Revision of Zelodia ( Hymenoptera , Braconidae , Agathidinae ) from Thailand

The species of Thai Zelodia (Hymenoptera: Braconidae: Agathidinae) are revised. Twenty-one species are treated, 19 new species are described, i.e. Zelodia charoeni, Zelodia chongkraii, Zelodia cholathorni, Zelodia idrisi, Zelodia nikomi, Zelodia nopadoli, Zelodia pahangensis, Zelodia panyaii, Zelodia poonsathii, Zelodia ratanae, Zelodia saksiti, Zelodia surachaii, Zelodia suyaneeae, Zelodia toyae, Zelodia uthaii, Zelodia wangi, Zelodia wichaii, Zelodia wirati, Zelodia wirotei. A dichotomous key to species is presented; links to electronic interactive keys and to distribution maps are also included.


Introduction
Agathidinae is a moderately large subfamily of Braconidae with 1,061 described species worldwide and 238 in the Oriental Region (Yu et al. 2005) though there are an estimated 2,000-3,000 species awaiting description worldwide (Sharkey et al. 2006).The subfamily has a worldwide distribution and members are found in most terrestrial habitats.The history of higher classification of the Agathidinae was summarized by Sharkey (1992) who also proposed a tribal level classification based on ground-plan coding.Sharkey et al. (2006) conducted phylogenetic analyses based on morphology and the D2-3 regions of 28S rDNA.The Oriental fauna of Agathidinae was first revised by Bhat and Gupta (1977) and they provided a detailed history of taxonomic research for the area.Sharkey et al. (2009) revised the Oriental genera of Agathidinae.Achterberg and Long (2010) revised the Vietnamese agathidine fauna, erected the genus Zelodia, described 5 new species of Zelodia and transferred 20 species to Zelodia, mostly from Coccygidium and Zelomorpha.Sharkey and Clutts (2011) revised the Thai species of Biroia, Braunsia, Camptothlipsis, Coccygidium, Cremnops, Cremnoptoides, Disophrys, Earinus, Gyrochus, Lytopylus, and Troticus, and included a key to the Agathidinae genera of the Oriental region.This paper is the second in a series to revise all Thai species of Agathidinae.The genera Aneurobracon, Bassus, Euagathis, and Therophilus will be dealt with in subsequent publications.

