Selective breeding produces increased occurrence of alleles with
intermediate frequencies on which natural selection can act
Selective breeding is one genetic intervention strategy that may quickly
increase adaptive genetic variation in corals, as such facilitating
adaptation to increasing sea surface temperatures (Chan, Hoffmann, &
van Oppen, 2019; van Oppen et al., 2015). This approach has been used
across a range of aquaculture and mariculture species to improve a
number of commercially important traits. Commercially important species
like oysters and mussels share similar life-history attributes with
corals, including the production of numerous, potentially low-quality
gametes, and high levels of genetic diversity (e.g. R-strategist)
(Ellegren & Galtier, 2016). In those systems, only a few generations of
selective breeding has resulted in 30% higher growth under elevated
pCO2 conditions compared to wild populations (Parker et
al., 2012) and 50% higher growth in redclaw crayfish (Stevenson, Jerry,
& Owens, 2013) without significantly eroding genetic diversity compared
to wild populations after pooling breeding combinations (O’Connor, Dove,
& Knibb, 2016). Selective
breeding over only one generation in corals has shown that significant
increases in heat tolerance of larvae (Dixon et al., 2015) and juveniles
(Quigley et al., 2020) are possible using these methods. Lessons learnt
here may therefore provide insights into the mechanisms underpinning the
success of these techniques in a wild marine context.
We found that alleles at intermediate frequencies were at greater
abundances than expected under HWE in WC and CW crosses relative to the
on average distribution of loci in WW crosses, suggesting an overall
increase in genetic diversity resulting from selective breeding of
corals from different regions of the GBR. Distributions of allele
frequencies typically follow that most loci nearly reach fixation at
either 0 or 1, where alleles at intermediate frequencies are less common
but are important given an increased abundance of intermediate alleles
are the raw material for selection (Jombart & Collins, 2015). Hence,
genetic material on which selective adaptive processes can operate
likely exists at greater abundances in the WC and CW relative to the WW
crosses. Hybridisation may lead to increased performance/fitness
relative to the parental generation, with this increased performance
having been linked to dominant, regulatory, single quantitative trait
nucleotides (Jakobson & Jarosz, 2019). Increases in genetic diversity,
hybrid vigour and genetic rescue
are well-known but inconsistent features of intra- and inter-specific
hybridization (Chan, Hoffmann & van Oppen, 2019; Flowers et al., 2019;
Hazzouri et al., 2019; Weeks et al., 2011) but has not yet been
demonstrated in the selective breeding of corals. For example, it was
unknown whether the breeding of divergent populations would cause a
decrease in genetic diversity due to processes like genome
incompatibility (Hogenboom, 1975). These results support that even in a
small number of crosses and contributing parents, genetic diversity
improves. Observed heterozygosity and alpha Diversity
(0Dα and2Dα at q = 0 and 2) was highest in the
interpopulation crosses (WC, CW). The0Dα metric is sensitive to the
presence of rare alleles, which is important given breeding across
populations may initially introduce novel mutants (Sherwin et al.,
2017), suggesting that WC, CW, and WW2 had an increased occurrence of
rare variants. 1Hα, also known as the
Shannon Information criteria, is informative as a natural measure of
evolvability, and was highest in WC and WW2. Finally, preliminary
analysis of adult colonies shows Tijou corals had on average higher
observed heterozygosity compared to Backnumbers corals
(Ho = 0.065 vs. 0.0217, unpublished data), suggesting
that there could have been a risk of reduced diversity in offspring.
Importantly, we did not observe a loss of genetic diversity by crossing
these two divergent populations.
Finally, we found that interpopulation selective breeding significantly
changed the resulting allele frequencies of offspring compared to
modelled HWE distributions. This is perhaps unsurprising given
assumptions of random mating, no gene flow, and infinite population
sizes were not met. However, this suggests the influence of breeding on
the genetic architecture of the resulting F1 generation.
Therefore, the interbreeding of
even a small number of corals from different reefs across the GBR may
result in extensive introgression and therefore accelerate the potential
for adaptation to warming, although the production of F2 generations
would be needed to confirm this.
We also demonstrate that variants associated with immune responses,
growth and cellular operating may be re-arranged during breeding but are
maintained within the next generation. This suggests that the important
functional diversity (i.e., at stress tolerance genes) associated with
focal populations can be maintained in the breeding process. In response
to heat stress, corals alter their gene and protein expression patterns,
as reflected in changes in their structural lipids, metabolism, and
immune responses (Barshis et al., 2013; Sogin et al., 2016). Differences
in proteins associated with collagen production and sodium bicarbonate
transport were important in differentiating the five families produced
in our study. Collagen is important for the production of the
extracellular matrix, required for multicellularity and the spatial
organization of functional units of cells (Helman et al., 2008) whereas
bicarbonate transporters are pivotal for coral calcification and hence
growth (Zoccola et al., 2015). Basic cellular functioning also
potentially varied through the differences in dTDP-glucose
4,6-dehydratase-like proteins detected and their involvement in the
non-oxidative pentose phosphate pathway (Buerger, Wood-Charlson,
Weynberg, Willis, & van Oppen, 2016; Yuyama, Watanabe, & Takei, 2011),
critical for glucose utilization. Differences between crosses in these
foundational processes like cellular organization and biomineralization
therefore suggests that even breeding across relatively few individuals
has the potential to substantially create distinct genetic combinations.
