Genomics

By Paul VanRaden and Daniel Null

Previous Brown Swiss and Jersey haplotypes BH1 and JH2 will be discontinued effective with the December 2018 run. Several European bulls are homozygous for BH1, and its fertility effect is no longer significant. Thus, US and European Brown Swiss associations have decided to discontinue reporting BH1.

Haplotypes BH1 and JH2 met the initial statistical tests for publication in 2011 and 2013, with no homozygous animals found at that time. The new ARS-UCD reference map makes JH2 very difficult to trace. The fertility losses from BH1 and from JH2 carrier matings were retested using the most current four years of data, and neither were significant. As causative mutations were not found for either BH1 or JH2, both haplotypes will no longer be reported.

A new recessive haplotype in Holsteins (HH6) was discovered in France on chromosome 16, and will now be listed. The fertility effects of HH6 were confirmed with a 9% drop in conception rate.

Haplotypes Affecting Fertility and their Impact on Dairy Cattle Breeding Programs

Dr. Kent A. Weigel, Professor and Chair, Department of Dairy ScienceUniversity of Wisconsin – Madison; August 5, 2011

New tools in genomics have provided an unprecedented look at the inner workings of the cattle genome and the relationship between variation in DNA sequences and differences in the conformation, health, fertility, and performance of dairy cattle. Genetic selection has become genomic selection, and terms such as single nucleotide polymorphism (SNP), imputation, and haplotype have become part of our everyday vocabulary. Many leading dairy countries have developed large reference populations of bulls and cows with extensive phenotypes and low‐density (3K), medium‐density (50K), or high‐density (777K) genotypes. Data from these reference populations can be used to identify associations between DNA sequence differences at various locations in the genome and performance of these animals or their progeny for traits such as milk yield, udder depth, or female fertility. These “SNP effects” form the basis of genomic breeding values, because they are matched with the genotypes of young bulls and heifers to facilitate accurate selection decisions among animals that are too young to have phenotypes of their own. Genomic selection is now common practice, and dairy cattle breeding will never be the same.

As genetic selection has given way to genomic selection, research scientists, industry workers, and dairy farmers have had to re‐think the ways in which genetic information is interpreted and used. Predicted transmitting ability (PTA), which was once for a mature animal with lactation records or milk‐recorded offspring, has been replaced by genomic predicted transmitting ability (GPTA), which can be assigned to a newborn calf. Inbreeding, which was once estimated from pedigree records provided by the farmer, has been replaced by homozygosity, which can be measured from a DNA sample. A sire analyst, who once carried production records and lists of genetic predictions, now carries envelopes of hair.

We are not only faced with the task of re‐thinking the way that information about an animal’s entire genome is interpreted and used in genomic selection, but also the way that information about specific genes, markers, or chromosomal segments is interpreted and used in breeding decisions. In the past, the process of identifying specific genes with adverse effects on an animal’s appearance, health, or performance was lengthy and expensive. Success occurred when years of field reports, veterinary examinations, planned matings, and studies of similar abnormalities in other species led to an understanding of the condition at the biological level, knowledge regarding its mode of inheritance, and a reliable test for identification of carriers. The strategy for using this information, which may have been reasonable given our limited understanding of the cattle genome, typically involved an attempt to eliminate the condition by eradicating animals that were known or suspected carriers. Even then, many realized that attempted eradication of such a condition was often accompanied by the culling of many animals that could have contributed significantly to genetic progress for other key traits. In fact, in a discussion of the weaver condition in Brown Swiss nearly two decades ago, one dairy geneticist argued strongly against efforts to remove all carriers of the weaver gene, recognizing that the collective economic value of their genetic superiority far exceeded the likely costs associated with this disorder.

Times have changed, and today we recognize that such inherited conditions are not rare anomalies that occur once in a decade in a handful of genetically unfit animals. Armed with a much deeper understanding of the genomes of cattle and other food animal species, scientists now believe that it is likely that every individual carries several genes that, if expressed in homozygous form, would lead to a severely impaired or lethal phenotype.

