Cuticular hydrocarbons for identifying Sarcophagidae (Diptera)

Abstract

The composition and quantity of insect cuticular hydrocarbons (CHCs) can be species-specific as well as sexually dimorphic within species. CHC analysis has been previously used for identification and ageing purposes for several insect orders including true flies (Diptera). Here, we analysed the CHC chemical profiles of adult males and females of eleven species of flesh flies belonging to the genus Sarcophaga Meigen (Sarcophagidae), namely Sarcophaga africa (Wiedemann), S. agnata Rondani, S. argyrostoma Robineau-Desvoidy, S. carnaria (Linnaeus), S. crassipalpis Macquart, S. melanura Meigen, S. pumila Meigen, S. teretirostris Pandellé, S. subvicina Rohdendorf, S. vagans Meigen and S. variegata (Scopoli). Cuticular hydrocarbons extracted from pinned specimens from the collections of the Natural History Museum, London using a customised extraction technique were analysed using Gas Chromatography-Mass Spectrometry. Time of preservation prior to extraction ranged between a few weeks to over one hundred years. CHC profiles (1) allowed reliable identification of a large majority of specimens, (2) differed between males and females of the same species, (3) reliably associated males and females of the same species, provided sufficient replicates (up to 10) of each sex were analysed, and (4) identified specimens preserved for up to over one hundred years prior to extraction.

Figure 1

Heat map of all 60 + compounds from the males and females of all Sarcophaga species, showing gender, time and species-dependent differences in the chromatograms. The historical samples cover a time period which ranges over 117 years. The x-axis represents the retention time and the chromatographs are grouped along the y-axis by species. The heat map is a visual aid, enabling multiple chromatographs to be efficiently stacked for comparison in a small vertical space, in which darker spots represent larger peak areas. For example, the most abundant compound, with a retention time of around 22.1 min on the heat map, is C27 (Table S1, peak number 23, Table S2, peak number 35). Some individual S. pumila female samples exhibited chromatograms that appeared different on the heat map. Nevertheless, SVM correctly classified all of these specimens as S. pumila. The compounds used for classifying are presented in Table S3.

Figure 2

A heat map of differences between cuticular hydrocarbon profiles of historical and recent (2018) specimens of three species of Sarcophaga (males only). The intensity of the red spots are proportional to the abundance of each compound with the observed retention time.

Figure 3

Principal component analysis for samples from the males of four species (Sarcophaga subvicina, S. variegata, S. crassipalpis and S. carnaria) collected and sampled in 2018. The first three principal components cover 43.4%, 23.5% and 14.3% of the variance, respectively, for a total of 81.2% variance. Factor loadings for the first three principal components are listed in the Supporting Data (Table S4).

Figure 4

Principal component analysis for male samples from three species (Sarcophaga subvicina, S. variegata and S. carnaria) collected recently (2018), compared to samples from males of the same three species collected historically (5 + years prior to extraction). The compounds used for PCA are given in Table S3. The first three principal components cover 39.3%, 23.3% and 14.4% of the variance, respectively, for a total of 77.0% variance. Factor loadings for the first three principal components are listed in the Supporting Data (Table S5).

Figure 5

Image showing three Sarcophaga agnata males (top row) and three S. agnata females (bottom row), 6 months after the specimens were extracted with hexane to examine their cuticular hydrocarbon profiles.