Wolbachia and DNA barcoding insects: patterns, potential and problems

Wolbachia is a genus of bacterial endosymbionts that impacts the breeding systems of their hosts. Wolbachia can confuse the patterns of mitochondrial variation, including DNA barcodes, because it influences the pathways through which mitochondria are inherited. We examined the extent to which these endosymbionts are detected in routine DNA barcoding, assessed their impact upon the insect sequence divergence and identification accuracy, and considered the variation present in Wolbachia COI. Using both standard PCR assays (Wolbachia surface coding protein – wsp), and bacterial COI fragments we found evidence of Wolbachia in insect total genomic extracts created for DNA barcoding library construction. When >2 million insect COI trace files were examined on the Barcode of Life Datasystem (BOLD) Wolbachia COI was present in 0.16% of the cases. It is possible to generate Wolbachia COI using standard insect primers; however, that amplicon was never confused with the COI of the host. Wolbachia alleles recovered were predominantly Supergroup A and were broadly distributed geographically and phylogenetically. We conclude that the presence of the Wolbachia DNA in total genomic extracts made from insects is unlikely to compromise the accuracy of the DNA barcode library; in fact, the ability to query this DNA library (the database and the extracts) for endosymbionts is one of the ancillary benefits of such a large scale endeavor – for which we provide several examples. It is our conclusion that regular assays for Wolbachia presence and type can, and should, be adopted by large scale insect barcoding initiatives. While COI is one of the five multi-locus sequence typing (MLST) genes used for categorizing Wolbachia, there is limited overlap with the eukaryotic DNA barcode region

Remarkable fly (Diptera) diversity in a patch of Costa Rican cloud forest : Why inventory is a vital science

Study of all flies (Diptera) collected for one year from a four-hectare (150 x 266 meter) patch of cloud forest at 1,600 meters above sea level at Zurqui de Moravia, San Jose Province, Costa Rica (hereafter referred to as Zurqui), revealed an astounding 4,332 species. This amounts to more than half the number of named species of flies for all of Central America. Specimens were collected with two Malaise traps running continuously and with a wide array of supplementary collecting methods for three days of each month. All morphospecies from all 73 families recorded were fully curated by technicians before submission to an international team of 59 taxonomic experts for identification. Overall, a Malaise trap on the forest edge captured 1,988 species or 51% of all collected dipteran taxa (other than of Phoridae, subsampled only from this and one other Malaise trap). A Malaise trap in the forest sampled 906 species. Of other sampling methods, the combination of four other Malaise traps and an intercept trap, aerial/hand collecting, 10 emergence traps, and four CDC light traps added the greatest number of species to our inventory. This complement of sampling methods was an effective combination for retrieving substantial numbers of species of Diptera. Comparison of select sampling methods (considering 3,487 species of non-phorid Diptera) provided further details regarding how many species were sampled by various methods. Comparison of species numbers from each of two permanent Malaise traps from Zurqui with those of single Malaise traps at each of Tapanti and Las Alturas, 40 and 180 km distant from Zurqui respectively, suggested significant species turnover. Comparison of the greater number of species collected in all traps from Zurqui did not markedly change the degree of similarity between the three sites, although the actual number of species shared did increase. Comparisons of the total number of named and unnamed species of Diptera from four hectares at Zurqui is equivalent to 51% of all flies named from Central America, greater than all the named fly fauna of Colombia, equivalent to 14% of named Neotropical species and equal to about 2.7% of all named Diptera worldwide. Clearly the number of species of Diptera in tropical regions has been severely underestimated and the actual number may surpass the number of species of Coleoptera. Various published extrapolations from limited data to estimate total numbers of species of larger taxonomic categories (e.g., Hexapoda, Arthropoda, Eukaryota, etc.) are highly questionable, and certainly will remain uncertain until we have more exhaustive surveys of all and diverse taxa (like Diptera) from multiple tropical sites. Morphological characterization of species in inventories provides identifications placed in the context of taxonomy, phylogeny, form, and ecology. DNA barcoding species is a valuable tool to estimate species numbers but used alone fails to provide a broader context for the species identified.Peer reviewe

