Hemogregarines from Thamnophis s. sirtalis, Coluber constrictor priapus, Elaphe obsoleta quadrivittata, and E. g. guttata in northern Florida appeared to be conspecific on the basis of similar gamonts from all the hosts and sporogonic stages obtained from 3 hosts. The resemblance of gamonts to those of Hepatozoon sauritus, described from T. sauritus sackenii in southern Florida, justified comparison of DNA isolates from the type infection of H. sauritus with samples from each of the northern Florida hosts and with a morphologically distinct species, H. sirtalis, from northern Florida. A nucleotide sequence (530 bp) alignment of the 18S ribosomal RNA gene revealed 2 hemogregarine haplotypes that varied at 15 sites (p distance = 2.8%), which included 10 transitions and 5 transversions. Two well-supported clusters (100% bootstrap support) were revealed by a neighbor-joining tree topology. One cluster included the type infection of H. sauritus and all 4 of the other samples from the northern Florida hosts, with samples of H. sirtalis comprising a second cluster. Hepatozoon sauritus, therefore, is a polytopic species in contrast to the 8 other Hepatozoon species thus far described from snakes in Florida, each of which appears to parasitize a single host species.