Development of an intraperitoneal implant chamber for the study of in vivo-grown Pasteurella haemolytica in cattle

Abstract

An intraperitoneal implant chamber was developed for the study of the in vivo growth of Pasteurella haemolytica in calves. The chamber had a volume of approximately 100 ml and featured an external sampling port which allowed multiple and sequential sampling of the chamber contents. A single polycarbonate diffusion membrane with a pore size of 0.22 μm allowed host peritoneal fluid to diffuse into the chamber and maintained the bacterial population free of white blood cells. Chambers were implanted into the peritoneal cavities of four five-month-old dairy-cross calves, demonstrated to be sero-negative by indirect haemagglutination assay. Three days later, four different P. haemolytica isolates, of serotypes A1 or A2, were inoculated into the chambers. In all cases, there was a slow decline in the viable bacterial numbers within the chambers. Western blot analysis of the antibody content of the chamber fluids revealed IgG antibodies to P. haemolytica OMPs in the fluid prior to inoculation and both 9 and 15 days after inoculation. Furthermore, there was no significant change in the IgG antibody content of the chamber fluid, either quantitatively or qualitatively, during the course of the experiment. Analysis of the bactericidal activity of pre-inoculation chamber fluid against the corresponding bacterial isolate suggested that an antibody-dependent complement-mediated process was not responsible for the decline in bacterial numbers. Overall, the chamber design was demonstrated to be extremely effective for in vivo studies of P. haemolytica in calves, allowing easy and regular sampling of the chamber contents and maintaining bacteria free of white blood cells. Although there was a slow decline in bacterial numbers over time, sufficient numbers of cells could be obtained for analysis of cell-surface antigens.