Tungstate ion (WO42-) confined in hydrophilic/hydrophobic nanomaterials functionalized br?nsted acidic ionic liquid as highly active catalyst in the selective aerobic oxidation of alcohols in water

Article abstract of DOI:10.1016/j.mcat.2020.111202

A Br?nsted acidic Ionic Liquid containing tungstate anion functionalized polysiloxane network (PMO-IL-WO₄²-) was synthesized by simple self-condensation of tungstic acid and zwitterionic organosilane precursor possessing both imidazolium and sulfonate groups. Characterization by scanning electron microscopy (SEM), Fourier-transform infrared spectroscopy (FT-IR), X-ray powder diffraction (XRD), thermal gravimetric analysis TGA, nitrogen porosimetry, solid-state NMR spectroscopy and elemental analysis confirmed that both imidazolium cation and tungstate anion of zwitterion are successfully incorporated inside the organosilica framework. The catalytic activity of resulting hybrid PMO-IL- WO₄^2- material was studied in the selective aerobic oxidation of primary and secondary alcohols using an atmospheric pressure of air in pure water. Due to the ionic liquid-based charged surface containing hydrophilic sulfonic acid and tungstate group, the synergistic hydrophilic/hydrophobic and redox effect of PMO-IL-WO₄²- as water-friendly catalyst facilitates and enhances the activity and selectivity toward the target oxidative products in water and proved to have a particularly broad substrate scope for reliable aerobic oxidation reaction. Furthermore, the catalyst showed outstanding stability and could be easily separated and reused at least ten reactions run under the same conditions as fresh catalyst without any loss of catalytic activity and product selectivity.

Full text of DOI:10.1016/j.mcat.2020.111202

J. Dairy Sci. 98:1286–1295
http://dx.doi.org/10.3168/jds.2014-8302
®
©
American Dairy Science Association , 2015.
Genome-wide association study for claw disorders
and trimming status in dairy cattle
1
D. van der Spek, J. A. M. van Arendonk, and H. Bovenhuis
Animal Breeding and Genomics Centre, Wageningen University, PO 338, 6700 AH, Wageningen, the Netherlands
ABSTRACT
or suggestive SNP associations were detected. For trim-
ming status, 1 significant and 1 suggestive SNP were
detected, both located close to each other on BTA15.
Some significant and suggestive SNP were located close
to SNP detected in studies on feet and leg conformation
traits. Genes with major effects could not be detected
and SNP associations were spread across the genome,
indicating that many SNP, each explaining a small pro-
portion of the genetic variance, influence claw disorders.
Therefore, to reduce the incidence of claw disorders by
breeding, genomic selection is a promising approach.
Key words: association study, hoof lesion, Holstein-
Friesian
Performing a genome-wide association study (GWAS)
might add to a better understanding of the develop-
ment of claw disorders and the need for trimming.
Therefore, the aim of the current study was to perform
a GWAS on claw disorders and trimming status and to
validate the results for claw disorders based on an in-
dependent data set. Data consisted of 20,474 cows with
phenotypes for claw disorders and 50,238 cows with
phenotypes for trimming status. Recorded claw disor-
ders used in the current study were double sole (DS),
interdigital hyperplasia (IH), sole hemorrhage (SH),
sole ulcer (SU), white line separation (WLS), a combi-
nation of infectious claw disorders consisting of (inter-)
digital dermatitis and heel erosion, and a combination
of laminitis-related claw disorders (DS, SH, SU, and
WLS). Of the cows with phenotypes for claw disorders,
INTRODUCTION
Breeding goals in dairy cattle focus not only on pro-
duction traits, but increasing emphasis is also on health
and durability traits (Miglior et al., 2005). Claw disor-
ders are common in dairy cattle with a prevalence of
more than 70% (e.g., Manske et al., 2002; van der Waaij
et al., 2005). Claw disorders are important because of
welfare issues (Enting et al., 1997) and economic im-
pact (Bruijnis et al., 2012a,b). A trait currently not
considered but of interest is the need for claw trim-
ming. Some cows need more claw trimming than others
and van der Spek et al. (2013) showed that the need
for trimming, or “trimming status,” is a heritable trait.
High scores for trimming status reflect that daughters
of a bull need more trimming, which is unfavorable and
which is positively correlated with increased occurrence
of claw disorders (van der Spek et al., 2013).
Genetic selection for reduced claw disorders is dif-
ficult because the disorders are not routinely recorded.
