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Proceedings of the SMBE Tri-National Young Investigators' Workshop 2005. Genome-wide associations between hybrid sterility QTL and marker transmission ratio distortion.

Marker transmission ratio distortion (TRD) in genetic mapping populations is frequently ascribed to selection against allelic combinations that cause hybrid incompatibility. Accordingly, genomic regions of TRD should be nonrandomly associated (colocated) with loci that underlie hybrid incompatibility. To directly test this hypothesis, we evaluated the genome-wide qualitative and quantitative agreement between chromosomal regions exhibiting marker TRD and those known to contain hybrid incompatibility quantitative trait locus (QTL). Incompatibility data came from a near-isogenic line (NIL) analysis of pollen and seed sterility in a cross between two Solanum (formerly Lycopersicon) species. We assessed (1) whether these incompatibility loci are colocated with markers that show significant TRD in two earlier generations preceding these introgression lines and (2) whether the magnitude of marker distortion quantitatively matches the estimated strength of selection against each incompatibility locus. We found evidence that TRD regions are chromosomally colocated with hybrid incompatibility loci more frequently than is expected by chance: pollen sterility QTLs were most closely associated with distorted heterozygote frequencies in later-generation backcrosses. Nonetheless, there was no evidence for an association between TRD and seed sterility and little evidence of a quantitative association between the magnitude of marker TRD and the fitness effects of heterospecific alleles at each chromosomal location. We propose and test a model (the "dance partner" model) to explain several cases where regions of TRD are not associated with hybrid incompatibility loci. Under this model, some NILs containing greater than one heterospecific introgression may not express hybrid incompatibility phenotypes because they carry both appropriate genetic dance partners required for a fully functional interaction. Accordingly, negative interactions expressed in earlier backcross generations are masked in these double-introgression NILs. Based on this model, we identify the location of several new putative pairwise interactors underlying hybrid incompatibility in this species cross.

Chromosome Mapping↗

Recent evolution of the human pathogen Cryptococcus neoformans by intervarietal transfer of a 14-gene fragment.

The availability of the whole-genome sequence from the 2 known varieties of the human pathogenic fungus Cryptococcus neoformans provides an opportunity to study the relative contribution of divergence and introgression during the process of speciation in a genetically tractable organism. At the genomic level, these varieties are nearly completely syntenic, share approximately 85-90% nucleotide identity, and are believed to have diverged approximately 18 MYA. Via a comparative genomic approach, we identified a 14-gene region (approximately 40 kb) that is nearly identical between the 2 varieties that resulted from a nonreciprocal transfer event from var. grubii to var. neoformans approximately 2 MYA. The majority of clinical and environmental var. neoformans strains from around the world contain this sequence obtained from var. grubii. This introgression event likely occurred via an incomplete intervarietal sexual cycle, creating a hybrid intermediate where mobile elements common to both lineages mediated the exchange. The subsequent duplication in laboratory strains of a fragment of this same genomic region supports evolutionary theories that instabilities in subtelomeric regions promote adaptive evolution through gene amplification and subsequent adaptation. Along with a more ancient predicted transfer event in C. neoformans and a recently reported example from Saccharomyces cerevisiae, these data indicate that DNA exchange between closely related sympatric varieties or species may be a recurrent theme in the evolution of fungal species. It further suggests that although evolutionary divergence is the primary force driving speciation, rare introgression events also play a potentially important role.

Chromosome Mapping↗

Interspecies transfer of female mitochondrial DNA is coupled with role-reversals and departure from neutrality in the mussel Mytilus trossulus.

