While it is widely agreed that the conservation of local breeds is key to the maintenance of livestock biodiversity, the implementation of such programmes is often carried out by amateur breeders and may be inadequate due to a lack of knowledge and financial resources. Bortoluzzi et al. (2024) clearly demonstrate the utility of whole-genome sequencing (WGS) data for this purpose, compare two scenarios that differ in the consistency of conservation efforts, and provide valuable recommendations for conservation programmes.

Genetic diversity in livestock is generally considered to be crucial to maintaining food security and ensuring the provision of necessary nutrients to humans (Godde et al. 2021). It is also important to recognise that the preservation of local breeds is a matter of cultural identity for certain regions, and that the products of these breeds are niche products which are in high demand. Especially today, as we face extreme weather conditions, drought and other consequences of global warming, modern breeds selected to perform under constant and temperate conditions are being challenged. The possibility of tapping into the reservoir of genetic variation held by traditional, locally adapted breeds offers an important option for breeding robust livestock. The best way to characterise genetic diversity is through modern molecular methods, based on whole genome sequencing and subsequent advanced population analyses, which has been demonstrated for domesticated and wild chicken (Qanbari et al. 2019).

But are local breed conservation programmes up to the task? In their article, Bortoluzzi and colleagues show that well-designed and professionally managed conservation programmes for local chicken breeds are effective in maintaining genetic diversity. Their article is based on a comparison of two examples of conservation programmes for local chicken breeds: the Barbezieux and the Gasconne, which originated from comparably sized founder populations and for which WGS data were available in a biobank at two timepoints, 2003 and 2013, representing 10 generations. While the conservation programme for the former was continuous, that for the latter was interrupted and later started from scratch with a small number of sires and dams.

The greater loss of genomic diversity in the Gasconne than in the Barbezieux shown in this article may therefore be unsurprising, but the authors provide a range of evidence for this using their population genomics toolbox. The less well-managed breed, Gasconne, shows a lower genome-wide heterozygosity, higher lengths of runs of homozygosity, higher levels of genomic inbreeding, a smaller effective population size and a higher genetic load in terms of predicted deleterious mutations. 

The sample sizes available for population genetic analyses are typically small for local breeds, which is difficult to change as the populations are very small at any given time. It is therefore all the more important to make the most out of it, and Bortoluzzi and co-authors approach the issue from several angles that help support their claim, using WGS data and the latest genomic resources.

In addition to their analyses, the authors provide clear and valuable advice for the management of such conservation programmes. Their analysis of signatures of selection suggests that, apart from adult fertility, not much selection has been taking place. However, the authors emphasise that clear selection objectives other than maintaining the breed, such as production and product quality, can help conservation efforts by providing better guidelines for managing the programme and increasing the availability of resources for conservation programmes when the products of these local breeds become successful. 

In summary, Bortoluzzi et al. (2024) have provided a clear, well-written account of the impact of conservation programme management on the genetic diversity of local chicken breeds, using the most up-to-date genomic resources and analysis methods. As such, this article makes a significant and valuable contribution to the maintenance of genomic resources in livestock, providing approaches and lessons that will hopefully be adopted by other such initiatives.

            
References

Bortoluzzi C, Restoux G, Rouger R, Desnoues B, Petitjean F, Bosse M, Tixier-Boichard M (2024) Trends in genome diversity of small populations under a conservation program: a case study of two French chicken breeds. bioRxiv, ver. 2 peer-reviewed and recommended by PCI Genomics. https://doi.org/10.1101/2024.02.22.581528

Godde CM, Mason-D’Croz D, Mayberry DE, Thornton PK, Herrero M (2021) Impacts of climate change on the livestock food supply chain; a review of the evidence. Global Food Security 28:100488. https://doi.org/10.1016/j.gfs.2020.100488

