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Abstract Background A core principle of antimicrobial stewardship (AMS) in veterinary settings is the need for engagement of all stakeholders; however, no studies have addressed the role of veterinary technicians in AMS specifically. The objective of this study was to qualitatively assess knowledge, opinions, and practices related to AMS among technicians. Semi-structured interviews were conducted with 33 veterinary technicians with varied backgrounds, experience and roles. Interviews centered on participants work experience and interactions with their employer, perceptions of antimicrobial resistance and overuse in veterinary medicine, observed application of AMS principles, opinions on potential opportunities for technicians to contribute to AMS and concomitant potential barriers to these opportunities. Transcripts of interviews were coded thematically by two authors, then organized into a hierarchical framework, and the characterization of codes was compared across different categories of respondents. Results Most veterinary technicians were knowledgeable about antimicrobial drugs but could not provide a complete definition of antimicrobial resistance or AMS. Most veterinary technicians could identify examples of antimicrobial misuse. Participants identified areas of client education and discussion with veterinarians as potential areas to contribute to AMS. Barriers identified included hierarchical structures of veterinary practices and time-constraints. Most participants expressed a personal interest in participating in AMS. Conclusions There is a possible appetite among some veterinary technicians to participate in AMS and they already play applicable roles in practices. Barriers such as educational needs, hierarchical structures of veterinary practices and time constraints will need to be addressed if technicians are included in AMS efforts.

Abstract California's Channel Islands are home to two endemic mammalian carnivores: island foxes (Urocyon littoralis) and island spotted skunks (Spilogale gracilis amphiala). Although it is rare for two insular terrestrial carnivores to coexist, these known competitors persist on both Santa Cruz Island and Santa Rosa Island. We hypothesized that examination of their gut microbial communities would provide insight into the factors that enable this coexistence, as microbial symbionts often reflect host evolutionary history and contemporary ecology. Using rectal swabs collected from island foxes and island spotted skunks sampled across both islands, we generated 16S rRNA amplicon sequencing data to characterize their gut microbiomes. While island foxes and island spotted skunks both harbored the core mammalian microbiome, host species explained the largest proportion of variation in the dataset. We further identified intraspecific variation between island populations, with greater differentiation observed between more specialist island spotted skunk populations compared to more generalist island fox populations. This pattern may reflect differences in resource utilization following fine‐scale niche differentiation. It may further reflect evolutionary differences regarding the timing of intraspecific separation. Considered together, this study contributes to the growing catalog of wildlife microbiome studies, with important implications for understanding how eco‐evolutionary processes enable the coexistence of terrestrial carnivores–and their microbiomes–in island environments.

If the recent turbulent times have shown educators anything, it is that we need to be prepared to address our own and our students’ social and emotional needs. However, moments of tension are not the time to start. Rather, students need to be prepared to engage in meaningful ways with skills and competencies. To achieve this state of readiness, teachers can use self-regulation strategies such as the one I call “CHILL.” CHILL is an easy-to-implement five-step process designed to reduce tension in moments of crisis and create the conditions whereby students are prepared to reengage with instruction, both with the teacher and with the class. C is for Calm down, H is for Hear yourself breathe, I is for Investigate your condition, L is for Let yourself know what you need, and the second L is for Let others know what you need. Based on the unique developmental aspects of the adolescent brains, CHILL is designed to support both pre- and in-service teachers as they seek to negotiate tense classroom situations and build skills in self-regulation and resiliency. Importantly, CHILL is a strategy for de-escalation that supports students and teachers in the face of student behavioral challenges.

Goal: The aim of the study herein reported was to review mobile health (mHealth) technologies and explore their use to monitor and mitigate the effects of the COVID-19 pandemic. Methods: A Task Force was assembled by recruiting individuals with expertise in electronic Patient-Reported Outcomes (ePRO), wearable sensors, and digital contact tracing technologies. Its members collected and discussed available information and summarized it in a series of reports. Results: The Task Force identified technologies that could be deployed in response to the COVID-19 pandemic and would likely be suitable for future pandemics. Criteria for their evaluation were agreed upon and applied to these systems. Conclusions: mHealth technologies are viable options to monitor COVID-19 patients and be used to predict symptom escalation for earlier intervention. These technologies could also be utilized to monitor individuals who are presumed non-infected and enable prediction of exposure to SARS-CoV-2, thus facilitating the prioritization of diagnostic testing.