Mussels of the genus Mytilus have distinct and highly diverged male and female mitochondrial DNA (mtDNA) genomes with separate routes of inheritance. Previous studies of European populations of Mytilus trossulus demonstrated that 33% of males are heteroplasmic for a second mtDNA genome of increased length and that hybridization with Mytilus edulis does not block mtDNA introgression, in contrast to reports for American populations. Here, we demonstrate that the female mtDNA type of M. edulis has replaced the resident female mtDNA type of European M. trossulus. This is supported by COIII sequence data indicating that the female mtDNA of European M. trossulus is very similar to that of M. edulis and that in phylogenetic trees, the mtDNAs of these two species cluster together but separately from American M. trossulus sequences, the latter not being disturbed by introgressive hybridization. We also provide evidence that the mtDNA genome of increased length found in heteroplasmic males of European M. trossulus derives from a recent partition of an introgressed M. edulis female type into the male route of transmission. Neutrality tests reveal that European populations of M. trossulus display an excess of replacement polymorphism within the female mtDNA type with respect to conspecific American populations, as well as a significant excess of rare variants, of a similar magnitude to those previously reported for the invading European M. edulis mtDNA. Results are consistent with a nearly neutral model of molecular evolution and suggest that selection acting on European M. trossulus mtDNA is largely independent of the nuclear genetic background.

Americas↗

Genetic mapping of a wide spectrum nematode resistance gene (Hero) against Globodera rostochiensis in tomato.

The Hero gene confers resistance to a wide spectrum of pathotypes of the potato cyst nematode Globodera rostochiensis. This gene has been introgressed from the wild tomato species Lycopersicon pimpinellifolium into the cultivated tomato. We have used RFLP and RAPD analysis for the targeted search of the L. pimpinellifolium into the cultivated tomato. We have used RFLP and RAPD analysis for the targeted search of the L. pimpinellifolium segment. The resistant line LA 1792 contains a single introgressed segment on chromosome 4, which is characterized by three RFLP markers from the high-density RFLP map of tomato. The map position of the Hero gene in large populations, four additional markers were identified in the introgressed region. After analyzing more than 800 gametes for recombination, we found that one marker is only 0.4 cM away from the Hero gene. YAC clones isolated from a region near the Hero gene indicate that in this area of the genome, the kb/cM ratio is relatively low (<450 kb/cM) and chromosome walking should be feasible in order to isolate this gene.

Animals↗

Origin, radiation, dispersion and allopatric hybridization in the chub Leuciscus cephalus.

The phylogenetic relationships of 492 chub (Leuciscus cephalus) belonging to 89 populations across the species' range were assessed using 600 base pairs of cytochrome b. Furthermore, nine species belonging to the L. cephalus complex were also analysed (over the whole cytochrome b) in order to test potential allopatric hybridization with L. cephalus sensu stricto (i.e. the chub). Our results show that the chub includes four highly divergent lineages descending from a quick radiation that took place three million years ago. The geographical distribution of these lineages and results of the nested clade analysis indicated that the chub may have originated from Mesopotamia. Chub radiation probably occurred during an important vicariant event such as the isolation of numerous Turkish river systems, a consequence of the uplift of the Anatolian Plateau (formerly covered by a broad inland lake). Dispersion of these lineages arose from the changes in the European hydrographic network and, thus, the chub and endemic species of the L. cephalus complex met by secondary contacts. Our results show several patterns of introgression, from Leuciscus lepidus fully introgressed by chub mitochondrial DNA to Leuciscus borysthenicus where no introgression at all was detected. We assume that these hybridization events might constitute an important evolutionary process for the settlement of the chub in new environments in the Mediterranean area.

Animals↗

Glacial refugia and reticulate evolution: the case of the Tasmanian eucalypts.

Tasmania is a natural laboratory for investigating the evolutionary processes of the Quaternary. It is a large island lying 40-44 degrees S, which was repeatedly glaciated and linked to southeastern continental Australia during the Quaternary. Climate change promoted both the isolation of species in glacial refugia, and an exchange between Tasmanian and mainland floras. Eucalyptus is a complex and diverse genus, which has increased in abundance in Australia over the past 100 kyr, probably in response to higher fire frequency. Morphological evidence suggests that gene flow may have occurred between many eucalypt species after changes in their distribution during the Quaternary. This paper summarizes recent genetic evidence for migration and introgressive hybridization in Tasmanian Eucalyptus. Maternally inherited chloroplast DNA reveals a long-term persistence of eucalypts in southeastern Tasmanian refugia, coupled with introgressive hybridization involving many species. Detailed analysis of the widespread species Eucalyptus globulus suggests that migration from mainland Australia was followed by introgression involving a rare Tasmanian endemic. The data support the hypothesis that changes in distribution of interfertile species during the Quaternary have promoted reticulate evolution in Eucalyptus.

