3.3 R27 exhibits a broad potential host range, but a low host
divergence and is less similar to the sewage community than pB10.
In order to investigate the phylogenetic host range of the two plasmids,
we conducted 16S rRNA gene profiling of the transconjugants originating
from conjugation assay challenging Ellinge sewage community with
plasmids R27 (sewage community/R27) and pB10 (sewage community/pB10).
Subsequently, we profiled the catalogue of recipients in the sewage
community to examine how widely the plasmids transfer within this pool
of recipients.
The plasmid transfer was initially evaluated in the two transconjugant
sewage communities by alpha diversity analyses: Faith’s phylogenetic
diversity (PD) (Faith, 1992)
measures the total phylogenetic distance among the hosts, which we
interpret as a metric of the distribution breath describing a plasmids
potential host range. Mean pairwise distance
(Webb et al., 2002)
describes the mean phylogenetic distance between hosts, which we
interpret as average pairwise host divergence
(Tucker et al., 2017). PD
was similar for R27 and pB10 in this sewage community (figure 3a),
revealing a broad potential host range for both plasmids. We observed a
significantly higher mean pairwise distance of transconjugants for
sewage community/pB10 compared to sewage community/R27
(P<0.05) (figure 3b). Thus, R27 has a broad potential host
range but a low host divergence, whereas pB10 has a broad potential host
range and a high host divergence.
Next, we compared the composition of the two transconjugant communities
to the entire recipient sewage community to investigate beta diversity.
The diversities between the sample groups: recipient sewage community,
sewage community/pB10, and sewage community/R27, were calculated using
Weighted Unifrac. Ordination by Principal Coordinates Analysis (PCoA)
with Weighted Unifrac distances between samples revealed that samples
cluster neatly according to the sample group, highlighting a strong
biological underpinning for the observed trends. Furthermore, axis 1 and
2 comprise/explain a total of 93.8 % of the variance (figure 3c). This
decisively shows less distance in-between replicates than between sample
groups and suggests that plasmid-specific host range drives the
divergence of transconjugant communities. Pairwise weighted Unifrac
distances between sample groups revealed a higher distance between the
recipient sewage community and sewage community/R27 than between the
recipient sewage community and sewage community/pB10 (figure 3d).
Additional PERMANOVA analyses of the weighted Unifrac distances found
higher R2 for sewage community/R27
(R2=0.71, p < 0.001) than for sewage
community/pB10 (R2= 0.49, p < 0.001) (figure
3d). These results imply that the transfer of R27 is a stronger driver
of phylogenetic distance between the resulting transconjugant sewage
community and the recipient sewage community than the transfer of pB10.