NGS is Good For the Earth
Next-generation sequencing (NGS) is an influential tool used in genomics research that enables cost-effective, high-throughput analysis of millions of DNA fragments to advance investigations related to human health and development, detect genetic variations associated with disease, and much more (Satam H 2023). Besides improving studies of the human genome and other model organisms, NGS is also a powerful tool harnessed in environmental investigations, having extensively been applied to many non-model organisms including plants, animals, and insects. In the spirit of Earth Day, we’d like to highlight some of our client publications where NGS was leveraged to progress their research. NGS improves studies relating to plant and animal conservation, the microbiome, agriculture, evolutionary research, infectious disease, population genetics, and much more.
Over the last century, genus Panthera, or the ‘big cats’ (comprised of lions, tigers, leopards, jaguars, and snow leopards) have suffered drastic declines in population due to human activity and slow maturity rates, pushing them into either “threatened” or “endangered” listings by the International Union for Conservation of Nature (IUCN) & the Convention on International Trade in Endangered Species (CITES). During this timeframe, the Panthera genus has undergone taxonomic changes as the number of species classified under Panthera has become increasingly more refined. In this study, Armstrong and collaborators constructed chromosome-level assemblies for 3 members of the genus including the African leopard (Panthera pardus pardus), the tiger (Panthera tigris), and the snow leopard (Panthera uncia) using a combination of short-read, long-read, and proximity ligation data from Hi-C (Armstrong EE 2022). The researchers found incorporating Hi-C data greatly improved the continuity of the genome assemblies to roughly chromosome level and for the long-read (PacBio HiFi) data, which also consistently improved overall assembly continuity. In agreement with previous studies, the phylogenetic data (the largest gene set for this clade or group) showed the Panthera clade was strikingly conserved at the karyotypic level and showed no chromosome level inversions when compared to the domestic cat & other genomes. These high-quality reference genomes will be useful for conservation and evolutionary research to enable these ‘big cats’ to thrive in the future (Armstrong EE 2022).
*Image provided by Ellie Armstrong
Understanding the causes of antimicrobial resistance (AMR) in food-producing animals is of utmost importance, as AMR bacteria and in some cases, antimicrobial resistance genes (ARGs), have been found to spread to humans and livestock. Previously, AMR has largely been attributed to antimicrobial usage (AMU) but is also associated with socio-economic factors. In this study, Munk et al. used shotgun metagenomic sequencing to analyze the resistomes (or total pool of ARGs) of 538 herds from pigs, veal calves, poultry, turkey, and rainbow trout, across 9 European countries (Munk P 2024). Notably, the clear core resistomes for each species were identified and confirmed previous associations between AMR, biosecurity, and AMU, indicating the observed trends were robust to the updated methodology and reference data. The resistomes saw clear clustering by species; one with pigs and veal calves, another cluster with broiler (chicken) and turkey, and a third cluster from the rainbow trout resistome. In total, 534 different ARGs were detected across all samples, with the greatest abundance discovered in pigs, veal calves, then broilers, turkey, and finally fish, where variants of the same ARG were discovered in different species. This gives the impression that although AMU is still a driver for AMR, future studies should also consider other ecological and epidemiological factors that may be associated with greater AMR load, such as examining bacterial diversity. Munk and colleagues provide a critical resource for future efforts to combat AMR and maintain the focus on biosecurity measures (Munk P 2024).
Single nucleotide polymorphisms (SNPs) are effective markers used in many genomic applications, including genome-wide association studies, population management, and breeding initiatives. However, many non-model organisms lack well-curated reference genomes making classification schemes difficult, particularly for organisms with complex genomes like the Nordmann fir (Abies nordmanniana), a leading Christmas tree species in Europe with over 40 million trees sold annually (Christensen, C.J. 2019). In this paper, Ousmael and colleagues combined both reference-free & reference-based approaches to detect SNPs in the Nordmann fir for a subsequent ad hoc breeding program (Ousmael K 2023). First, the authors identified >2M SNPs using a genome assembly (reference-based) from a closely related species (Silver fir, Abies alba), of which only 250 SNPs were detected when combined with a de novo assembly (reference-free) constructed from low-copy regions of the Nordmann fir. Next, a subset of 200 SNPs shared between the two approaches was validated and used to genotype 342 trees from commercial seed lots, correctly identifying all copies of each clone within the population. Also of note was the discovery of 5 mislabeled individuals when using 113 highly reliable SNPs following identity analysis. In general, the researcher’s approach retained 56.5% of the tested SNPs, which could be used for future research related to genome selection, pedigree reconstruction, and clonal identification (Ousmael K 2023).