Qanbari S, Rubin C-J, Maqbool K, Weigend S, Weigend A, Geibel J, Kerje S, Wurmser C, Peterson AT, IL Brisbin Jr., Preisinger R, Fries R, Simianer H, Andersson L (2019) Genetics of adaptation in modern chicken. PLOS Genetics, 15, e1007989. https://doi.org/10.1371/journal.pgen.1007989

Ever since the introduction of the first widely usable assemblers for transcriptomic reads (Huang and Madan 1999; Schulz et al. 2012; Simpson et al. 2009; Trapnell et al. 2010, and many more), it has been a technical challenge to compare different methods and to choose the “right” or “best” assembly. It took years until the first widely accepted set of benchmarks beyond raw statistical evaluation became available (e.g., Parra, Bradnam, and Korf 2007; Simão et al. 2015)⁠⁠. However, an approach to find the right balance between the number of transcripts or isoforms vs. evolutionary completeness measures has been lacking. This has been particularly pronounced in the field of non-model organisms (i.e., wild species that lack a genomic reference). Often, studies in this area employed only one set of assembly tools (the most often used to this day being Trinity, Haas et al. 2013; Grabherr et al. 2011)⁠. While it was relatively straightforward to obtain an initial assembly, its validation, annotation, as well its application to the particular purpose that the study was designed for (phylogenetics, differential gene expression, etc) lacked a clear workflow. This led to many studies using a custom set of tools with ensuing various degrees of reproducibility.

TransPi (Rivera-Vicéns et al. 2021)⁠ fills this gap by first employing a meta approach using several available transcriptome assemblers and algorithms to produce a combined and reduced transcriptome assembly, then validating and annotating the resulting transcriptome. Notably, TransPI performs an extensive analysis/detection of chimeric transcripts, the results of which show that this new tool often produces fewer misassemblies compared to Trinity. TransPI not only generates a final report that includes the most important plots (in clickable/zoomable format) but also stores all relevant intermediate files, allowing advanced users to take a deeper look and/or experiment with different settings. As running TransPi is largely automated (including its installation via several popular package managers), it is very user-friendly and is likely to become the new "gold standard" for transcriptome analyses, especially of non-model organisms.  

References

Grabherr MG, Haas BJ, Yassour M, Levin JZ, Thompson DA, Amit I, Adiconis X, Fan L, Raychowdhury R, Zeng Q, Chen Z, Mauceli E, Hacohen N, Gnirke A, Rhind N, di Palma F, Birren BW, Nusbaum C, Lindblad-Toh K, Friedman N, Regev A (2011) Full-length transcriptome assembly from RNA-Seq data without a reference genome. Nature Biotechnology, 29, 644–652. https://doi.org/10.1038/nbt.1883

Haas BJ, Papanicolaou A, Yassour M, Grabherr M, Blood PD, Bowden J, Couger MB, Eccles D, Li B, Lieber M, MacManes MD, Ott M, Orvis J, Pochet N, Strozzi F, Weeks N, Westerman R, William T, Dewey CN, Henschel R, LeDuc RD, Friedman N, Regev A (2013) De novo transcript sequence reconstruction from RNA-seq using the Trinity platform for reference generation and analysis. Nature Protocols, 8, 1494–1512. https://doi.org/10.1038/nprot.2013.084

Huang X, Madan A (1999) CAP3: A DNA Sequence Assembly Program. Genome Research, 9, 868–877. https://doi.org/10.1101/gr.9.9.868

Parra G, Bradnam K, Korf I (2007) CEGMA: a pipeline to accurately annotate core genes in eukaryotic genomes. Bioinformatics, 23, 1061–1067. https://doi.org/10.1093/bioinformatics/btm071

Rivera-Vicéns RE, Garcia-Escudero CA, Conci N, Eitel M, Wörheide G (2021) TransPi – a comprehensive TRanscriptome ANalysiS PIpeline for de novo transcriptome assembly. bioRxiv, 2021.02.18.431773, ver. 3 peer-reviewed and recommended by Peer Community in Genomics. https://doi.org/10.1101/2021.02.18.431773

