In Vivo Properties of Pseudomonas Aeruginosa: Outer Membrane Components and their Antigenicity

Abstract

This study involved an investigation of bacterial properties in vivo
with particular reference to the outer membrane (OM) antigens of such
bacteria and their recognition by host immunoglobulins during infection.
Pseudomonas aeruginosa was recovered directly, without subculture, from the
infected wounds of burn patients and the bacterial OM antigens analysed by
sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE). The
results indicated that the in vivo bacteria expressed high molecular weight
OM proteins not present in cells of the same isolate cultivated in complex
media, unless depleted of iron. Immunoblotting techniques further
demonstrated recognition of the iron-regulated membrane proteins (IRMPs) by
antibodies present in the patient serum and locally in wound tissue fluid,
suggesting their potential as vaccine candidates. Differences between
lipopolysaccharide (LPS) antigens from in vivo and in vitro grown
bacteria were additionally observed following analysis by SDS-PAGE.

Methods were devised to purify the IRMPs in a form relatively free
from LPS. These included detergent extraction to solubilize the proteins
and gel filtration procedures to separate the IRMPs from other OM proteins
and subsequently to remove LPS.

An investigation was made of the rabbit humoral immune response to P.
aeruginosa growing as an adherent microcolony on the surface of inert
materials (used in medical prostheses) and implanted into the peritoneum.
The resulting localized infection was chronic in nature and involved large
numbers of bacteria . Electron microscopy studies revealed that the
adherent cells were embedded in an extensive glycocalyx partly composed of
exopolysaccharide. Crossed immunoelectrophoresis demonstrated that in this
biofilm mode of growth the bacteria avoided evoking a strong immune
response, perhaps due to masking of key antigens by the glycocalyx. This
was in contrast to bacteria in an acute disseminated peritonitis model,
which elicited a high titre of antibodies, notably to the LPS antigen.
Moreover, antibodies to the IRMPs were produced at an early stage of the
disseminated infection. The results may help to explain the persistence of
infections associated with medical prostheses such as peritoneal dialysis
catheters.

Date of Award	1987
Original language	English