Comprehensive inventory of true flies (Diptera) at a tropical site

Peer reviewe

Comprehensive inventory of true flies (Diptera) at a tropical site

Estimations of tropical insect diversity generally suffer from lack of known groups or faunas against which extrapolations can be made, and have seriously underestimated the diversity of some taxa. Here we report the intensive inventory of a four-hectare tropical cloud forest in Costa Rica for one year, which yielded 4332 species of Diptera, providing the first verifiable basis for diversity of a major group of insects at a single site in the tropics. In total 73 families were present, all of which were studied to the species level, providing potentially complete coverage of all families of the order likely to be present at the site. Even so, extrapolations based on our data indicate that with further sampling, the actual total for the site could be closer to 8000 species. Efforts to completely sample a site, although resource-intensive and time-consuming, are needed to better ground estimations of world biodiversity based on limited sampling

Natural history specimens collected and/or identified and deposited.

Natural history specimen data collected and/or identified by D. Monty Wood, http://www.wikidata.org/entity/Q21772597. Claims or attributions were made on Bionomia, https://bionomia.net using specimen data from the Global Biodiversity Information Facility, https://gbif.org.http://www.wikidata.org/entity/Q2177259

FIGURES 1 – 4. 1 in A new species of Axymyiidae (Diptera) from western North America and a key to the Nearctic species

FIGURES 1 – 4. 1. Ideal Protaxymyia thuja habitat; waterlogged Western Red Cedar in mature forest. Figs. 2 – 4. P. thuja. 2. Adult female, dorsal. 3. Pupa, dorsal. 4. Larva, lateral

Ceratopogonidae

CERATOPOGONIDAE <p> <i>Description (Fig. 7C)</i></p> <p> <i>Testis:</i> Each testis is narrow, spindle-shaped, tapered anteriorly, and organized into zones.</p> <p> <i>Epididymis:</i> Not differentiated.</p> <p> <i>Vas deferens:</i> The vasa deferentia are packed with mature spermatozoa, even at emergence (Linley, 1981). The ducts extend posteriorly to near segment 9, where they bend abruptly back, become fused to the midline of the accessory gland complex, and run juxtaposed to the anterior apex of the gland. The ducts enter their respective half of the gland through a sphincter (Linley, 1981).</p> <p> <i>Accessory gland and seminal vesicle:</i> The accessory gland complex comprises three pairs of chambers in Ceratopogonidae, united along the midline. The most anterior chamber was referred to as the seminal vesicle by Linley (1981), and is lined with an inner layer of narrow secretory cells with spherical vacuoles containing secretions. A narrow aperture separates the seminal vesicle from the middle chamber, referred to as the glutinous gland (Linley, 1981). The secretory cells of this chamber produce a viscid material, which forms the envelope of the spermatophore. The third chamber is not easily observed in whole mounts and is not lined with glandular cells. The entire accessory gland complex is covered in a layer of circular muscle.</p> <p> <i>Ejaculatory duct:</i> This duct comprises two sections. In the anterior half, the ducts remain separate, whereas the ducts unite near the base of the genitalia and in this section the duct is lined with columnar glandular cells (Linley, 1981). The ejaculatory duct is sheathed in a layer of circular muscles.</p> <p> <i>Ejaculatory apodeme, sperm pump, and aedeagus:</i> The ejaculatory apodeme is absent and the true aedeagus is membranous. The ventral plate with its recurved tip, present in most Ceratopogonidae, is pressed into the opening of the dorsal wall of the common female spermathecal duct and may aid in stretching the opening (Linley, 1981). The ventral plate (termed aedeagus by authors in this family) is present throughout the family, including <i>Austroconops</i> Wirth & Lee, an early lineage of Ceratopogonidae (Borkent, Wirth & Dyce, 1987; Borkent & Craig, 2004).</p> <p> <i>Remarks:</i> The male genital tract of Ceratopogonidae was first described by Pomerantzev (1932) and the method of spermatophore formation in <i>Culicoides</i> Latreille was thoroughly described and excellently illustrated by Linley (1981). Two packages of sperm are prepared in the long, narrow, tapering glutinous gland (middle chamber) and are pushed into paired, secreted sheaths upon exiting, which fuse medially further down the ejaculatory duct. The spermatophore is then passed to the female during copulation. As observed by Linley (1981), spermatozoa remain in the vasa deferentia until ejaculation. In the present study, spermatozoa-packed vasa deferentia were observed in freshly collected, nonmated males of Bibionidae and Thaumaleidae (Figs 3B, D, 7A), suggesting a similar process of spermatophore production as is present in Ceratopogonidae.</p> <p> <i>Remarks – features of Chironomoidea.</i> The accessory gland complex of Chironomoidea is similar in form among all four families. The elongate second chamber with its glandular cells is similar in Ceratopogonidae, Simuliidae (Fig. 5C; Rubtsov, 1989: fig. 10A, epsd), and Chironomidae (Fig. 5F). Only in Thaumaleidae do the vasa deferentia enter the accessory/seminal gland subapically (Fig. 7A).</p> <p>The genital tract of the Chironomoidea is modified for the passage of a preformed spermatophore (Wood, 1978; Wood & Borkent, 1989), the existence of which has been confirmed in the families Simuliidae, Ceratopogonidae, and Chironomidae, and presumably also the Thaumaleidae, given the configuration of the internal genital tract (Sinclair, 1992a). The enlarged accessory gland complex is quite stout and generally easily identified in whole mount dissections, whereas the vasa deferentia and testes are often more fragile, the latter often distinctively clothed in pigment/fat granules.</p>Published as part of <i>Sinclair, Bradley J., Borkent, Art & Wood, D. Monty, 2007, The male genital tract and aedeagal components of the Diptera with a discussion of their phylogenetic significance, pp. 711-742 in Zoological Journal of the Linnean Society 150 (4)</i> on pages 729-730, DOI: 10.1111/j.1096-3642.2007.00314.x, <a href="http://zenodo.org/record/5432328">http://zenodo.org/record/5432328</a&gt