Schulz MH, Zerbino DR, Vingron M, Birney E (2012) Oases: robust de novo RNA-seq assembly across the dynamic range of expression levels. Bioinformatics, 28, 1086–1092. https://doi.org/10.1093/bioinformatics/bts094

Simão FA, Waterhouse RM, Ioannidis P, Kriventseva EV, Zdobnov EM (2015) BUSCO: assessing genome assembly and annotation completeness with single-copy orthologs. Bioinformatics, 31, 3210–3212. https://doi.org/10.1093/bioinformatics/btv351

Simpson JT, Wong K, Jackman SD, Schein JE, Jones SJM, Birol İ (2009) ABySS: A parallel assembler for short read sequence data. Genome Research, 19, 1117–1123. https://doi.org/10.1101/gr.089532.108

Trapnell C, Williams BA, Pertea G, Mortazavi A, Kwan G, van Baren MJ, Salzberg SL, Wold BJ, Pachter L (2010) Transcript assembly and quantification by RNA-Seq reveals unannotated transcripts and isoform switching during cell differentiation. Nature Biotechnology, 28, 511–515. https://doi.org/10.1038/nbt.1621

Biological systems depend on finely tuned interactions of their components. Thus, regulating these components is critical for the system's functionality. In prokaryotic cells, riboswitches are regulatory elements controlling transcription or translation. Riboswitches are RNA molecules that are usually located in the 5′-untranslated region of protein-coding genes. They generate secondary structures leading to the regulation of the expression of the downstream protein-coding gene (Kavita and Breaker, 2022). Riboswitches are very versatile and can bind a wide range of small molecules; in many cases, these are metabolic byproducts from the gene’s enzymatic or signaling pathway. Their versatility and abundance in many species make them attractive for synthetic biological circuits. One class that has been drawing the attention of synthetic biologists is toehold switches (Ekdahl et al., 2022; Green et al., 2014). These are single-stranded RNA molecules harboring the necessary elements for translation initiation of the downstream gene: a ribosome-binding site and a start codon. Conformation change of toehold switches is triggered by an RNA molecule, which enables translation.

To exploit the most out of toehold switches, automation of their design would be highly advantageous. Cisneros and colleagues (Cisneros et al., 2022) developed a tool, “Toeholder”, that automates the design of toehold switches and performs in silico tests to select switch candidates for a target gene. Toeholder is an open-source tool that provides a comprehensive and automated workflow for the design of toehold switches. While web tools have been developed for designing toehold switches (To et al., 2018), Toeholder represents an intriguing approach to engineering biological systems by coupling synthetic biology with computational biology. Using molecular dynamics simulations, it identified the positions in the toehold switch where hydrogen bonds fluctuate the most. Identifying these regions holds great potential for modifications when refining the design of the riboswitches. To be effective, toehold switches should provide a strong ON signal and a weak OFF signal in the presence or the absence of a target, respectively. Toeholder nicely ranks the candidate toehold switches based on experimental evidence that correlates with toehold performance (based on good ON/OFF ratios).

Riboswitches are highly appealing for a broad range of applications, including pharmaceutical and medical purposes (Blount and Breaker, 2006; Giarimoglou et al., 2022; Tickner and Farzan, 2021), thanks to their adaptability and inexpensiveness. The Toeholder tool developed by Cisneros and colleagues is expected to promote the implementation of toehold switches into these various applications.

References

Blount KF, Breaker RR (2006) Riboswitches as antibacterial drug targets. Nature Biotechnology, 24, 1558–1564. https://doi.org/10.1038/nbt1268

Cisneros AF, Rouleau FD, Bautista C, Lemieux P, Dumont-Leblond N, ULaval 2019 T iGEM (2022) Toeholder: a Software for Automated Design and In Silico Validation of Toehold Riboswitches. bioRxiv, 2021.11.09.467922, ver. 3 peer-reviewed and recommended by Peer Community in Genomics. https://doi.org/10.1101/2021.11.09.467922