This brings us to a recent research study by scientists at the USDA‐ARS Animal Improvement Programs Laboratory who, along with their colleagues at the USDA‐ARS Bovine Functional Genomics Laboratory, are at the forefront of genomics research in dairy cattle. In this study, VanRaden and colleagues (2011) describe a method for using genotypes from the 50K SNP array to identify regions of the genome that may be associated with failed conception or embryonic loss in dairy cattle. This approach starts with conversion of SNP genotypes, which represent the number of alleles of a particular form (A, C, T, or G) at a given location in the genome, to haplotypes, which represent strings of adjacent SNP alleles that are inherited together as a group from the sire or dam. For example, an animal’s SNP genotype at some point in the genome might be 0, 1, or 2, corresponding to the number of copies of the “A” allele that were inherited from its parents. On the other hand, an animal’s haplotype in a small region of the genome might be “AATCCATCGTTGACG” from its sire and “TATCCGATTCAAAGC” from its dam. Thousands of different haplotypes are represented on each chromosome, and using advanced computer software scientists can track the inheritance of these haplotypes across generations in an animal’s pedigree. The SNP markers that make up these haplotypes are not in genes, and changes in the SNP genotype or haplotype don’t directly cause changes in the phenotype. However, these markers are evenly spaced throughout the genome, and between them are the actual genes that affect key traits. Because these markers and genes are usually inherited together, we can follow the inheritance of different forms (good, bad, or neutral) of the gene by tracking inheritance of the corresponding marker.

Haplotypes can have positive, neutral, or negative effects on any trait, depending on the genes that are located within a given chromosomal region and the effects that changes in DNA sequence within these genes have on the biological processes underlying each trait. In an earlier project in the same laboratory, Cole (2011) showed that a hypothetical animal with favorable haplotypes at every location in the genome would have a breeding value 3.5‐fold greater than the best animal alive today – a result that would take 77 years to achieve through genetic selection. Obviously no animal will have all of the favorable haplotypes, so in practice we try to select bulls and cows that have inherited more good haplotypes than bad. For example, when selecting animals with superior genetic merit for protein yield, we might choose one for which two‐thirds of the haplotypes are favorable and one‐third are unfavorable. Because we are trying to improve several traits simultaneously, we might choose an animal with many favorable haplotypes for protein yield but fewer favorable haplotypes for udder depth. Over time, this process of balanced selection increases the frequency of favorable haplotypes, and the genes that are inherited with them, and the performance of the population is enhanced.

In the VanRaden et al. (2011) study, haplotypes represented strings of 17 to 74 consecutive SNP markers at various locations in the genome. More than 75,000 Holstein, Jersey, and Brown Swiss animals that had been genotyped with the 50K SNP array were included, and the objective was to identify haplotypes that are never found in a homozygous state (homozygous means that the same haplotype was inherited from both the sire and dam). There are many rare haplotypes for which we wouldn’t expect to find an animal with two copies, but in this study the scientists found several haplotypes that were common in the population but not in a homozygous state, even when present on the maternal and paternal sides of the pedigree. Next, they compared the average conception rate for matings in which the sire and maternal grandsire carried the same haplotype with that of other matings in the national database. Five haplotypes were identified that were missing in homozygous state and associated with below average conception rate and 60‐day non‐return rate. They repeated this analysis for stillbirth rate, and no differences were found. Based on this evidence, the authors concluded that they had identified five “haplotypes affecting fertility”, one in Brown Swiss, three in Holsteins, and one in Jerseys. The specific genes located within these chromosomal regions and their roles in biological pathways related to fertilization and embryo development are unknown, but it appears that the union of an egg and sperm carrying identical haplotypes results in failed conception or early embryonic loss. These haplotypes were labeled as Brown Swiss haplotype 1 (BH1), Holstein haplotypes 1, 2, and 3 (HH1, HH2, and HH3), and Jersey haplotype 1 (JH1). This labeling system reflects our lack of knowledge about the underlying biological mechanisms associated with these haplotypes, as well as our expectation that more haplotypes will be identified each year. A summary of these haplotypes, including their locations, frequencies among North American animals that have been genotyped to date, estimated effects on conception rate (CR) and 60‐day non‐return rate (NR60) when present in the sire and maternal grandsire, and the earliest known ancestor in which they were found, is given below:

Because these are common haplotypes, they are known to exist in thousands of bulls, cows, heifers, and calves that have been genotyped with the 50K SNP array. In addition, they are suspected to exist in thousands of animals that have been genotyped with the 3K SNP array – this tool is about 95% as accurate as the 50K array and can lead to inconclusive results for some animals. More importantly, these haplotypes are carried by millions of bulls, cows, heifers, and calves around the world that have not been genotyped, as well as thousands of animals that have been genotyped in countries that don’t yet report this information. Lack of a genotype does not equate to absence of these haplotypes, just as throwing away the pedigree of an inbred animal does not make it an outcross.