Protein NLRC3-like, which has been previously implicated in acroporid
immune suppression in response to heat stress by acting in Toll-like
receptor modification (Zhou et al., 2017), also varied significantly
across families. Other proteins involved in immunity and stress were
also detected (lysosomal-trafficking regulator-like proteins), and these
have also been linked to the mounting of innate immune responses through
Toll-like receptor activity in mice (Westphal et al., 2017). Additional
immunity related proteins were detected, including CEPU-1-like protein
(Spaltmann & Brummendorf, 1996), E3 ubiquitin-protein ligase
RNF213-like (Iguchi et al., 2019), NACHT (Hamada et al., 2013), and
spondin (Palmer & Traylor-Knowles, 2012). Interestingly, NFX1-type zinc
finger-containing protein was found here and downregulated in resistant
corals exposed to disease (Polato et al., 2010). Expression of nascent
polypeptide-associated complex subunit alpha protein (Bellantuono,
Hoegh-Guldberg, & Rodriguez-Lanetty, 2012) was similarly downregulated
in resistant corals, suggesting that these proteins provide an important
role in protective immune responses.
Genotyping individual aposymbiotic
larva from the five families reared under ambient conditions (27.5°C)
provides foundational knowledge as to how selective breeding influences
underlying genetic architecture. It also sheds light on the underlying
molecular origins and mechanisms of heritability, a long-standing goal
of quantitative genetics (Jakobson & Jarosz, 2019). Broad and
narrow-sense heritability has been quantified for a range of traits, but
the underlying mechanisms have rarely been described (Dixon et al.,
2015; Dziedzic, Elder, & Meyer, 2017). The majority of alleles fixed at
the extremes of allele frequency distributions (“U-shaped”sensu Hill, Goddard, & Visscher, 2008) are likely driven by the
small number of parents from which the larvae in each cross were derived
(5 unique parental colonies used across the families, Supplementary
Table 1), but may also suggest selection against heterozygotes during
the aquarium rearing period. Interpopulation hybrids displayed greater
genetic diversity relative to within population purebreds, which may be
the result of the varying effects of selection on purebreds versus
hybrids in the aquarium environment. U-shaped distributions may arise
under strong cases of artificial selection (e.g. aquariums) combined
with rare mutations (Hill et al., 2008).
Functional variation
associated with selectively bred families
How does genomic variation lead to phenotypic differences between
corals? The location and effect size of the SNP difference are important
to determining its eventual phenotypic effect, and simplistically,
differences in coding vs. non-coding regions are predicted to cause
phenotypic effects through a variety of mechanisms (Cavallo & Martin,
2005). Assigning function to SNP differences is challenging given the
majority of SNPs detected by association studies are non-coding
(Nishizaki & Boyle, 2017), whereas the majority of key changes are
coding (Cavallo & Martin, 2005), setting up a situation of difficult
detection and classification. Analysis of population structure using
DAPC links the genomic patterns seen in the multilocus genotypes with
the underlying biological processes quantified in the heritability
models. Using this approach, we identified alleles contributing to the
separation of these selectively bred families, revealing that breeding
of the selected populations targets changes to the immunity and stress
responses and growth, likely important processes in survival generally.
Assigning differences in SNPs to phenotypic differences between
individuals will be key to understanding and increasing thermal
tolerance for intervention methods.
Selective breeding
influences the level of admixture and population discontinuity
Admixture events affect members of species, populations and individuals
differently (Lawson, Van Dorp, & Falush, 2018). We saw this in the
extent of admixture and its associated variance across the five
families, in which some families exhibited very little admixture
relative to others (especially CW). Although this can be somewhat
dependent on the k structure of the model used, both patterns
were explored independently using two techniques (DAPC and PCoA) and in
conjunction with AIC, confirmed statistically the likelihood of
population differentiation. Therefore, this may suggest that the shared
ancestry of the colony sourced from Backnumbers2 may be limited between
the other Backnumbers and Tijou corals or whose origins were from few or
divergent founders (Lawson et al., 2018). This would suggest that adult
colony Backnumbers2 is not highly related to other Backnumbers or Tijou
colonies. Furthermore, PCoA and DAPC both demonstrated that the five
families separate out in multidimensional space given the magnitude of
allelic covariance between individuals. Irrespective of population
labels, DAPC analysis also recapitulated the number of selectively bred
families produced, although interestingly, the purebred families were
not assigned to single population clusters but instead retained hybrid
clustering structure in which the proportion of ancestry was shared
between multiple two to three clusters simultaneously. The discontinuity
between populations was also surprising, as demonstrated by the reduced
spread of individuals between clusters, especially in WW1, suggestive
that selective breeding produces discrete differences in the underlying
genomic architecture of F1 individuals, even in populations likely
undergoing some underlying level of gene flow (Lukoschek, Riginos, &
van Oppen, 2016).