Climate↗

Sexual selection on plumage and behavior in an avian hybrid zone: experimental tests of male-male interactions.

In western Panama, an unusual hybrid zone exists between white-collared manakins, Manacus candei, and golden-collared manakins, M. vitellinus. Unidirectional introgression of plumage traits from vitellinus into candei has created a region in which all definitively plumaged males have a collar that is lemon-colored. These males are nearly indistinguishable from white-collared candei genetically and morphometrically, but strongly resemble golden-collared vitellinus due to the introgression of secondary sexual plumage traits, particularly the lemon-colored collar. The introgression could be explained by sexual selection for golden-collared traits or by a series of mechanisms that do not invoke sexual selection (e.g., neutral diffusion, dominant allele). Sexual selection on male-male interactions implies behavioral differences among the plumage forms--specifically that golden- and lemon-collared males should be more aggressive than white-collared males. In contrast, the nonsexual hypotheses predict behavioral similarity between lemon- and white-collared males, based on their nearly identical genetics. We tested the sexual selection hypothesis experimentally, by presenting males with taxidermic mounts of the three forms. As response variables, we monitored vocalizations and attacks on the mounts by replicate subject males. Both golden-collared and lemon-collared males were more likely to attack than were white-collared males, as predicted under sexual selection but not by the nonsexual hypotheses. Lemon-collared males were more vocally reactive than either parental form, contrary to the prediction of the nonsexual hypotheses. Our study demonstrates that sexual selection on male-male interactions may play an important role in the dynamics of character evolution and hybrid zones.

Aggression↗

Accumulating Dobzhansky-Muller incompatibilities: reconciling theory and data.

Theoretical models of the accumulation of Dobzhansky-Muller incompatibilities (DMIs) are studied, and in particular, the framework introduced by Orr (1995) and a verbal model introduced by Kondrashov et al. (2002). These models embody very different assumptions about the relationship between the substitution process underlying evolutionary divergence and the formation of incompatibilities. These differences have implications for our ability to make inferences about the divergence from patterns in the relevant data. With this in mind, the models are investigated for their ability to account for three patterns evident in this data: (1) the asymmetrical nature of incompatibilities under reciprocal introgression; (2) the finding that multiple concurrent introgressions may be necessary for an incompatibility to form; and (3) the finding that the probability of obtaining an incompatibility by introgressing a single amino acid remains roughly constant over a wide range of genetic distances. None of the models available in the literature can account for all of the empirical patterns. However, modified versions of the models can do so. Ways of discriminating between the different models are then discussed.

Alleles↗

Hybrid zone structure and the potential role of selection in hybridizing populations of native westslope cutthroat trout (Oncorhynchus clarki lewisi) and introduced rainbow trout (O. mykiss).