Indicator traits for claw disorders, which may be more
accurate and easier to obtain, are lameness (Laursen et
al., 2009; Weber et al., 2013) and feet and leg confor-
mation traits (van der Waaij et al., 2005; van der Linde
et al., 2010). Scores for feet and leg conformation are
routinely collected in most breeding schemes. Previous
studies have detected QTL for lameness and feet and
leg conformation (Ashwell et al., 1998a,b; Schrooten
et al., 2000; Boichard et al., 2003; Buitenhuis et al.,
2007). However, to the best of our knowledge, no link-
1
,771 cows were genotyped and these cow data were
used for the GWAS on claw disorders. A SNP was con-
sidered significant when the false discovery rate ≤ 0.05
and suggestive when the false discovery rate ≤ 0.20. An
independent data set of 185 genotyped bulls having at
least 5 daughters with phenotypes (6,824 daughters in
total) for claw disorders was used to validate significant
and suggestive SNP detected based on the cow data. To
analyze the trait “trimming status” (i.e., the need for
claw trimming), a data set with 327 genotyped bulls
having at least 5 daughters with phenotypes (18,525
daughters in total) was used. Based on the cow data,
in total 10 significant and 45 suggestive SNP were de-
tected for claw disorders. The 10 significant SNP were
associated with SU, and mainly located on BTA8. The
suggestive SNP were associated with DS, IH, SU, and
laminitis-related claw disorders. Three of the sugges-
tive SNP were validated in the data set of 185 bulls,
and were located on BTA13, BTA14, and BTA17. For
infectious claw disorders, SH, and WLS, no significant
Received April 29, 2014.
Accepted October 28, 2014.
1
Corresponding author: dianne.vanderspek@wur.nl
1286

GENOME-WIDE ASSOCIATION STUDY OF CLAW DISORDERS
1287
age or GWAS have been reported on claw disorders. Phenotypic Data on Claw Disorders
The bovine genome sequencing and the emergence of
After removing records of cows with both parents
unknown or with 2 different trimming records on the
same date (n = 6,374 records), the data set contained
high-throughput genotyping technologies have made
it possible to perform genome-wide association stud-
ies (GWAS, e.g., Tellam et al., 2009). Genome-wide
association studies enable the detection of genetic
variants associated with a particular trait or disease,
using dense genome-wide markers, also known as SNP
5
0,238 cows. The cows descended from 3,603 sires with
an average of 14 daughters per sire. Phenotypes on
claw disorders were collected by 6 professional claw
trimmers, from January 2007 through February 2012,
during routine visits on 574 dairy farms in France. The
farmer decided which cows were to be trimmed, and as
a result, not all cows present in a herd were trimmed.
Information on parity, stage of lactation, and pedigree
was available on all cows present in a herd at the mo-
ment of trimming (including the nontrimmed cows).
Of the 50,238 cows, 20,474 had one or more claw trim-
ming records and in total 29,994 claw trimming records
were available. Trimming records were repeated within
and across lactations; 69% of the cows had 1 trimming
record, 20% had 2 trimming records, and 11% had 3 or
more trimming records. Claw disorders were recorded
for the hind legs and scored as a binary trait: 0 = no
claw disorder, 1 = claw disorder in at least one hind leg.
Recorded claw disorders used in the current study were
double sole (DS), interdigital hyperplasia (IH), sole
hemorrhage (SH), sole ulcer (SU), white line separation
(
Hirschhorn and Daly, 2005; Matukumalli et al., 2009).
Performing a GWAS might add to a better understand-
ing of the development of claw disorders and the need
for trimming, when the underlying genetic background
can be identified. A GWAS is a good method to de-
tect SNP associations, but some of the results can
be false positives. False-positive associations occur
especially due to population structure (Goddard and
Hayes, 2009). Even if population structure is accounted
for in the analysis and stringent significance thresholds
are used, false-positive results might occur due to the
high chance of some unaccounted for data structure in
livestock populations given the large number of tested
SNP (Hayes, 2013). To eliminate false positives, asso-
ciations detected in a GWAS study should therefore
be validated in an independent population (Chanock
et al., 2007; Hayes, 2013). Therefore, the aim of the
current study was to perform a GWAS on several claw
disorders in dairy cows and to validate the results for
claw disorders based on an independent data set. In
addition, a GWAS will be performed on the trait trim-
ming status using daughter yield deviations (DYD) of
bulls.
(
WLS), and a combination of infectious lesions (DER)
consisting of (inter-)digital dermatitis and heel erosion.
The trimming status trait indicates whether a cow was
trimmed (score 1) or not trimmed (score 0) during a
visit by the claw trimmer on a specific date. The claw
disorders and the trait trimming status are explained
in more detail by van der Spek et al. (2013). van der
Spek et al. (2013) showed moderate to high genetic cor-
relations between laminitis-related claw disorders (DS,
SH, SU, and WLS). Therefore, 4 laminitis-related claw
disorders were combined by adding up the scores for
the individual traits, resulting in a trait LAMIN, with
scores ranging from 0 (no claw disorder) to 4 (all 4 claw
disorders present). For trimming status, 50,238 cows
with phenotypes were available, and for claw disorders
MATERIALS AND METHODS
Analyses were performed based on 2 data sets: one
with genotyped cows and one with genotyped bulls. The
data set with cows was based on genotyped cows, which
also have phenotypes for claw disorders. These data
will be referred to as the cow data. The data set with
bulls was based on genotyped bulls that have daughters
with phenotypes for claw disorders. Phenotypes of cows
adjusted for systematic environmental effects were
used to calculate the DYD for bulls. Phenotypes from
genotyped cows used in the cow data were dropped
from calculating DYD for bulls. In this case, no overlap
exists in phenotypes between the 2 data sets. The DYD
2
0,474 cows with phenotypes were available.