The next step is to determine how to effectively use information about these haplotypes, along with knowledge regarding the identity of their carriers, in practical breeding programs. The strategy of eradicating all animals that carry these haplotypes is neither feasible nor desirable. Imagine the genetic progress in milk yield, milk composition, conformation, health, and even fertility that would be lost by discarding thousands of haplotypes that are favorable for these traits while trying to eliminate the five aforementioned haplotypes affecting fertility! An attempt to fully eradicate these haplotypes would be no different from an attempt to eradicate all haplotypes that lead to low milk production, undesirable milk composition, unattractive conformation, or suboptimal health. If such a process were undertaken, we would soon have no animals left in our breeding programs, genetic progress would slow to a halt, and consumers would face a shortage of dairy products. Plus, just as we prepared to congratulate ourselves for eradicating one unfavorable haplotype, another would be discovered. For these reasons, breeding companies should not remove outstanding genome‐tested bulls or progeny tested bulls that carry these haplotypes from their programs, and farmers who have valuable cows, heifers, and calves with these haplotypes should neither cull these animals nor be afraid to use them as breeding stock.

A more proactive approach is to use the information about undesirable haplotypes to make informed breeding decisions. Two types of decisions must be made: selection of the best males and females to serve as parents of the next generation, and identification of the optimal mating combinations among these males and females. With regard to selection decisions, one must recognize that national genetic evaluations already exist for male fertility, in the form of service sire conception rate (SCR), and female fertility, in the form of daughter pregnancy rate (DPR). Bulls with these haplotypes affecting fertility have already been mated to thousands of cows that possess the same haplotypes. Because 25% of these matings resulted in failed conception or early embryonic loss, this information is reflected in lower SCR evaluations. Bulls with an undesirable haplotype that is common in the population will have greater reductions in SCR than bulls with an uncommon haplotype, because the chance that these bulls will be mated to cows with the same haplotype is greater. Likewise, Holstein bulls with two of the five undesirable haplotypes will have poorer SCR evaluations than bulls with one undesirable haplotype, because their data will include failed matings to cows with either (or both) haplotype(s). The impact of these haplotypes on a bull’s DPR is smaller than for SCR, because the breeding events of his daughters occur one generation later. A bull with one such haplotype will transmit it to 50% of his daughters, and they will transmit it to 25% of their eggs. Thus, when the daughter of a bull with BH1, HH1, HH2, HH3, or JH1 is mated to a bull that is known to have the same haplotype, 12.5% of the matings will end in failed conception or embryonic loss. Again, this information is already reflected in the bull’s genetic evaluation for DPR, and if we try to avoid purchasing semen of bulls that carry these haplotypes, we will be double‐counting their effects.

The USDA‐ARS Animal Improvement Programs Laboratory routinely publishes the Lifetime Net Merit (LNM$) index, which weights every trait according to its economic value; bulls that carry BH1, HH1, HH2, HH3, or JH1 have already been penalized in LNM$. The magnitude of this penalty depends on the frequency of the haplotype within the breed. For example, let’s assume that the cost of one extra day open is $2, homozygous embryos are lost at 5 to 10 days of gestation, and 20% of cows in the population have a given haplotype. If we make 100 matings, 20 will be to cows with this haplotype, 10 of their eggs will carry the haplotype, 5 will encounter a sperm with the haplotype, and each of those 5 cows will have an increase of roughly 30 days open. The total economic loss for all 100 matings will be 5 cows x 30 days per cow x $2 per day = $300, or about $3 per mating. Now assume that this bull’s Lifetime Net Merit evaluation is +$600, and we decided to buy semen from another bull that was +$500 instead – we just gave up $97 in our attempt to save $3. Plus, that $3 was already included in the first bull’s LNM$ evaluation.

