Formicine ants swallow their highly acidic poison for gut microbial selection and control

Abstract

12Animals continuously encounter microorganisms that are essential for health or cause disease. 13 They are thus challenged to control harmful microbes while allowing acquisition of beneficial 14 microbes, a challenge that is likely especially important concerning microbes in food and in 15 animals such as social insects that exchange food among colony members. Here we show that 16 formicine ants actively swallow their antimicrobial, highly acidic poison gland secretions 17 after feeding. The ensuing creation of an acidic environment in the stomach, the crop, 18 improves individual survival in the face of pathogen contaminated food and limits disease 19 transmission during mutual food exchange. At the same time, crop acidification selectively 20 allows acquisition and colonization by known bacterial gut associates. The results of our 21 study suggest that swallowing of acidic poison gland secretions acts as a microbial filter in 22 formicine ants and indicate a potentially widespread but so far underappreciated dual role of 23 antimicrobials in host-microbe interactions. 24 25 85 5 experimental manipulation of poison gland access, and behavioural observations. In loss of 86 poison gland function experiments, we then investigate whether analogous to acidic stomachs 87 of higher vertebrates and acidic midgut regions in the fruit fly, swallowing of poison gland 88 substances can serve gut microbial control and prevent bacterial pathogen infection and 89 transmission. Finally, we explore whether swallowing of poison gland substances acts as a 90 microbial filter that is permissible to gut colonization of bacteria from the family 91 Acetobacteracea. 92 93 Results and Discussion 94To reveal whether poison gland secretions are swallowed during acidopore grooming, we 95 monitored acidity levels in the crop lumen of the Florida carpenter ant Camponotus floridanus 96 at different time points after feeding them 10% honey water (pH = 5). We found that over 97 time, the crop lumen became increasingly acidic, reaching highly acidic values 48h after 98 feeding (median pH = 2; 95% CI: 1.5-3.4), whilst renewed access to food after 48h restored 99 the pH to levels recorded after the first feeding trial ( Fig. 1a; LMM, LR-test, χ 2 = 315.18, df = 100 3, P < 0.001; Westfall corrected post-hoc comparisons: 0+4h vs. 48h+4h: P = 0.317, all other 101 comparisons: P < 0.001). This acidification was limited to the crop and did not extend to the 102 midgut (Fig. 1figure supplement 1; pH-measurements at four points along the midgut 24h 103 after access to 10% honey-water; mean ± se; midgut position 1 = 5.08 ± 0.18, midgut position 104 2 = 5.28 ± 0.17, midgut position 3 = 5.43 ± 0.16, midgut position 4 = 5.31 ± 0.19). Prevention 105 of acidopore grooming in C. floridanus ants for 24h after feeding resulted in a significantly 106 diminished acidification of the crop lumen (Fig. 1b; LMM, LR-test, χ 2 = 44.68, df = 1, P < 107 0.001), a result that was invariably obtained in a comparative survey across seven formicine 108 ant species (g...

Prophylactic Avoidance of Hazardous Prey by the Ant Host Myrmica rubra

“…Since extensive self-grooming by M. rubra ants did not differ with the infectiousness of the retrieved prey, it rather appears as a non-specific prophylactic behaviour similar to the “washing hands before dinner” hygienic measure. Our observations are in line with Tragust et al 2020 study [ 92 ] who found a higher proportion of acidopore grooming after fluid ingestion, irrespective of the fluid nutritional value.…”

“…Furthermore, ant workers performed intensive self-groomings interspersed by an anal-mouth contact through the bending of their gaster. A similar behaviour has been reported as a so-called acidopore grooming in several formicinae species (Lasius neglectus: [91]; Camponotus floridanus: [92]) or as a faecal fluid grooming in Atta ants [93]. In formicinae ants, acidopore grooming was related to the release of poison secretions, which limit the growth of pathogens on the ant's cuticle [91].…”

“…In formicinae ants, acidopore grooming was related to the release of poison secretions, which limit the growth of pathogens on the ant’s cuticle [ 91 ]. Ants swallow acidic poison gland secretions which act as microbial control agents in the gut and that sanitize ingested food [ 92 ]. In Atta spp ants, faecal droplets collected while grooming were used by the foundress to facilitate the removal or decontamination of particles remaining on the ant’s body or in the fungus garden [ 93 ].…”

Behaviours indicating cannibalistic necrophagy in ants are modulated by the perception of pathogen infection level

“…During auto-and allogrooming, which are the most common hygienic behaviours, antimicrobial secretions from the metapleural and venom glands are spread on the surface of the cuticle, which suppresses the development of fungi and bacteria 19,24,62,66,67 . It seems that formicine ants also create an acidic environment in the stomach by actively ingesting their venom gland secretions, thereby improving individual survival in the face of pathogen contaminated food and limits disease transmission during mutual food exchange (trophallaxis) 68 . This way the infected corpses can elicit only a low-intensity infection in the grooming individuals, inducing the production of specific protective peptides and enzymes that achieve the promptness of the immune system 34,35 .…”

Competition-based screening secures the evolutionary stability of a defensive microbiome

“…Screening has also been suggested to act in animal-microbe symbioses. For instance, Tragust et al 14 showed that carpenter ants acidify their own stomachs by swallowing acidopore secretions. Entomopathogenic bacteria are rapidly killed off, whereas the gut bacterial symbiont Asiaia sp.…”