The rise of global temperatures and the increase in human population will require innovations in plant breeding and agricultural strategies to guarantee worldwide food security. To combat these effects, plant breeders have turned to crop wild relatives (CWRs), known to harbor genetic diversity not present in cultivated relatives (or cultivars) and may also possess more desirable traits (e.g. resistance to disease or pests). However, CWRs face many human-induced threats, like hybridization with cultivated crops that can lead to introgression—or the stable incorporation of ‘foreign’ genetic material from the cultivars to the CWRs. Hybridization poses a threat to wild coffee species like Coffea canephora (cultivated as Robusta coffee), found in the Yangami region in the Democratic Republic of Congo. In this study, Verleysen and colleagues used genotyping-by-synthesis (GBS) to evaluate the geographical distribution of cultivated C. canephora and the prevalence of hybridization in 83 wild C. canephora samples from either home gardens or the tropical rainforest and compared the genomic fingerprints with existing GBS data of C. canephora (Verleysen L 2024). The researchers discovered clear patterns of human activity in cultivated & cultivated-wild hybrid plants suggesting that cultivated C. canephora may act as a source for crop-to-wild gene flow, due to its proximity to wild gene pools. Still, hybridization analysis showed minimal evidence of admixture and hybridization between the wild and cultivated C. canephora samples, suggesting that if hybridization were to occur, it would likely be of genetic material already part of the wild gene pool. In general, the large genetic similarity between the wild and cultivated plants highlights the importance of monitoring habitat integrity to ensure global food security (Verleysen L 2024).
Grapevine trunk diseases (GTDs) pose a threat to viticulture or winegrowing. GTDs are caused by wood pathogenic fungi belonging to different genera, including Phaeomoniella, Fomitiporia, Phaeoacremonium, and members of the family Botryosphaeriaceae. However, the co-occurrence of these fungi has been found in both symptomatic and asymptomatic vines suggesting a complete understanding of GTD etiology is lacking. Here, Bekris and collaborators used amplicon sequencing to study the fungal and bacterial microbiome in wood tissues from three different Greek cultivars, or plant sites (Vidiano, Agiorgitiko, & Xinomavro), that were both symptomatic and asymptomatic to GTDs (Bekris F 2021). They discovered that cultivar (or biogeography) was the strongest predictor of the wood fungal microbiome, with strong positive correlations found with several fungi in symptomatic vines including Kalmusia variispora, Phaemoniella chlamydospore, Alternaria alternata & Cladosporium sp., which was also dependent on cultivar location. In contrast, GTD symptoms also showed a significant effect, albeit weaker, and were prominent only in the Xinomavro cultivar. Similarly, cultivar location was a stronger determinant of the wood bacterial microbiome (vs. GTD status), where Streptomyces & Bacillus amplicon sequence variants (ASVs) were found to be positively correlated with asymptomatic vines and formed a negative co-occurrence network with several fungi (Phaeomoniella, Phaeoacremonium, and Seimatosporium). This study provides further evidence for the role of the wood fungal microbiome in GTD, but also highlights the strong interaction between the bacterial and fungal microbiome in asymptomatic vines, which could be leveraged for future efforts to differentiate healthy and diseased vines. More broadly, this research can aid in the future exploration of novel biocontrol agents (Bekris F 2021).
NGS With Admera Health
NGS is a powerful technique used in many genomic research applications including plant and animal conservation, the microbiome, agriculture, evolutionary research, infectious disease, population genetics, and much more. We are grateful to work with many groundbreaking researchers whose dedication leads to actionable change and improves the world for all living organisms. Admera Health offers a suite of NGS services to propel your environmental studies. Our team will help with your project’s experimental design and provide you with high-quality data to adhere to your project’s timeline.
References
1. Armstrong, E.A. et al. Genome report: chromosome-level draft assemblies of the snow leopard, African leopard, and tiger (Panthera uncia, Panthera pardus pardus, and Panthera tigris), G3 (2022). https://doi.org/10.1093/g3journal/jkac277
2. Bekris, F. et al. Grapevine wood microbiome analysis identifies key fungal pathogens and potential interactions with the bacterial community implicated in grapevine trunk disease appearance. Envi Microb (2021). https://doi.org/10.1186/s40793-021-00390-1
3. Christensen, C. J. Eksporten af juletræer og klippegrønt i (2019). Nåledrys (In Danish). https://christmastree.dk/formidling/naaledrys/110/eksporten-af-juletraeer-og-klippegroent-i-2019/
4. Ellstrand, N. Current knowledge of gene flow in plants: implications for transgene flow. Philosophical Transactions of the Royal Society of London, Series B: Biological Sciences (2003). https://doi.org/10.1098/rstb.2003.1299
5. Munk, P. et al. The European livestock resistome. mSystems (2024). https://doi.org/10.1128/msystems.01328-23
6. Ousmael, K. et al. Identification and high-throughput genotyping of single nucleotide polymorphism markers in a non-model conifer (Abies nordmanniana (Steven) Spach). Sci Rep (2023). https://doi.org/10.1038/s41598-023-49462-x
7. Satam, H. et al. Next-generation sequencing technology: current trends and advancements. Biology (2023). https://doi.org/10.3390/biology12070997
8. Verleysen, L. et al. Crop-to-wild gene flow in wild coffee species: the case of Coffea canephora in the Democratic Republic of the Congo. Annals of Botany (2024). https://doi.org/10.1093/aob/mcae034