Additional Breeding Research Articles

Season‐long characterization of high‐cannabinoid hemp (Cannabis sativa L.) reveals variation in cannabinoid accumulation, flowering time, and disease resistance

AbstractGiven the dramatic rise in high‐cannabinoid hemp (Cannabis sativa L.) production in the last decade, there is an increasingly urgent need to characterize available germplasm and develop knowledge to accelerate the breeding of uniform and stable cultivars. Despite persistent cultivation of hemp cultivars for grain and or fiber around the world, the diversity and genetic underpinning of key traits for breeding and cultivation are poorly understood. For 30 high‐cannabidiol hemp cultivars replicated on two field sites, we sought to evaluate yield, agronomic performance, and disease resistance while also conducting a detailed study of cannabinoid accumulation over the course of floral maturation. We observed significant variation in both within and among cultivars. During the growing season, the plants clustered into five groups by growth rate and varied in flowering time from photoperiod insensitive to photoperiod sensitive with very short critical photoperiods. Based on the observed ratio of total potential cannabidiol (CBD) to total potential tetrahydrocannabinol (THC), there was segregation for cannabinoid chemotype in some seeded cultivar populations. Analysis of cannabichromene (CBC) production revealed that some cultivars had a discretely lower CBD:CBC ratio than the others. There was a continuous range of powdery mildew severity by cultivar, with one that had little to no observed powdery mildew suggesting it might have genetic resistance. Biomass production at harvest was strongly influenced by location and cultivar, and there was variation by cultivar in the relative cannabinoid production in shoot tip samples compared with whole plant samples. While our results provide preliminary guidance regarding relative performance of current cultivars, our analyses indicate a need for additional hemp breeding to provide stable, uniform, and legally compliant cultivars with improved disease resistance and flowering times optimized for the latitudes of different growing locations.

Identification of Target Chicken Populations by Machine Learning Models Using the Minimum Number of SNPs.

Simple SummaryClassifying a target population at the genetic level can provide important information for the preservation and commercial use of a breed. In this study, the minimum number of markers was used in combination, to distinguish target populations based on high-density single nucleotide polymorphism (SNP) array data. Subsequently, a genome-wide association study for filtering target-population-specific SNPs between the case and control groups and principal component analysis with machine learning algorithms could be used to explore various combinations with the minimum number of markers. In addition, the optimal combination of SNP markers was able to produce stable results for the target population in verification studies, in which samples were analyzed.A marker combination capable of classifying a specific chicken population could improve commercial value by increasing consumer confidence with respect to the origin of the population. This would facilitate the protection of native genetic resources in the market of each country. In this study, a total of 283 samples from 20 lines, which consisted of Korean native chickens, commercial native chickens, and commercial broilers with a layer population, were analyzed to determine the optimal marker combination comprising the minimum number of markers, using a 600 k high-density single nucleotide polymorphism (SNP) array. Machine learning algorithms, a genome-wide association study (GWAS), linkage disequilibrium (LD) analysis, and principal component analysis (PCA) were used to distinguish a target (case) group for comparison with control chicken groups. In the processing of marker selection, a total of 47,303 SNPs were used for classifying chicken populations; 96 LD-pruned SNPs (50 SNPs per LD block) served as the best marker combination for target chicken classification. Moreover, 36, 44, and 8 SNPs were selected as the minimum numbers of markers by the AdaBoost (AB), Random Forest (RF), and Decision Tree (DT) machine learning classification models, which had accuracy rates of 99.6%, 98.0%, and 97.9%, respectively. The selected marker combinations increased the genetic distance and fixation index (Fst) values between the case and control groups, and they reduced the number of genetic components required, confirming that efficient classification of the groups was possible by using a small number of marker sets. In a verification study including additional chicken breeds and samples (12 lines and 182 samples), the accuracy did not significantly change, and the target chicken group could be clearly distinguished from the other populations. The GWAS, PCA, and machine learning algorithms used in this study can be applied efficiently, to determine the optimal marker combination with the minimum number of markers that can distinguish the target population among a large number of SNP markers.

