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A total of 363 Ayoreo Indians living in three localities of southern Bolivia and northern Paraguay were studied in relation to 33 genetic systems which are expressed in blood and one in saliva. 3 he degree of intratribal diversity observed is not large, in agreement with the considerable rate of exogamy found among them. They are unusual among South American Indians in showing no variation at the Esterase D locus (Es D\. 1.00), very high frequencies of TMS (MNSs blood groups) and ACPA (acid phosphatase system), as well as a low frequency of P'. No Diego (a) or Cm (x) positives were found among them. The frequencies of the other markers do not differ appreciably from those commonly encountered and no rare variants were observed. A six-locus comparison of the Ayoreo with 21 other Middle or South American tribes shows genetic distances that are generally larger than those obtained with any other of these groups.