Genotypic Data
The DNA was extracted from blood or semen samples.
data of bulls were used to validate significant or sugges- Herds with at least 10% of the cows having claw disor-
tive SNP detected for claw disorders using the cow data ders in previous years were identified and all cows on
and will be referred to as the bull validation data. The these herds trimmed in the second half of 2012 or first
trait trimming status was analyzed with the bull data half of 2013 were sampled. Subsequently, all cows with
without removing phenotypes of genotyped daughters. trimming records available were genotyped. Genotypes
The DYD were calculated and used as a phenotype for of bulls were already available and were included when
bulls. These data will be referred to as the trimming the bull had daughters with trimming records. In total,
status data.
1,771 Holstein-Friesian cows and 506 Holstein-Friesian
Journal of Dairy Science Vol. 98 No. 2, 2015

1288
VAN DER SPEK ET AL.
bulls were genotyped with the Illumina BovineSNP50
BeadChip (Illumina Inc., San Diego, CA) for 54,609
SNP. All genotyped cows have phenotypic data avail-
able as well, and all genotyped bulls have daughters
2
mental effects ~N(0, Iσ ), and e is a vector of residual
pe
2
2
effects ~N(0, Iσ ), where I is the identity matrix, σ is
the permanent environmental variance and σ is the
e
pe
2
e
with phenotypic data. The genotyped cows were kept in residual variance; Z
is the incidence matrix relating
is the incidence
1
8
7 herds and descended from 434 sires of which 214 had observations to animal effects, and Z
2
genotypic data available. One bull had a call rate <95% matrix relating observations to permanent environmen-
and was eliminated. Genotypes were analyzed using the tal effects.
Illumina GenomeStudio software. Quality control was
A linear model was used for the binary traits as it
performed on the genotypic data and a SNP was only is computationally more feasible as compared with a
included when the following criteria were met: (1) the threshold model. Also, Pirinen et al. (2013) showed
minor allele frequency (MAF) >2%; (2) the percent- that for a GWAS, the logistic regression model can be
age of missing genotypes across all samples <5%; (3) no accurately approximated by the linear model. The heri-
2
strong deviation from Hardy-Weinberg equilibrium (χ tabilities for claw disorders were fixed at the estimates
values < 600). The last criterion was included as a way obtained from the variance component analysis as given
to filter out poor-quality SNP. Extreme deviations from by van der Spek et al. (2013). Variance components
Hardy-Weinberg equilibrium are expected to be due to for LAMIN, combining the 4 laminitis-related claw
poorly called genotypes. Genotypes of bulls with fewer disorders, were estimated using the model described by
than 5 daughters with records on claw disorders were van der Spek et al. (2013) and resulted in a heritabil-
eliminated; the number of daughters per bull ranged ity of 0.07 (±0.01) and a phenotypic variance of 0.34.
from 5 to 905. A total of 41,761 SNP for 1,771 cows, Analyses were performed using ASReml v3.0 (Gilmour
3
27 bulls with phenotypes for trimming status of 6,824 et al., 2009).
daughters, and 185 bulls with phenotypes for claw dis-
The significance threshold for the GWAS was ad-
orders of 18,525 daughters were retained and available justed for multiple testing using the false discovery rate
for analyses.
(FDR). The qvalue package in R statistical software
Storey and Tibshirani, 2003) was used to obtain the
(
FDR. A FDR ≤ 0.20 will be referred to as suggestive
and a FDR ≤ 0.05 as significant in our study.
Association Study for Claw Disorders—Cow Data
The association of an individual SNP with a claw
When a SNP genotype contained less than 5 cows,
disorder was estimated using the following linear ani- the records for this genotype were omitted and the
mal model:
association for this SNP was reevaluated. When the
P-value is ≥ 0.05, the SNP will be omitted from further
analysis.
y = Xb + Z a + Z pe + e,
1
2
Association Study—Bull Data
where y is a vector of observations of the trait; b is a
vector of fixed effects, including herd, year-season of
trimming (season is defined as spring = March to May;
summer = June to August; autumn = September to
November; winter = December to February), parity at
trimming (consisting of 4 classes; 1, 2, 3, and ≥4), lac-
tation stage at trimming (consisting of 10 classes; class
The association of an individual SNP with a claw
disorder or trimming status was estimated using the
following linear animal model:
y = Xb + Z a + e,
1
1
to 9 are 50 d each, with the first class from 1 to 50 d,
the second class from 50 to 100 d, and so on, and cows where y is a vector of observations of the trait; b is a
with lactation stage ≥450 d were assigned to class 10), vector of the fixed effect SNP (SNP is treated as a class
and SNP (SNP is treated as a class variable with 2 or variable with 2 or 3 classes, depending on the number
3
classes, depending on the number of genotypes); X is of genotypes); X is the incidence matrix for the fixed
the incidence matrix for the fixed effects; a is a vector effect; a is a vector of animal additive genetic effects
of animal additive genetic effects and is assumed to and is assumed to follow a multivariate normal distri-
2
a
2
a
follow a multivariate normal distribution ~N(0, Aσ ), bution ~N(0,Aσ ), where A is the additive genetic rela-
where A is the additive genetic relationships matrix tionships matrix, which consisted of 2,172 animals, and
2
a
2
which consisted of 56,867 animals and σ is the additive σ is the additive genetic variance; e is a vector of re-
a
2
e
genetic variance; pe is a vector of permanent environ- sidual effects ~N(0,Rσ ), where R is a diagonal matrix
Journal of Dairy Science Vol. 98 No. 2, 2015

GENOME-WIDE ASSOCIATION STUDY OF CLAW DISORDERS
1289
with the reciprocal of the reliabilities as diagonal ele- of SU. The AA genotype had an effect size of 0.28 and
2
e
ments and σ is the residual variance; Z is the incidence the BB had an effect size of 0.01, corresponding to an
1
increase in incidence of SU with 27% when a cow has
genotype AA as compared with genotype BB. For other
significant SNP, the difference between both homozy-
gote genotypes corresponded to an increase in incidence
of SU ranging from 10 to 38%.
