Jeffrey Powell | Search USGS

USGS2022

Haploid gynogens facilitate disomic marker development in paleotetraploid sturgeons

Acipenseriformes (sturgeons and paddlefishes) are of substantial conservation concern, and development of genomic resources for these species is difficult due to past whole genome duplication. Development of disomic markers for polyploid organisms can be challenging due to difficulty in resolving alleles at a single locus from those among duplicated loci. In this study, we detail the development of disomic markers for the endangered pallid sturgeon ( Scaphirhynchus albus ) found in North America. One of the strategies for pallid sturgeon conservation is to stock U.S. rivers with offspring of pure pallid sturgeon, but introgression with the sympatric shovelnose sturgeon ( S. platorynchus ) threatens pallid sturgeon genetic integrity. Currently, 19 microsatellite loci are used to differentiate between both species and their hybrids, but the markers are insufficient to robustly identify backcrosses. We performed double digest restriction site-associated DNA sequencing (ddRADseq) on shovelnose sturgeon haploid gynogens to produce a reduced-representation genomic reference. Contiguous sequences that were heterozygous within a haploid individual were flagged as potentially encompassing multiple loci. Approximately 60 individuals of each species from two management units were sequenced, and reads were mapped to the haploid reference to identify single nucleotide polymorphisms (SNPs) at individual loci. The final data set contained 11,082 microhaplotyped loci which offer at least an order of magnitude greater resolution for species discrimination than the current panel of 19 microsatellites. These markers will be used to examine a larger sample of Scaphirhynchus individuals throughout their ranges to determine the extent and trajectory of hybridization.

Molecular Ecology Resources↗

USGS2021

Production of haploid gynogens to inform genomic resource development in the paleotetraploid pallid sturgeon (Scaphirhynchus albus)

Order Acipenseriformes (sturgeons and paddlefishes) is an ancient lineage of osteichthyan fishes (>200 million years old) with most extant species at conservation risk. A relatively basal species, the pallid sturgeon, Scaphirhynchus albus, is a federally endangered species native to the Mississippi and Missouri River basins. Hybridization with sympatric shovelnose sturgeon, S. platorynchus, is one of several threats to pallid sturgeon. Current molecular markers cannot reliably distinguish among pure species and multigenerational backcrosses. This information is critical for implementation of management strategies to increase populations through natural reproduction and artificial propagation. Genotypes from a large panel of unlinked single-nucleotide polymorphisms (SNPs) may provide greater resolution of the two species; however, paralogous sequence variants (PSVs) within individuals resulting from an ancient whole genome duplication event confound SNP development. The aim of this study was to produce pallid sturgeon gynogens that contain 100% homozygous DNA contributed by only the maternal parent and have enough DNA for future SNP marker development. When homozygous gynogens are sequenced, heterozygosity at a locus within an individual indicates the presence of incorrectly aligned sequences that contain PSVs; accurate identification of these multi-locus contigs can facilitate their exclusion when developing disomic markers. In this study, we attempted to produce two types of pallid sturgeon gynogens: a) haploid gynogens produced from the activation of pallid sturgeon eggs with ultraviolet-irradiated sperm from the distantly related paddlefish (Polyodon spathula), and b) doubled haploids produced from the activation of pallid sturgeon eggs with irradiated paddlefish milt followed by thermal shock to suppress the first mitotic division. Production of doubled haploids, gynogens with 100% homozygous DNA and double the genome content of haploid gynogens, was pursued because it was originally unknown if haploid gyongens would survive long enough to attain enough genetic material for SNP marker development. We performed flow cytometry and microsatellite genotyping on the specimens in order to confirm haploid and doubled haploid status. Our study was unable to successfully yield doubled haploids; however, we successfully produced haploid gynogens that contained enough nuclear DNA for our future SNP marker development study. Interestingly, this study also produced paddlefish × pallid sturgeon hybrids in the control groups in two separate years; this is the first study to report viable offspring between the paddlefish and a Scaphirhynchus sturgeon species and reflects on the malleability of the genomes of the species in this order.

Louisiana, Arkansas, Missouri, Nebraska, North Dak↗