Understanding the distribution of biodiversity along environmental gradients allows us to predict how communities respond to natural and anthropogenic impacts. In fjord ecosystems, the overlap of strong salinity and temperature gradients provides us with the opportunity to assess the spatial variation of biodiversity along abiotic environmental gradients. However, in Northern Chilean Patagonia (NCP), a unique and at the same time threatened fjord system, the variation of macrobenthic communities along abiotic environmental gradients is still poorly known. Here, we tested whether macrobenthic species diversity and community structure followed systematic patterns of variation according to the spatial variation in salinity and temperature in Comau Fjord, NCP. A spatially extensive nested sampling design was used to quantify the abundance of subtidal macrobenthic species along the fjord axis (fjord sections: head, middle, and mouth) and a depth gradient (0–21 m). The vertical structure of the water column was strongly stratified at the head of the fjord, characterized by a superficial (depth to ca. 5 m) low-salinity and relatively colder layer that shallowed and decayed toward the mouth of the fjord. The biotic variation followed, in part, this abiotic spatial pattern. Species richness peaked at high salinities (>27 psu) between 5 and 10 m in the head section and between 15 and 21 m in the middle and mouth sections. Diversity and evenness were also highest at these salinities and depth ranges in the head and middle sections, but at shallower depth ranges in the mouth. Information theory-based model selection provided a strong empirical support to the depth- and section-dependent salinity, but not temperature, effects on the three biodiversity metrics. Erect algae and the edible mussel Aulacomya atra numerically dominated in shallow water (0–3 m) at the head and the middle of the fjord, coinciding with the horizontal extension of the low-density water layer—these taxa were further replaced by the crustose algae Lithothamnion sp. and deep-dwelling suspension filters (e.g., corals, polychaetes, and sponges) along depth gradient. Macrobenthic biodiversity correlated, therefore, with the influence of freshwater inputs and the density-driven stratification of the water column in this ecosystem. The spatially variable (across both, horizontal and vertical fjord axes) thresholds observed in our study question the widely accepted pattern of increasing biodiversity with increasing distance from the head of estuarine ecosystems. Finally, non-linear environmental stress models provide us a strong predictive power to understand the responses of these unique ecosystems to natural and anthropogenic environmental changes.

Purpose. Research shows that smoking during pregnancy is related to mental health diagnoses. The purpose of this study was to assess whether current general mental health status is related to current smoking status in pregnant women after controlling for other factors related to both mental health and tobacco use during pregnancy. Methods. This cross-sectional analysis used 2017 Behavioral Risk Factor Surveillance Survey (BRFSS) data for 621 pregnant women aged 18-38 from Florida (N=136), Kansas (N=116), Minnesota (N=105), Nebraska (N=90), New York (N=78), and Utah (N=96). Multiple logistic regression analysis was used to assess the relationship between current mental health status and current tobacco use, while controlling for state, depression diagnosis, routine checkup, healthcare plan, age, marital status, ethnicity/race, education level, income level, and employment status. Results. Overall, very few participants reported current smoking (6%) and about one-third reported low or moderate mental health status in the past 30 days. Adjusted results indicated that those who reported high mental health status were about 3 times less likely (OR=0.29, 95% CI=0.09, 0.88) to report current smoking status compared to those who reported low mental health status. Conclusions. Overall, current mental health status was highly related to current smoking status in pregnant women. Clinicians in obstetrics may expect a very low proportion of pregnant women to report smoking and up to one-third to report low or moderate current general mental health status. Given that current mental health issues and current tobacco use may harm both mother and child, be highly related in pregnant women, and change throughout the pregnancy, pregnant women should be screened automatically for both at each visit.

Hydrocarbon (i.e. oil) extraction, transport, consumption, and pollution occur daily in marine environments, threatening vulnerable natural resources, habitats, and biodiversity. However, threats of episodic oil spills to imperiled marine species are not as well-studied as more acute, readily apparent threats such as incidental capture in fisheries and habitat degradation. We performed a global review of spill incident databases to evaluate reported effects of oil spills on sea turtles, which are flagship species of the world’s oceans. Our goals were to (1) summarize available information about oil spills and their effects on sea turtles; (2) identify major knowledge gaps; and (3) provide recommendations related to oil spills and sea turtles for managers, researchers, and conservation groups around the world. Over 2000 oil spill incidents of variable magnitude (range: ~20 to >1 million barrels) have occurred worldwide in the past 60 yr in areas where sea turtles are found, but resulting effects on sea turtles have been reported in less than 2% of those incidents. Further, evidence indicates that most effects are related to heavy external oiling, while chemical effects of oil exposure have not been well defined. We recommend that, where available, resources be prioritized to document co-occurrence of turtles and oil, the degree and nature of oil exposure, and mortality and reproductive losses to individual sea turtles and their populations caused by spills and spill response activities. These data will best inform assessments of the extent and magnitude of adverse effects of oil spills on sea turtles.