The 10 significant SNP were located on BTA8,
BTA10, BTA11, BTA18, and BTA22. Most significant
SNP (n = 5) were located on BTA8. The 45 suggestive
SNP were located on 20 different chromosomes: BTA1,
BTA5 to BTA15, BTA17, BTA18, BTA20 to BTA24,
and BTA26. Most suggestive SNP were detected on
BTA8 (n = 10), BTA9 (n = 5), and BTA20 (n = 5).
Estimated SNP effects of the 10 significant SNP are
presented in Table 1, and estimated SNP effects of the
matrix relating observations to animal effects.
Variance components for trimming status and claw
disorders were fixed at the estimates obtained from
analyses with a weighted univariate linear animal
model, with DYD as the dependent variable and animal
as a random additive genetic effect, using reliabilities
as weights.
Validation of Suggestive SNP for Claw Dis-
orders. In the bull validation set for claw disorders,
the effect of a suggestive or significant SNP detected
in the cow data was considered validated if it showed
a significant effect (P ≤ 0.05) and if the allele with the
favorable effect was identical in both analyses.
Trimming Status. Based on the trimming status
data a SNP association was significant if FDR ≤ 0.05
and suggestive if FDR ≤ 0.20. When a SNP genotype
4
5 suggestive SNP are presented in Appendix Table A1.
contained fewer than 5 bulls, the records for this geno- SNP Analyses on DYD of Bulls
type were omitted and the association for this SNP was
reevaluated. When the P-value is ≥ 0.05, the SNP will
Validation of Suggestive SNP for Claw Dis-
orders. Three of the suggestive SNP detected in the
GWAS based on cow data were confirmed based on
the bull validation data. These SNP were ARS-BFGL-
NGS-113540 on BTA13 (P = 0.02), ARS-BFGL-
NGS-4929 on BTA14 (P = 0.02), and BTB-00678060
on BTA17 (P = 0.02). The favorable allele was identical
be omitted from further analysis.
RESULTS
SNP Analysis on Claw Disorders with the Cow Data
The results of the genome-wide association studies for in the cow data as in the bull validation data.
the different claw disorders are shown in Figure 1. In to-
SNP Associations with Trimming Status. The
tal, 17 significant and 77 suggestive SNP were detected. GWAS results for trimming status are shown in Figure
When a genotype contained less than 5 cows, the SNP 2. Four significant and 6 suggestive SNP associations
was reevaluated by omitting the records for this geno- were detected for the trait trimming status. Eight SNP
type and reevaluating the association for this SNP. Of had one genotype containing 5 or less bulls and there-
the 94 significant and suggestive associations, 39 were fore these associations were reevaluated by omitting
reevaluated. None of the reevaluated associations had the smallest genotype. None of the reevaluated asso-
a P-value ≤ 0.05 and were consequently omitted from ciations had a P-value ≤ 0.05 and were consequently
further analysis; they are also not shown in Figure 1. omitted from further analysis. Two suggestive SNP
In total, 10 significant and 45 suggestive SNP remained associations, both located on BTA15, remained. Fig-
and will be discussed in more detail. Ten significant ure 2 shows the results for trimming status after the
and 20 suggestive SNP associations were detected for reevaluation. The −log10 P-values were 5.13 for SNP
SU. One suggestive association was detected for DS, 17 UA-IFASA-6898 (32713410 bp) and 4.99 for SNP ARS-
for IH, and 7 for LAMIN, whereas DER, WLS and SH BFGL-NGS-57210 (32637662 bp).
did not show any suggestive SNP association (Figure
1
). For 1 significant and 1 suggestive SNP, no records
DISCUSSION
were present for 1 of the 3 genotypes due to low MAF.
The chromosome number, position, SNP name, and the
number of cows per genotype for the 10 significant SNP
with associated claw disorder are shown in Table 1 and
SNP Associations with Claw Disorders in Cows
In the cow data set, 10 significant and 45 suggestive
for the 45 suggestive SNP in Appendix Table A1. The SNP were detected for DS, IH, SU, and LAMIN. The
total number of cows per SNP differs slightly due to 55 significant and suggestive SNP were located on 20
missing SNP genotypes. The −log10 P-values for the different chromosomes. This suggests that claw disor-
significant SNP associations range from 4.67 to 6.79 ders are influenced by many genes dispersed across the
(
Table 1). The most significant SNP (−log10 P-value entire genome, each explaining a small part of the ge-
of 6.79) was associated with an increase in incidence netic variance. Caution must be taken because it might
Journal of Dairy Science Vol. 98 No. 2, 2015

1290
VAN DER SPEK ET AL.
