Molecular characterization of a marine turtle tumor epizootic, profiling external, internal and postsurgical regrowth tumors

Sea turtle populations are under threat from an epizootic tumor disease (animal epidemic) known as fibropapillomatosis. Fibropapillomatosis continues to spread geographically, with prevalence of the disease also growing at many longer-affected sites globally. However, we do not yet understand the precise environmental, mutational and viral events driving fibropapillomatosis tumor formation and progression. Here we perform transcriptomic and immunohistochemical profiling of five fibropapillomatosis tumor types: external new, established and postsurgical regrowth tumors, and internal lung and kidney tumors. We reveal that internal tumors are molecularly distinct from the more common external tumors. However, they have a small number of conserved potentially therapeutically targetable molecular vulnerabilities in common, such as the MAPK, Wnt, TGFβ and TNF oncogenic signaling pathways. These conserved oncogenic drivers recapitulate remarkably well the core pan-cancer drivers responsible for human cancers. Fibropapillomatosis has been considered benign, but metastatic-related transcriptional signatures are strongly activated in kidney and established external tumors. Tumors in turtles with poor outcomes (died/euthanized) have genes associated with apoptosis and immune function suppressed, with these genes providing putative predictive biomarkers. Together, these results offer an improved understanding of fibropapillomatosis tumorigenesis and provide insights into the origins, inter-tumor relationships, and therapeutic treatment for this wildlife epizootic

Evolutionary comparisons of chelonid alphaherpesvirus 5 (ChHV5) genomes from fibropapillomatosis-afflicted green (chelonia mydas), Ooive ridley (lepidochelys olivacea) and kemp’s ridley (lepidochelys kempii) sea turtles

peer-reviewedThe spreading global sea turtle fibropapillomatosis (FP) epizootic is threatening some of Earth’s ancient reptiles, adding to the plethora of threats faced by these keystone species. Understanding this neoplastic disease and its likely aetiological pathogen, chelonid alphaherpesvirus 5 (ChHV5), is crucial to understand how the disease impacts sea turtle populations and species and the future trajectory of disease incidence. We generated 20 ChHV5 genomes, from three sea turtle species, to better understand the viral variant diversity and gene evolution of this oncogenic virus. We revealed previously underappreciated genetic diversity within this virus (with an average of 2035 single nucleotide polymorphisms (SNPs), 1.54% of the ChHV5 genome) and identified genes under the strongest evolutionary pressure. Furthermore, we investigated the phylogeny of ChHV5 at both genome and gene level, confirming the propensity of the virus to be interspecific, with related variants able to infect multiple sea turtle species. Finally, we revealed unexpected intra-host diversity, with up to 0.15% of the viral genome varying between ChHV5 genomes isolated from different tumours concurrently arising within the same individual. These findings offer important insights into ChHV5 biology and provide genomic resources for this oncogenic viru

Finishing the euchromatic sequence of the human genome

The sequence of the human genome encodes the genetic instructions for human physiology, as well as rich information about human evolution. In 2001, the International Human Genome Sequencing Consortium reported a draft sequence of the euchromatic portion of the human genome. Since then, the international collaboration has worked to convert this draft into a genome sequence with high accuracy and nearly complete coverage. Here, we report the result of this finishing process. The current genome sequence (Build 35) contains 2.85 billion nucleotides interrupted by only 341 gaps. It covers ∼99% of the euchromatic genome and is accurate to an error rate of ∼1 event per 100,000 bases. Many of the remaining euchromatic gaps are associated with segmental duplications and will require focused work with new methods. The near-complete sequence, the first for a vertebrate, greatly improves the precision of biological analyses of the human genome including studies of gene number, birth and death. Notably, the human enome seems to encode only 20,000-25,000 protein-coding genes. The genome sequence reported here should serve as a firm foundation for biomedical research in the decades ahead

Inadvertent human genomic bycatch and intentional capture raise beneficial applications and ethical concerns with environmental DNA