Accounting for competition in multi-environment tree genetic evaluations: a case study with hybrid pines

Key message A novel multi-environment competition individual-tree mixed model resulted in better fit, and greater individual narrow- and broad-sense heritabilities than the model without competition, notably for traits showing competition. In multi-environment tests, the proposed model would allow tree breeders to select genotypes with the best performance in both additive direct and competition breeding values, increasing forest productivity. ContextGenetic merit of trees is known to be affected and interact with local competition effects as well as changes across environmental conditions. Recent studies showed that competition genetic effects can affect the genetic variance and bias the tree breeding values, and its covariance with direct breeding values has been variable across traits.AimsThe present paper extends a mixed-model methodology to the problem of accounting for competition in a multi-environment set of forest genetic trials and exploring its impact on genetic variances as well as the multi-environment genetic correlation.MethodsThe proposed model is illustrated using data from two full-sib trials of Pinus elliottii var. elliottii × Pinus caribaea var. hondurensis F1. Dispersion parameters and (co)variance of total breeding values were estimated for diameter at breast height, total tree height, and stem straightness at age 10.ResultsFor traits showing competition effects (diameter at breast height and total tree height), the proposed multi-environment competition model gave better fit than the simpler model. Accounting for competition increased the direct additive variance, reduced the residual variances, and did not change significantly the across-site additive genetic correlation. However, for diameter at breast height, top 5% best genetic rankings showed differences.ConclusionWhen traits are strongly affected by inter-tree competition, the use of the proposed model in multi-environment analyses can efficiently identify the phenomenon with general benefits in the fitting of genetic components and open the door to select on the basis of competitiveness.

Nasljeđivanje porodne mase kod križanaca Istarske, Awassi, Istočnofrizijske i Travničke pasmine ovaca u Hrvatskoj: studija sa jedne farme

The purpose of the study was to estimate the influence of the additive and non-additive genetic effects on the birth weight, using two different crossbreed models. Data on 1176 birth weights were collected. The animals were crosses, with different ratios of Awassi, East-Friesian, Istrian, and Travnik Pramenka sheep breed. The focal breed in this study was Istrian, which is an autochthonous breed in Croatia. In both models mixed procedure was used with random effect of sire and fixed effects of sex, year and month of birth, litter size, and direct inbreeding. The first model contained the additive breed effects, while the second model additionally included non-additive effects of heterosis and recombination loss. Sex, litter size, year and month of birth significantly affected birth weight in both models. Effect of direct inbreeding was negative and significant only in second model. Significant positive additive breed effect was estimated for Awassi breed in both models. The East-Friesian additive breed effect was positive in both models, but it was significant for the first one only. The positive and significant heterosis effect for the birth weight was estimated for the crosses between East-Friesian and Travnik Pramenka, as well as between Travnik Pramenka and Istrian breed. Same crosses showed significant and positive effect of recombination loss. The negative effect of recombination loss was estimated for the crosses of Awassi breed.

DDB2 Genetic Risk Factor for Ocular Squamous Cell Carcinoma Identified in Three Additional Horse Breeds.

Squamous cell carcinoma (SCC) is the most common cancer affecting the equine eye. A missense variant within the gene damage-specific DNA binding protein 2 (DDB2 c.1013C>T, p.Thr338Met) was previously identified as a causal recessive genetic risk factor for the development of ocular SCC within Haflingers, Belgian Draft horses, and Rocky Mountain Horses, but not in the Appaloosa or Arabian breeds. This study aimed to evaluate three cases of ocular SCC in additional breeds and determine if DNA testing for the DDB2 variant in warmblood horses and Connemara ponies is warranted. Histopathology confirmed ocular SCC in all three cases and DNA testing confirmed each horse was homozygous for the DDB2 risk factor. The DDB2 risk allele frequency was estimated to be 0.0043 for Holsteiners (N = 115), 0.014 for Belgian Warmbloods (N = 71), and 0.22 for Connemara Ponies (N = 86). Taken together these data support using DNA testing for DDB2 in Connemara Ponies to assist in mate selection and clinical management. Given the low observed allele frequencies in both the Holsteiner and Belgian Warmblood breeds and that the case under investigation was a warmblood cross-bred, evaluating additional SCC affected warmbloods is warranted to fully determine the importance of DDB2 genotyping as a risk factor in warmblood breeds.

