In Australia, the endemic fungus Plectosporium alismatis (syn. Rhynchosporium alismatis) has potential use as a mycoherbicide for several species in the Alismataceae, a family of aquatic and semi-aquatic marsh herbs, which are considered to be important weeds in rice crops. Of five species identified in south-eastern Australia where rice is grown, two species, Sagittaria graminea and Sagittaria montevidensis are resistant (non-hosts), and no records of P. alismatis on these species have been reported. To better understand the interactions that lead to resistance in these pathosystems, the infection process of the fungus was studied on these species and also on the host Alisma plantago-aquatica, using light, fluorescent and scanning electron microscopy. On all three species both conidial germination and appressorium formation commenced within 6 h of inoculation with greater than 50% of conidia elongating to form germ tube structures and associated appressoria 12–18 h post inoculation. Germ tube elongation and appressorium formation occurred randomly over the leaf surface. Direct host penetration was facilitated by the production of penetration hyphae that emerged from beneath appressoria. Penetration sites were clearly identified by the presence of spherical holes 0.25–0.5 μm in diam, and were frequently accompanied by resistance reactions in non-host species. Visible symptoms of disease occurred 4–6 d after inoculation of susceptible (host) species.