The field of environmental DNA (eDNA) is advancing rapidly, yet human eDNA applications remain underutilized and underconsidered. Broader adoption of eDNA analysis will produce many well-recognized benefits for pathogen surveillance, biodiversity monitoring, endangered and invasive species detection, and population genetics. Here we show that deep-sequencing-based eDNA approaches capture genomic information from humans (Homo sapiens) just as readily as that from the intended target species. We term this phenomenon human genetic bycatch (HGB). Additionally, high-quality human eDNA could be intentionally recovered from environmental substrates (water, sand and air), holding promise for beneficial medical, forensic and environmental applications. However, this also raises ethical dilemmas, from consent, privacy and surveillance to data ownership, requiring further consideration and potentially novel regulation. We present evidence that human eDNA is readily detectable from 'wildlife' environmental samples as human genetic bycatch, demonstrate that identifiable human DNA can be intentionally recovered from human-focused environmental sampling and discuss the translational and ethical implications of such findings.Funding for the initial HGB eDNA study was generously provided by the National Save the Sea Turtle Foundation under project name Fibropapillomatosis Training and Research Initiative (D.J.D.), a Welsh Government Sêr Cymru II and the European Union Horizon 2020 research and innovation programme under the Marie Skłodowska-Curie grant agreement no. 663830-BU115 (D.J.D.). This research was also supported by Gumbo Limbo Nature Center d/b/a Friends of Gumbo Limbo (a 501c3 non-profit organization) through a generous donation through their Graduate Research Grant programme (J.A.F.) and by an Irish Research Council Government of Ireland Postgraduate Scholarship, under project no. GOIPG/2020/1056 (L.W.). Intentional human eDNA research was funded by D.J.D.’s University of Florida start-up funds. M.R.S. was supported by an EMBO long-term fellowship (ALTF 544-2021). We thank M. Q. Martindale, N. Condron, K. Yetsko, S. Creer, A. Pacetti, E. Ryan, C. Eastman and all of our generous co-authors on the wildlife and pathogen research papers for which these HGB samples were originally generated8,17. We thank P. Murphy and R. Rolfe for facilitating eDNA extraction of Irish samples in their lab in the Zoology Department, Trinity College Dublin, and A. Krstic, W. Kolch, A. G. Munoz and the Conway Core Facilities staff for facilitating qPCR of the Irish samples at Systems Biology Ireland and the Conway Institute of Biomolecular and Biomedical Research at University College Dublin. We also thank F. Duffy and I. Duffy for assistance with sampling; M. ten Cate and A. Whilde for gifting supplies; K. Foote, A. Rich and the NPS staff of the Fort Matanzas National Monument for valuable assistance with permitting and facilitating access to Rattlesnake Island and site selection; and S. Loesgen (University of Florida) for the SH-SY5Y cells. Finally, we thank the anonymous study participants who permitted us to collect their footprints and room air for human eDNA analysis, with full ethical approval and informed consent

Inadvertent human genomic bycatch and intentional capture raise beneficial applications and ethical concerns with environmental DNA

The feld of environmental DNA (eDNA) is advancing rapidly, yet human  eDNA applications remain underutilized and underconsidered. Broader  adoption of eDNA analysis will produce many well-recognized benefts  for pathogen surveillance, biodiversity monitoring, endangered and  invasive species detection, and population genetics. Here we show that  deep-sequencing-based eDNA approaches capture genomic information  from humans (Homo sapiens) just as readily as that from the intended  target species. We term this phenomenon human genetic bycatch (HGB).  Additionally, high-quality human eDNA could be intentionally recovered  from environmental substrates (water, sand and air), holding promise for  benefcial medical, forensic and environmental applications. However,  this also raises ethical dilemmas, from consent, privacy and surveillance  to data ownership, requiring further consideration and potentially novel  regulation. We present evidence that human eDNA is readily detectable from  ‘wildlife’ environmental samples as human genetic bycatch, demonstrate  that identifable human DNA can be intentionally recovered from  human-focused environmental sampling and discuss the translational and  ethical implications of such fndings. </p

Plastic ingestion in post-hatchling sea turtles: assessing a major threat in Florida near shore waters

Pollution from anthropogenic marine debris, particularly buoyant plastics, is ubiquitous across marine ecosystems. Due to the persistent nature of plastics in the environment, their buoyancy characteristics, degradation dynamics, and ability to mimic the behavior of natural prey, there exists significant opportunity for marine organisms to ingest these man-made materials. In this study we examined gastrointestinal (GI) tracts of 42 post-hatchling loggerhead (Caretta caretta) sea turtles stranded in Northeast Florida. Necropsies revealed abundant numbers of plastic fragments ranging from 0.36 to 12.39 mm in size (length), recovered from the GI tracts of 39 of the 42 animals (92.86%), with GI burdens ranging from 0 to 287 fragments with a mass of up to 0.33 g per turtle. Post-hatchlings weighed from 16.0 to 47.59 g yielding a plastic to body weight percentage of up to 1.23%. Several types of plastic fragments were isolated, but hard fragments and sheet plastic were the most common type, while the dominant frequency of fragment color was white. Fragment size and abundance mixed with natural gut contents suggests significant negative health consequences from ingestion in animals at this life stage. Gaining greater insight into the prevalence of plastic ingestion, the types of plastic and the physiological effects of plastic consumption by multiple life-stages of sea turtles will aid the prioritization of mitigation efforts for the growing marine debris problem. This report demonstrates that plastic ingestion is a critical issue for marine turtles from the earliest stages of life

Environmental DNA monitoring of oncogenic viral shedding and genomic profiling of sea turtle fibropapillomatosis reveals unusual viral dynamics