Figure 1. Genome-wide association study of claw disorders and the combination of laminitis-related claw disorders (double sole, sole hemor-
rhage, sole ulcer, and white line separation). The false discovery rate was set at 0.05 for significant SNP (dashed line) and 0.20 for suggestive
SNP (solid line). Color version available online.
Journal of Dairy Science Vol. 98 No. 2, 2015

GENOME-WIDE ASSOCIATION STUDY OF CLAW DISORDERS
1291
Figure 2. Genome-wide association study for trimming status using daughter-yield deviations of bulls. The false discovery rate was set at a
threshold of 0.05 for significant SNP (dashed line) and 0.20 for suggestive SNP (solid line). Color version available online.
be that not all relevant SNP were detected and some equal to 74% and with a proportion of 2.5% of the phe-
SNP might be false positives due to the low number notypic variance explained by the QTL, the detection
of animals and many SNP suffering from low MAF, power is equal to 28%. The power calculations show a
as will be discussed later. No suggestive or significant high probability of identifying SNP explaining at least
SNP were detected for claw disorders DER, SH, and 5% of the phenotypic variance in our study. Therefore,
WLS. Associations with these claw disorders were ap- if a gene with a major effect on claw disorders would be
parently too small to be detected in the present data segregating in the current population, it is likely that
set. A larger number of animals will increase the power it would have been detected. The power was too low to
to detect associations, especially for SNP explaining a detect genes with a moderate or small effect.
small proportion of the genetic variance of the trait
e.g., Visscher, 2008; Goddard and Hayes, 2009).
The GWAS signals detected in the current study
are different from what has been reported in other
(
We chose to calculate a trait-based FDR instead of GWAS. The first issue is that in several cases, only a
an experiment-based FDR. Adjusting the significance single significant or suggestive SNP was detected in a
threshold to account for testing multiple traits has region, whereas often several SNP in a region show a
some disadvantages because it would penalize studies significant association. This could be due to QTL with
that report on multiple traits and it would encourage a low MAF: 7 of the 10 significant SNP and 23 of the
authors to write papers for each trait separately.
45 suggestive SNP had one genotype with less than
Claw disorders have a low heritability (ranging from 50 cows and therefore have a low MAF (<3%). The
.02 to 0.14, van der Spek et al., 2013). With 1,771 QTL with low MAF in general have a low detection
0
genotyped cows and assuming a squared correlation power as they explain a small part of the variance
2
(
r ) between marker and QTL of 0.2, allele frequencies (e.g., Pritchard, 2001; Cirulli and Goldstein, 2010).
of both marker and QTL of 0.5, and a type I error equal The QTL with low MAF might also be in low LD with
to 0.001, the power to detect a QTL can be calculated SNP present on the SNP array due to ascertainment
[
using the function luo.ld.power (Luo, 1998) from the bias (Wray, 2005). Better coverage using SNP with low
package ldDesign for the statistical software R (Ball, MAF might be needed to identify stronger associations
2
010)]. With a proportion of 5% of the phenotypic (Lee et al., 2013). The second issue is that effects of
variance explained by the QTL, the detection power is SNP detected are large. Although studies have shown
Table 1. Single nucleotide polymorphisms significantly associated with sole ulcer [false discovery rate (FDR) ≤ 0.05]
No. of cows
SNP effect
AB (SE)
Position
(bp)
BTA
SNP name
−log10
P
FDR
AA
AB
BB
992
AA (SE)
BB (SE)
8
8
8
8
8
5437200
5490554
BTA-81490-no-rs
5.20
5.82
5.85
5.44
6.26
4.67
4.79
5.42
4.70
6.79
0.02
0.01
101 677
101 668 1,002
0.07 (0.03)
0.08 (0.03)
−0.01 (0.02)
0.38 (0.06)
0.15 (0.03)
−0.08 (0.02)
−0.02 (0.01)
−0.03 (0.01)
0.004 (0.01)
0.28 (0.03)
0
0
0
0
0
0
0
0
0
0
−0.03 (0.01)
−0.03 (0.01)
0.28 (0.06)
0.001 (0.02)
−0.02 (0.01)
—
0.16 (0.02)
0.24 (0.04)
0.25 (0.04)
0.01 (0.01)
ARS-BFGL-NGS-4062
Hapmap43489-BTA-121857
ARS-BFGL-NGS-108587
ARS-BFGL-NGS-34028
BTB-01321748
74093999
78533314
100873051
44065746
48179532
23878651
14300175
15276355
0.01 1,549 209
13
142 1,623
51 518 1,202
0.02
0.01
6
10
11
18
22
22
0.05 1,667 104
0.05 1,263 464
0.02 1,492 264
0.05 1,512 241
0
44
15
13
Hapmap38795-BTA-97039
ARS-BFGL-NGS-15436
BTB-00834995
ARS-BFGL-NGS-3298
0.01
15 249 1,507
Journal of Dairy Science Vol. 98 No. 2, 2015

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VAN DER SPEK ET AL.
