Characterisation of CD4+ T cells in healthy and diseased koalas (Phascolarctos cinereus) using cell-type-specific monoclonal antibodies

The koala (Phascolarctos cinereus) is an arboreal herbivorous marsupial that is an Australian icon. Chlamydia pecorum and C. pneumoniae are obligate intracellular Gram-negative bacteria that infect koalas and can cause, infertility, arthritis, conjunctivitis, respiratory disease, and in some cases death. Due to a lack of koala-specific immune reagents and assays there is currently no way to adequately analyse the immune response in diseased, healthy or vaccinated animals. Using the extracellular domain of koala CD4+ this paper reports the production and characterisation of the first anti-koala CD4 monoclonal antibody (mAb). Fluorochrome-conjugated anti-CD4 mAb was used to investigate the levels of CD4+ T helper cells collected from koala spleens, lymph nodes and peripheral blood by flow cytometry. Biotin-conjugated anti-CD4 mAb was also used for Western Blot and immunohistochemistry to identify CD4+ T helper cells in the lymph node. The results indicate that CD4+ specific T cells could be identified using flow cytometry of cells from both tissues and peripheral blood as well as visualised using immunohistochemistry techniques. Further analysis of the results also showed an increased level of CD4+ cells in animals with current signs of chlamydial infection/disease as opposed to those showing no signs of infection or clinical disease. Since CD4+ T cells have been shown to play a pivotal role in clearing chlamydial infection in both human and mouse infections, using this novel antibody will help determine the role CD4+ T cells play in protection against chlamydial infection in koalas