Phenotypic modulation of gonococcal lipooligosaccharide in acidic and alkaline culture

Abstract

Neisseria gonorrhoeae infects a diverse array of niches in its human host, which expose the organism to dramatic variations in pH. We examined growth and lipooligosaccharide expression of two gonococcal strains in liquid and solid cultures under acidic, neutral, and alkaline conditions. Growth rates in broth were similar under the three conditions, and the pH remained fairly constant throughout the growth cycle. Altered lipooligosaccharide expression at the different pHs was noted in both plate-and broth-grown organisms. Neisseria gonorrhoeae will colonize almost any mucosal surface in its human host, including the urethra, vagina, rectum, conjunctiva, and pharynx (1). In addition, viable organisms are often located within neutrophils and epithelial cells (1, 3). These diverse environments expose the gonococcus to dramatic variations in oxygen tension, available nutrients, reactive oxygen intermediates, and pH. A number of studies have been published concerning the response of N. gonorrhoeae to thermal stress (14), nutrient stress (9), and anaerobic stress (2). A particularly relevant environmental stress for N. gonorrhoeae is pH fluctuation, because this organism can be cultured from host tissues exhibiting remarkable differences in pH. For example, the average vaginal pH in infected and uninfected women is 5.5 (range, 4.1 to 7.4), and the average pH of male urethral exudates is 7.3 (range, 6.2 to 8.4) (5). We investigated growth and lipooligosaccharide (LOS) expression of two gonococcal strains in cultures ranging from pH 5.8 to 8.2. Because liquid and solid cultures likely reflect different in vivo conditions, growth and LOS expression were evaluated with both types of culture. Strains FA19 (serum resistant, protein 1A bearing [11, 12]) and F62 (serum sensitive, protein 1B bearing [11, 12]) were maintained on gonococcal typing agar (pH 7.0). Nonpiliated, non-Opa-bearing organisms were selected (13). Medium buffer was modified for pH experiments with a final concentration of 30 mM MES [2-(N-morpholino)ethanesulfonic acid; pK a , 6.1; useful pH range, 5.5 to 6.7 (SigmaUltra)] for pH 5.8 experiments and 30 mM HEPES (N-2-hydroxyethylpiperazine-NЈ-2-ethanesulfonic acid; pK a , 7.5; useful pH range, 6.8 to 8.2 [SigmaUltra]) for pH 7.2 and 8.2 experiments. Final pH adjustment was made by addition of NaOH to media at 37ЊC. The pH of all preparations was verified after sterilization. pH 7.0-grown organisms were used to inoculate pH 5.8, 7.2, and 8.2 broth cultures, and growth curves of shaken cultures were obtained. Cell densities over time under the three conditions were remarkably similar (Fig. 1). In contrast, Morse and Hebeler (8) found the generation time of gonococcal strain CS-7 at pH 6.0 was almost twice that of pH 7.5-grown CS-7. One obvious difference in their methods was the use of HEPES for broth cultures prepared at pHs 6 through 8. pH was monitored during growth in broth (Table 1). Under

Acid‐induced expression of an LPS‐associated gene in Helicobacter pylori

“…However, environmental regulation of LPS expression among Neisseria species occurs; differences in growth medium, glucose concentration and oxygen availability each influence the LPS phenotype in both gonococci and meningococci (Tsai and Civin, 1991; Van Putten and Robertson, 1995). Gonococcal cells grown at pH 5.8 produce an additional lipooligosaccharide band compared with cells grown at pH 8.2, indicating pH regulation of LPS profiles (Pettit et al ., 1995). In Rhizobium leguminosarum , LPS also is modified in response to several environmental stresses, including low pH (Tao et al ., 1992).…”

Regulation of gonococcal sialyltransferase, lipooligosaccharide, and serum resistance by glucose, pyruvate, and lactate

“…Alteration of LOS structure by growing GC in vitro at different pHs (43) and by using glucose-limiting conditions in continuous culture (33) have been reported. Thus, the expression of LOS appears to be altered by numerous conditions (anaerobiosis, limiting or nonlimiting carbon source, pH).…”

Alteration of gonococcal protein expression in acidic culture

“…These results are extremely relevant given that host cell attachment requires pili (22) and given the immune protective nature of sialylated gonococcal lipooligosaccharide (17). We recently described altered lipooligosaccharide expression in two gonococcal strains in acidic and alkaline culture (19).…”

“…Nonpiliated, non-Opa-bearing organisms were selected (23). Medium buffer was modified for pH experiments as we have previously described (19). The pHs of all preparations were verified after sterilization.…”