Development of a genomics-based PCR assay for detection of Mycoplasma pneumoniae in a large outbreak in New York State

A genomics-based PCR method was developed and used to test specimens from p atients involved in a large outbreak of Mycoplasma pneumoniae in a closed r eligious community in Ne rv York State. New Fl adhesin gene primers were de signed to bind to 9 of 10 target sequences in the repetitive-element sequen ces obtained from the whole genome sequence of M. pneumoniae. This PCR meth od had a sensitivity of 0.006 CFU and a specificity of 100% for Al. pneumon iae. The PCR was validated by testing a subset of patient samples by cultur e and comparing the results to those obtained by PCR, Of the initial 280 sa mples tested, 73 were positive by PCR and 22 were positive by culture. All samples positive by culture were also positive by PCR, Follow-up testing of selected patients 3 to h weeks after antibiotic treatment revealed that ei ght samples remained positive by PCR and that three samples remained positi ve by culture. Additionally, no nonspecific PCR inhibition was detected as a result of the specimen type, transport medium, or sample preparation meth odology. The study demonstrates that the PCR described here is a rapid, sen sitive, and specific method for the identification of M. pneumoniae and was helpful for the detection and monitoring of the outbreak.