СРАВНИТЕЛЬНАЯ ОЦЕНКА КОСТРОМСКОЙ И ЯРОСЛАВСКОЙ ПОРОД КРУПНОГО РОГАТОГО СКОТА ПО ГЕНУ БЕТА-КАЗЕИНА

Abstract. The article presents the results of a comparative assessment of the frequency of alleles and genotypes for the gene of beta-casein (β-CN) in cattle of the Kostroma and Yaroslavl breeds. The data on 137 bulls-producers of comparable and related breeds, presented on the official sites of “OOO Head Center for the Reproduction of Farm Animals” and the breeding farm “OOO Yaroslavskoe”, have been analyzed. The blood of cows from the Kostroma and Yaroslavl breeds in the breeding farms of the Kostroma region served as the object of DNA research. A total of 80 animals were genotyped. Evaluation of the distribution of the desired genotype β-CNА2/А2 in stud bulls showed that two breeds are in the lead – the Schwyz and the Kostroma breed, with the frequency of occurrence – 0.9090 and 0.6250, respectively. Moreover, in the Kostroma breed, there are no bulls carriers of the homozygous β-CNА1/А1 genotype. If we take into account the frequency of distribution of the β-CNА2 allele in bulls of the analyzed breeds, then it is higher – in the Schwyz breed – 0.9091, Kostroma – 0.8125 and Holstein – 0.6776. In bulls of the Yaroslavl breed there is 0.2083 frequency of the β-CNА2/А2 genotype and 0.4167 frequency of the β-CNА2 allele. A comparative assessment of the polymorphism data of the beta-casein locus in cows of the Yaroslavl and Kostroma breeds showed that in the Yaroslavl breed there were least of all animals with the β-CNА2/А2 genotype – 0.0333 and more – with the β-CNА1/А1 genotype – 0.6667. In the Kostroma breed, the genetic situation is completely different. So cows with genotypes β-CNА2/А2 – 0.5490, and with genotype β-CNА1/А1 – 0.0785. However, it is necessary to take into account the main indicators of cattle productivity in order to consider the beta-casein genotype as an additional selection trait. It is not possible to study the milk productivity of Yaroslavl breed cows with different genotypes, since only one cow with the CNА2/А2 genotype has been identified. However, cows of the Kostroma breed homozygous for the CNА2 allele, for the third and older lactations, had a milk yield of 1534 kg higher than cows homozygous for the CNА1 allele. Consequently, the beta-casein genotype can be considered an additional promising breeding trait.

Population structure, genetic diversity, and selective signature of Chaka sheep revealed by whole genome sequencing

