Attachment and ingestion of gonococci human neutrophils

Abstract

Previous studies have indirectly shown that type 1 gonococci are more resistant to phagocytosis by human neutrophils (PMN) than type 3 gonococci. Using phase contrast, fluorescent, and light microscopy, we directly quantitated PMN-gonococcal interaction, with emphasis on separating ingestion from attachment. PMN monolayers were incubated on slides with type 1 or type 3 gonococci. The slides were iced to stop phagocytosis and stained with antigonococcal fluorescent antibody (FA). After methanol fixation, the FA-stained gonococci associated with PMN were counted. Since the live PMN excludes FA, the FA-stained gonococci represent only extracellular gonococci. Methylene blue was then added to the same slide to stain both ingested and surface attached gonococci. Using these methods, intracellular and extracellular cell-associated gonococci were quantitated under varying conditions. The numbers of methylene blue-stained cell-associated gonococci were similar for types 1 and 3. The proportions of cell-associated gonococci that were ingested were: with normal serum, 3.7 4.1% for type 1 and 56.2 i 3.5% for type 3 (P < 0.001); with heat-inactivated serum, 1.0 + 3.0% for type 1 and 52.6 3.7% for type 3 (P < 0.001); with high-titer anti-gonococcal antibody serum, 4.8 + 4.3% for type 1 and 64.0 1.6% for type 3 (P < 0.001). Thus, most type 3 organisms were ingested, but most type 1 gonococci were bound on the PMN surface.

Infection of HeLa cells with Salmonella typhimurium 395 MS and MR10 bacteria

“…No simple method exists for the distinction between extra-and intracellular bacteria. Antibodies do not penetrate the cell membrane of intact mammalian cells (3,29), whereas pretreatment with acetone, for example, allows penetration and binding to intracellularly localized bacteria (20). By using the indirect immunofluorescence technique, this effect can be used to estimate the proportion of intracellular bacteria.…”

“…Fixation was used because it is considered to facilitate the passage of antibodies through the cell membrane (20) and consequently allows the quantitation of the total number of HeLa cell-associated bacteria. With unfixed HeLa cells only the adhering extracellular bacteria are observed because of the inability ofthe antibodies to penetrate the cell membrane (3,29). The following controls were included: (i) noninfected cells plus specific antibody plus FITC-labeled antiglobulin; (ii) infected cells plus normal rabbit serum plus FITC-labeled antiglobulin; and (iii) infected and noninfected cells plus FITC-labeled antiglobulin (20).…”

“…In these experiments, however, it was not demonstrated whether the associated bacteria gained intracellular access. A variety of techniques have been devised to assay the association ofbacteria with mammalian cells (26), but few differentiate between attachment and intemalization (1, 3,8). The present work employs two different techniques with the aim of distinguishing quantitatively the attached from the internalized bacteria when S. typhimurium 395 MS and MR10 infect HeLa cells.…”

Pili of Neisseria meningitidis. Analysis of structure and investigation of structural and antigenic relationships to gonococcal pili.

“…In vitro studies indicate that pili facilitate the attachment of meningococci and gonococci to human cells and to human mucosa in organ culture (6,7). Piliation is also correlated with resistance of gonococci to phagocytosis by human polymorphonuclear leukocytes (8). Antibodies to pili are detected after fulminant meningococcal disease (9) and in sera and mucosal secretions after natural gonococcal infection (10).…”

Mannose-inhibitable adhesins and T3-T7 receptors of Klebsiella pneumoniae inhibit phagocytosis and intracellular killing by human polymorphonuclear leukocytes

“…Possible interference of adhesins with host defenses has been studied in some cases (4,10,23,28,31,37,38). So far, a role of adhesins in the protection of adhesive organisms against phagocytosis has been discussed only for gonococci (10,18,28,31,32,41,42) and has been questioned by others (39). On the contrary, several reports have appeared showing that pili and other adhesins facilitate adherence to phagocytic cells and the subsequent phagocytosis of the microorganisms (4,37,38).…”