P1–165: Isolates of <i>Chlamydia pneumoniae</i> from the Alzheimer's brain infecting cultured human astrocytes and microglia display an active, not a persistent, growth phenotype

The intracellular bacterial pathogen C pneumoniae can cause persistent infections, during which its pathogenic characteristics differ from those of active infection. This organism was identified in astrocytes and microglia in the late–onset Alzheimer's disease (AD) brain. We cultured this organism from AD brain samples from two North American locations. Because the mechanisms of pathogenesis differ between active and persistent infection, we asked whether infection of the two common human CNS host cell types with the brain isolates displays an active or persistent phenotype. The human astrocytoma and microglioma cell lines U–87 MG and CHME–5 (respectively), and the human epithelial line HEp–2, were infected at MOI 1:1 with C pneumoniae index strain AR–39 as control, and with the Tor–1 (from a Toronto sample) and the Phi–1 (from a Philadelphia sample) AD brain isolates. Cultures were harvested at 24, 48, and 72 hr post–infection and subjected to analyses of inclusion number, size, morphology, and recovery of new organisms. DNA/RNA were prepared from infected cultures and analyzed for chromosome accumulation and presence/absence of specific bacterial mRNAs. Glial cells infected with C pneumoniae of the control strain and with both brain–derived isolates displayed inclusions indistinguishable from those characteristic of active infection of standard HEp–2 host cells. Real time PCR showed that accumulation of chlamydial DNA over time during infection of the two glial cell lines was similar to that in actively–infected HEp–2 cells. RT–PCR showed that mRNA from ftsK, pyk, and other chlamydial genes whose expression is attenuated during persistent infection were easily identifiable in Tor–1 and Phi–1 infected CHME–5 and U–87 MG cells. Recovery of new organisms at the termination of the chlamydial developmental cycle was equivalent for the AR–39 control–infected U87 and CHME–5 cells, and for those cells infected with the Tor–1 and Phi–1 brain isolates. In cultured human astrocytes and microglia, C pneumoniae isolated from the AD brain displays an active, rather than a persistent, growth phenotype. This observation indicates normal passage through the developmental cycle, demonstrating the destruction by lysis of some portion of host cells at the termination of that cycle in the CNS.

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