Rudolf Meier at the National University of Singapore) who are collaborating on three current biodiversity-related projects.

Publications

• Type: Journal Articles Status: Published Year Published: 2016 Citation: Tucker EM, Sharkey MJ. 2016. Deserters of Cremnops desertor (Hymenoptera: Braconidae: Agathidinae): Delimiting species boundaries in the C. desertor species-complex. Systematics and Biodiversity 2016: 1-9.
• Type: Journal Articles Status: Published Year Published: 2016 Citation: Li Q, Wei S-j, Tang P, Wu Q, Shi M, Sharkey MJ, Chen X-x. 2016. Multiple lines of evidence from mitochondrial genomes resolve phylogenetic relationships of parasitic wasps in Braconidae. Genome Biology and Evolution, advance Access published August 8, 2016 doi:10.1093/gbe/evw184
• Type: Journal Articles Status: Published Year Published: 2016 Citation: Sharkey MJ, Chapman EG. 2016. Revision of Aerophilus Sz�pligeti (Hymenoptera, Braconidae, Agathidinae) from eastern North America, with a key to the Nearctic species. Contributions in Science 524: 51-110.
• Type: Journal Articles Status: Awaiting Publication Year Published: 2017 Citation: Pook V, Athey KJ, Chapman EG, Clutts-Stoelb S, Sharkey MJ. 2017. New PCR primers enhance investigation of host- parasitoid food-webs. Entomologia Experimentalis et Applicata (in press).
• Type: Journal Articles Status: Awaiting Publication Year Published: 2017 Citation: Sharkey MJ, Chapman EG. 2017. Ten new genera of Agathidini (Hymenoptera: Braconidae: Agathidinae) from Southeast Asia. Zootaxa (in press).
• Type: Journal Articles Status: Published Year Published: 2016 Citation: Pook V, Sharkey MJ, Wahl D. 2016. Key to the species of Megarhyssa (Hymenoptera, Ichneumonidae, Rhyssinae) in America, north of Mexico. Deutsche Entomologische Zeitschrift 63(1): 137-148.

Progress 10/01/14 to 09/30/15

Outputs
Target Audience: Hymenoptera systematists/phylogeneticists who might use the DNA sequences generated in this study in other studies, or who are interested in using the relationships uncovered in this study to infer evolutionary and biological processes and patterns. Scientists interested in the use of parasitic wasps for biological control. Land managers interested in monitoring and maintaining biodiversity and understanding trophic interactions between plants, herbivores and their parasites occurring on the lands they manage. Changes/Problems: Nothing Reported What opportunities for training and professional development has the project provided?In 2015, we graduated one Ph.D. student who went on as a post-doctoral scholar at the University of New Hampshire. We currently have 1 PhD and 2 Master's students in our lab who are making significant progress. Hatch funds have directly contributed to their research. We have also hosted 2 international Ph.D. students in our lab in the past year from China and Brazil. These students learned DNA techniques by helping others in the lab with their projects that were supported in part by Hatch funds. Morphological studies of braconid wasps are made possible by the many thousands of specimens in the Hymenoptera Institute Collection available for study. Furthermore, the connections that PI Sharkey has to museums around the world allows students in our lab to borrow specimens critical for their studies. Our Ph.D. students accounted for 3 of our papers published this year. Furthermore, our students presented their research at the annual meeting of the Entomological Society of America - this research is made better by the available Hatch funds. Our PhD student was a finalist in the university-wide 3-minute Dissertation competition. How have the results been disseminated to communities of interest?We have published the results of our research in 11 publications in 2015. Many DNA sequences have been submitted to GenBank and are now publically available (referenced in said papers). Our Hymenoptera database is continually updated and available to the public at http://symbiota4.acis.ufl.edu/scan/portal/index.php - hundreds of hours were spent this year updating the information in this database - thousands of specimens were added, including digital images of many. Six presentations were given by our lab members at regional and national meetings. Most of our papers are available for download (depending on journal permissions) from our website: http://sharkeylab.org/sharkeylab/sharkeyCV.html. What do you plan to do during the next reporting period to accomplish the goals?We have numerous ongoing studies in Hymenoptera systematics, at least 2 of which are close to being submitted for publication, including (1) a paper on a group of agathidines from Thailand in which new species are being described (2) a revision of the genus Crassomicrodus, again with multiple new species described and (3) a high-throughput sequencing project to determine biodiversity from bulk insect samples. We are continuing to develop our methods for molecular determination of biodiversity: we have two ongoing projects on this front - one determining biodiversity from a comprehensive set of samples from a forest in central Kentucky and another in French Guiana. PI Sharkey is preparing an NSF grant proposal to study the parasitoid-host trophic interactions of the Lepidoptera of eastern North America and their ichneumonoid and tachinid parasites, to be submitted January 2016. Hatch funds continue to play an integral role towards obtaining preliminary data to support our grant proposals.