With respect to mating decisions, this is where we can use our new information powerfully. Hundreds of thousands of cows (maybe millions) are already mated with computerized programs every year, for the purpose of correcting faults in their physical conformation and avoiding inbreeding depression in their offspring. In the previous example, the cost of the undesirable haplotype was $3 per mating, because the bull was mated to a random group of cows whose haplotypes were unknown. If the bull were mated to 100 genotyped cows that were known to carry the same haplotype, 50 eggs would carry the haplotype, and 25 would encounter sperm that would lead to failed conception or early embryonic loss, for a total cost of $1500, or $15 per mating. If the bull were mated to 100 daughters of other genotyped bulls that carried the same haplotype, 50 of his mates would have the same haplotype, 25 eggs would carry the haplotype, and 12.5 matings would be affected, for a total cost of $750, or $7.50 per mating.

Few cows on commercial farms have been genotyped, so we can rarely foresee the mating of a bull and cow that are known to carry the same haplotype. However, nearly every sire whose semen is marketed for artificial insemination (AI) has been genotyped, so the genotypes of the service sire and sire of the cow are usually known. Therefore, in herds that rely heavily on AI, it is possible to foresee almost every potential mating of a daughter of a bull with a given undesirable haplotype to a service sire with the same haplotype. Right now, breeding companies and breed associations are modifying their mating programs to include an adjustment for the approximate economic loss associated with mating service sires that are carriers of BH1, HH1, HH2, HH3, or JH1 to genotyped cows (or daughters of genotyped bulls) that carry the same haplotype. As we learn more about these conditions, we can calculate precise estimates of the actual costs incurred when each haplotype is expressed in the homozygous state.

ReferencesCole, J. B., and P. M. VanRaden. 2011. Use of haplotypes to estimate Mendelian sampling effects and selection limits. Journal of Animal Breeding and Genetics (doi:10.1111/j.1439‐0388.2011.00922.x).

VanRaden, P. M., K. M. Olson, D. J. Null, and J. L. Hutchison. 2011. Detection of harmful recessive effects on fertility and stillbirth by absence of homozygous haplotypes. Journal of Dairy Science (in review).

A haplotype is a combination of alleles (DNA sequences) at adjacent locations on a chromosome that are inherited together. When a calf inherits a haplotype from both the sire and dam as a homozygous recessive, it may be stillborn or die soon after birth. In Europe, two calves that died shortly after birth have been discovered as homozygous for BH2. A list of BH2 carrier bulls is now available to breeders. The list is also published in this (November) issue of the Brown Swiss Bulletin and breeders are encouraged to utilize it when making mating decisions as to avoid mating two carrier animals. The BH2 Carrier list will be updated with each sire evaluation run.

Further research is being gathered in the U.S. and tests are in process to determine if stillborn calves that have been found as having carrier parents are homozygous for BH2. The ultimate goal is to identify this gene by working with USDA and worldwide research centers as to not duplicate research. Making efforts to avoid inbreeding when selecting matings will also assist in avoiding stillbirth, SMA, SDM, BH1, BH2 and other potential genetic defects that have not yet been discovered.

From New Fertility and stillbirth haplotypes and changes in haplotype status

By Paul Van Raden, Dan Null, Jana Hutchison and Tabatha Cooper (USDA-AIPL):

“The BH2 carrier frequency gradually increased from 4% before 1980 to 20% today. Nearly all carriers trace to U.S. Brown Swiss bull 144488 Rancho Rustic My Design born in 1963. Schwarzenbacher et al. (2012 EAAP meeting) discovered BH2 (which they labeled 19-1) on chromosome 19 at a range of 10.6-11.7 Mbase. The effect of BH2 is significant in both U.S. and Austrian data for stillbirth (not fertility), and the rate of calf loss is similar to the previously known defect SMA. No homozygotes for BH2 were found in U.S., Austrian, or Intergenomic Brown Swiss genotypes even though 29 were expected. Swiss and Austrian researchers recently identified 2 BH2 homozygous calves born with low birth weight that died from poor immune response, and are now using sequence data to locate the mutation causing the calf loss.”

