Although DNA sequences of linear mitochondrial genomes are available for a wide variety of species, sequence and conformational data from the extreme ends of these molecules (i.e., the telomeres) are limited. Data on the telomeres is important because it can provide insights into how linear genomes overcome the end-replication problem. This study explores the evolution of linear mitochondrial DNAs (mtDNAs) in the green-algal genus Polytomella (Chlorophyceae, Chlorophyta), the members of which are non-photosynthetic. Earlier works analyzed the linear and linear-fragmented mitochondrial genomes of Polytomella capuana and Polytomella parva. Here we present the mtDNA sequence for Polytomella strain SAG 63-10 [also known as Polytomella piriformis (Pringsheim 1963)], which is the only known representative of a mostly unexplored Polytomella lineage. We show that the P. piriformis mtDNA is made up of two linear fragments of 13 and 3 kb. The telomeric sequences of the large and small fragments are terminally inverted, and appear to end in vitro with either closed (hairpin-loop) or open (nicked-loop) structures as also shown here for P. parva and shown earlier for P. capuana. The structure of the P. piriformis mtDNA is more similar to that of P. parva, which is also fragmented, than to that of P. capuana, which is contained in a single chromosome. Phylogenetic analyses reveal high substitution rates in the mtDNA of all three Polytomella species relative to other chlamydomonadalean algae. These elevated rates could be the result of a greater number of vegetative cell divisions and/or small population sizes in Polytomella species as compared with other chlamydomonadalean algae.