The use of ruminal metabolic information to select microbial genes for microbiome-driven breeding to mitigate methane emissions from beef cattle

Application
The identified animal genetic effects on ruminal metabolites concentrations and on the
abundances of microbial genes together with their genetic correlations with methane
emissions are expected to improve the accuracy of microbiome-driven breeding to reduce
this highly potent greenhouse gas efficiently and cost-effectively by reliable selection of low
emitting cattle.
Introduction
Volatile fatty acids (VFAs) in the rumen are the primary energy source for cattle and are
known to be phenotypically related to CH4 emissions. Our research aimed to investigate how
these ruminal metabolites are animal genomically influenced and genetically correlated with
CH4 emissions. Additionally, we were interested in identifying microbial genes that are closely
genetically correlated with both metabolites and CH4 emissions. The identification of the most
informative biomarkers (VFAs, microbial genes) is essential for the microbiome-driven
breeding strategy (Roehe et al., 2016; Martinez-Alvaro et al., 2022) and for improving our
understanding of the functional regulation of the ruminal metabolite metabolism and CH4
production.
Materials and Methods
The animal trials were conducted following the UK Animals Act 1986 and were approved by
the Animal Experiment Committee of SRUC. The data comprised of 363 steers that were
deeply phenotyped (including CH4 emissions measured using respiration chambers) and
genotyped using a 50k SNP chip. The animals were tested at SRUC’s Beef Research Centre
across five trials and represented four breed types, with two basal diets (480:520 and 920:80
forage:concentrate ratios). In two of the trials, the feed additives nitrate and rapeseed oil
were investigated. In addition, whole metagenome sequencing data of microbial DNA from
rumen fluid samples taken at slaughter were available. Aligning the ruminal metagenomic
sequence reads to the Kyoto Encyclopedia of Genes and Genomes (KEGG) database resulted
in the identification of 3362 microbial genes. For a subset of these animals (n = 137), VFA
concentrations in the rumen fluid, collected at slaughter, were determined using HPLC.
Bayesian genomic analyses were applied to estimate the heritabilities of the ruminal
metabolites and their genetic correlations with CH4 emissions as well as with functional
microbial KEGG genes. The genomic model included fixed environmental effects and the
animal’s random genomic effects, considering 36780 SNPs. Fixed effects in the model for
metabolites included trial, breed, basal diet, feed additives and as a covariable age at
slaughter, whereas for CH4 emissions and microbial KEGG genes, fixed effects were the
combined trial-breed-diet effects with additional consideration of a covariable, either the age
entering the respiration chambers or age at slaughter, respectively.
Results
Estimated heritabilities of molar proportion of (iso)butyrate were at high magnitude, whereas
those of acetate, propionate and (iso)valerate were at moderate level, indicating a host
genomic influence on the ruminal microbial metabolism of VFAs (Table 1). The genetic
correlations of the main VFAs with daily CH4 emissions (CH4d) were moderate in magnitude
and associated with probabilities (P0) of more than 80% to be different from zero. The
direction of the correlations indicates that higher molar proportions of acetate and butyrate
in the rumen were genetically associated with increased CH4d. In contrast, higher proportions
of propionate and valerate were genetically correlated with decreased CH4d. The genetic
correlations of isobutyrate and isovalerate were close to zero, indicating that they did not
relate to CH4 metabolism. The magnitude of the genetic correlations between VFAs and CH4
emissions per kg dry matter intake (CH4y) were similar to those emissions obtained on a daily
basis.
One interesting microbial gene group that was moderately to highly genetically correlated
with the concentration of the main ruminal metabolites was anaerobic sulphite reductase
(asr) subunits. The abundance of the microbial KEGG gene asrC showed genetic correlations
with acetate, propionate, butyrate, and valerate of -0.74, 0.87, -0.67 and 0.77, respectively,
which were associated with P0 in the range of 0.95 to 0.98. Genetic correlations of equal
direction and similar magnitude were also found for asr subunits A and B. The abundance of
the asr genes were negatively genetically correlated with CH4d between -0.23 to -0.41 with
P0 ranging from 0.77 to 0.90. These results indicate that selection for increased abundances
of the asr genes will decrease CH4 emissions by favouring ruminal propionate and valerate
metabolism compared to acetate and butyrate production.

Table 1.
Heritabilities (h2) of ruminal metabolites and their genetic correlations (rg) with daily CH4
emissions and CH4 yield
Trait h2 SD1 rg with
CH4d2
P0 rg with
CH4d
rg with
CH4y3
P0 rg with
CH4y4
Acetate 0.22 0.18 0.51 0.84 0.37 0.75
Propionate 0.34 0.24 -0.57 0.88 -0.62 0.89
Butyrate 0.51 0.26 0.43 0.82 0.52 0.87
Isobutyrate 0.46 0.27 -0.09 0.58 0.24 0.67
Valerate 0.36 0.26 -0.50 0.83 -0.41 0.77
Isovalerate 0.32 0.24 -0.01 0.50 0.26 0.68
1Standard deviation of the posterior distribution of h2 (SD); 2daily methane emissions (g/d)
(CH4d); 3methane yield (g/kg dry matter intake) (CH4y); 4probability that the genetic
correlation is different from zero (P0). Heritabilities of CH4d and CH4y were 0.46 (±0.19) and
0.43 (±0.20), respectively.
Conclusions
The molar proportions of ruminal VFAs were found to be heritable and genetically correlated
with CH4 emissions. These VFAs could be combined with the microbial gene-based
microbiome-driven breeding strategy to improve its accuracy to estimate breeding values for
CH4 emissions. In addition, key microbial genes (asrsubunits), genetically correlated with both
VFAs and CH4 emissions were identified, which are of high value to be directly included into
microbiome driven breeding to mitigate CH4 emissions. The asr genes are involved in the
reduction of sulphite to sulphide and might compete with methanogenic archaea for
molecular hydrogen (H2).
Acknowledgments
This research was funded by the Scottish Government and based on data generated from
experiments funded by the Scottish Government, BBSRC (BB/N01720X/1, BB/N016742/1,
BB/S006567/1, and BB/S006680/1), AHDB, and QMS.
References
Martínez-Álvaro, M., Auffret, M.D., Duthie, C.-A., Dewhurst, R.J., Cleveland, M.A., Watson,
M., and Roehe, R. 2022. Communications Biology 5, 350.
Roehe, R., Dewhurst, R.J., Duthie, C.A., Rooke, J.A., McKain, N., Ross, D.W., Hyslop, J.J.,
Waterhouse, A., Freeman, T.C., Watson, M. and Wallace, R.J. 2016. PLOS Genetics 12,
e1005846.