During the BP Deepwater Horizon (DWH) oil spill in 2010, 319 live oiled sea turtles were rescued and admitted to rehabilitation centers for decontamination and veterinary care. Most turtles were small, surfaceñpelagic juveniles that were collected from oiled habitat distant from shore. Serial hematology, plasma biochemistry, and blood gas analyses were reviewed to characterize abnormalities relative to observed degree of oiling. Clinicopathological abnormalities upon admission indicated acute, nonspecific metabolic and osmoregulatory derangements that were attributable to a combination of events including stress, exertion, physical exhaustion, and dehydration related to oiling, capture, and transport. Specific toxicological effects reported in other taxa were not observed. Initial point-of-care blood data from one rescue center were evaluated using clinical assessment of physiological status for all turtles of all species with available data for pH, pCO2, sodium, and potassium. In addition, a prognostic model that was specifically developed for cold-stunned Kempís ridley sea turtles Lepidochelys kempii was applied to oiled Kempís ridley turtles from one center. Thirty-six percent of oiled turtles were identified as physiologically deranged based on a clinical assessment of their physiological status, and 25% of oiled Kempís ridley sea turtles exceeded the mortality risk threshold of the prognostic model. These results indicate that the physiological derangements in these animals were relatively severe and clinically relevant. Based on observations during the DWH spill, adverse physiological effects in sea turtles may be an important consequence of stress, exertion, physical exhaustion, and dehydration secondary to oiling, capture, and transport.

The Deepwater Horizon (DWH) oil spill was unprecedented in extent and duration, and affected marine natural resources, including sea turtles, throughout the northern Gulf of Mexico. Consequently, US federal and state Trustees documented and quantified oil exposure and resulting injuries to sea turtles under the DWH Natural Resource Damage Assessment. At-sea rescue operations focused on surface-pelagic juvenile sea turtles, which were especially at risk to oil exposure within oceanic convergence zones, and provided direct observations of the degree that turtles in this young life stage were exposed to DWH oil. In contrast, locations of larger neritic juvenile and adult turtles were documented during aerial surveys, but because these turtles were not captured, their oiling status could not be directly evaluated. Both the rescue operations and aerial surveys were able to observe only a small fraction of sea turtles within the vast spill footprint. We developed a spatio-temporally explicit approach that used direct observations of oiled surface-pelagic juvenile sea turtles and satellite-derived surface oil distributions to statistically estimate the probabilities of oil exposure for all sea turtles that were present within the area of the DWH spill, but whose oiling status was unknown. Our results enabled an expansion of exposure and injury quantification across the entire DWH spill area and period. This approach was conceptually straightforward and used common geospatial and statistical techniques, making it applicable to other situations in which the full extent of oil exposure for marine natural resources must be estimated from an incomplete sample.

Documentation of exposure of threatened and endangered sea turtles to petroleum and dispersant released into the northern Gulf of Mexico (GoM) during the Deepwater Horizon (DWH) oil spill was a critical component of the Natural Resource Damage Assessment (NRDA) process. Substances collected from the skin of oiled and suspected oiled turtles were analyzed for petroleum hydrocarbons to determine oiling status and oil sources. Tissue, gastroenteric and bile samples from a subset of visibly oiled and unoiled turtles that died during the spill in 2010 and in 2011 were analyzed for evidence of internal exposure and absorption of polycyclic aromatic hydrocarbons (PAHs) and the dispersant component dioctyl sodium sulfosuccinate (DOSS). The volume of external oil collected from sea turtles was sufficient to confirm the presence of petroleum on 61% of turtles, and oil from the DWH spill was identified as the source in 97% of those turtles in which conclusive comparison was possible. Visibly oiled turtles had higher concentrations of tissue PAH or biliary fluorescent PAH metabolites compared to those determined in unoiled animals. Findings in most of the unoiled turtles were suggestive of low-level PAH exposure from various sources that may represent background values for sea turtles from the northern GoM. DOSS levels were below the limit of quantitation in all samples analyzed except in an esophagus sample of a heavily oiled sea turtle. Overall, the results for petroleum or petroleum-derived compounds of both external and internal samples of sea turtles supported visual observations of oiling.

