FIG. 7.

Infection of cultured keratinocytes with C. pneumoniae TW-183. Normal human neonatal keratinocytes were infected with 5,000 IFU₅₀ and cultured for 3 days in T-25 flasks. RNA was extracted followed by DNase treatment, and then cDNA was prepared with 1 to 2 μg of RNA by using MMLV reverse transcriptase. cDNA was subjected to PCR with primers annealing with DNA of the 16S rRNA gene of C. pneumoniae or the ompA gene (ompA primer set 1; see Materials and Methods). (A) Lane 1, DNA ladder; lane 2, cDNA from uninfected cells; lane 3, PCR product for 16S rRNA gene from infected cells; lane 4, DNA from TW-183 (positive control). (B) Lane 1, DNA ladder; lane 2, cDNA from the uninfected culture; lane 3, PCR product for the ompA gene from the infected culture (light 340-bp band); lane 4, PCR product for ompA gene from TW-183. Keratinocytes in chamber slides were infected simultaneously with the T-25 flasks with an equal concentration of C. pneumoniae (200 IFU₅₀ per cm²) (C and D), and uninfected cultures were established (E and F). After 3 days, cells were washed and fixed with STF and then reacted with anti-LPS (C and E) or anti-OMP (D and F). Bars = 50 μm (C to F).