BackgroundChaka sheep, named after Chaka Salt Lake, are adapted to a harsh, highly saline environment. They are known for their high-grade meat quality and are a valuable genetic resource in China. Furthermore, the Chaka sheep breed has been designated a geographical symbol of agricultural products by the Chinese Ministry of Agriculture.ResultsThe genomes of 10 Chaka sheep were sequenced using next-generation sequencing, and compared to that of additional Chinese sheep breeds (Mongolian: Bayinbuluke and Tan; Tibetan: Oula sheep) to explore its population structure, genetic diversity and positive selection signatures. Principle component analysis and a neighbor-joining tree indicated that Chaka sheep significantly diverged from Bayinbuluke, Tan, and Oula sheep. Moreover, they were found to have descended from unique ancestors (K = 2 and K = 3) according to the structure analysis. The Chaka sheep genome demonstrated comparable genetic diversity from the other three breeds, as indicated by observed heterozygosity (Ho), expected heterozygosity (He), runs of homozygosity (ROH), linkage disequilibrium (LD) decay. The enrichment analysis revealed that in contrast to Mongolian or Tibetan lineage groups, the genes annotated by specific missense mutations of Chaka sheep were enriched with muscle structure development (GO:0061061) factors including insulin-like growth factor 1 (IGF1), growth differentiation factor 3 (GDF3), histone deacetylase 9 (HDAC9), transforming growth factor beta receptor 2 (TGFBR2), and calpain 3 (CAPN3), among others. A genome-wide scan using Fst and XP-CLR revealed a list of muscle-related genes, including neurofibromin 1 (NF1) and myomesin 1 (MYOM1), under potential selection in Chaka sheep compared with other breeds.ConclusionsThe comprehensive genome-wide characterization provided the fundamental footprints for breeding and management of the Chaka sheep and confirmed that they harbor unique genetic resources.

Intrabreed and interbreed variation of the BOLA-DRB3.2 gene in the Kostroma and Yaroslavl indigenous Russian cattle breeds

Intrabreed and interbreed variation of BOLA-DRB3 exon 2 (BOLA-DRB3.2) was for the first time studied in the Kostroma and Yaroslavl cattle breeds by PCR-RFLP. These breeds are among the best Russian breeds and were developed as dairy-beef and dairy cattle, respectively. Twenty-nine alleles were observed in five Kostroma samples, and 14 of them proved unique in comparison with two Yaroslavl samples, in which 25 alleles were detected, and 10 of them were unique. The total frequency of bovine leukemia virus (BLV) resistance alleles (*11, *23, and *28) was 23.2% in the Kostroma, while the total frequency of BLV susceptibility alleles (*8, *16, *22, *24) was low, 8.4%. The frequencies were 25.8 and 30.1%, respectively, in Yaroslavl cattle. Testing Hardy-Weinberg equilibrium revealed a significant deficit of heterozygotes: the observed (Ho) and expected (He) heterozygosities were, respectively, 0.734 and 0.859 in Kostroma cattle and 0.613 and 0.886 in Yaroslavl cattle. The intrabreed differentiation (FST) in the Kostroma (4.5%, P = 0.001) was substantially higher than in the Yaroslavl (0.5%, P = 0.158), between the two breeds was 8.2% (P = 0.001). The Bayesian clustering approach showed an intrabreed structure for each of the breeds, with the most probable number of clusters being 2 in the Kostroma and 3 in the Yaroslavl. The structure observed in the Kostroma remained the same when the breed was analyzed together with six additional breeds. Our data provide important clues toward the understanding of the genetic structure of indigenous breeds.

Spatial and competition models increase the progeny testing efficiency of Japanese larch

The main purpose of this study was to examine spatial and competition effects on estimates of genetic parameters, as well as on selection options for growth traits, including height (H), diameter at breast height (DBH), and volume (V), in a progeny test of Japanese larch (Larix kaempferi (Lam.) Carrière) at age 20 years. We compared performances among the individual-tree additive genetic base model (B) with design factors only, the spatial effect model (AR1), the competition model (C), and the combined competition and spatial model (CS). We found that spatial heterogeneity had significant effects on growth traits and that plot variance decreased by more than 80% in the AR1 model relative to the B model. Competition had significant effects on DBH and V but a smaller effect on H. In the C model, direct additive genetic variances ([Formula: see text]) for DBH and V increased by 205% and 93%, respectively, whereas residual variances ([Formula: see text]) decreased by 8% and 6%, respectively. In the CS model, the correlations between direct and competitive genetic effects were 0.83, −0.97, and −0.98 for H, DBH, and V, respectively. Competition significantly affected the forward selection. The proportions of selected elite trees were only 39% and 25% common between the B and CS models for DBH and V, respectively, when selection intensity was 5%. For breeding selection, depending on thinning regimes planned, trees of high additive breeding values but low competitive breeding values are preferable for plantation.

