Mutualism, whereby species interact to their mutual benefit, is extraordinary in a competitive world. To recognize general patterns of origin and maintenance from the plethora of mutualistic associations proves a persisting challenge. The simplest situation is believed to be that of a single mutualist specific to a single host, vertically transmitted from one host generation to the next. We characterized ascomycete fungal associates cultured for nest architecture by the ant subgenera Dendrolasius and Chthonolasius. The ants probably manage their fungal mutualists by protecting them against fungal competitors. The ant subgenera display different ant-to-fungus specificity patterns, one-to-two and many-to-one, and we infer vertical transmission, in the latter case overlaid by horizontal transmission. Possible evolutionary trajectories include a reversal from fungiculture by other Lasius subgenera and inheritance of fungi through life cycle interactions of the ant subgenera. The mosaic indicates how specificity patterns can be shaped by an interplay between host life-cycles and transmission adaptations.