Chronic, culture-negative septic arthritis represents a major clinical dilemma. It often results in prolonged courses of antimicrobial therapy, which may or may not have activity against the organism responsible for the infection. It is not possible to cultivate all types of pathogens in vitro with use of traditional laboratory testing methods, but emerging techniques can be helpful to define the etiology of these infections in some cases. We describe a case of a periprosthetic joint infection in a thirty-year-old woman with juvenile rheumatoid arthritis (JRA) and hypogammaglobulinemia, who developed septic arthritis after prolonged pneumonia. She received multiple courses of antimicrobial therapy without improvement of the joint symptoms. The microbiologic diagnosis was made with use of 16S ribosomal RNA (rRNA) gene amplification and sequencing from synovial fluid and tissue. The patient gave permission for this information to be submitted for publication.