Seven new species of Spathidexia Townsend (Diptera: Tachinidae) reared from caterpillars in Area de Conservación Guanacaste, Costa Rica

We describe seven new species of Spathidexia (Diptera: Tachinidae) reared from Area de Conservación Guanacaste (ACG), northwestern Costa Rica. All were reared from ­various species of ACG caterpillars during an ongoing inventory of caterpillars, their food plants and their parasitoids. By coupling morphology, photographic documentation, life history and molecular data, we provide a clear and concise description of each species. All are known to be previously undescribed as a result of a comprehensive study of the genus by DMW. Spathidexia atripalpus sp. n., Spathidexia juanvialesi sp. n., Spathidexia marioburgosi sp. n., Spathidexia luisrobertogallegosi sp. n., Spathidexia luteola sp. n., Spathidexia hernanrodriguezi sp. n. and Spathidexia aurantiaca sp. n. are all authored and described by Fleming and Wood. Minthodexiopsis Townsend is proposed by Wood as a new synonym of Spathidexia. A new combination proposed by Wood as a result of the new synonymy is S. flavicornis (Brauer & Bergenstamm) comb. n

Three new species of Ametadoria Townsend (Diptera: Tachinidae) from Area de Conservación Guanacaste, Costa Rica

We describe three new species in the genus Ametadoria Townsend from Area de Conservación Guanacaste (ACG), Costa Rica. All three were reared from wild-caught Zygaenidae and Lacturidae caterpillars. We provide a concise description of each species using morphology, life history and molecular data, with photographic documentation. The new species are authored and described by Fleming and Wood: Ametadoria karolramosae sp. nov., Ametadoria leticiamartinezae sp. nov., and Ametadoria mauriciogurdiani sp. nov. The following are proposed by Wood as new synonyms of Ametadoria Townsend: Adidyma Townsend syn. nov., and Abolodoria Townsend syn. nov. The following new combinations occur as a result of these new synonymies: Ametadoria abdominalis (Townsend) comb. nov., Ametadoria austrina (Coquillett) comb. nov., Ametadoria humilis (Wulp) comb. nov., Ametadoria misella (Wulp) comb. nov. Ametadoria adversa (Townsend) is proposed as a junior synonym of ​Ametadoria unispinosa Townsend, syn. nov​