The genetic diversity of Australian bat lyssavirus (ABL) was investigated by comparing 24 ABL isolate glycoprotein (G) gene nucleotide sequences with those of 37 lyssaviruses representing Lyssavirus genotypes 1-6. Phylogenetic analyses indicated that ABL forms a monophyletic group separate from other lyssaviruses. This group differentiates into two clades: one associated with Pteropus (flying fox) species, the other with the insectivorous bat Saccolaimus flaviventris. Calculation of percentage nucleotide identities between isolates of the two clades revealed up to 18.7 % nucleotide sequence divergence between the two ABL variants. These observations suggest that ABL is a separate lyssavirus species with a similar epidemiology to chiropteran rabies virus (RV), where two distinct ABL variants co-exist in Australia in bat species with dissimilar ecology. Analyses of selection pressures in ABL G gene sequences provided some evidence of weak positive selection within the endodomain at amino acids 499 and 501, although in general the dominant evolutionary process observed was purifying selection. This intimates that, in nature, isolates of ABL, like those of RV, are subject to relatively strong selective constraints, suggesting a stability of host species, cell tropisms and ecological conditions.