Volatiles from the Mandibular Gland Reservoir Content of Colobopsis explodens Laciny and Zettel, 2018, Worker Ants (Hymenoptera: Formicidae).

Forty-five volatile organic compounds (VOCs) were identified or annotated in the mandibular gland reservoir content (MGRC) of the Southeast Asian ant Colobopsis explodens Laciny and Zettel, 2018 (Hymenoptera: Formicidae), using headspace solid-phase microextraction (HS-SPME) coupled to gas chromatography mass spectrometry (GC-MS) and liquid extraction combined with GC-MS. In extension of previous reports on VOCs of C. explodens, members of different compound classes, such as alkanes, aliphatic and aromatic carboxylic acids, and phenolics, were detected. The ketone 2-heptanone and the biochemically related phenolics benzene-1,3,5-triol (phloroglucinol, PG), 1-(2,4,6-trihydroxyphenyl)ethanone (monoacetylphloroglucinol, MAPG), 5,7-dihydroxy-2-methylchromen-4-one (noreugenin), and 1-(3-Acetyl-2,4,6-trihydroxyphenyl)ethanone (2,4-diacetylphloroglucinol, DAPG) dominated the GC-MS chromatograms. The identities of the main phenolics MAPG and noreugenin were further verified by liquid chromatography-high resolution-tandem mass spectrometry (LC-HRMS/MS). A comparative study of MGRC samples originating from three distinct field expeditions revealed differences in the VOC profiles, but the presence and relative abundances of the dominating constituents were largely consistent in all samples. Our study considerably extends the knowledge about the number and type of VOCs occurring in the MGRC of C. explodens. Based on the type of the detected compounds, we propose that the likely irritant and antibiotic phenolic constituents play a role in defense against arthropod opponents or in protection against microbial pathogens.

Pathogen evasion of social immunity.

Treating sick group members is a hallmark of collective disease defence in vertebrates and invertebrates alike. Despite substantial effects on pathogen fitness and epidemiology, it is still largely unknown how pathogens react to the selection pressure imposed by care intervention. Using social insects and pathogenic fungi, we here performed a serial passage experiment in the presence or absence of colony members, which provide social immunity by grooming off infectious spores from exposed individuals. We found specific effects on pathogen diversity, virulence and transmission. Under selection of social immunity, pathogens invested into higher spore production, but spores were less virulent. Notably, they also elicited a lower grooming response in colony members, compared with spores from the individual host selection lines. Chemical spore analysis suggested that the spores from social selection lines escaped the caregivers' detection by containing lower levels of ergosterol, a key fungal membrane component. Experimental application of chemically pure ergosterol indeed induced sanitary grooming, supporting its role as a microbe-associated cue triggering host social immunity against fungal pathogens. By reducing this detection cue, pathogens were able to evade the otherwise very effective collective disease defences of their social hosts.

Colobopsis explodens sp. n., model species for studies on "exploding ants" (Hymenoptera, Formicidae), with biological notes and first illustrations of males of the Colobopsis cylindrica group.

A taxonomic description of all castes of Colobopsis explodens Laciny & Zettel, sp. n. from Borneo, Thailand, and Malaysia is provided, which serves as a model species for biological studies on "exploding ants" in Southeast Asia. The new species is a member of the Colobopsis cylindrica (COCY) group and falls into a species complex that has been repeatedly summarized under the name Colobopsis saundersi (Emery, 1889) (formerly Camponotus saundersi). The COCY species group is known under its vernacular name "exploding ants" for a unique behaviour: during territorial combat, workers of some species sacrifice themselves by rupturing their gaster and releasing sticky and irritant contents of their hypertrophied mandibular gland reservoirs to kill or repel rivals. This study includes first illustrations and morphometric characterizations of males of the COCY group: Colobopsis explodens Laciny & Zettel, sp. n. and Colobopsis badia (Smith, 1857). Characters of male genitalia and external morphology are compared with other selected taxa of Camponotini. Preliminary notes on the biology of C. explodens Laciny & Zettel, sp. n. are provided. To fix the species identity of the closely related C. badia, a lectotype from Singapore is designated. The following taxonomic changes within the C. saundersi complex are proposed: Colobopsis solenobia (Menozzi, 1926), syn. n. and Colobopsis trieterica (Menozzi, 1926), syn. n. are synonymized with Colobopsis corallina Roger, 1863, a common endemic species of the Philippines. Colobopsis saginata Stitz, 1925, stat. n., hitherto a subspecies of C. badia, is raised to species level.

Isolation of Mandibular Gland Reservoir Contents from Bornean 'Exploding Ants' (Formicidae) for Volatilome Analysis by GC-MS and MetaboliteDetector.

The aim of this manuscript is to present a protocol describing the metabolomic analysis of Bornean 'exploding ants' belonging to the Colobopsis cylindrica (COCY) group. For this purpose, the model species C. explodens is used. Ants belonging to the minor worker caste possess distinctive hypertrophied mandibular glands (MGs). In territorial combat, they use the viscous contents of their enlarged mandibular gland reservoirs (MGRs) to kill rival arthropods in characteristic suicidal 'explosions' by voluntary rupture of the gastral integument (autothysis). We show the dissection of worker ants of this species for the isolation of the gastral portion of the wax-like MGR contents as well as listing the necessary steps required for solvent-extraction of the therein contained volatile compounds with subsequent gas chromatography-mass spectrometry (GC-MS) analysis and putative identification of metabolites contained in the extract. The dissection procedure is performed under cooled conditions and without the use of any dissection buffer solution to minimize the changes in the chemical composition of the MGR contents. After solvent-based extraction of volatile metabolites contained therein, the necessary steps for analyzing the samples via liquid-injection-GC-MS are presented. Lastly, data processing and putative metabolite identification with the use of the open-source software MetaboliteDetector is shown. With this approach, the profiling and identification of volatile metabolites in MGRs of ants belonging to the COCY group via GC-MS and the MetaboliteDetector software become possible.