Ekdahl AM, Rojano-Nisimura AM, Contreras LM (2022) Engineering Toehold-Mediated Switches for Native RNA Detection and Regulation in Bacteria. Journal of Molecular Biology, 434, 167689. https://doi.org/10.1016/j.jmb.2022.167689

Giarimoglou N, Kouvela A, Maniatis A, Papakyriakou A, Zhang J, Stamatopoulou V, Stathopoulos C (2022) A Riboswitch-Driven Era of New Antibacterials. Antibiotics, 11, 1243. https://doi.org/10.3390/antibiotics11091243

Green AA, Silver PA, Collins JJ, Yin P (2014) Toehold Switches: De-Novo-Designed Regulators of Gene Expression. Cell, 159, 925–939. https://doi.org/10.1016/j.cell.2014.10.002

Kavita K, Breaker RR (2022) Discovering riboswitches: the past and the future. Trends in Biochemical Sciences. https://doi.org/10.1016/j.tibs.2022.08.009

Tickner ZJ, Farzan M (2021) Riboswitches for Controlled Expression of Therapeutic Transgenes Delivered by Adeno-Associated Viral Vectors. Pharmaceuticals, 14, 554. https://doi.org/10.3390/ph14060554

To AC-Y, Chu DH-T, Wang AR, Li FC-Y, Chiu AW-O, Gao DY, Choi CHJ, Kong S-K, Chan T-F, Chan K-M, Yip KY (2018) A comprehensive web tool for toehold switch design. Bioinformatics, 34, 2862–2864. https://doi.org/10.1093/bioinformatics/bty216

When discussing the Earth BioGenome Project with scientists and potential funding agencies, one common question is: why sequence everything? Whether sequencing a subset would be more optimal is not an unreasonable question given what we know about the mathematics of importance and Pareto’s 80:20 principle, that 80% of the benefits can come from 20% of the effort. However, one must remember that this principle is an observation made in hindsight and selecting the most effective 20% of experiments is difficult. As an example, few saw great applied value in comparative genomic analysis of the archaea Haloferax mediterranei, but this enabled the discovery of CRISPR/Cas9 technology (1). When discussing whether or not to sequence all life on our planet, smaller countries such as the Faroe Islands are seldom mentioned. 
 
Mikalsen and co-authors (2) provide strong arguments to appreciate, investigate and steward genetic diversity, from a Faroese viewpoint, a fishery viewpoint, and a global viewpoint. As readers, we learn to cherish the Faroe Islands, the Faroese, and perhaps by extension all of nature and the people of the world. The manuscript describes the proposed Faroese participation in the European Reference Genome Atlas (ERGA) consortium through Gen@FarE – the Genome Atlas of Faroese Ecology. Gen@FarE aims to: i) generate high-quality reference genomes for all eukaryotes on the islands and in its waters; ii) establish population genetics of all species of commercial or ecological interest; and iii) establish a “databank” for all Faroese species with citizen science tools for participation.

In the background section of the manuscript, the authors argue that as caretakers of the earth (and responsible for the current rapid decrease in biodiversity), humanity must be aware of the biodiversity and existing genetic diversity, to protect these for future generations. Thus, it is necessary to have reference genomes for as many species as possible, enabling estimation of population sizes and gene flow between ecosystem locations. Without this the authors note that “…it is impossible to make relevant management plans for a species, an ecosystem or a geographical area…”. Gen@FarE is important. The Faroe nation has a sizable economic zone in the North Atlantic and large fisheries. In terms of biodiversity and conservation, the authors list some species endemic to other Faroe islands, especially sea birds. The article discusses ongoing marine environmental-DNA-based monitoring programs that started in 2018, and how new reference genome databases will help these efforts to track and preserve marine biodiversity. They point to the lack of use of population genomics information for Red List decisions on which species are endangered, and the need for these techniques to inform sustainable harvesting of fisheries, given collapses in critical food species such as Northwest Atlantic cod and herring. In one example, they highlight how the herring chromosome 12 inversion contains a “supergene” collection of tightly linked genes associated with ecological adaptation. Genetic tools may also help enable the identification and nurturing of feeding grounds for young individuals. Critically, the Faroe Islands have a significant role to play in protecting the millions of tons of seafood caught annually upon which humanity relies. As the authors note, population genomics based on high-quality reference sequences is “likely the best tool” to monitor and protect commercial fisheries. There is an important section discussing the role of interactions between visible and “invisible" species in the marine ecosystem on which we all depend. Examples of “invisible” species include a wide range of morphologically similar planktonic algae, and invasive species transported by ballast water or ship hulls.​ As biologists, I believe we forget that our population studies of life on the earth have so far been mostly in the dark. Gen@FarE is but one light that can be switched on. 