Impacts
What was accomplished under these goals? We have increased in the specimen holdings at the Hymenoptera Institute and other museums for comparative purposes and morphological study. This year, we returned thousands of borrowed specimens to many museums that we identified to species - by providing identification services, we benefit by being able to catalog specimen information and the museums benefit by having their specimens identified by a world-recognized expert. We have added digital information on many hundreds of braconid specimens to an ever-growing, web-accessible Integrated Digitized Biocollections (IDigBio)-supported database (Symbiota; http://symbiota4.acis.ufl.edu/scan/portal/index.php). We have developed a better understanding of the relationships among braconid wasps and their host-associations. This knowledge has been published (see publications) and could aid in biological control efforts in cases where one or more hosts become pests, as species-specific parasitoids have been among the most successful biological control agents. Our papers have led to a better understanding of the identification and phylogenetic relationships of the parasitoid species contained within, which is essential information when developing biological control programs. A better understanding of the biodiversity in selected study areas is becoming increasingly important as we try to understand how to maintain and manage our natural resources. We are developing molecular techniques to identify arthropods in bulk field samples. These techniques will allow land managers to assess the arthropod diversity in natural areas at a very low cost (under $10k), whereas studies involving hundreds of taxonomists and years of study can cost millions of dollars. Consequently, comprehensive biodiversity studies are currently thought to be highly impractical, and our studies aim to make such studies not only feasible, but an essential tool for evaluating the value of natural areas and the ecosystem services provided within. We have developed collaborative relationships with two recognized experts in this field (Matthieu Leray, a post-doc at the Smithsonian Institution and Rudolf Meier at the National University of Singapore) who are collaborating on three of our current biodiversity-related projects.