Methods
As part of the inventory of Thai insects, we ran three Malaise traps at 30 different localities throughout Thailand from 2007Thailand from -2010, comprising , comprising approximately 90 Malaise trap years.The specimens dealt with here are primarily from these traps.
Species concepts are based on morphological data and cytochrome c oxidase (COI) data.Phenetic and phylogenetic trees, using 558 base pairs of COI data, were constructed using neighbor-joining (NJ), maximum parsimony (MP) and Bayesian methods.MP was performed using TNT (Goloboff et. al, 2008) [traditional search with 100 random addition sequences followed by branch-swapping, saving 100 trees per replication; 1000 bootstrap replications were used to estimate branch reliability].The Bayesian analysis was performed using MrBayes v3.1.2(Ronquist and Huelsenbeck 2003).Best-fitting DNA substitution models were determined using MrModeltest2.2(Nylander 2004).The general time reversible model of evolution with a parameter for invariant sites and rate heterogeneity modeled under a gamma distribution (GTR+I+G) was determined as the best-fitting model.The Bayesian analysis consisted of two independent Bayesian MCMC runs initiated from different random starting trees.The analysis ran for 2,000,000 generations, reaching a topological similarity criterion of 0.01; trees were sampled every 500 generations.25% of the trees from each run were removed as burn-in upon topological convergence.The NJ tree was produced from PAUP* (Swofford 2002) using default settings.Figure 2 presents the NJ tree, which was much more resolved than were the phylogenetic trees produced by MP and Bayesian analyses.We mapped the support values of the Bayesian and MP analyses on the NJ tree.Branches without values are those that collapsed in the phylogenetic analyses.
Morphological terms follow Sharkey and Wharton (1997) except for the following: measurements are given for the length and apical width of the first metasomal mediotergite (MT1).Measurement of the apical width is straightforward, however since the base of the tergite is usually hidden from view it is difficult to measure the total length.Instead we measure from the apex of the large tendon that emanates from the propodeum and inserts near the base of the median tergite.Abbreviations used in text: S1, S2, S3; metasomal mediosternite 1, 2, 3; MT1, MT2, MT3: metasomal mediotergite 1, 2, 3; LT1, LT2, LT3: metasomal laterotergite 1, 2, 3.
Morphological terms used in this revision were matched to the Hymenoptera Anatomy Ontology (HAO, Yoder et al. 2010) (Appendix 4).Identifiers (URIs) in the format http://purl.obolibrary.org/obo/HAO_XXXXXXXrepresent anatomical concepts in HAO version http://purl.obolibrary.org/obo/hao/2011-05-18/hao.owl.They are provided to enable readers to confirm their understanding of the anatomical structures being referenced.To find out more about a given structure, including, images, references, and other metadata, use the identifier as a web-link, or use the HAO:XXXXXXX (note colon replaces underscore) as a search term at http://glossary.hymao.org.
All 19 species are treated with a diagnosis and distributional data.They are illustrated with color photos using a JVC digital camera mounted on a Leica MZ16 microscope and Automontage® stacking software.Distributional data are listed for all species and a Google map via Berkeley Mapper is included for all species.The descriptions are of the holotype and variation is given in parentheses.
The source files for the keys, descriptions, illustrations, DNA sequence and distributional data are all freely available to future researchers who may wish to build on this beginning.DNA trace files and primer information are available through the Barcode of Life Data system (BOLD) [Ratnasingham and Hebert 2007] at http:// www.boldsystems.org.Sixteen of the twenty five Zelodia COI sequences were generated by BOLD (project ASTRK Revisions of Thai Agathidinae Braconids), the remaining nine were generated in the Sharkey lab.All sequences have been deposited in GenBank database (JQ763436-JQ763460).All twenty five Zelodia COI sequences are characterized by a -1 frameshift mutation.A majority of Agathidinae CO1 sequences are distinguished by a series of 1 bp deletions which are not restricted to one portion of the barcode region.Codon composition in sequences with 1bp deletions remains highly biased towards AT and substitutions remain biased towards 3 rd codon position (M.Alex Smith pers.comm.).It is suggested that the genes are correctly decoded by a programmed frameshift during translation (Beckenbach et al. 2005) and are functional.
Distribution data, pdf's of non-copyright references, images, notes, and host and type information can be found by searching TaxaBank (a combined specimen and taxonomic database; http://purl.org/taxabank).Codes beginning with an "H" and followed by numbers are unique identifiers used for specimens in the Sharkey lab at the University of Kentucky, and in the specimen database TaxaBank (e.g., H647).

BMNH
The Natural History Museum, London, England.

FSCA
Florida State Collection of Arthropods, Gainesville, Florida, USA.

HIC
Hymenoptera Institute Collection, University of Kentucky, Department of Entomology, Lexington, Kentucky, USA.

Results
Refer to the tree in Sharkey and Clutts (2010) for generic level placement.The host lepidopterans of the genus are unknown.The neighbor joining branching diagram in Figure 2, based on COI mtDNA, was used to help in determining species limits although we did not devise any cutoff threshold to delimit species.We conducted Bayesian and parsimony analyses and where these agreed with the NJ tree the support values are given in Figure 2.An examination of this figure shows that the NJ diagram and the phylogenetic analyses were in close agreement.We also used morphological differences to delimit species.For example, Z. saksiti and Z. charoeni are very similar (1/558 bp difference) in COI sequences but are very different morphologically.Contrastingly, Zelodia wangi is a widespread species and COI sequences show some variation (2/558 bp difference), however we could find no morphological differences.The complex may represent several species.
Distribution.Widespread throughout western Thailand.Distribution map can be found at http://purl.org/thaimap/saksitiEtymology.Dedicated to Saksit Poonsapsiri, one of the chiefs of Phataem National Park.
Distribution.Known only from the type locality in northwestern Thailand.Distribution map can be found at http://purl.org/thaimap/toyaeEtymology.Dedicated to Ms. Chayanit (Toy) Satatha.Toy was the sorter of Diptera for the TIGER project and is currently a technician at QSBG.The species name reflects her nick-name, Toy.
Distribution.Widespread throughout Thailand.Distribution map can be found at http://purl.org/thaimap/wangiEtymology.Dedicated to Mr. Wang Saeyang, who was the sorter for Hymenoptera for the TIGER project.