Inbreeding affects racing performance negatively in the Standardbred trotter

ABSTRACT We estimated the effect of inbreeding on racing performance in the Standardbred trotter. A random sample of 1,217 horses with organized pedigree was utilized to calculate individual proportions of French and American ancestry and their heterozygosity and inbreeding coefficients. Average earnings per start for the 807 raced horses summarized between three and five years of age were analysed, after first applying a fourth-root transformation followed by standard-normalization within their birth-year group (1988 and 2006/2007). The resulting variable was analysed using a linear model with fixed effects of sex, additive breed effect and a regression on the squared inbreeding coefficient. The model was expanded with a fixed effect of birth-year and a random effect of animal. In both models, the regression coefficient for the squared inbreeding coefficient was found negative (P < 0.04). Breed effect was not significant. Heritability was estimated as 0.227 ± 0.106.

Western Canadian producers’ attitudes towards wheat breeding funding

AbstractIn 2017, the federal government initiated national consultations for two new crop royalty systems that could be used to support additional crop breeding. In this study, we examine wheat growers’ attitudes towards breeding research and assess their inclination to contribute more to wheat variety development through checkoffs or enhanced royalties. We report a random effect probit estimation for a survey of 877 western Canadian wheat producers that took place from November 2018 to January 2019. We found at least 26% of survey respondents were willing to pay more to support additional wheat breeding. However, this support is contingent on the model for revenue collection and where additional revenue is invested. Producers generally favored increased checkoffs over enhanced royalty collection. Among the royalty options presented, the farm saved seed royalties mechanism had less support than the simpler to implement end‐point royalties mechanism. We also found support is much higher if new royalty mechanisms are used to support university or government programs versus private breeding programs. This result suggests developing widespread producer support for enhanced royalty collection may require broader commitments for funding, ownership, and control of crop breeding programs.

Identification of the novel polymorphisms and potential genetic features of the prion protein gene (PRNP) in horses, a prion disease-resistant animal

Prion diseases, a protein misfolded disorder (PMD) caused by misfolded prion protein (PrPSc), present in a wide variety of hosts, ranging from ungulates to humans. To date, prion infections have not been reported in horses, which are well-known as prion disease-resistant animals. Several studies have attempted to identify distinctive features in the prion protein of horses compared to prion disease-susceptible animals, without the study on polymorphisms of the horse prion protein gene (PRNP). Since single nucleotide polymorphisms (SNPs) of PRNP in prion disease-susceptible animals are major susceptibility factors, the investigation of SNPs in the horse PRNP gene is important; however, only one study investigated a single horse breed, Thoroughbred. Thus, we investigated genetic polymorphisms and potential characteristics of the PRNP gene in 2 additional horse breeds. To this end, we performed amplicon sequencing of the horse PRNP gene and investigated SNPs in Jeju and Halla horses. We compared genotype, allele and haplotype frequencies among three horse breeds, namely, Thoroughbred, Jeju and Halla horses. In addition, we evaluated the potential influence of the identified nonsynonymous SNPs on the prion protein using PolyPhen-2, PROVEAN, and PANTHER. Furthermore, we measured the aggregation propensity of prion proteins using AMYCO and analyzed linkage disequilibrium (LD) between PRNP and prion-like protein gene (PRND) SNPs. A total of 4 SNPs were found, including two nonsynonymous SNPs (c.301 T > A, c.525 C > A) and three novel SNPs (c.-3A > G, c.301 T > A and c.570 G > A). There were significant differences in genotype, allele and haplotype frequencies among the three horse breeds. The nonsynonymous SNP, c.301 T > A (W101R), was predicted to be benign, deleterious, and possibly damaging by PolyPhen-2, PROVEAN and PANTHER, respectively. In addition, the amyloid propensity of horse prion protein according to 4 haplotypes of nonsynonymous SNPs was predicted to be benign by AMYCO. Finally, we identified weak LD between PRNP and PRND SNPs.