Publications

• Type: Journal Articles Status: Published Year Published: 2015 Citation: Letourneau DK, Bothwell Allen SG, Kula RR, Sharkey MJ, Stireman JO, III. 2015. Habitat eradication and cropland intensification may reduce parasitoid diversity and natural pest control services in annual crop fields. Elementa: Science of the Anthropocene DOI 10.12952/journal.elementa.000069
• Type: Journal Articles Status: Published Year Published: 2015 Citation: Azevedo CO, Dal Molin A, Penteado-Dias A, Macedo ACC, Rodriguez-V B, Dias BZK, Waichert C, Aquino D, Smith DR, Shimbori EM, Noll FB, Gibson G, Onody HC, Carpenter JM, Lattke JE, Ramos KdS, Williams K, Masner L, Kimsey LS, Tavares MT, Olmi M, Buffington ML, Ohl M, Sharkey MJ, Johnson NF, Kawada R, Gon�alves RB, Feitosa RM, Heydon S, Guerra TM, da Silva TSR, Costa V.2015. Checklist of the genera of Hymenoptera (Insecta) from Esp�rito Santo state, Brazil. Bol. Mus. Biol. Mello Leit�o (N. S�r.) 37(3):313-343.
• Type: Journal Articles Status: Published Year Published: 2015 Citation: Wang M, Rasnitsyn AP, Shih C, Sharkey MJ, Ren A. 2015. New fossils from China elucidating the phylogeny of Praesiricidae (Insecta: Hymenoptera). Systematic Entomology doi: 10.1111/syen.12142
• Type: Journal Articles Status: Published Year Published: 2015 Citation: Fisher JR, Tucker EM, Sharkey MJ. 2015. Colemanus keeleyorum (Braconidae, Ichneutinae s. l.): a new genus and species of Eocene wasp from the Green River Formation of western North America. Journal of Hymenoptera Research 44: 57-67.
• Type: Journal Articles Status: Published Year Published: 2015 Citation: Pook VG, Chapman EG, Janzen DH, Hallwachs W, Smith MA, Sharkey MJ. 2015. Polydnavirus gene provides accurate identification of species in the genus Hyposoter (Hymenoptera: Ichneumonidae). Insect Conservation and Diversity doi: 10.1111/icad.12118.
• Type: Journal Articles Status: Published Year Published: 2015 Citation: Tucker EM, Chapman EG, Sharkey MJ. 2015. A revision of the New World species of Cremnops F�rster (Hymenoptera: Braconidae: Agathidinae). Zootaxa 3916(1): 001-083.
• Type: Other Status: Published Year Published: 2015 Citation: Tucker EM, Chapman EG, Sharkey MJ. 2015. Interactive key, DELTA data matrix, images, and other files to species descriptions for A revision of the New World species of Cremnops F�rster (Hymenoptera: Braconidae: Agathidinae). DOI: 10.13140/2.1.1322.0649. Dataset published in: Zootaxa 3916(1): 001-083.
• Type: Journal Articles Status: Published Year Published: 2015 Citation: Wang M, Rasnitsyn AP, Li H, Shih C, Sharkey MJ, Ren A. 2015. Phylogenetic analyses elucidate the inter-relationships of Pamphilioidea (Hymenoptera, Symphyta). Cladistics (2015): 1-22.
• Type: Journal Articles Status: Published Year Published: 2015 Citation: Deans AR, Lewis SE, Huala E, Anzaldo SS, Ashburner M, Balhoff JP, Blackburn DC, Blake JA, Burleigh JG, Chanet B, Cooper LD, Courtot M, Cs�sz S, Cui H, Dahdul W, Das S, Dececchi TA, Dettai A, Diogo R, Druzinsky RE, Dumontier M, Franz NM, Friedrich F, Gkoutos GV, Haendel M, Harmon LJ, Hayamizu TF, He Y, Hines HM, Ibrahim N, Jackson LM, Jaiswal P, James-Zorn C, K�hler S, Lecointre G, Lapp H, Lawrence CJ, Le Nov�re N, Lundberg JG, Macklin J, Mast AR, Midford PE, Mik� I, Mungall CJ, Oellrich A, Osumi-Sutherland D, Parkinson H, Ram�rez MJ, Richter S, Robinson PN, Ruttenberg A, Schulz KS, Segerdell E, Seltmann KC, Sharkey MJ, Smith AD, Smith B, Specht CD, Squires RB, Thacker RW, Thessen A, Fernandez-Triana J, Vihinen M, Vize PD, Vogt L, Wall CE, Walls RL, Westerfeld M, Wharton RA, Wirkner CS, Woolley JB, Yoder MJ, Zorn AM, Mabee P. 2015. Finding Our Way through Phenotypes. PLOS Biology 13: e1002033.
• Type: Journal Articles Status: Submitted Year Published: 2016 Citation: Sharkey MJ, Chapman EG, Iza de Campos GY. (Submitted to Contributions in Science, Natural History Museum of Los Angeles County, on Nov 10, 2015). Revision of Aerophilus Sz�pligeti (Hymenoptera, Braconidae, Agathidinae) from eastern North America, with a key to the Nearctic species.
• Type: Journal Articles Status: Published Year Published: 2015 Citation: Sharkey MJ, Chapman EG, Janzen DJ, Hallwachs W, Smith MA. 2015. Revision of Aphelagathis(Hymenoptera, Braconidae, Agathidinae, Agathidini). Zootaxa 4000 (1): 073-089.
• Type: Journal Articles Status: Published Year Published: 2015 Citation: Sharkey MJ, Chapman EG. 2015. The Nearctic genera of Agathidinae (Hymenoptera: Braconidae)with a phylogenetic analysis, illustrated generic key, and the description of three new genera. Zootaxa 4000 (1): 049072.

