Stefanie Kautz

Highly similar microbial communities are shared among related and trophically similar ant species.

Ants dominate many terrestrial ecosystems, yet we know little about their nutritional physiology and ecology. While traditionally viewed as predators and scavengers, recent isotopic studies revealed that many dominant ant species are functional herbivores. As with other insects with nitrogen‐poor diets, it is hypothesized that these ants rely on symbiotic bacteria for nutritional supplementation. In this study, we used cloning and 16S sequencing to further characterize the bacterial flora of several herbivorous ants, while also examining the beta diversity of bacterial communities within and between ant species from different trophic levels. Through estimating phylogenetic overlap between these communities, we tested the hypothesis that ecologically or phylogenetically similar groups of ants harbor similar microbial flora. Our findings reveal: (i) clear differences in bacterial communities harbored by predatory and herbivorous ants; (ii) notable similarities among communities from distantly related herbivorous ants and (iii) similar communities shared by different predatory army ant species. Focusing on one herbivorous ant tribe, the Cephalotini, we detected five major bacterial taxa that likely represent the core microbiota. Metabolic functions of bacterial relatives suggest that these microbes may play roles in fixing, recycling, or upgrading nitrogen. Overall, our findings reveal that similar microbial communities are harbored by ants from similar trophic niches and, to a greater extent, by related ants from the same colonies, species, genera, and tribes. These trends hint at coevolved histories between ants and microbes, suggesting new possibilities for roles of bacteria in the evolution of both herbivores and carnivores from the ant family Formicidae.

Divergent investment strategies of Acacia myrmecophytes and the coexistence of mutualists and exploiters.

Ant-plant interactions represent a diversity of strategies, from exploitative to mutualistic, and how these strategies evolve is poorly understood. Here, we link physiological, ecological, and phylogenetic approaches to study the evolution and coexistence of strategies in the Acacia–Pseudomyrmex system. Host plant species represented 2 different strategies. High-reward hosts produced significantly more extrafloral nectar (EFN), food bodies, and nesting space than low-reward hosts, even when being inhabited by the same species of ant mutualist. High-reward hosts were more effectively defended against herbivores and exploited to a lower extent by nondefending ants than low-reward hosts. At the phenotypic level, secretion of EFN and ant activity were positively correlated and a mutualistic ant species induced nectar secretion, whereas a nondefending exploiter did not. All of these mechanisms contribute to the stable association of high-reward hosts with defending ant species. However, exploiter ants are less dependent on the host-derived rewards and can colonize considerable proportions of the low-reward hosts. Mapping these strategies onto phylogenetic trees demonstrated that the low-reward hosts represent the derived clade within a monophyletic group of obligate ant plants and that the observed exploiter ant species evolved their strategy without having a mutualistic ancestor. We conclude that both types of host strategies coexist because of variable net outcomes of different investment–payoff regimes and that the effects of exploiters on the outcome of mutualisms can, thus, increase the diversity within the taxa involved.

Surveying the Microbiome of Ants: Comparing 454 Pyrosequencing with Traditional Methods To Uncover Bacterial Diversity

ABSTRACT We are only beginning to understand the depth and breadth of microbial associations across the eukaryotic tree of life. Reliably assessing bacterial diversity is a key challenge, and next-generation sequencing approaches are facilitating this endeavor. In this study, we used 16S rRNA amplicon pyrosequencing to survey microbial diversity in ants. We compared 454 libraries with Sanger-sequenced clone libraries as well as cultivation of live bacteria. Pyrosequencing yielded 95,656 bacterial 16S rRNA reads from 19 samples derived from four colonies of one ant species. The most dominant bacterial orders in the microbiome of the turtle ant Cephalotes varians were Rhizobiales, Burkholderiales, Opitutales, Xanthomonadales, and Campylobacterales, as revealed through both 454 sequencing and cloning. Even after stringent quality filtering, pyrosequencing recovered 445 microbe operational taxonomic units (OTUs) not detected with traditional techniques. In comparing bacterial communities associated with specific tissues, we found that gut tissues had significantly higher diversity than nongut tissues, and many of the OTUs identified from these groups clustered within ant-specific lineages, indicating a deep coevolutionary history of Cephalotes ants and their associated microbes. These lineages likely function as nutritional symbionts. One of four ant colonies investigated was infected with a Spiroplasma sp. (order Entomoplasmatales), a potential ant pathogen. Our work shows that the microbiome associated with Cephalotes varians is dominated by a few dozen bacterial lineages and that 454 sequencing is a cost-efficient tool to screen ant symbiont diversity.

