Until recently it was assumed that the healthy mammalian lung did not harbour a microbiota, unlike
other body sites. However, through the use of sequencing based technologies this has been shown to
not be the case. Low biomass communities of microbes can be identified in the healthy lung and the
lung microbiota in various diseases states has been shown to differ form these 'healthy' communities. The sheep respiratory microbiota is of interest from both an animal health perspective and due to the
potential use of the sheep as a large animal model for studying the lung microbiota. In this thesis I
seek to characterise the composition and variability of the sheep lung microbiota; the differences
between the sheep upper and lower respiratory tract bacterial communities and to assess whether
exhaled breath condensate collection can be used as a non-invasive lung microbiota sampling method.
To study the bacterial communities present in samples I have used 16S rRNA gene sequencing and
analysis. In Chapter 3 I examine the inter-individual and spatial variability present within the sheep lung
microbiota. Protected specimen brushings were collected from three lung segments in six animals at
three time-points. In a separate sheep a greater number of brushings was taken (n=16) in order to
examine the amount of variability over a smaller spatial scale. I find that there can be large differences
between the bacterial communities isolated from different locations within the lung, even over short
distances. Samples also cluster by the sheep from which they were taken, indicating a host specific
influence on the lung microbiota. In Chapter 4 I compare whole lung washes and oropharyngeal swabs
from 40 lambs in order to examine the differences between the upper and lower respiratory tract
microbiotas. I find that oropharyngeal swabs separate into rumen-like or upper respiratory tract-like
bacterial communities. Despite the fact that in humans the upper and lower respiratory microbiotas
have been shown to have similar compositions, the sheep lung microbiota samples in this study do not
resemble either oropharyngeal samples or reagent only controls. In my first two results chapters, lung sampling methods were used which involved either anaesthesia
combined with a bronchoscopic procedure (Chapter 3) or samples being taken from dead animals
(Chapter 4). In Chapter 5 I assess whether there is a less invasive way of taking lung microbiota
samples from a living individual, both to minimise the procedural stress on animals used as models
and to increase the pool of potential volunteers for human lung microbiota studies. I compared
samples taken via protected specimen brushings to samples taken via exhaled breath condensate
collection, a less invasive sampling technique. I find that condensate samples contain less bacterial
DNA and different bacteria than brushing samples, indicating that it is unlikely they could be used as
a replacement for invasive sampling methods. In my final results chapter I compare the results across
Chapters 3, 4 and 5 to identify bacteria which occur consistently in the sheep lung and could therefore
potentially be described as core lung microbiota members. In conclusion, while I have found that there are large differences between the sheep lung microbiota
and that which has previously been described in humans, the sheep can still be of use as a model in
studies where these differences would not have a significant impact, such as in Chapter 5 of this
thesis. I have identified several bacterial members of the core sheep lung microbiota which in future it
would be interesting to better characterise and to assess whether they play a role in sheep health