that alleles with low frequencies can have large effects region of SNP ARS-BFGL-NGS-113540 on BTA13, the
on complex traits (e.g., Mackay et al., 2012; Weber et genes C1QL3 and PTER were located. In the region
al., 2012), the SNP detected in our cow data set are of SNP ARS-BFGL-NGS-4929 on BTA14, the genes
likely overestimated. The significant SNP associations AZIN1, TRNAE-UUC, KLF10, ODF1, and UBR5 were
in our study in general have a low MAF and explain a located, and in the region of SNP BTB-00678060 on
small part of the genetic variance. When the explained BTA17, the genes SFSWAP, MMP17, ULK1, PUS1,
genetic variance is low, the detection power is low and EP400, LOC101906627, and LOC100138728 were
effect sizes are likely overestimated (Lynch and Walsh, located. However, none of the genes has an apparent
1
998). This relates to the effect known as the winner’s function related to claw disorders. Eight suggestive
curse (Ioannidis, 2008; Kraft, 2008). The effect sizes and 3 significant SNP detected based on the cow data
for the SNP that are validated based on the bull data set were located close to associations with feet and leg
are on average almost 4 times smaller as compared conformation detected in previous research. The SNP
with the effect sizes in the cow data. Although SNP ARS-BFGL-NGS-12807 on BTA5 at 71.8 Mbp was
effects estimated based on the bull data are allele sub- suggestively associated with SU in our study. At al-
stitution effects and therefore not fully comparable, most the same location (71.7 Mbp), Cole et al. (2011)
it illustrates that the effects reported in Table 1 are detected an association with general feet and leg score.
likely overestimates. The above-mentioned reasons for Furthermore, in the same region (Lysmic-ETH10 at 72
both issues might explain why only one significantly cM) Hiendleder et al. (2003) detected a QTL associated
or suggestively associated SNP with large effect was with foot angle. Buitenhuis et al. (2007) detected a
detected in a region.
QTL on BTA8 (marker MCM64-CSSM047 at 92.0 cM)
associated with foot angle and this appears to be in a
region associated with SU in our study: 3 significant
and 5 suggestive SNP in the region 67.8 to 106.6 Mbp.
Literature
No other genome-wide association studies on claw A linkage study by Hiendleder et al. (2003) showed
disorders have been published. However, linkage stud- an association of marker MILSTS077 on BTA13 (54
ies (e.g., Ashwell et al., 2005; Buitenhuis et al., 2007) cM) with rear leg side view. This marker appears to be
and a GWAS (Cole et al., 2011) have been published close to SNP ARS-BFGL-NGS-113236 (29.6 Mbp) as-
on traits which have been shown to be genetically cor- sociated with LAMIN in the current study. On BTA6,
related to claw disorders (van der Waaij et al., 2005; Hiendleder et al. (2003) detected marker FBN14 (88
Laursen et al., 2009; van der Linde et al., 2010; Weber cM), which is associated with foot angle and appears
et al., 2013), for example, rear leg rear view, rear leg to be close to SNP BTA-77057-no-rs (87.4 Mbp) as-
side view, foot angle, general feet and legs score, hock sociated with IH in the current study. Another linkage
quality, and lameness. First we will discuss how the study detected marker BP7 on BTA6 (85 cM), which
validated SNP in our study relate to findings in litera- is also located close to SNP BTA-77057-no-rs, and in
ture and subsequently other significant and suggestive their study it is associated with rear leg set (Schrooten
results will be discussed.
The 3 validated SNP were located in chromosomal
et al., 2000).
The SNP on BTA18 position 58.7 Mbp detected by
regions which have been associated with feet and leg Cole et al. (2011) and associated with rear leg side view
conformation traits. The validated SNP ARS-BFGL- was also detected in our study. In our study, this SNP
NGS-113540 (31.3 Mbp) associated with SU in our was associated with IH, but one genotype had only 3
study is located in the same region of BTA13 as marker observations. After removing this genotype, the SNP
UWCA25-BL42 (20–83 cM) associated with foot angle was no longer significant (P > 0.05). A recent study of
(
Ashwell et al., 2005). Another marker detected by Ash- Swalve et al. (2014) using a preselected set of 384 SNP
well et al. (2005) associated with foot angle, BMS1899- detected a strong association on BTA21 with SH. The
BM4513 (0–76 cM), was located in the same region SNP detected by Swalve et al. (2014), SNP rs29017173,
of BTA14 as validated SNP ARS-BFGL-NGS-4929 had a nominal P-value of 0.06 for SU in our study.
(64.0 Mbp) associated with IH in our study. The third The favorable SNP allele in our study is identical to
validated SNP, BTB-00678060 (46.3 Mbp) associated Swalve et al. (2014). For other laminitis-related traits
with SU in our study, was located in the same region the nominal P-value of this SNP was higher than 0.20.
of BTA17 as marker CSSM9-OARFCB48 (32.0 cM) as- Even though we did not find a significant (FDR ≤ 0.05)
sociated with bone quality detected by Buitenhuis et or suggestive SNP (FDR ≤ 0.20) on BTA21 associ-
al. (2007).
ated with SH or laminitis-related claw disorders, this
We searched for candidate genes by taking a 200,000- result seem to confirm the SNP detected in the study
bp window surrounding the 3 validated SNP. In the of Swalve et al. (2014).