Engineered Durum Wheat Germplasm with Multiple Alien Introgressions: Agronomic and Quality Performance

If genetic gains in wheat yield are to be achieved in today’s breeding, increasing the genetic variability of cultivated genotypes is an essential requisite to meet. To this aim, alien gene transfer through chromosome engineering (CE) is a validated and sound strategy. Attempts to incorporate more than one alien segment into cultivated wheat have been rare, particularly for tetraploid durum wheat. Here, we present the agronomic and quality performance of the first successful CE-mediated multiple introgression into the latter species. By assembling into 7AL, 3BS, and 1AS arms of a single genotype homoeologous segments of Thinopyrum ponticum 7el1L, Aegilops longissima 3SlS, and Triticum aestivum 1DS arms, respectively, we have stacked several valuable alien genes, comprising Lr19+Sr25+Yp (leaf and stem rust resistance and a gene increasing semolina yellowness), Pm13 (powdery mildew resistance), and Gli-D1/Glu-D3 (genes affecting gluten properties), respectively. Advanced progenies of single, double, and triple recombinants were field-tested across three years in a typical durum wheat growing area of central Italy. The results showed that not only all recombinants had normal phenotype and fertility, but also that one of the triple recombinants had the highest yield through all seasons compared with all other recombinants and control cultivars. Moreover, the multiple introgressions enhanced quality traits, including gluten characteristics and semolina yellow index. The presence of effective disease resistance genes confers additional breeding value to the novel and functional CE products, which can greatly contribute to crop security and safety.

Gamevar.f90: a software package for calculating individual gametic diversity

BackgroundTraditional selection in livestock and crops focuses on additive genetic values or breeding values of the individuals. While traditional selection utilizes variation between individuals, differences between gametes within individuals have been less frequently exploited in selection programs. With the successful implementation of genomic selection in livestock and crops, estimation and selection for gametic variation is becoming possible.ResultsThe gamevar.f90 software is designed to estimate individual-level variance of genetic values of gametes for complex traits in large populations. The software estimates the (co)variances of gametic diversity as well as other diversity parameters that are useful for selection programs and mating designs. The calculation is carried out chromosome by chromosome and can be easily parallelized. The gamevar.f90 program is written in Fortran with efficient computing algorithms in a user-friendly software package with easily-handled input and output files. Finally, we applied the program to estimate gametic variance for hundreds of bulls for lifetime net merit, productive life, and livability. The RPTA (relative predicted transmitting ability), assuming a future selection intensity (if) of 1.5, showed larger variance than GEBV/2, indicating that greater future genetic gains can be obtained using an index that includes gametic variances. We also used the relative coefficient of variation to estimate with 95% confidence the sample sizes required to observe 90% variability of the progeny for lifetime net merit (or to allow at maximum 10% of change in the EBV predicted from progeny data).ConclusionsCollectively, we develop an efficient computer program package, gamevar.f90, for estimating gametic variance for large numbers of individuals. The novel information on gametic variation will be useful in future animal and crop breeding programs.

Nine months in the Simpson Desert: The anatomy of a letter-winged kite breeding irruption

Highly mobile animals in the world's drylands can have distinct core and irruptive ranges with the latter being occupied only during population outbreaks driven by resource pulses. Breeding events occur within the irruptive range during these periods of high resource availability. Here we quantify such an event involving the letter-winged kite (Elanus scriptus) in the Simpson Desert, Australia; a region that experiences major rainfall-driven pulses in primary productivity. The irruption of kites lasted for a minimum of nine months (12 July 2016 to 24 May 2017). Breeding was confined to July–October 2016. Population size rose from an initial count of 22 adults in July to a peak of 38 adults and 19 juveniles in October 2016. The pattern of aggregation during 2016 consisted of dispersed breeding individuals whereas by 2017 this pattern was replaced by larger non-breeding aggregations. Kites arrived in the area when small mammal abundance was relatively low, but was increasing, and bred as the abundance of small mammals reached its peak. Unexpectedly, kites departed the area when abundance of small mammals was still high and sufficient to support an additional breeding event in winter-spring 2017. Our study provides insights in to the duration and size of breeding events in the irruptive range of nomadic birds. From a management perspective it identifies the importance of the keystone tree species, Acacia peuce, as a nesting site.

