Interaction of pneumolysin-sufficient and -deficient isogenic variants of Streptococcus pneumoniae with human respiratory mucosa

Abstract

Streptococcus pneumoniae is the most common cause of community-acquired pneumonia, and pneumolysin, a hemolytic toxin, is thought to be an important virulence factor. We have studied the interaction of a pneumolysin-sufficient type II S. pneumoniae strain (PL؉) and an otherwise identical pneumolysin-deficient derivative (PL؊) with human respiratory mucosa in an organ culture with an air interface for up to 48 h. Ciliary beat frequency (CBF) was measured by a photometric technique, and adherence to and invasion of the epithelium were assessed by scanning and transmission electron microscopy. PL؉ and PL؊ caused a progressive fall in CBF compared with the control which became significant (P < 0.01) at 24 h for PL؉ and at 48 h for PL؊. At 24 h, there was a significant increase in the percentage of the mucosa of the organ culture that was damaged for PL؉ compared with the control (P < 0.01) and PL؊ (P < 0.02). At 48 h, there was a significant increase in mucosal damage for both PL؉ (P < 0.005) and PL؊ (P < 0.05) compared with the control. At 24 and 48 h, PL؉ and PL؊ adhered predominantly to mucus and damaged cells. PL؉ infection alone caused separation of tight junctions between epithelial cells, and at 48 h PL؉ cells were adherent to the separated edges of otherwise healthy unciliated cells. PL؉ and PL؊ both caused damage to the epithelial cell ultrastructure. S. pneumoniae infection caused patchy damage to the respiratory mucosa and a lowered CBF. These changes were more severe and occurred earlier with the pneumolysin-sufficient variant.

Use of green fluorescent protein in visualisation of pneumococcal invasion of broncho-epithelial cells in vivo

“…These observations are in keeping with previous ¢ndings [11], but some di¡erences were observed. S. pneumoniae were shown to cause the separation of tight junctions in an organ culture of human adenoid ciliated and unciliated epithelium [13]. In this study we did not ¢nd pGFP1 pneumococci within cell junctions in vivo, indicating di¡erences between the in vivo and in vitro culture systems.…”

The role of streptococcal toxins in disease

“…If the pneumolysin-negative mutant is co-infected with the wild type, the mutant exhibits wild type growth kinetics in the blood suggesting that the toxin can exert its effect at a distance (Benton et al 1995). Use of the pneumolysin-negative mutant in an organ culture system with human respiratory epithelium showed that the toxin mediated loosening of the tight junctions between epithelial cells (Rayner et al 1995). The wild-type organism then adhered preferentially to the newly exposed areas.…”

Disruption of nasopharyngeal epithelium by pneumococci is density‐linked

“…The ability of pneumococci to cause a separation of tight junctions in in vitro organ cultures of adenoid tissue was shown by Rayner et al . [3] using scanning electron microscopy.…”