Introgressive hybridization is a common feature of many zones of contact between divergent lineages of fishes. This is particularly common when taxa that are normally allopatric come into artificial (human-induced) secondary contact. We examined 18 native populations of westslope cutthroat trout (Oncorhynchus clarki lewisi, WCT) to determine the extent of introgressive hybridization with introduced rainbow trout (O. mykiss, RBT) and the genetic structure of hybridizing populations in the upper Kootenay River, southeastern British Columbia, Canada. Using four diagnostic nuclear loci we calculated a hybrid index, inbreeding coefficient, FIS, and the linkage disquilibrium correlation coefficient, Rij, for each locality to determine the distribution of genotypes in each population. We also categorized the 142 hybrid individuals found across localities into four hybrid classes based on their genotypes. The majority of localities (11/18) showed a unimodal distribution of genotypes skewed towards genotypes of WCT. Two localities, however (lower Gold Creek and Lodgepole Creek) showed a flat to bimodal distribution and one site (lower Bull River) showed a unimodal distribution skewed towards RBT genotypes. The majority of hybrid individuals were classified genotypically as WCT backcrosses (59%) and post-F1 individuals (24%). We found a skewed ratio of pure WCT to pure RBT (17:1) and only four F1 hybrids (3%), suggesting that the spread of RBT alleles may be facilitated by hybrids straying to neighbouring populations. We also tested for the action of selection in one population using cohort analyses, but found little evidence of differential selection between pure WCT and hybrid individuals. Pooled across age classes there were significant differences in genotypic frequencies among loci suggesting differential introgression. There was no asymmetry to the hybridization between rainbow trout and westslope cutthroat trout because both species' mitochondrial DNA haplotypes were observed at similar frequencies in the hybrids. Our analyses suggest that hybrid swarms are likely to form in the upper Kootenay River drainage and that certain native WCT populations in British Columbia are at risk of local genomic extinction.

Age Factors↗

Fertility, segregation at a herbicide-resistance locus, and genome structure in BC hybrids from two important weedy Amaranthus species.

Field studies have established high potential for hybridization between two important and often coexisting weedy species, Amaranthus hybridus and Amaranthus tuberculatus. Prezygotic reproductive barriers between these species are believed to be limited to pollen competition and availability. A greenhouse study showed that a herbicide-resistance gene (ALS) from A. hybridus could be introgressed into an advanced A. tuberculatus background (BC2). However, evidence is lacking in support of such transfer in nature. Postzygotic reproductive barriers may minimize, if not preclude, natural introgression. Indeed, A. hybridus xA. tuberculatus hybrids are characterized by reduced fertility and even floral neuterism. The purpose of this study was to assess hybrid fertility in the BC1 generation and its relationship with genome structure and segregation at ALS. Fertility was assessed by measuring seed output and by pollen evaluation, and segregation at ALS was determined via a molecular marker system. The two parental species have the same ploidy (2n = 32) but differ in DNA content (2C) values, with A. tuberculatus chromosomes being on average 29% greater than those of A. hybridus. Given that most (98%) BC(1)s were homoploid, 2C values were used as indicators of relative genomic constitution. Fertility in the BC1 generation was greater than that of F1s, and 3% of BC1s had seed output similar to that of the parental species. Fertility in the BC1 did not correlate (in a strict way) with reconstitution of parental genomes. Hybrid sterility appeared to be controlled by relatively few loci. Heterozygosity at ALS was negatively correlated with fertility. Also, the A. tuberculatus ALS allele was not observed in the A. hybridus sexual condition, monoecism. Linkage of ALS to a locus associated (directly or via epistasis) with hybrid sterility may explain the fertility penalty observed with ALS introgression. Moreover, this linkage might explain why sequenced herbicide-resistance ALS alleles from sympatric A. tuberculatus and A. hybridus populations show independent evolution.

Amaranthaceae↗

Hybridization and reproductive isolation among syntopic populations of the topminnows Fundulus notatus and F. olivaceus.

Fundulus notatus and Fundulus olivaceus are two closely related topminnow species that exhibit similar ecological niches and broad, largely overlapping, North American ranges extending throughout much of the Mississippi River drainage as well as the coastal drainages of the Gulf of Mexico. Previous studies have suggested that these two species are reproductively compatible despite cytogenetic differences and will hybridize when syntopic. We used nuclear and mtDNA loci to assess levels of hybridization and test for introgression in syntopic populations of these two species in four drainages in southern Illinois. Although hybridization was detected in all syntopic populations, an assessment of the proportion of hybrid individuals indicated a deficiency of hybrids relative to expectations under random mating. We determined that, although mtDNA introgression was prevalent and extended beyond the zones of contact, evidence of nuclear introgression was limited to the zone of sympatry.