Unexpectedly high genetic diversity in a rare and endangered seabird in the Hawaiian Archipelago.

Seabirds in the order of Procellariiformes have one of the highest proportions of threatened species of any avian order. Species undergoing recovery may be predicted to have a genetic signature of a bottleneck, low genetic diversity, or higher rates of inbreeding. The Hawaiian Band-rumped Storm Petrel (‘Akē‘akē; Hydrobates castro), a long-lived philopatric seabird, suffered massive population declines resulting in its listing under the Endangered Species Act in 2016 as federally Endangered. We used high-throughput sequencing to assess patterns of genetic diversity and potential for inbreeding in remaining populations in the Hawaiian Islands. We compared a total of 24 individuals, including both historical and modern samples, collected from breeding colonies or downed individuals found on the islands of Kaua‘i, O‘ahu, Maui, and the Big Island of Hawai‘i. Genetic analyses revealed little differentiation between breeding colonies on Kaua‘i and the Big Island colonies. Although small sample sizes limit inferences regarding other island colonies, downed individuals from O‘ahu and Maui did not assign to known breeding colonies, suggesting the existence of an additional distinct breeding population. The maintenance of genetic diversity in future generations is an important consideration for conservation management. This study provides a baseline of population structure for the remaining nesting colonies that could inform potential translocations of the Endangered H. castro.

Runs of Homozygosity and NetView analyses provide new insight into the genome-wide diversity and admixture of three German cattle breeds.

Angler (RVA) and Red-and-White dual-purpose (RDN) cattle were in the past decades crossed with influential Red Holstein (RH) sires. However, genome-wide diversity studies in these breeds are lacking. The objective of the present study was to elucidate the genome-wide diversity and population structure of the three German cattle breeds. Using 40,851 single nucleotide polymorphism markers scored in 337 individuals, runs of homozygosity (ROH) were analysed in each breed. Clustering and a high-resolution network visualisation analyses were performed on an extended dataset that included 11 additional (outgroup) breeds. Genetic diversity levels were high with observed heterozygosity above 0.35 in all three breeds. Only RVA had a recent past effective population size (Ne) estimate above 100 at 5 generations ago. ROH length distribution followed a similar pattern across breeds and the majority of ROH were found in the length class of >5 to 10 Mb. Estimates of average inbreeding calculated from ROH (FROH) were 0.021 (RVA), 0.045 (RDN) and 0.053 (RH). Moderate to high positive correlations were found between FROH and pedigree inbreeding (FPED) and between FROH and inbreeding derived from the excess of homozygosity (FHOM), while the intercept of the regression of FROH on FPED was above zero. The population structure analysis showed strong evidence of admixture between RVA and RH. Introgression of RDN with RH genes was minimally detected and for the first time, the study uncovered Norwegian Red Cattle ancestry in RVA. Highly heterogeneous genetic background was found for RVA and RH and as expected, the breeds of the extended dataset effectively differentiated mostly based on geographical origin, validating our findings. The results of this study confirm the impact of RH sires on RVA and RDN populations. Furthermore, a close monitoring is suggested to curb further reduction of Ne in the breeds.

