Chlamydia abortus is an obligate intracellular bacterial pathogen and is one of
the most common causes of infectious abortion in sheep world-wide. The
organism is responsible for the disease known as ovine enzootic abortion
(OEA), which manifests in late pregnancy. Development of a safe and effective
vaccine deployed alongside existing molecular diagnostic tests remains a
long-term ambition. This approach is dependent on improving our knowledge
of disease pathogenesis that leads to tissue damage (pathogen and immune
cell-mediated) in the sheep placenta. Immune responses that control disease
and those responses that facilitate pregnancy are not always compatible or
completely understood. Interferon-gamma (IFN-ɣ), the signature cytokine of
T-helper (Th)-1 cells, is known to control C. abortus growth. Little is known
about other cytokines co-expressed during OEA that might contribute to
control or disease progression. Incidentally, knowledge of the existence of
other cluster of differentiation (CD)4+ve T cells that produce interleukin-(IL)-4,
IL-10 or IL-17A in sheep are limited or non-existent. These capability gaps
need to be addressed first, to enable measurement of the quality of immune
responses during pregnancy or OEA.
This thesis addressed the hypothesis:
Systemic immune responses of sheep uniquely correlate with pregnancy
outcome following OEA.
Three major research aims were undertaken:
1. To expand the range of assays to characterise CD4+ve T cell immune
responses in sheep.
2. To develop and apply tools to investigate local immune regulation in the
ovine placenta.
3. To investigate immunological correlates of protection against OEA.
Specific immunological assays developed in this project enabled single cell
identification, multi-parameter cytokine assessment and quantification of
specific immunomodulatory molecule transcripts. Studies revealed that IFN-ɣ
and IL-10 were initiated upon antigen or mitogen stimulation and their kinetics
are as reported for other mammals. Presumptive T-regulatory cells were
identified in peripheral blood mononuclear cells (PBMC). IFN-ɣ and
IL-17A-expressing cells include Th-1 and Th-17 cells which are differentially
represented in mitogen-activated PBMC.
Analysis of placental immunity involved collection of placental tissues and
associated lymph nodes at full term of pregnancy. Selective predominance of
classical major histocompatibility complex Class I transcripts were identified
but no indoleamine-2, 3 dioxygenase-1 transcripts within the placental tissues
were found and no natural killer cell staining observed. Collectively, this
expression pattern of ‘immunological parameters’ are unique to date to
placental mammals.
Refined and new assays were fundamental to the experimental design to
investigate immune responses to OEA. New patterns of Chlamydia-specific
responses were identified. Selective elevation of IFN-ɣ and IL-10 during the
latent stage of disease prior to pregnancy was observed. These cytokines
remained elevated during and after active phases of OEA. Measurement of
targeted immunological parameters failed to identify unique responses
correlated with protection and do therefore not support the PhD hypothesis.
This work has significantly extended the knowledge of placental immunity and
cellular immunity during OEA utilizing species-validated immunological
assays. Several new areas of investigation have been identified. Concurrent
progress in antigen discovery, transcriptomic analysis and tissue organoid
cultures have now provided an environment to accelerate innovative research
for reproductive disease vaccinology. Defined protective immune signatures
will inform on the design of new safe vaccines to OEA, by refinement of
protective antigen selection and mechanisms of vaccine delivery. Placental
tissue-associated organoids may facilitate antigen discovery for other sheep
reproductive pathogens. Together, these proposed areas of further
investigation may enable development of novel multi-pathogen vaccines to
prevent reproductive diseases of small ruminants