Progress 01/22/14 to 09/30/14

Outputs
Target Audience: Hymenoptera systematists/phylogeneticists who might use the DNA sequences generated in this study in other studies, or who are interested in using the relationships uncovered in this study to infer evolutionary and biological processes and patterns. Scientists interested in the use of parasitic wasps for biological control. Land managers interested in monitoring and maintaining biodiversity and understanding trophic interactions between plants, herbivores and their parasites occurring on the lands they manage. Changes/Problems: Nothing Reported What opportunities for training and professional development has the project provided? We currently have 2 PhD students in our lab who are making significant progress towards their dissertations. Hatch funds have directly contributed to their research. We have also had 3 international students visit our lab in the past year (2 PhD and 1 undergraduate) from China and Brazil. These students learned some DNA techniques by helping others in the lab with their projects that were supported in part by Hatch funds. Morphological studies of braconid wasps are made possible by the many thousands of specimens in the Hymenoptera Institute Collection available for study. Furthermore, the connections that PI Sharkey has to museums around the world allows students in our lab to borrow specimens critical for their studies. Our students present their research at national and international meetings - and this research is made better by the available Hatch funds. One of our PhD students was a finalist in the university-wide 3-minute Dissertation competition and also won an award in the student competition at the national meeting of the Entomological Society of America in Austin, Texas. Our other PhD student won an award for her poster presentation at the meeting of the International Society of Hymenopterists in Cuzco, Peru. How have the results been disseminated to communities of interest? We have published the results of our research in numerous papers published in the last year. Many DNA sequences have been submitted to GenBank and are now publically available (referenced in said papers). Our Hymenoptera database is continually updated and available to the public. What do you plan to do during the next reporting period to accomplish the goals? We plan to stay the course. We have numerous ongoing studies in Hymenoptera systematics, at least 4 of which are close to being submitted for publication, including (1) a paper on southwestern US agathidines in which new species are being described and will include the most comprehensive agathidine phylogeny published to date and (2) a revision of the genus Lytopylus, again with multiple new species described. We are continuing to develop our methods for molecular determination of biodiversity, with one project nearing publication-ready status.

Impacts
What was accomplished under these goals? We have increased in the specimen holdings at the Hymenoptera Institute and other museums for comparative purposes and morphological study. We have added digital information on many hundreds of braconid specimens to an ever-growing, web-accessible Integrated Digitized Biocollections (IDigBio)-supported database (Symbiota). We have developed a better understanding of the relationships among braconid wasps and their host associations. This knowledge has been published (see publications by Tucker et al. 2015; Pook et al. 2015; van Acherberg et al. 2015). This knowledge could aid in biological control efforts in cases where one or more hosts become pests, as species-specific parasitoids have been among the most successful biological control agents. Our papers have led to a better understanding of the identification and phylogenetic relationships of the parasitoid species contained within, which is essential information when developing biological control programs. A better understanding of the biodiversity in selected study areas is becoming increasingly important as we try to understand how to maintain and manage our natural resources. We are developing molecular techniques to identify arthropods in field samples. These techniques will allow land managers to assess the arthropod diversity in natural areas at a very low cost (under $10k), whereas studies involving hundreds of taxonomists and years of study can cost millions of dollars. Consequently, comprehensive biodiversity studies are currently thought to be highly impractical, and our studies aim to make such studies not only feasible, but an essential tool for evaluating the value of natural areas and the ecosystem services provided within.