Brown Swiss fertility haplotype BH14

Switzerland last year began reporting a new lethal haplotype in Brown Swiss (BH14) causing early pregnancy loss. Brown Swiss USA received a list of 1,254 carrier bulls from Switzerland and the Animal Genomics and Improvement Lab (AGIL) compared those to haplotype calls for BH14 from CDCB data for the same bulls. In both analyses the haplotype traced primarily to BSUSA175545 VENTURES ESP BABARAY (W) born in 1978, and no homozygotes were detected in CHE or USA analyses. Of the carrier bulls from the CHE file, 94% were also carriers from CDCB data, and 6% of the carriers from CHE data were non-carriers here. Of those 73 non-matching bulls, only 5 were USA bulls and those had no daughters. Likely almost all 43,000 non-carrier bulls matched, and if so, the concordance is very high for BH14. For some bulls, the CHE analysis may have included a gene test for the MRPL55 mutation on chromosome 7 at ARS-UCD1 location 2,996,436. Carrier frequency was 6.1% reported by Häfliger et al. (2021). In CDCB data, the number of genotyped carriers was 1,113 of 16,207 USA animals (6.9%) and across all countries was 2,960 animals of 59,127 genotyped (5.0%) as of April 2022. CDCB conception rate data included less than 100 matings of carrier sires to carrier maternal grandsires (the dam’s sire for the potential embryo being created). A conception loss of about 0.04 or 4% is expected if an extra 1 / 8 of embryos do not survive. The significant effect on fertility reported in CHE was not significant in USA because so few observed matings gave a standard error > 5%. Haplotype or gene tests from CHE may be reported initially, but routine publication of BH14 status in USA will allow selection and mating of males and females to further improve BS fertility. CDCB should begin reporting haplotype BH14 in the April 2023 Genetic Evaluations. The Board has directed that we will go forward with labeling of BH14 carriers.

by Sophie Eaglen for Progressive Dairy, Published on 24 February 2021

Do you ever feel like the age of genomics has made dairy cattle breeding more complicated? Or perhaps more scientific? While genomics brought us accelerated progress, it has also introduced us to a range of terms previously used by scientists alone.

“Haplotype” has become a household name used in bull catalogs and communication by evaluation centers. But it is probably one of the most misunderstood scientific terms in the industry. Often used in the same breath as “recessive,” “genetic defect” or “genetic condition,” haplotypes have gotten a bad reputation, which is not completely fair.

To correctly use this new information in genetic herd management, it’s good to explore what haplotypes are, what they are used for and if they really are all bad.

What is a haplotype?

The definition of a haplotype is very simple, and it has little to do with genetic defects. DNA consists of base pairs. A haplotype is simply a short section of DNA, or a sequence of those base pairs, passed on to the next generation. If you would cut up the strand of DNA into bits, each bit would be a haplotype if it is more than one base pair and the whole segment is inherited from either the mother or father (Figure 1).

In genomic selection, we do not read individual base pairs; instead we use genetic markers – most often single-nucleotide polymorphisms (SNPs) scattered across the genome. Therefore, when we refer to a haplotype for a genomically tested animal, we are talking about a sequence of SNPs, which signifies a piece of DNA inherited from either sire or dam.

Therefore, a haplotype is not a gene or even part of a gene. A haplotype can contain one or multiple genes – or none at all.

Are all haplotypes genetic defects?

No, a haplotype can contain a positive or negative genetic variant.

Our industry has inadvertently made the word “haplotype” synonymous to genetic defect, while it actually isn’t. This has given the word “haplotype” its negative reputation.

We use haplotypes as a handy tool to trace genes or genetic variants that cause the expression of a particular phenotype. In some cases, this is a desired trait, and in other cases it is detrimental or lethal.

Because genetic defects have such a large impact, they are more often discussed than desirable traits and, thereby, haplotypes are most often related to genetic defects. But haplotypes are also used to identify desirable traits such as polled and coat color.

So why do we talk about haplotypes and not genes?

There are multiple answers to this question, but one of the main reasons is: We simply do not know what genetic variant is causing the phenotype.

Our commercial genotypes mainly consist of SNPs, which means that unless the causal genetic variant is a specific SNP we have mapped, we do not know what causes the observed phenotype. When we have enough animals with the distinct phenotype, we can track where on the genome the variant likely is and dial in to a particular part of related DNA – the haplotype. We then know that this haplotype likely contains the causal variant, but we do not know what it looks like without genome sequencing and additional research.

This is often the reason why a genetic variant is first presented as a haplotype. Testing for the particular haplotype allows breeders to get a good sense of whether their animal may be a carrier for the new genetic variant.

