Fungus-growing termites rely on mutualistic fungi of the genus Termitomyces and gut microbes for plant biomass degradation. Due to a certain degree of symbiont complementarity, this tripartite symbiosis has evolved as a complex bioreactor, enabling decomposition of nearly any plant polymer, likely contributing to the success of the termites as one of the main plant decomposers in the Old World. In this study, we evaluated which plant polymers are decomposed and which enzymes are active during the decomposition process in two major genera of fungus-growing termites. We found a diversity of active enzymes at different stages of decomposition and a consistent decrease in plant components during the decomposition process. Furthermore, our findings are consistent with the hypothesis that termites transport enzymes from the older mature parts of the fungus comb through young worker guts to freshly inoculated plant substrate. However, preliminary fungal RNA sequencing (RNA-seq) analyses suggest that this likely transport is supplemented with enzymes produced in situ. Our findings support that the maintenance of an external fungus comb, inoculated with an optimal mixture of plant material, fungal spores, and enzymes, is likely the key to the extraordinarily efficient plant decomposition in fungus-growing termites.IMPORTANCE Fungus-growing termites have a substantial ecological footprint in the Old World (sub)tropics due to their ability to decompose dead plant material. Through the establishment of an elaborate plant biomass inoculation strategy and through fungal and bacterial enzyme contributions, this farming symbiosis has become an efficient and versatile aerobic bioreactor for plant substrate conversion. Since little is known about what enzymes are expressed and where they are active at different stages of the decomposition process, we used enzyme assays, transcriptomics, and plant content measurements to shed light on how this decomposition of plant substrate is so effectively accomplished.

In the vast majority of sexual life cycles, fusion between single-celled gametes is directly followed by nuclear fusion, leading to a diploid zygote and a lifelong commitment between two haploid genomes. Mushroom-forming basidiomycetes differ in two key respects. First, the multicellular haploid mating partners are fertilized in their entirety, each cell being a gamete that simultaneously can behave as a female, i.e. contributing the cytoplasm to a zygote by accepting nuclei, and a male gamete, i.e. only donating nuclei to the zygote. Second, after gamete union, the two haploid genomes remain separate so that the main vegetative stage, the dikaryon, has two haploid nuclei per cell. Only when the dikaryon produces mushrooms, do the nuclei fuse to enter a short diploid stage, immediately followed by meiosis and haploid spore formation. So in basidiomycetes, gamete fusion and genome mixing (sex) are separated in time. The ‘living apart together’ of nuclei in the dikaryon maintains some autonomy for nuclei to engage in a relationship with a different nucleus. We show that competition among the two nuclei of the dikaryon for such ‘extramarital affairs’ may lead to genomic conflict by favouring genes beneficial at the level of the nucleus, but deleterious at that of the dikaryon.This article is part of the themed issue ‘Weird sex: the underappreciated diversity of sexual reproduction’.

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Although mutualistic symbioses per definition are beneficial for interacting species, conflict may arise if partners reproduce independently. We address how this reproductive conflict is regulated in the obligate mutualistic symbiosis between fungus-growing termites and Termitomyces fungi. Even though the termites and their fungal symbiont disperse independently to establish new colonies, dispersal is correlated in time. The fungal symbiont typically forms mushrooms a few weeks after the colony has produced dispersing alates. It is thought that this timing is due to a trade-off between alate and worker production; alate production reduces resources available for worker production. As workers consume the fungus, reduced numbers of workers will allow mushrooms to ‘escape’ from the host colony. Here, we test a specific version of this hypothesis: the typical asexual structures found in all species of Termitomyces —nodules—are immature stages of mushrooms that are normally harvested by the termites at a primordial stage. We refute this hypothesis by showing that nodules and mushroom primordia are macro- and microscopically different structures and by showing that in the absence of workers, primordia do, and nodules do not grow out into mushrooms. It remains to be tested whether termite control of primordia formation or of primordia outgrowth mitigates the reproductive conflict.

The asexual spore balls formed by Termitomyces are not the primordia of mushrooms (this thesis). 2. There is no such thing as a standard life cycle for basidiomycete fungi (this thesis) 3. If the outcome of a student's project is vital to her or his supervisor, a conflict of interest is inevitable. 4. Society is the customer of science. 5. Selection for soundbites and catchy headlines in science communication is in conflict with the credibility of science as a whole. 6. The core issue for the acceptance of evolutionary theory is the question whether evolutionary theory applies to humans. 7. Differences in payed parental-leave drive the observed patterns of paternal and maternal childcare. 8. Men in top positions are often not hired for their qualities, but rather because they are men.

Background The termite-fungus symbiosis is an ancient stable mutualism of two partners that reproduce and disperse independently. With the founding of each termite colony the symbiotic association must be re-established with a new fungus partner. Complementarity in the ability to break down plant substrate may help to stabilize this symbiosis despite horizontal symbiont transmission. An alternative, non-exclusive, hypothesis is that a reduced rate of evolution may contribute to stabilize the symbiosis, the so-called Red King Effect. Methods To explore this concept, we produced the first linkage map of a species of Termitomyces, using genotyping by sequencing (GBS) of 88 homokaryotic offspring. We constructed a highly contiguous genome assembly using PacBio data and a de-novo evidence-based annotation. This improved genome assembly and linkage map allowed for examination of the recombination landscape and its potential effect on the mutualistic lifestyle. Results Our linkage map resulted in a genome-wide recombination rate of 22 cM/Mb, lower than that of other related fungi. However, the total map length of 1370 cM was similar to that of other related fungi. Conclusions The apparently decreased rate of recombination is primarily due to genome expansion of islands of gene-poor repetitive sequences. This study highlights the importance of inclusion of genomic context in cross-species comparisons of recombination rate.

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