Primary pneumonic vs cultured Y. pestis

Although pneumonic plague is the deadliest manifestation of disease caused by the bacterium Yersinia pestis, there is surprisingly little information on the cellular and molecular mechanisms responsible for Y. pestis-triggered pathology in the lung. Therefore, to understand the progression of this unique disease, we developed an intranasal mouse model of primary pneumonic plague. Mice succumbed to a purulent multifocal severe exudative bronchopneumonia that closely resembles the disease observed in humans. In order to assess the adaptation of Y. pestis to a mammalian environment, we employed DNA microarray technology to analyze the transcriptional responses of the bacteria during interaction with the mouse lung as compared to bacteria cultured in vitro. Keywords: Infectious expression analysis A Y. pestis genespecific microarray, consisting of 70-mer oligonucleotides representing 100% of the ORFs of strain CO92, was constructed. Two groups of 10 mice each were independently infected with Y. pestis strain CO92 as described above. After 48 hours, mice were sacrificed and RNA derived from infecting bacteria was extracted, amplified, and hybridized to the arrays. Each experiment consisted of a pair-wise competitive hybridization of amplified RNA (aRNA) samples from mouse and culture derived RNA and a reciprocal dye-flip replicate. Since 4 biological duplicates were tested, a total of eight DNA microarrays were used for comparison for each set of RNA amplifications. The scanner photomultiplier tube (PMT)values were set for optimal intensity with minimal background. An additional scan was done for each slide (low PMT) with the PMT such that <1% of the elements are saturated in order to characterize spots which were saturated at the higher PMT setting.