Effects of in vitro and in vivo growth conditions on expression of type 8 capsular polysaccharide by Staphylococcus aureus

Abstract

Type 8 capsular polysaccharide (CP8) is widely prevalent among clinical isolates of Staphylococcus aureus, but the role that the capsule plays in the pathogenesis of staphylococcal infections is unclear. This study was performed to identify growth conditions that would optimize the production of CP8 and to determine whether enhanced CP8 expression would influence staphylococcal virulence. S. aureus Becker grown in a chemically defined broth medium with < 1 microM ferric nitrate produced up to eightfold more CP8 per milligram of biomass than did bacteria cultivated in the same medium containing 20 microM ferric nitrate. The bacteria produced > 350-fold more cell-associated CP8 per milligram of biomass when grown on the surface of Columbia agar than when grown in Columbia broth. Most of the CP8 produced by broth-grown cells was secreted into the culture medium. S. aureus cultivated on the surface of nitrocellulose membranes floating on Columbia broth produced levels of CP8 similar to those produced by cells grown on Columbia agar. Similarly, bacteria harvested from endocardial vegetations of rabbits infected with S. aureus produced high levels of CP8. These results indicate that staphylococci grown on surfaces, both in vitro and in vivo, produce larger quantities of cell-associated CP8 than those grown in liquid cultures. However, no differences were observed in the 50% lethal dose for mice of strain Becker grown on solid medium (high levels of capsule expression) or in liquid medium (low levels of capsule expression).

Encapsulated Staphylococcus aureus strains vary in adhesiveness assessed by atomic force microscopy

“…Several studies have demonstrated the capability to detect variations in the surface morphology of bacteria, using AFM. 18,[26][27][28] The effect of growth conditions on CP expression has been investigated 20,21 but not exhaustively. However, the presence of intercellular material and surface protrusions for serotype 2 and 8 suggest that the culturing method used in this investigation does not inhibit the expression of CP.…”

“…Bacteria were cultured on Columbia blood agar plates for 24 h at 378C and subsequently refrigerated for 24 h to ensure that the bacteria were in the stationary growth phase, a period of maximum CP production. [19][20][21] Preparation for atomic force microscopy One to three colonies were transferred to 100 lL of deionized water and gently pipetted to suspend the colonies. S. aureus were immobilized by transferring 20 lL of the suspension to a freshly cleaved piece of muscovite mica, and only preparations in which the drop spread uniformly were used for experimentation.…”

Identification of a gene essential for O‐acetylation of the Staphylococcus aureus type 5 capsular polysaccharide

“…Capsule analysis and quantitation CP5 was purified as described by Fournier et al (1987), except that teichoic acid was eliminated by oxidation with sodium metaperiodate (Lee et al, 1987a). Capsular extracts were prepared as described previously (Lee et al, 1993) from S. aureus cultivated on Columbia agar plates (Difco Laboratories) supplemented with 2% NaCl. Capsule expression was evaluated by immunoprecipitation of bacterial extracts with CP5-specific monoclonal or polyclonal antibodies.…”

“…An ELISA inhibition method was used to quantitate the relative degree of CP5 acetylation by strain Reynolds, mutant JL232 and JL387(pJCL85). This assay, described previously (Albus et al, 1991;Lee et al, 1993), is based on the ability of native CP5 or whole bacteria (trypsinized to remove protein A) to adsorb mAbs that specifically recognize O-acetylated CP5.…”

Effect of pharynx epithelial cells surface desialylation on receptor-mediated adherence ofStaphylococcus aureus

“…Staphylococcus aureus produces a wide variety of molecules that influence its interaction with the human host. It expresses adherence factors such as teichoic acid (Baddiley et al 1962), capsule (Lee et al 1993), protein A (Van Belkum et al 1997), receptors for fibronectin and fibrinogen, collagen-binding proteins and an extracellular adherence protein (eap) (Palma et al 1999), all of which regulate its adherence to host cells. Bacterial attachment, the first step in this complex interaction, may dictate the tropism of the bacteria to distinct host tissues.…”