Computational protocol: A Formal Re-Description of the Cockroach Hebardina concinna Anchored on DNA Barcodes Confirms Wing Polymorphism and Identifies Morphological Characters for Field Identification

Similar protocols

Protocol publication

[…] Our terminology follows McKittrick 1964 , Grandcolas 1996 , Anisyutkin 2010 and Anisyutkin 2013 . The genital segments of the examined specimens were macerated in 10% KOH and observed in glycerin with a Zeiss Discovery V12 stereomicroscope. Wings were floated in hot water until fully spread, embedded in neutral balsam, then mounted on slides and covered with coverslips. Drawings were made using a Zeiss Discovery V12 stereomicroscope fitted with a Canon PowerShot G1X digital camera and drawn using Adobe Illustrator CS6. All images of specimens were photographed using a Canon 60D plus a Canon EF 100 mm f/2.8L IS USM Macro lens combined with Helicon Focus software. All specimens studied were pinned in a natural posture and deposited in the medical vector collections of the Zhongshan Entry-Exit Inspection and Quarantine Bureau (ZSCIQ). The specimens we collected in China fully match other published morphological descriptions of H. concinna individuals from other geographic locations , , . [...] Sequences from H. concinna and other species were submitted to the International Nucleotide Sequence Database Collaboration via NCBI GenBank. As noted above we used two primer sets: one amplified a section of the mitochondrial COI gene and a second amplified a short stretch of the 3′ terminus of the COI gene, the tRNA-leu gene, and the 5′ 306 bp of the COII gene. For this second amplicon the tRNA-leu and 3′ COI sequences have little variation, and we used only the COII sequences from this amplicon for our analysis.We also identified nine additional cockroaches in GenBank for which either or both the COI and COII regions were available and created separate COI and COII data sets for phylogenetic analysis (). Unfortunately there were not enough individuals for which both regions had been sequenced to assemble a combined dataset using both genes so we analyzed the two data sets separately.We estimated maximum likelihood and neighbor-joining phylogentic trees for both the COI and COII data sets using Mega 5.2 , and tested robustness of the results using non-parametric bootstrapping. For both data sets the most appropriate maximum likelihood model (TN+I) was identified using the model testing function of Mega and this model was used to estimate the ML tree for each data set. Support for each branch was assessed using the same model for 1000 bootstrap replicates. Neighbor joining trees were constructed for each data set using the same Tamura-Nei model and also tested using 1000 non-parametric bootstrap replicates. […]

Pipeline specifications

Software tools Adobe Illustrator, MEGA
Databases INSDC
Applications Miscellaneous, Phylogenetics