If possible, researchers then sequence the DNA of affected animals and try to find the causal gene. When found, it is then a matter of getting the variant on the DNA chips used for genotyping, which can be a complicated process.

However, even when the genetic variant is known and a gene test is available, haplotype tests often remain offered because they present a much lower-cost way to test for carrier status. Also, if we were to add gene tests as soon as they were discovered by researchers, that would require frequent updates to commercial genotyping tests, which is not practical.

Why are haplotype tests not 100% accurate?

Especially for new genetic variants, haplotype tests are not 100% accurate.

Not all animals are genotyped, and not all animals are genotyped using high-density SNP chips. We use mathematics and pedigree data to predict and fill in SNP markers that we are missing in our haplotype of interest. Our haplotypes and the result of a haplotype test thereby depend partly on the quality of pedigree data and how many genotyped animals exist in that pedigree data. Data is added continuously, which means that a haplotype result can change when there is pedigree added or parentage was corrected in the lineage of the specific individual. In addition, new genetic variants may not have a large number of known carriers, which means the identified haplotype may be relatively long and thereby less exactly pinpointing the causal variant. As more carriers are discovered, we can dial in the exact coordinates of the haplotype and make our prediction more accurate.

Ideally, we would test every genetic variant with a gene test. This would remove much of the insecurity and frustration breeders experience. That said, this would also be a lot more costly for breeders. The haplotype test allows us to detect carriers fast and at almost no cost at a reasonable reliability level of 95% to 99%. As more animals are genomic tested and DNA arrays improve, reliability levels of haplotypes and the potential to detect genetic variants improves as well.

The slightly lower reliability level of haplotype tests is also the reason some breed associations choose not to place haplotype results on any official export documents. And it is the reason why it is possible that a haplotype test result differs from the gene test result when a gene test can be performed.

What is a lethal haplotype?

A lethal haplotype is most often a lethal recessive we are testing using a haplotype.

The HH1 to HH6 haplotypes in Holstein, JH1 in Jerseys and AH1 are examples of lethal recessive haplotypes. These genetic defects cause the embryo to die when two carriers are mated. Most often, this happens so early in gestation it simply seems that the animal has come back in heat. We therefore say these lethal recessive haplotypes affect fertility.

Are all lethal haplotypes equally bad?

All detected lethal recessive haplotypes have a proven effect on fertility. However, this effect is not equally bad, and the chance of this effect occurring is not equally high. The chance of a carrier mating depends on the frequency of the gene in the population. Each of the published lethal recessive haplotypes have a different allele frequency. For example, 1.92% of U.S. animals carry a copy of HH1. This frequency is 2.22% for HH5 and 0.37% for HH4. All lethal haplotype frequencies are published by the USDA and can be found online.

Should I stop using bulls with a carrier status for lethal haplotypes or recessives?

No, not necessarily. The chance of carrier matings is low for the majority of lethal haplotypes. To continue with the example of HH4, only 0.37% of U.S. Holstein animals carry a copy for HH4, and those animals have to be mated with each other for HH4 to be expressed. With the availability of genomic testing and a multitude of mating programs, carrier matings can easily be avoided.

Any genetic defects with large effect and high frequency get quickly taken care of by breeding programs from A.I. providers. Bulls are tested at a very young age and culled when carrying detrimental genetic defects. Pretty much all U.S. A.I. companies publish carrier statuses on every bull page for your information.

Unfortunately, not all A.I. companies worldwide publish carrier statuses of their bulls, and there is no international standard with how carrier statuses should be expressed either. This includes the denotation whether the result comes from a haplotype or gene test. When you are missing information or information is not clear, ask your A.I. provider how and for what the bull was tested.

The bottom line

Scientific jargon has entered our industry at a rate that makes it difficult to keep up on what everything means. When it comes to managing your herd, don’t be too scared of “haplotypes,” as their bad reputation is not entirely justified. Know that not all haplotypes are bad, and not all bad haplotypes are equally bad.

In addition, feel safe in knowing there are measures in place to detect any new detrimental genetic defects. Data on genetic variants is publicly available, and mating programs allow for easy avoidance of carrier matings of genetic defects. Also, feel free to ask if a genetic variant was tested using a haplotype test or gene test. When it comes to genetic management, it’s always good to know what exactly you are looking at and how it can be managed to get the best result for your herd.