Erythrocyte Pyruvate Kinase Deficiency (PK deficiency) is an inherited hemolytic anemia that has been documented in the Abyssinian and Somali breeds as well as random bred domestic shorthair cats. The disease results from mutations in PKLR, the gene encoding the regulatory glycolytic enzyme pyruvate kinase (PK). PK catalyzes the transformation of phosphoenolpyruvate to pyruvate and ATP. This reaction is the major source of energy for erythrocytes. Multiple isozymes are produced by tissue-specific differential processing of PKLR mRNA. Perturbation of PK decreases erythrocyte longevity resulting in anemia. Additional symptoms include: severe lethargy, weakness, weight loss, jaundice, and abdominal enlargement. In domestic cats, PK deficiency has an autosomal recessive mode of inheritance with high variability in onset and severity of clinical symptoms. Sequence analysis of PKLR revealed an intron 5 SNP concordant with the disease phenotype in Abyssinian and Somali cats. Located 52 nucleotides 5’ of the exon 6 splice site, cats with this SNP produce liver and blood processed mRNA with a 13 bp deletion at the 3’ end of exon 5. The resultant frame-shift mutation creates a stop codon at amino acid position 252 in exon 6. Samples from over 12,000 cats representing 40 breeds (minimum 10 unrelated samples) were assayed to determine the presence and frequency of the concordant SNP. In addition to Abyssinian and Somali cats, 14 additional breeds possess the diagnostic PK deficiency SNP. The frequency of the affected allele across the entire data set is 4.79% and 7.72% when restricted to the 16 breeds carrying the concordant SNP.