The B allele of the bovine alpha (S2)-casein gene (CSN1S2) was characterized at the molecular level and the distribution of zebu-specific milk protein alleles was determined in 26 cattle breeds originating from 3 continents. The CSN1S2*B allele is characterized by a C --> T transition affecting nucleotide 17 of exon 3, which leads to a change in the eighth amino acid of the mature protein, from Ser to Phe (i.e., TCC --> TTC). DNA-based methods were developed to identify carriers of CSN1S2*B and the other alleles (CSN1S2*A, C, and D) at the same locus. CSN1S2*B and other zebu-specific milk protein alleles and casein haplotypes are widely distributed in European cattle breeds, particularly those of southeastern origin. Alleles CSN1S2*B and CSN3*H are important in searching for zebu imprints in European cattle breeds. Diversity estimates at the milk protein loci were highest in the zebus followed by southeastern European taurines. Anatolian Black had the highest number of zebu alleles among European taurines. Common, group, and intergroup haplotypes occurred in the breeds and demonstrated relationships that concurred with developmental histories, genetic makeup, and, in particular, exposed the extent of zebu influence on southeastern European cattle.