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A polymerase chain reaction (PCR)-based restriction fragment-length polymorphism (RFLP) assay was developed that discriminates among the 4 mitotypes found in North, Central, and South American honey bee racial groups—eastern European (Apis mellifera ligustica Spinola, caucasica Gorbachev, and carnica Pollman), western European (A. m. mellifera Linnaeus), Egyptian (A. m. lamarckii Cockerell Lepeletier), and sub-Saharan African (A. m. scutellata). Before the development of this assay, 13% of southern Californian feral bees collected before the arrival of the Apis mellifera scutellata (Africanized) race were found to contain a non-European mitochondrial genotype that could not be distinguished from that of A. m. scutellata. DNA sequence analysis suggests the unusual mitotype to be that of A. m. lamarckii. An RFLP polymorphism was identified that distinguished this subspecies from all others present in North America. This polymorphism was not found in any of 96 bees collected primarily in Mexico and Central America. Thus, the Egyptian mitochondrial type is either absent or extremely rare in these regions. The PCR assay also distin- guishes A. m. lamarckii from 2 other north African racial types, A. m. intermissa Buttel-Reepen and A. m. sahariensis Baldensperger.

We propose an Africanized honey bee identification strategy using morphometrics and an improved polymerase chain reaction (PCR)-based mitotyping procedure that distinguishes between feral and commercial bees maternally descendent from 4 racial groups-Eastern European (Apis mellifera ligustica, caucasica, and carnica), Western European (A. m. mellifera), Egyptian (A. m. lamarckii), and other African origins. Mitochondrial genotype is highly correlated with morphology. Ninety-five percent of morphometrically determined Africanized feral colonies collected in Texas, Arizona, California, and Mexico also contained African mitochondria. Sixty-two percent of colonies from commercial or minimally managed apiaries in Mexico and Central America, with Africanized forewing lengths and 17% of colonies with intermediate forewing lengths, had African mitochondria. The strong correlation between non-European morphology and African mitotype, as well as the speed and accuracy of mitotype determination, suggest a 3-step Africanized bee identification procedure. This identification procedure first examines forewing length. Bees with lengths above a given threshold (9.12 mm) have a very high probability of being pure European in origin and are not examined further. Those bees with wing lengths below the threshold are subjected to mitochondrial analysis (mitotyping). Samples having African mitochondria are not examined further. Those bees with small forewing lengths, but European mitotypes, are then identified using detailed morphometric discriminant function analysis. By performing these steps in sequence, the number of bees requiring full morphometric analysis is reduced, saving time and improving the accuracy of Africanized honey bee identification.