Animals↗

Marker-assisted pyramiding of brown planthopper (Nilaparvata lugens Stål) resistance genes Bph1 and Bph2 on rice chromosome 12.

Brown planthopper (BPH) (Nilaparvata lugens Stål) is a significant insect pest of rice (Oryza sativa L.). We constructed a gene-pyramided japonica line, in which two BPH resistance genes Bph1 and Bph2 on the long arm of chromosome 12 independently derived from two indica resistance lines were combined through the recombinant selection. The gene-pyramiding was achieved based on the previously constructed high-resolution linkage maps of the two genes. Two co-dominant and four dominant PCR-based markers flanking the loci were used to select for a homozygous recombinant line in a segregating population that was derived from a cross between the parental homozygous single-gene introgression lines. BPH bioassay showed that the resistance level of the pyramided line was equivalent to that of the Bph1-single introgression line, which showed a higher level of resistance than the Bph2-single introgression line. The pyramid line should provide a useful experimental means for studying the fine structure of the chromosomal region covering these two major BPH resistance genes.

Alleles↗

Recent Adaptation in a Threatened Salmonid Revealed by Museum Genomics.

Steelhead/rainbow trout (Oncorhynchus mykiss) is an imperilled salmonid with two main life history strategies: migrate to the ocean or remain in freshwater. Domesticated hatchery forms of this species have been stocked into almost all California waterways, possibly resulting in introgression into natural populations and altered population structure. We compared whole-genome sequence data from contemporary populations against a set of museum population samples of steelhead from the same locations that were collected prior to most hatchery stocking. We observed minimal introgression and few steelhead-hatchery trout hybrids despite a century of extensive stocking. Our historical data show signals of introgression with a sister species and indications of an early hatchery facility. Finally, we found that migration-associated haplotypes have become less frequent over time, a likely adaptation to decreased opportunities for migration. Since contemporary migration-associated haplotype frequencies have been used to guide species management, we consider this to be a rare example of shifting baseline syndrome that has been validated with historical data. We suggest cautious optimism that a century of hatchery stocking has had minimal impact on California steelhead population genetic structure, but we note that continued shifts in life history may lead to further declines in the ocean-going form of the species.

Animals↗

Cytogenetics and genomics analysis of cold-hardy perennial wheatgrass: insights into agronomic performance, chromosome composition, and gene expression.

Intermedium wheatgrass (Thinopyrum intermedium), a perennial species with extensive root systems and high tolerance to cold, drought, and salinity, is a valuable genetic resource for the development of perennial crops. Over a decade-long selection process, two cold-hardy perennial wheatgrass lines were developed by crossing wheat-Thinopyrum partial amphiploids with Th. intermedium. These lines inherited key traits from Th. intermedium, including plant stature, spike morphology, and postharvest regrowth. Transcriptome-based single-nucleotide polymorphism tracing and sequential multicolor genomic in situ hybridization analyses revealed variations in the chromosome compositions of the perennial wheatgrass lines. The introgression of wheat chromosomes enhanced grain weight and size, while preserving the cold-hardy, perennial characteristics of the wheatgrass lines compared to Th. intermedium. Genome-wide gene expression was generally suppressed in the wheatgrass lines relative to Th. intermedium, particularly in conserved genes. This suppression was especially pronounced in genes involved in cell division and DNA repair pathways. In contrast, genes associated with cold tolerance and the water stress response were upregulated. We identified eight cold-tolerance genes in the Th. intermedium chromosomes and validated three of them, Thint.J05G452200, Thint.J05G452300, and Thint.V05G408900, using qRT-PCR. These genes encode proteins associated with cold tolerance and are potential candidates for further functional validation. Additionally, three chromosomes from homoeologous group 6 were introgressed, carrying six genes potentially associated with superior grain traits. Among them, TraesCS6D02G287800, which encodes a specific protein, exhibited high expression levels in both wheatgrass lines, suggesting its critical role in enhancing grain traits. Our results indicate that the suppression of grass gene expression, likely due to the introgression of wheat chromosomes and the upregulation of pathways related to cold tolerance and overwintering ability, contributes to the adaptive features of the wheatgrass lines. This study provides a genomic foundation for understanding gene expression regulation in distant hybrid progeny and offers valuable insights for designing new breeding strategies for perennial wheat or wheatgrass.

