METHODS OF TARGETED ANTIBIOTIC SUSCEPTIBILITY TESTING

摘要

Methods are provided for performing antibotic susceptibility testing based on the detection of RNA, such as tmRNA, from microbial cells after exposure to antibiotics. In some embodiments, aliquots are obtained from a sample, one of which contains a selected antibiotic. The aliquots, which include growth media, are incubated under conditions suitable for microbial growth, and the microbial cells in each aliquot are removed and lysed, and the lysate is subjected to reverse transcription and amplification in infer the effect of the selected antibiotic on the microbial cells. In one embodiment, a sample containing microbial cells is incubated in the pressence of a selected antibiotic and a stimulus is provided to induce the production on m RNA within the microbial cells. The microbial cells are subsequently lysed without substantial degradation of the m RNA within the lysate, and the m RNA is detected to determine the effect of the antibiotic on the microbial cells.

主权项

1. A method of performing rapid antibiotic susceptibility testing, comprising:
performing a multiplexed identification test panel on a first lysate, the first lysate having been obtained from a first sample that is suspected of containing microbial cells; obtaining, from a second sample that is suspected of containing the microbial cells, at least a primary aliquot and a reference aliquot, wherein the primary aliquot and the reference aliquot comprise growth media, and wherein the first sample and the second sample are derived from a common subject; adding, to the primary aliquot, at least one selected antibiotic, the selected antibiotic having been selected, at least in part, based on the results of the multiplexed identification test panel; incubating the primary aliquot and the reference aliquot under conditions suitable for promoting microbial growth for testing the effectiveness of the selected antibiotic; separating microbial cells from the primary aliquot and resuspending the separated microbial cells to obtain a primary suspension; separating microbial cells from the reference aliquot and resuspending the separated microbial cells to obtain a reference suspension; lysing the microbial cells in the primary suspension and the reference suspension, thereby obtaining a primary lysate and a reference lysate; performing reverse transcription and amplification on the primary lysate and to detect nucleic acids associated with microbial cells detectable by the multiplexed identification test panel, thereby obtaining a primary assay signal; performing reverse transcription and amplification on the reference lysate to detect nucleic acids associated with microbial cells detectable by the multiplexed identification test panel identification panel, thereby obtaining a reference assay signal; comparing the primary assay signal and the reference assay signal to obtain a measure of the effectiveness of the selected antibiotic against the microbial cells.