Publications

• Type: Journal Articles Status: Published Year Published: 2015 Citation: Deans, A.R., Lewis, S.E., Huala, E., Anzaldo, S.S., Ashburner, M., Balhoff, J.P., Blackburn, D.C., Blake, J.A., Burleigh, J.G., Chanet, B., Cooper, L.D., Courtot, M., Cs�sz, S., Cui, H., Dahdul, W., Das, S., Dececchi, T.A., Dettai, A., Diogo, R., Druzinsky, R.E., Dumontier, M., Franz, N.M., Friedrich, F., Gkoutos, G.V., Haendel, M., Harmon, L.J., Hayamizu, T.F., He, Y., Hines, H.M., Ibrahim, N., Jackson, L.M., Jaiswal, P., James-Zorn, C., K�hler, S., Lecointre, G., Lapp, H., Lawrence, C.J., Le Nov�re, N., Lundberg, J.G., Macklin, J., Mast, A.R., Midford, P.E., Mik�, I., Mungall, C.J., Oellrich, A., Osumi-Sutherland, D., Parkinson, H., Ram�rez, M.J., Richter, S., Robinson, P.N., Ruttenberg, A., Schulz, K.S., Segerdell, E., Seltmann, K.C., Sharkey, M.J., Smith, A.D., Smith, B., Specht, C.D., Squires, R.B., Thacker, R.W., Thessen, A., Fernandez-Triana, J., Vihinen, M., Vize, P.D., Vogt, L., Wall, C.E., Walls, R.L., Westerfeld, M., Wharton, R.A., Wirkner, C.S., Woolley, J.B., Yoder, M.J., Zorn, A.M., Mabee, P. 2015. Finding Our Way through Phenotypes. PLOS Biology 13: e1002033.
• Type: Book Chapters Status: Published Year Published: 2014 Citation: Nieves-Aldrey, J.L., Sharkey, M.J. 2014. Ants, Bees, Wasps and the Majority of Insect Parasitoids. pp 395-408. (In) Vargas P. & R. Zardoya (eds) The Tree of Life, Sinauer, Sunderland, MA.
• Type: Journal Articles Status: Published Year Published: 2014 Citation: van Achterberg, C., Sharkey, M.J., Chapman, E.G. 2014. Revision of the genus Euagathis Sz�pligeti (Hymenoptera, Braconidae, Agathidinae) from Thailand, with description of three new species. Journal of Hymenoptera Research 36: 1-25.
• Type: Journal Articles Status: Published Year Published: 2015 Citation: Pook, V.G., Chapman, E.G., Janzen, D.H., Hallwachs, W., Smith, M.A., Sharkey, M.J. 2015. Polydnavirus gene provides accurate identification of species in the genus Hyposoter (Hymenoptera: Ichneumonidae). Insect Conservation and Diversity. Accepted January, 2015.
• Type: Journal Articles Status: Published Year Published: 2015 Citation: Tucker, E.M., Chapman, E.G., Sharkey, M.J. 2015. A revision of the New World species of Cremnops F�rster (Hymenoptera: Braconidae: Agathidinae). Zootaxa 3916(1): 001-083.
• Type: Other Status: Published Year Published: 2015 Citation: Tucker, E.M., Chapman, E.G., Sharkey, M.J. 2015. Interactive key, DELTA data matrix, images, and other files to species descriptions for A revision of the New World species of Cremnops F�rster (Hymenoptera: Braconidae: Agathidinae). DOI: 10.13140/2.1.1322.0649. Dataset published in: Zootaxa 3916(1): 001-083.