Figure 1 .
Figure 1.Map showing collection sites in Thailand.

Figure 2 .
Figure 2. NJ phylogram of the COI barcode region for 17 of the 21 Zelodia species treated here.Wherever Bayesian and parsimony analyses agreed with the NJ tree branch support values are included in the figure i.e., Bayesian posterior probabilities / parsimony bootstrap (bootstrap values less than 60 are not illustrated).

Figure 3 .
Figure 3. Zelodia brevifemoralis Achterberg and Long a lateral habitus b wings c dorsal head d lateral head e lateral mesosoma f dorsal mesosoma g dorsal propodeum and MT1-MT2.

Figure 4 .
Figure 4. Zelodia charoeni sp.n. a lateral habitus b wings c dorsal head d lateral head e lateral mesosoma f dorsal mesosoma g dorsal propodeum and MT1.

Figure 6 .
Figure 6.Zelodia chongkraii sp.n. a lateral habitus b wings c dorsal head d lateral head e lateral mesosoma f dorsal mesosoma g dorsal propodeum and MT1.

Figure 7 .
Figure 7. Zelodia idrisi sp.n. a lateral habitus b dorsal head c lateral head and mesosoma d wings with buffered light source e wings with normal light source f dorsal head and mesosoma g dorsal propodeum h dorsal MT1-MT3.

Figure 8 .
Figure 8. Zelodia longidorsata a lateral habitus b dorsal head c lateral head and mesosoma d wings e dorsal mesosoma f dorsal propodeum and metasomal terga 1 and 2.

Figure 10 .
Figure 10.Zelodia nopadoli sp.n. a lateral habitus b wings c dorsal head d lateral head e lateral mesosoma f dorsal mesosoma g dorsal propodeum and MT1.

Figure 11 .
Figure 11.Zelodia pahangensis sp.n. a lateral habitus b dorsal head c lateral head and mesosoma d apex of fore wing e wings f dorsal head and mesosoma g dorsal propodeum h dorsal MT1-MT3.

Figure 12 .
Figure 12.Zelodia panyaii sp.n. a lateral habitus b wings c dorsal head d lateral head e lateral mesosoma f dorsal mesosoma g dorsal propodeum and MT1 and MT2.

Figure 13 .
Figure 13.Zelodia poonsathiti sp.n. a lateral habitus b wings c dorsal head d lateral head e lateral mesosoma f dorsal mesosoma g dorsal propodeum and MT1.
Molecular data.TaxaBank#/Genbank Accession: H273 /JQ763452.Distribution.Found only at the type locality in southern peninsular Thailand.Distribution map can be found at http://purl.org/thaimap/ratanaeEtymology.Dedicated to Ms. Ratana Luckanawarakul.She is currently the director of the Recreation and Interpretation Division of the Thai Department of National Parks.Ratana was of great assistance to us on numerous field trips and was always the best of company.Material examined.Holotype ♂.H0273 [QSBG], Thailand, Khao Pu-Khao YaNP, 75m, 7.551°N, 99.789°E, MT, 8-10.v.2006.

Figure 14 .
Figure 14.Zelodia ratanae sp.n. a lateral habitus b dorsal head c lateral head and mesosoma d wings e dorsal head and mesosoma f dorsal propodeum g dorsal metasoma.

Figure 17 .
Figure 17.Zelodia suyaneeae sp.n. a lateral habitus b wings c dorsal head d dorsolateral head e lateral mesosoma f dorsal mesosoma g dorsal propodeum and metasoma.

Figure 18 .
Figure 18.Zelodia toyae sp.n. a lateral habitus b wings c dorsal head d lateral head and mesosoma e dorsal head and mesosoma f dorsal propodeum g dorsal metasoma.

Figure 19 .
Figure 19.Zelodia uthaii sp.n. a lateral habitus b wings c anterior head d dorsal head e lateral head and mesosoma f dorsal head and mesosoma g dorsal propodeum h dorsal MT1-MT3.