Journal of Dairy Science Vol. 98 No. 2, 2015

GENOME-WIDE ASSOCIATION STUDY OF CLAW DISORDERS
1293
Whether a cow was trimmed or not, the so-called Also, these 3 SNP were located in the same region on
trimming status of a cow reflects the need for trimming. the genome as QTL detected in previous research as-
One significant and one suggestive SNP association was sociated with feet and leg conformation traits. Genes
detected, both located close to each other on BTA15. with a major effect were not detected. Likely, many
Ashwell et al. (2005) detected a QTL (BMS2684-HBB) genes each explaining a small proportion of the genetic
located in the same region, associated with stature. The variance influence the development of claw disorders
SNP ARS-BFGL-NGS-73559 on BTA26 position 44.7 but the power of the current study was too low to verify
Mbp, was associated with trimming status but had one this.
genotype with only 2 observations, and when this geno-
type was removed the SNP was no longer significant (P
0.05). This SNP might still be interesting because
it was situated within a region with 7 significant SNP
ACKNOWLEDGMENTS
>
This study was financially supported by Genes Diffu-
detected by Cole et al. (2011), spanning from 39.3 to sion, Douai, France. The authors thank the claw trim-
9.2 Mbp. mers, technicians, farmers, and all the people involved
One candidate gene (SORL1) for the trimming status for their major effort in collecting data and making it
4
trait was detected for both UA-IFASA-6898 and ARS- available for us. Amélie Vallée (Genes Diffusion, Douai,
BFGL-NGS-57210 within a 200,000-bp window, but France) is acknowledged for her contribution in obtain-
does not have an apparent function related to the trait. ing the genotypes.
Even though we indicated that many significant and
suggestive SNP were located in close proximity to SNP
identified in previous studies on claw and leg conforma-
tion traits, the overlap is not very high. This is most
likely due to the rather low to moderate genetic cor-
relations between claw and leg conformation traits and
claw disorders, ranging from −0.07 to 0.69 (van der
Waaij et al., 2005; Laursen et al., 2009).
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Journal of Dairy Science Vol. 98 No. 2, 2015

APPENDIX
Table A1. Single nucleotide polymorphism associations for claw disorders and the combination of laminitis-related claw disorders (double sole, sole hemorrhage, sole ulcer, and
white line separation), that were suggestively [false discovery rate (FDR) ≤ 0.20] associated
No. of cows
AB
SNP effect
AB (SE)
Position
(bp)
BTA
SNP name
Claw disorder
−log10
P
FDR
AA
BB
28
AA (SE)
BB (SE)
1
1
5
6
7
7
7
8
8
8
8
8
8
8
8
8
8
9
9
9
9
9
1
1
1
1
1
1
1
1
1
1
1
1
18285474
93639215
71778071
87365840
61196630
68156093
68281468
8953354
24869885
25012621
25035564
43241581
67822026
67846594
100537840
101044054
106648392
1333028
79256833
83904725
85494205
90283190
31162777
44082049
14134626
62307557
62615139
29647662
31304022
6203679
63972246
52875986
67131730
46296022
25022927
25046677
37721846
48937082
64084187
67899804
71498820
7526820
Hapmap33466-BTA-107178
BTA-48243-no-rs
ARS-BFGL-NGS-12807
BTA-77057-no-rs
Sole ulcer
Laminitis related
Sole ulcer
Interdigital hyperplasia
Interdigital hyperplasia
Interdigital hyperplasia
Sole ulcer
3.87
4.41
3.81
5.17
4.13
3.87
3.94
3.81
4.17
4.17
4.17
4.37
4.40
4.34
4.25
4.22
3.67
3.94
3.94
4.97
4.07
4.18
4.18
4.07
3.84
4.34
4.27
4.30
4.61
4.37
4.69
5.08
4.31