Short communication: A splice site mutation in CENPU is associated with recessive embryonic lethality in Holstein cattle

A genome scan for homozygous haplotype deficiency coupled with whole-genome sequence data analysis is a very effective method to identify embryonic lethal mutations in cattle. Among other factors, the power of the approach depends on the availability of a greater amount of genotyping and sequencing data. In the present study, we analyzed the largest known panel of Illumina BovineSNP50 (Illumina Inc., San Diego, CA) genotypes, comprising 401,896 Holstein animals, and we report the mapping of a new embryonic lethal haplotype on chromosome 27, called HH7. We fine mapped the locus in a 2.0-Mb interval using an identical-by-descent approach and analyzed genome sequence data from 4 carrier and 143 noncarrier Holstein bulls to identify the causative mutation. We detected a strong candidate variant in the gene encoding centromere protein U (CENPU), a centromere component essential for proper chromosome segregation during mitosis. The mutant allele is a deletion of 4 nucleotides located at position +3 to +6 bp after the splicing donor site of exon 11. Cross-species nucleotide alignment revealed that the nucleotide at position +3 is entirely conserved among vertebrates, suggesting that it plays an important role in the regulation of CENPU splicing. For verification, we genotyped the candidate variant in 232,775 Holstein individuals and did not observe any homozygotes, whereas 16 were expected (Poisson P-value = 1.1 × 10-7; allele frequency = 0.8%). In addition, genotyping of 250,602 animals from 19 additional breeds revealed that the mutant allele is restricted to animals of Holstein descent. Finally, we estimated the effect of the candidate variant on 2 fertility traits in at-risk mating (i.e., between carrier bulls and daughters of carrier bulls) versus non-risk mating. In agreement with a recessive lethal inheritance pattern, we observed a marked reduction in both conception rate and 56-d nonreturn rate in heifers and cows. The effect on 56-d nonreturn rate suggests that a substantial proportion of homozygous mutants die before 35 d after insemination, which is consistent with the early embryonic death previously reported in CENPU-/- mouse embryos. In conclusion, we demonstrate that with more than 400,000 genotypes, we can map very rare recessive lethal mutations segregating at a frequency below 1% in the population. We recommend performing new analyses regularly as data are accumulating.

Frameshift Variant in MFSD12 Explains the Mushroom Coat Color Dilution in Shetland Ponies.

Mushroom is a unique coat color phenotype in Shetland Ponies characterized by the dilution of the chestnut coat color to a sepia tone and is hypothesized to be a recessive trait. A genome wide association study (GWAS), utilizing the Affymetrix 670K array (MNEc670k) and a single locus mixed linear model analysis (EMMAX), identified a locus on ECA7 for further investigation (Pcorrected = 2.08 × 10−10). This locus contained a 3 Mb run of homozygosity in the 12 mushroom ponies tested. Analysis of high throughput Illumina sequencing data from one mushroom Shetland pony compared to 87 genomes from horses of various breeds, uncovered a frameshift variant, p.Asp201fs, in the MFSD12 gene encoding the major facilitator superfamily domain containing 12 protein. This variant was perfectly concordant with phenotype in 96 Shetland Ponies (P = 1.15 × 10−22), was identified in the closely related Miniature Horse for which the mushroom phenotype is suspected to occur (fmu = 0.02), and was absent in 252 individuals from seven additional breeds not reported to have the mushroom phenotype. MFSD12 is highly expressed in melanocytes and variants in this gene in humans, mice, and dogs impact pigmentation. Given the role of MFSD12 in melanogenesis, we propose that p.Asp201fs is causal for the dilution observed in mushroom ponies.

Limpkin, Aramus guarauna (L., 1766) (Gruiformes, Aramidae), extralimital breeding in Louisiana is associated with availability of the invasive Giant Apple Snail, Pomacea maculata Perry, 1810 (Caenogastropoda, Ampullariidae)

We document the first breeding record of Limpkin, Aramus guarauna (Linnaeus, 1766) (Gruiformes, Aramidae), for Louisiana, describe an additional unpublished breeding record from Georgia, as well as a possible record from Alabama, and associate these patterns with the concurrent establishment of the invasive Giant Apple Snail, Pomacea maculata Perry, 1810 (Caenogastropoda, Ampullariidae). We predict that an invasive prey species may facilitate range expansion by native predator species, which has ramifications for conservation and management.