Cyanogenesis of Wild Lima Bean (Phaseolus lunatus L.) Is an Efficient Direct Defence in Nature

A broad range of chemical plant defenses against herbivores has been studied extensively under laboratory conditions. In many of these cases there is still little understanding of their relevance in nature. In natural systems, functional analyses of plant traits are often complicated by an extreme variability, which affects the interaction with higher trophic levels. Successful analyses require consideration of the numerous sources of variation that potentially affect the plant trait of interest. In our recent study on wild lima bean (Phaseolus lunatus L.) in South Mexico, we applied an integrative approach combining analyses for quantitative correlations of cyanogenic potential (HCNp; the maximum amount of cyanide that can be released from a given tissue) and herbivory in the field with subsequent feeding trials under controlled conditions. This approach allowed us to causally explain the consequences of quantitative variation of HCNp on herbivore-plant interactions in nature and highlights the importance of combining data obtained in natural systems with analyses under controlled conditions.

How to Prevent Cheating: A Digestive Specialization Ties Mutualistic Plant-Ants to Their Ant-Plant Partners

Mutualisms often involve reciprocal adaptations of both partners. Acacia ant-plants defended by symbiotic Pseudomyrmex ant mutualists secrete sucrose-free extrafloral nectar, which is unattractive to generalists. We aimed to investigate whether this extrafloral nectar can also exclude exploiters, that is nondefending ant species. Mutualist workers discriminated against sucrose whereas exploiters and generalists with no affinity toward Acacia myrmecophytes preferred sucrose, because mutualist workers lacked the sucrose-cleaving enzyme invertase, which is present in workers of the other two groups. Sucrose uptake induced invertase activity in workers of parasites and generalists, but not mutualists, and in larvae of all species: the mutualists loose invertase during their ontogeny. This reduced metabolic capacity ties the mutualists to their plant hosts, but it does not completely prevent the mutualism from exploitation. We therefore investigated whether the exploiters studied here are cheaters (i.e., have evolved from former mutualists) or parasites (exploiters with no mutualistic ancestor). A molecular phytogeny demonstrates that the exploiter species did not evolve from former mutualists, and no evidence for cheaters was found. We conclude that being specialized to their partner can prevent mutualists from becoming cheaters, whereas other mechanisms are required to stabilize a mutualism against the exploitation by parasites.

Induced plant defense via volatile production is dependent on rhizobial symbiosis

Nitrogen-fixing rhizobia can substantially influence plant–herbivore interactions by altering plant chemical composition and food quality. However, the effects of rhizobia on plant volatiles, which serve as indirect and direct defenses against arthropod herbivores and as signals in defense-associated plant–plant and within-plant signaling, are still unstudied. We measured the release of jasmonic acid (JA)-induced volatiles of rhizobia-colonized and rhizobia-free lima bean plants (Fabaceae: Phaseolus lunatus L.) and tested effects of their respective bouquets of volatile organic compounds (VOCs) on a specialist insect herbivore (Mexican bean beetle; Coccinellidae: Epilachna varivestis Mulsant) in olfactometer choice trials. In a further experiment, we showed that VOC induction by JA reflects the plant responses to mechanical wounding and insect herbivory. Following induction with JA, rhizobia-colonized plants released significantly higher amounts of the shikimic acid-derived compounds, whereas the emission of compounds produced via the octadecanoid, mevalonate and non-mevalonate pathways was reduced. These changes affected the choice behavior of beetles as the preference of non-induced plants was much more pronounced for plants that were colonized by rhizobia. We showed that indole likely represents the causing agent for the observed repellent effects of jasmonic acid-induced VOCs of rhizobia-colonized lima bean plants. Our study demonstrates a rhizobia-triggered efficacy of induced plant defense via volatiles. Due to these findings, we interpret rhizobia as an integral part of legume defenses against herbivores.

6-MSAS-like polyketide synthase genes occur in lichenized ascomycetes

Lichenized and non-lichenized filamentous ascomycetes produce a great variety of polyketide secondary metabolites. Some polyketide synthase (PKS) genes from non-lichenized fungi have been characterized, but the function of PKS genes from lichenized species remains unknown. Phylogenetic analysis of keto synthase (KS) domains allows prediction of the presence or absence of particular domains in the PKS gene. In the current study we screened genomic DNA from lichenized fungi for the presence of non-reducing and 6-methylsalicylic acid synthase (6-MSAS)-type PKS genes. We developed new degenerate primers in the acyl transferase (AT) region to amplify a PKS fragment spanning most of the KS region, the entire linker between KS and AT, and half of the AT region. Phylogenetic analysis shows that lichenized taxa possess PKS genes of the 6-MSAS-type. The extended alignment confirms overall phylogenetic relationships between fungal non-reducing, 6-MSAS-type and bacterial type I PKS genes.

