The objectives of this study was to compare morphological characteristics and to establish evolutionary relationship of 14 Thai dwarf buffaloes and 21 swamp buffaloes based on mitochondrial DNA (mtDNA) D-loop variations. Morphological characteristics could be constructed phylomorphologic tree and showed clearly classified between dwarf and swamp buffaloes. Most of morphologic traits were highly significant difference adult contest swamp buffaloes (P LT 0.01) from 9 contest swamp buffaloes. Only morphological traits of length between eye, length between base of horn, horn length and horn width were not significant difference (P GT 0.05). Average shoulder height of Thai dwarf buffaloes were 108.33+-2.08 cm, whereas average shoulder height of contest swamp buffaloes were 156.78+-4.21 cm. Thai dwarf buffaloes would be defined as disproportional dwarfism. Sequences of mtDNA D-loop (374 bp) of dwarf and normal swamp buffaloes with reference sequences showed 84 polymorphic sites and defined as 12 and 11 haplotypes, respectively. It was noticed that transversion in Thai dwarf buffaloes occurred twice time of normal swamp buffaloes. Phylogenetic tree showed 2 clades of water buffaloes. Swamp buffaloes were classified into lineage A and lineage B, in which lineage A was more predominant than lineage B. Median joining network showed 2 clades of river and swamp buffaloes. Swamp buffalo (SaenCP) in lineage A1 seem to be ancestral node of some Thai swamp buffaloes, Chinese swamp buffaloes and Philippines carabao. Most of dwarf swamp buffaloes were in lineage A2 and dwarf swamp buffalo with no horn, black color- coat was latter evolution.