Rapid Damage to Membranes of Neisseria gonorrhoeae Caused by Human Neutrophil Granule Extracts

Abstract

Neisseria gonorrhoeae were exposed to extracts of human neutrophil granules and effects on gonococcal growth and membranes were determined. Enumeration of gonococci by phasecontrast microscopy at 0 and 60 min revealed that they underwent very limited cell division after exposure to granule extract. At 60 min, treated gonococci tended to clump, and some lost their refractivity under phase-contrast optics, indicating membrane damage. Treated and untreated gonococci utilized oxygen at similar rates at time 0; treated gonococci utilized oxygen at a relatively constant rate for 60 min, even though colony-forming ability (i.e. viability) decreased by 90 %, whereas untreated gonococci showed a steadily increasing rate of oxygen consumption over the same period, which essentially paralleled increase in colony-forming ability. Membrane ultrastructure of untreated and treated gonococci was compared in thin section by transmission electron microscopy. Extract treatment resulted in a time-related increase in disruption of the bacterial outer membrane, which became apparent almost immediately after treatment. This was accompanied by increasingly aberrant septum structure. Extract treatment also increased the resolution of peptidoglycan by electron microscopy, as early as 10 min after treatment. These data suggest that extract treatment of gonococci caused a rapid loss of the ability to form colonies on agar concomitant with alteration of gonococcal peptidoglycan and outer-membrane structure, but with little alteration of inner-membrane function.

Interactions of Neisseria gonorrhoeae with human neutrophils

“…Addition of sodium nitrite, however, permitted growth in anaerobic broth and resulted in a level of granule protein susceptibility similar to that in aerobically grown gonococci. This is consistent with the earlier work of Rest and coworkers (14,81,88,89), who showed that granule proteins kill actively growing gonococci better than they kill stationary-phase gonococci.…”

“…Nevertheless, some progress has been made in determining the mechanism by which gonococci succumb to antimicrobial proteins. Gonococci incubated in the presence of lethal amounts of unfractionated granule extracts continue protein and nucleic acid synthesis (14) and retain respiratory activity, but are impaired in PG synthesis (88) and show a limited capacity to divide (89). This was evidenced by a reduced incorporation of radiolabeled glucosamine into PG during incubation of gonococci with granule extracts.…”

A Thermonuclease ofNeisseria gonorrhoeaeEnhances Bacterial Escape From Killing by Neutrophil Extracellular Traps

“…We hypothesize that NETs kill N. gonorrhoeae by placing the bacteria in proximity to NET-associated cationic antimicrobial proteins. These components include LL-37 and cathepsin G, which have antigonococcal activity in vitro [32][33][34][35] and, for cathepsin G, in neutrophil phagolysosomes [13].…”

Resistance of Neisseria Gonorrhoeae to Neutrophils

“…Seminal research from the Rest and Shafer laboratories indicated that components found inside PMN granules display oxygen-independent antigonococcal activity (Rest, 1979; Casey et al, 1985; Rock and Rest, 1988). These components include the bactericidal/permeability-increasing protein (“hCAP57”), cathepsin G protease, and LL-37 antimicrobial peptide (Casey et al, 1985; Shafer et al, 1986, 1998).…”