The authors conclude by discussing Gen@FarE plans for citizen science and education, perhaps the most important part of this project if humanity is to learn to cherish and care for the earth. Where initiatives such as the Human Genome Project did not need the collaborative efforts of the world for sample access, the Earth BioGenome Project most certainly does. In the same way, at a smaller scale, Gen@FarE requires the support and determination of the Faroese. 
 

References    

1          Mojica, F. J., Díez-Villaseñor, C. S., García-Martínez, J. & Soria, E. Intervening sequences of regularly spaced prokaryotic repeats derive from foreign genetic elements. J Mol Evol 60, 174-182 (2005).

2          Mikalsen, S-O., Hjøllum, J. í., Salter, I., Djurhuus, A. & Kongsstovu, S. í. The need of decoding life for taking care of biodiversity and the sustainable use of nature in the Anthropocene – a Faroese perspective. EcoEvoRxiv (2024), ver. 3 peer-reviewed and recommended by Peer Community in Genomics. https://doi.org/10.32942/X21S4C

The European Reference Genome Atlas (ERGA) (Mc Cartney et al, 2024, Mazzoni et al, 2023) demonstrates the collaborative spirit and intellectual abilities of researchers from 33 European countries. This ambitious project, which is part of the Earth BioGenome Project (Lewin et al., 2018) Phase II, has embarked on an unprecedented mission: to decipher the genetic makeup of 150,000 species over a span of four years. At the heart of ERGA is a decentralized pilot infrastructure specifically built to assist the production of high-quality reference genomes. This structure acts as a scaffold for the massive task of genome sequencing, giving the necessary framework to manage the complexity of genomic research. The research paper under consideration offers a comprehensive narrative of ERGA's evolution, outlining both successes and challenges encountered along the road. 

One of the most significant issues addressed in the manuscript is the equitable distribution of resources and expertise among participating laboratories and countries. In a project of this magnitude, it is critical to leverage the pooled talents and capacities of researchers from across Europe. ERGA's pan-European network promotes communications and collaboration, creating an environment in which knowledge flows freely and barriers are overcome. This adoption of strong coordination and communication tactics will be essential to ERGA's success. Scientific collaboration depends on efficient communication channels because they allow researchers to share resources, collaborate on new initiatives, and exchange ideas. Through a diverse range of gatherings, courses, and virtual discussion boards, ERGA fosters an environment of transparency and cooperation among members, enabling scientists to overcome challenges and make significant discoveries. The importance ERGA places on training and information transfer programmes is a pillar of its strategy. Understanding the importance of capacity development, ERGA invests in providing researchers with the knowledge and abilities necessary for effectively navigating the complicated terrain of genomic research. A wide range of subjects are covered in training programmes (Larivière et al. 2023), from sample preparation and collection to data processing methods and sequencing technology. Through the development of a group of highly qualified experts, ERGA creates the foundation for continued advancement and creativity in the genomics sector.