Chromosomes, Plant↗

fs3.1: a major fruit shape QTL conserved in Capsicum.

fs3.1 is a major fruit shape (defined as the ratio of fruit length to fruit width) quantitative trait locus (QTL) originally detected in an intraspecific cross of Capsicum annuum between the blocky and elongated-fruited inbreds 'Maor' and 'Perennial', respectively. In addition to increasing fruit shape index, the 'Perennial' allele at fs3.1 increased fruit elongation and decreased fruit width and pericarp thickness. We verified the effect of fs3.1 in backcross inbred lines (BILs) derived from crossing 'Perennial' with 'Maor' and with a second blocky-type inbred line of C. annuum. To determine the effect of the fs3.1 region in additional Capsicum species, we constructed an advanced backcross population from the cross of 'Maor' and the oval-fruited Capsicum frutescens BG 2816 and an F2 of the introgression line IL 152 that contains an introgression of the fs3.1 region from Capsicum chinense PI 152225. QTLs for fruit shape, fruit width, and pericarp thickness, but not for fruit length, were detected in both crosses, indicating the conservation of the fs3.1 region as a QTL affecting fruit shape in pepper. We also tested tomato (Lycopersicon spp.) introgression lines containing the corresponding fs3.1 region from L. pennellii and L. hirsutum, but we did not detect a significant fruit shape QTL in these lines. The effect of fs3.1 on the growth of fruit dimensions varied with the genetic background. By measuring the length and width of ovaries and fruits of near-isogenic C. annuum lines that differ in fs3.1 during fruit development, we determined that fs3.1 controls shape predominantly by increasing the growth rate of the longitudinal axis in the first 2 weeks after pollination. However, in the crosses of C. annuum with C. frutescens and C. chinense, fs3.1 predominantly exerted its effect on the width dimension.

Capsicum↗

Development and characterization of recombinant chromosome substitution lines (RCSLs) using Hordeum vulgare subsp. spontaneum as a source of donor alleles in a Hordeum vulgare subsp. vulgare background.

The ancestor of barley (Hordeum vulgare subsp. spontaneum) may be a source of novel alleles for crop improvement. We developed a set of recombinant chromosome substitution lines (RCSLs) using an accession of H. vulgare subsp. spontaneum (Caesarea 26-24, from Israel) as the donor and Hordeum vulgare subsp. vulgare 'Harrington' (the North American malting quality standard) as the recurrent parent via two backcrosses to the recurrent parent, followed by six generations of selfing. Here we report (i) the genomic architecture of the RCSLs, as inferred by simple sequence repeat (SSR) markers, and (ii) the effects of H. vulgare subsp. spontaneum genome segment introgressions in terms of three classes of phenotypes: inflorescence yield components, malting quality traits, and domestication traits. Significant differences among the RCSLs were detected for all phenotypes measured. The phenotypic effects of the introgressions were assessed using association analysis, and these were referenced to quantitative trait loci (QTL) reported in the literature. Hordeum vulgare subsp. spontaneum, despite its overall inferior phenotype, contributed some favorable alleles for agronomic and malting quality traits. In most cases, the introgression of the ancestral genome resulted in a loss of desirable phenotypes in the cultivated parent. Although disappointing from a plant breeding perspective, this finding may prove to be a useful tool for gene discovery.

Alleles↗

Pervasive context-dependent effects in the genetic architecture of complex and quantitative traits revealed by a powerful multiparent mapping population in yeast.