Quantitative variability of cyanogenesis in Cathariostachys madagascariensis—the main food plant of bamboo lemurs in Southeastern Madagascar

Giant bamboo (Cathariostachys madagascariensis) is a major food plant for three sympatric species of bamboo‐eating lemurs (Hapalemur aureus, H. griseus, and Prolemur simus) in the rain forests of southeastern Madagascar. This plant species is strongly cyanogenic. However, quantitative data on cyanide concentration in C. madagascariensis are scarce. Previous studies reported 15 mg cyanide per 100 g fresh shoot material (corresponding to approx. 57 µmol cyanide per gram dry weight). However, we found mean concentrations (±SE) ranging from 139.3±19.32 in ground shoots to 217.7±16.80 µmol cyanide per gram dry weight in branch shoots. Thus, cyanogenesis of C. madagascariensis was up to four times higher than reported before. In contrast to the strongly cyanogenic shoots no cyanide could be detected in differently aged leaves of C. madagascariensis confirming earlier studies. Within individual shoots fine‐scaled analysis revealed a characteristic ontogenetic pattern of cyanide accumulation. Highest concentrations were found in youngest parts near the apical meristem, whereas concentrations decreased in older shoot parts. Beyond the general intra‐individual variability of cyanogenic features analyses indicated site‐specific variability of both, the ontogenetic pattern of cyanide concentration as well as the total amount of cyanide accumulated in shoots. Additionally, analyses of soluble proteins—one important nutritive measure affecting food plant quality—demonstrated a converse quantitative relation of protein concentrations in leaves to cyanide concentration in shoots at the site‐specific level. We, thus, suggest integrative analyses on quantitative variation of cyanogenesis together with nutritive plant parameters in future studies. This approach would allow obtaining more detailed insights into spatial variability of giant bamboos overall browse quality and its impact on lemur herbivores. Am. J. Primatol. 71:305–315, 2009.

Bacterial Infections across the Ants: Frequency and Prevalence of Wolbachia, Spiroplasma, and Asaia

Bacterial endosymbionts are common across insects, but we often lack a deeper knowledge of their prevalence across most organisms. Next-generation sequencing approaches can characterize bacterial diversity associated with a host and at the same time facilitate the fast and simultaneous screening of infectious bacteria. In this study, we used 16S rRNA tag encoded amplicon pyrosequencing to survey bacterial communities of 310 samples representing 221 individuals, 176 colonies and 95 species of ants. We found three distinct endosymbiont groups—Wolbachia (Alphaproteobacteria: Rickettsiales), Spiroplasma (Firmicutes: Entomoplasmatales), and relatives of Asaia (Alphaproteobacteria: Rhodospirillales)—at different infection frequencies (at the ant species level: 22.1%, 28.4%, and 14.7%, resp.) and relative abundances within bacterial communities (1.0%–99.9%). Spiroplasma was particularly enriched in the ant genus Polyrhachis, while Asaia relatives were most prevalent in arboreal ants of the genus Pseudomyrmex. While Wolbachia and Spiroplasma have been surveyed in ants before, Asaia, an acetic acid bacterium capable of fixing atmospheric nitrogen, has received much less attention. Due to sporadic prevalence across all ant taxa investigated, we hypothesize facultative associations for all three bacterial genera. Infection patterns are discussed in relation to potential adaptation of specific bacteria in certain ant groups.

Distance and sex determine host plant choice by herbivorous beetles.

Background Plants respond to herbivore damage with the release of volatile organic compounds (VOCs). This indirect defense can cause ecological costs when herbivores themselves use VOCs as cues to localize suitable host plants. Can VOCs reliably indicate food plant quality to herbivores? Methodology We determined the choice behavior of herbivorous beetles (Chrysomelidae: Gynandrobrotica guerreroensis and Cerotoma ruficornis) when facing lima bean plants (Fabaceae: Phaseolus lunatus) with different cyanogenic potential, which is an important constitutive direct defense. Expression of inducible indirect defenses was experimentally manipulated by jasmonic acid treatment at different concentrations. The long-distance responses of male and female beetles to the resulting induced plant volatiles were investigated in olfactometer and free-flight experiments and compared to the short-distance decisions of the same beetles in feeding trials. Conclusion Female beetles of both species were repelled by VOCs released from all induced plants independent of the level of induction. In contrast, male beetles were repelled by strongly induced plants, showed no significant differences in choice behavior towards moderately induced plants, but responded positively to VOCs released from little induced plants. Thus, beetle sex and plant VOCs had a significant effect on host searching behavior. By contrast, feeding behavior of both sexes was strongly determined by the cyanogenic potential of leaves, although females again responded more sensitively than males. Apparently, VOCs mainly provide information to these beetles that are not directly related to food quality. Being induced by herbivory and involved in indirect plant defense, such VOCs might indicate the presence of competitors and predators to herbivores. We conclude that plant quality as a food source and finding a potentially enemy-free space is more important for female than for male insect herbivores, whereas the presence of a slightly damaged plant can help males to localize putative mating partners.