Abstract. The 16S ribosomal RNA (rRNA) gene of Eperythrozoon suis was amplified using gene-specific primers developed from GenBank sequence accession U88565. The gene was subsequently cloned and sequenced. Based on these sequence data, 3 sets of E. suis-specific primers were designed. These primers selectively amplified 1394, 690, and 839 base-pair (bp) fragments of the 16S rRNA gene from DNA of E. suis extracted from the blood of an experimentally infected pig during a parasitemic episode. No polymerase chain reaction (PCR) products were amplified from purified DNA of Haemobartonella felis, Mycoplasma genitalium, or Bartonella bacilliformis using 2 of these primer sets. When the primer set amplifying the 690-bp fragment was used, faint bands were observed with H. felis as the target DNA. No PCR products were amplified from DNA that had been extracted from the blood of a noninfected pig or using PCR reagents without target DNA. The detection limits for E. suis by competitive quantitative PCR were estimated to range from 57 and 800 organisms/assay. This is the first report of the utility of PCR-facilitated diagnosis and quantitation of E. suis based on the 16S rRNA gene. The PCR method developed will be useful in monitoring the progression and significance of E. suis in the disease process in the pig.Eperythrozoon suis is an extracellular, rod-shaped parasite that attaches to and causes deformity and damage to the red blood cells of pigs. 27 The disease, eperythrozoonosis, is characterized by 4 syndromes: 1) decreased reproductive efficiency of sows, 2) weakness and anemia in baby pigs with increased incidence of enteric and respiratory infections, 3) delayed production gains in feeder pigs, and 4) acute hemolytic anemia in feeder pigs. 5,14,35 This disease was first reported in the United States in 1934. 16 Nonetheless, our understanding of this organism and the disease(s) it causes remains largely incomplete.Eperythrozoon suis has not been successfully grown in agar or cell cultures. Diagnosis of acute infection with E. suis is usually based on direct microscopic observation of organisms attached to red blood cells on Giemsa-stained peripheral blood smears. However, this method has limitations because the parasitemia is transient and parasites are not often seen on blood smears until clinical signs develop. 27 The diagnosis of eperythrozoonosis is also complicated by the lack of a readily identifiable parasitemia in latent and chronic infections. 27 variability in antibody response and failure to identify acutely infected pigs. The lack of an efficient test for diagnosing acute and chronic E. suis infection has resulted in tremendous controversy concerning the true impact of this disease in pig populations. The refinement of the polymerase chain reaction (PCR) during the past decade has presented new opportunities for the development of diagnostic tests that have the potential to be highly sensitive and specific. In this respect, there are reports of a PCR protocol for detection of E. suis infection in pigs ba...