This manuscript also covers in detail the technological workflows and sequencing techniques used in ERGA's pilot infrastructure. With the aid of cutting-edge sequencing technologies based on both long-read and short-read sequencing, they are working to unravel the complex structure of the genetic code with a level of accuracy and precision never before possible. To guarantee the accuracy of genetic data and prevent mistakes and flaws that can jeopardize the findings' integrity, quality control methods are put in place. Despite having a focus on genome sequencing due to its technological complexities, ERGA also remains firm in its dedication to metadata collection and sample validation. Metadata serves as a critical link between raw genetic data and useful scientific insights, giving necessary context and allowing researchers to draw practical findings from their investigations. Sample validation approaches improve the reliability and reproducibility of the results, providing users confidence in the quality of the genetic data provided by ERGA.​

Looking ahead, ERGA envisions its decentralized infrastructure serving as a model for global collaborative research efforts. By embracing diversity, encouraging cooperation, and pushing for open access to data and resources, ERGA hopes to catalyze scientific discovery and generate positive change in the field of biodiversity genomics. ERGA aims to promote a more equitable and sustainable future for all by ongoing interaction with stakeholders, intensive outreach and education activities, and policy change advocacy. In addition to its immediate goals, ERGA considers the long-term implications of its work. As genomic technology progresses, the potential application of high-quality reference genomes will continue to grow. From informing conservation efforts and illuminating evolutionary histories to revolutionizing healthcare and agriculture, it is likely that ERGA's contributions will have far-reaching consequences for people and the planet as a whole.​

Furthermore, ERGA understands the importance of interdisciplinary collaboration in addressing the difficult challenges of the twenty-first century. ERGA aims to integrate genetic research into larger initiatives to promote sustainability and biodiversity conservation by forming relationships with stakeholders from other areas, such as policymakers, conservationists, and indigenous groups. Through shared knowledge and community action, ERGA seeks to create a future in which mankind coexists peacefully with the natural world, guided by a thorough grasp of its genetic legacy and ecological interconnectivity.

Finally, the manuscript exemplifies ERGA's collaborative ambitions and achievements, capturing the spirit of creativity and collaboration that defines this ground-breaking effort. As ERGA continues to push the boundaries of genetic research, it remains dedicated to scientific excellence, inclusivity, and the quest of knowledge for the benefit of society. I wholeheartedly recommend the publication of this groundbreaking initiative, offering my enthusiastic endorsement for its valuable contribution to the scientific community.​​

References
Larivière, D., Abueg, L., Brajuka, N. et al. (2024). Scalable, accessible and reproducible reference genome assembly and evaluation in Galaxy. Nature Biotechnology 42, 367-370. https://doi.org/10.1038/s41587-023-02100-3

Lewin, H. A., Robinson, G. E., Kress, W. J., Baker, W. J., Coddington, J., Crandall, K. A., Durbin, R., Edwards, S. V., Forest, F., Gilbert, M. T. P., Goldstein, M. M., Grigoriev, I. V., Hackett, K. J., Haussler, D., Jarvis, E. D., Johnson, W. E., Patrinos, A., Richards, S., Castilla-Rubio, J. C., … Zhang, G. (2018). Earth BioGenome Project: Sequencing life for the future of life. Proceedings of the National Academy of Sciences, 115(17), 4325–4333. https://doi.org/10.1073/pnas.1720115115

Mazzoni, C. J., Claudio, C.i, Waterhouse, R. M. (2023). Biodiversity: an atlas of European reference genomes. Nature 619 : 252-252. https://doi.org/10.1038/d41586-023-02229-w

Mc Cartney, A. M., Formenti, G., Mouton, A., Panis, D. de, Marins, L. S., Leitão, H. G., Diedericks, G., Kirangwa, J., Morselli, M., Salces-Ortiz, J., Escudero, N., Iannucci, A., Natali, C., Svardal, H., Fernández, R., Pooter, T. de, Joris, G., Strazisar, M., Wood, J., … Mazzoni, C. J. (2024). The European Reference Genome Atlas: piloting a decentralised approach to equitable biodiversity genomics. bioRxiv, ver. 4 peer-reviewed and recommended by Peer Community in Genomics. https://doi.org/10.1101/2023.09.25.559365​