4.09
4.08
4.08
5.25
4.27
4.38
4.32
4.58
3.94
3.94
4.15
4.91
0.15 1,335 405
−0.02 (0.01)
−0.15 (0.03)
0.11 (0.03)
−0.01 (0.01)
0.02 (0.01)
−0.01 (0.01)
0.08 (0.03)
0.18 (0.04)
0.29 (0.06)
−0.03 (0.02)
0.29 (0.06)
0.06 (0.01)
0.26 (0.05)
0.26 (0.05)
0.13 (0.02)
−0.02 (0.01)
0.32 (0.06)
−0.03 (0.01)
0.23 (0.04)
0.15 (0.05)
−0.02 (0.01)
−0.06 (0.02)
0.16 (0.03)
−0.09 (0.02)
0.05 (0.01)
0.10 (0.09)
0.12 (0.09)
−0.16 (0.08)
0.02 (0.01)
0.03 (0.01)
0.17 (0.05)
0.51 (0.09)
0.27 (0.05)
0.44 (0.09)
0.04 (0.02)
−0.02 (0.01)
−0.01 (0.03)
−0.02 (0.03)
0.03 (0.02)
0.29 (0.05)
−0.05 (0.11)
−0.01 (0.01)
0.36 (0.06)
0.02 (0.02)
−0.01 (0.01)
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0.15 (0.03)
−0.02 (0.07)
0.02 (0.01)
0.16 (0.04)
0.08 (0.02)
0.22 (0.05)
0.03 (0.01)
0.04 (0.01)
−0.03 (0.02)
0.29 (0.06)
−0.03 (0.02)
0.02 (0.02)
0.02 (0.02)
0.02 (0.02)
−0.01 (0.01)
0.07 (0.02)
−0.01 (0.02)
0.07 (0.04)
0.004 (0.01)
−0.02 (0.01)
0.15 (0.02)
0.21 (0.12)
−0.02 (0.01)
—
0.07 (0.03)
0.14 (0.03)
0.13 (0.03)
−0.02 (0.04)
0.28 (0.05)
0.10 (0.03)
−0.00 (0.01)
0.03 (0.02)
−0.02 (0.02)
0.01 (0.01)
−0.02 (0.01)
0.04 (0.02)
0.27 (0.06)
0.22 (0.05)
0.23 (0.04)
0.02 (0.02)
0.14 (0.03)
0.06 (0.02)
0.03 (0.02)
0.35 (0.07)
0.09 (0.03)
0.17
0.16
497 898
68 520 1,183
379
0.06 1,204 506
0.12 726 823
0.19 1,362 389
172 822
22 404 1,345
0.12 1,500 261
57
222
20
ARS-BFGL-NGS-5134
Hapmap54765-rs29012520
ARS-BFGL-NGS-65419
ARS-BFGL-NGS-69316
BTB-01052583
0.13
0.20
777
Interdigital hyperplasia
Interdigital hyperplasia
Interdigital hyperplasia
Interdigital hyperplasia
Interdigital hyperplasia
Sole ulcer
10
BTB-01052539
0.12
0.12
0.12
0.08
0.09
0.10
10 261 1,500
10 261 1,500
564 888
ARS-BFGL-NGS-5492
ARS-BFGL-NGS-4797
ARS-BFGL-NGS-115977
ARS-BFGL-NGS-40613
ARS-BFGL-NGS-79358
ARS-BFGL-NGS-112179
Hapmap27876-BTA-147931
Hapmap41734-BTA-103209
Hapmap26214-BTA-163696
ARS-BFGL-NGS-35130
Hapmap59396-rs29023560
BTB-00404361
319
9
9
204 1,558
203 1,558
Sole ulcer
Sole ulcer
46 510 1,215
Sole ulcer
0.10 1,013 657
0.20 217
0.13 1,125 578
101
1545
68
Sole ulcer
9
Sole ulcer
Sole ulcer
0.13
0.06
15 255 1,501
52 494 1,225
0.13 1,244 459
Interdigital hyperplasia
Interdigital hyperplasia
Interdigital hyperplasia
Sole ulcer
47
6
0.12
0.11
1494 270
25 399
0.11 1,662 109
0.19 1,027 655
0
0
1
2
2
3
3
4
4
5
5
7
BTB-00416921
BTB-01413108
Hapmap31534-BTA-26223
BTA-85739-no-rs
Hapmap27169-BTA-141662
ARS-BFGL-NGS-113236
ARS-BFGL-NGS-113540
Hapmap32486-BTC-056478
ARS-BFGL-NGS-4929
BTB-00600391
BTB-01649233
BTB-00678060
ARS-BFGL-NGS-24004
ARS-BFGL-NGS-107737
BTA-50515-no-rs
ARS-BFGL-NGS-59274
BTB-00795527
ARS-BFGL-NGS-6648
ARS-BFGL-NGS-13702
Hapmap44255-BTA-52888
ARS-BFGL-NGS-23572
ARS-BFGL-NGS-107083
BTA-111275-no-rs
1347
0
Sole ulcer
Interdigital hyperplasia
Laminitis related
Laminitis related
Laminitis related
Sole ulcer
89
0.17
0.17
0.17
124 721
110 685
424 908
926
976
439
13
0.06 1,447 311
0.08 1,136 576
Sole ulcer
59
Interdigital hyperplasia
Double sole
0.07
0.07
0.09
0.11
0.11
0.11
0.07
0.17
53 497 1,221
6
8
6
239 1,521
196 1,567
290 1,475
Sole ulcer
Sole ulcer
18
18
20
20
20
20
20
21
23
24
26
Sole ulcer
316 857
598 856
970 678
878 748
598
316
123
144
19
Sole ulcer
Laminitis related
Laminitis related
Interdigital hyperplasia
Interdigital hyperplasia
Laminitis related
Sole ulcer
0.12 1,440 312
0.12
0.16
0.13
0.13
15 359 1,398
70 619 1,082
655 889
227
15725194
8435459
9195089
Sole ulcer
Interdigital hyperplasia
Interdigital hyperplasia
6
88 1,677
0.12 1,513 242
0.06 780 805
8
186