The genetic dissection of complex traits remains a major challenge in basic and biomedical research, but is essential for understanding the molecular pathways that shape phenotypic variation and for developing predictive models of trait and disease susceptibility. Here, we leverage a novel multiparent mapping population of budding yeast, CYClones, comprising 9,344 haploid strains derived from eight genetically diverse founders (~270,000 SNVs,&#x2009;~&#x2009;1 per 44 bp, capturing 56% of common variants and 32% of all variants with a minor allele frequency greater than 0.005 in the global population), to identify quantitative trait loci (QTL) and systematically investigate the genetic architecture of growth rates across ten environmental conditions. In total, we identified 349 QTL (ranging from 18 to 49 QTL per growth condition) that explained between 60% and 100% of narrow sense heritability across traits. The high power and resolution of CYClones revealed that growth traits exhibited distinct, condition-specific genetic architectures with extensive allelic heterogeneity, where a QTL was the result of multiple tightly linked causal variants. We also observed pleiotropy among QTL with complex, trait-dependent allele effects that are also consistent with allelic heterogeneity. Genetic complexity varied widely, with some traits showing nearly Mendelian architectures, while others were highly polygenic. Introgressed loci played a prominent role in the landscape of growth rate QTL, including a QTL localized to a 2.4 kb interval in the PCA1 cadmium transporter that explains 72% of variation in cadmium resistance and is largely driven by an introgression, and a non-additive interaction between the GAL3 regulator and introgressed GAL1/7/10 alleles, extending a previously described three-locus GAL-pathway incompatibility to a four-locus interaction. In both cadmium and galactose conditions, we show that allelic variation at a small number of loci stratifies the population into regulatory or physiological subgroups, each with distinct genetic architectures, a specific manifestation of epistasis we term allele-dependent stratification. Collectively, our results provide novel insights into the genetics of growth rates in budding yeast, the architectural features of genetic complexity, and demonstrate that CYClones is a powerful platform for revealing the molecular basis of complex trait variation.

Quantitative Trait Loci↗

Sources of variation in the morphological characteristics of sperm subpopulations assessed objectively by a novel automated sperm morphology analysis system.

There is evidence that the mammalian ejaculate contains distinct subpopulations of spermatozoa and that the variability among these subpopulations may have adaptive and functional significance. This study investigated the precision, reproducibility and operating characteristics of a novel automated sperm morphology analysis system, the Hobson Morphology package, establishing protocols to investigate boar sperm characteristics. Five ejaculates were collected from each of three boars from different genetic lines: Landrace-Meishan introgression, Sireline Large White and Damline Large White. Five semen smears per ejaculate were stained with haematoxylin and eosin. Two hundred spermatozoa per slide were analysed. No significant differences among slides within an ejaculate were detected for sperm tail length (P = 0.770), head width (P = 0.736) and head length (P = 0.615), indicating that both staining and morphology analysis were precise and reproducible. Among the boars, variability in tail length was detected (P = 0.001), but head width (P = 0.114) and length (P = 0.069) did not differ significantly. Multivariate pattern analysis (PATN computer package) highlighted three sub-populations of spermatozoa objectively on the basis of tail length (10.0-22.0 microns, 22.1-73.0 microns and 73.1-130.0 microns). The Landrace-Meishan introgression boar possessed more spermatozoa (P < 0.0001) with tails 73.1-130 microns long. Subsequent analysis of morphology parameters in a pure-bred Meishan boar showed similar measurements for tail length (mean +/- SD; 66.36 +/- 24.70 microns) to the Landrace-Meishan introgression boar (mean +/- SD; 67.09 +/- 21.80 microns). Sperm subpopulations originate during spermatogenesis, when heterogeneous genotypic effects determine the structural features of spermatozoa. The findings of this study confirm that tail length differs between boars and that subpopulations of spermatozoa can be detected within a single ejaculate.

Animals↗