Pathogen-induced cancers account for 15% of human tumors and are a growing concern for endangered wildlife. Fibropapillomatosis is an expanding virally and environmentally coinduced sea turtle tumor epizootic. Chelonid herpesvirus 5 (ChHV5) is implicated as a causative virus, but its transmission method and specific role in oncogenesis and progression is unclear. We applied environmental (e)DNA-based viral monitoring to assess viral shedding as a direct means of transmission, and the relationship between tumor burden, surgical resection and ChHV5 shedding. To elucidate the abundance and transcriptional status of ChHV5 across early, established, regrowth and internal tumors we conducted genomics and transcriptomics. We determined that ChHV5 is shed into the water column, representing a likely transmission route, and revealed novel temporal shedding dynamics and tumor burden correlations. ChHV5 was more abundant in the water column than in marine leeches. We also revealed that ChHV5 is latent in fibropapillomatosis, including early stage, regrowth and internal tumors; higher viral transcription is not indicative of poor patient outcome, and high ChHV5 loads predominantly arise from latent virus. These results expand our knowledge of the cellular and shedding dynamics of ChHV5 and can provide insights into temporal transmission dynamics and viral oncogenesis not readily investigable in tumors of terrestrial species

Ordering and arrangement of deformed red blood cells in flow through microcapillaries

The shapes and alignment of elastic vesicles similar to red blood cells (RBCs) in cylindrical capillary flow are investigated by mesoscopic hydrodynamic simulations. We study the collective flow behavior of many RBCs, where the capillary diameter is comparable to the diameter of the RBCs. Two essential control parameters are the RBC volume fraction (the tube hematocrit, H-T), and the suspension flow velocity. Depending on H-T, flow velocity and capillary radius, the RBC suspension exhibits a disordered phase and two distinct ordered phases, consisting of a single file of parachute-shaped cells and a zigzag arrangement of slipper-shaped cells, respectively. We argue that thermal fluctuations, included in the simulation method, coupled to hydrodynamic flows are important contributors to the RBC morphology. We examine the changes to the phase structures when the capillary diameter and the material properties (bending rigidity kappa and stretching modulus mu) of the model RBCs are varied, constructing phase diagrams for each case. We focus on capillary diameters, which range from about 1.0 to about 1.4 times the RBC long diameter. For the smallest capillary diameter, the single-file arrangement dominates; for the largest diameter, the ordered zigzag arrangement begins to loose its stability and alternates with an asymmetric structure with two lanes of differently oriented cells. In simulations with long capillaries, the coexistence of different phases can be observed

Fibropapillomatosis and chelonid alphaherpesvirus 5 infection in kemp’s ridley sea turtles (lepidochelys kempii)

Fibropapillomatosis (FP), a debilitating, infectious neoplastic disease, is rarely reported in endangered Kemp’s ridley sea turtles (Lepidochelys kempii). With this study, we describe FP and the associated chelonid alphaherpesvirus 5 (ChHV5) in Kemp’s ridley turtles encountered in the United States during 2006–2020. Analysis of 22 case reports of Kemp’s ridley turtles with FP revealed that while the disease was mild in most cases, 54.5% were adult turtles, a reproductively valuable age class whose survival is a priority for population recovery. Of 51 blood samples from tumor-free turtles and 12 tumor samples from turtles with FP, 7.8% and 91.7%, respectively, tested positive for ChHV5 DNA via quantitative polymerase chain reaction (qPCR). Viral genome shotgun sequencing and phylogenetic analysis of six tumor samples show that ChHV5 sequences in Kemp’s ridley turtles encountered in the Gulf of Mexico and northwestern Atlantic cluster with ChHV5 sequences identified in green (Chelonia mydas) and loggerhead (Caretta caretta) sea turtles from Hawaii, the southwestern Atlantic Ocean, and the Caribbean. Results suggest an interspecific, spatiotemporal spread of FP among Kemp’s ridley turtles in regions where the disease is enzootic. Although FP is currently uncommon in this species, it remains a health concern due to its uncertain pathogenesis and potential relationship with habitat degradatio

Molecular characterization of a marine turtle tumor epizootic, profiling external, internal and postsurgical regrowth tumors

Sea turtle populations are under threat from an epizootic tumor disease (animal epidemic) known as fibropapillomatosis. Fibropapillomatosis continues to spread geographically, with prevalence of the disease also growing at many longer-affected sites globally. However, we do not yet understand the precise environmental, mutational and viral events driving fibropapillomatosis tumor formation and progression. Here we perform transcriptomic and immunohistochemical profiling of five fibropapillomatosis tumor types: external new, established and postsurgical regrowth tumors, and internal lung and kidney tumors. We reveal that internal tumors are molecularly distinct from the more common external tumors. However, they have a small number of conserved potentially therapeutically targetable molecular vulnerabilities in common, such as the MAPK, Wnt, TGFβ and TNF oncogenic signaling pathways. These conserved oncogenic drivers recapitulate remarkably well the core pan-cancer drivers responsible for human cancers. Fibropapillomatosis has been considered benign, but metastatic-related transcriptional signatures are strongly activated in kidney and established external tumors. Tumors in turtles with poor outcomes (died/euthanized) have genes associated with apoptosis and immune function suppressed, with these genes providing putative predictive biomarkers. Together, these results offer an improved understanding of fibropapillomatosis tumorigenesis and provide insights into the origins, inter-tumor relationships, and therapeutic treatment for this wildlife epizootic