The 2010 Deepwater Horizon (DWH) oil spill posed a severe threat to surface-pelagic sea turtles because the surface convergence zones, which provide vital habitat by aggregating pelagic Sargassum and other floating material, also aggregated floating oil. Following the DWH spill, turtle rescue operations between 17 May and 9 September 2010 documented 937 juvenile sea turtles in the spill area and examined 574 captured turtles. Of the captured turtles, 81% were visibly oiled. Transect searches in convergence zones found Kempís ridleys (51% of individuals), green turtles (37%), loggerheads (7%), hawksbills (2%), and unidentified sea turtles (2%). Line-transect methods estimated the density of all surface-pelagic sea turtles in surface convergence zones to be 3.32 km-2 (95% CI = 2.82ñ3.88), and the density of heavily oiled turtles to be 0.24 km-2 (95% CI = 0.15ñ0.39). Turtle densities and the areal extent of heavy oiling probability were used to estimate total number of turtles exposed to DWH oil. We estimate approximately 402000 surface-pelagic sea turtles were exposed, and of those, 54800 were likely to have been heavily oiled. Our estimates formed the basis of surface-pelagic juvenile sea turtle mortality estimates included in the DWH natural resource damage assessment.

Objective: To describe the presentation, clinical findings, diagnosis, treatment, and outcome of cases of trismus (lockjaw) in cold-stunned sea turtles., Animals: 4 Kemp's ridley (Lepidochelys kempii) and 1 loggerhead (Caretta caretta) sea turtle., Methods: Cold-stunned sea turtles that presented with difficulty or inability to open their jaw between 2009 and 2023 were included. Information retrieved from medical records included signalment, physical exam findings, diagnostic information, definitive diagnosis via either advanced imaging or histopathology, treatment, and clinical outcome., Results: Turtles presented between 4 and 48 days into rehabilitation. Three were diagnosed by advanced imaging (CT or MRI), and 2 were diagnosed based on clinical signs and postmortem histopathology. Treatment was multimodal and consisted of antibiotics (5/5), nonsteroidal anti-inflammatories (5/5), vitamin E (3/5), intralesional steroid therapy (3/5), acupuncture (3/5), antifungals (2/5), anti-inflammatory parenteral steroids (2/5), physical therapy (2/5), therapeutic laser (2/5), and supportive feeding via either total parenteral nutrition (1/5), or tube feedings (2/5). Two animals were released, 2 died naturally, and 1 was euthanized., Clinical Relevance: Trismus (lockjaw) is an uncommon finding in stranded cold-stunned sea turtles that can have a significant impact on animal welfare if not diagnosed and treated. This report describes the condition to aid clinical case management and resource allocation in rehabilitation facilities.

Viruses from a new species of piscichuvirus were strongly associated with severe lymphocytic meningoencephalomyelitis in several free-ranging aquatic turtles from 3 coastal US states during 2009-2021. Sequencing identified 2 variants (freshwater turtle neural virus 1 [FTuNV1] and sea turtle neural virus 1 [STuNV1]) of the new piscichuvirus species in 3 turtles of 3 species. In situ hybridization localized viral mRNA to the inflamed region of the central nervous system in all 3 sequenced isolates and in 2 of 3 additional nonsequenced isolates. All 3 sequenced isolates phylogenetically clustered with other vertebrate chuvirids within the genus Piscichuvirus. FTuNV1 and STuNV1 shared ≈92% pairwise amino acid identity of the large protein, which narrowly places them within the same novel species. The in situ association of the piscichuviruses in 5 of 6 turtles (representing 3 genera) with lymphocytic meningoencephalomyelitis suggests that piscichuviruses are a likely cause of lymphocytic meningoencephalomyelitis in freshwater and marine turtles.

An adult male Kemp's ridley turtle was found dead on the coast of Kenedy County, Texas, in August 2019 with bilateral severe, diffuse granulomatous nephritis. Pan-bacterial 16S rRNA gene polymerase chain reaction (PCR) and amplicon sequencing of affected tissue indicated the presence of a Neorickettsia. Neorickettsia is a genus of obligate intracellular Alphaproteobacteria that are transmitted by digenean trematodes. For further characterization, primers were designed to amplify and sequence the groEL gene. Phylogenetic analysis found that the organism was distinct from other known species to a degree consistent with a novel species. Immunohistochemistry using an antibody directed against a Neorickettsia surface protein showed bacterial clusters within the renal granulomas. A species-specific quantitative PCR was designed and detected the organism within the liver and colon of the index case. A quantitative PCR survey of grossly normal kidneys opportunistically collected from additional stranded sea turtle kidneys detected this organism in five of 15 Kemp's ridley turtles, two of nine green turtles, and neither of two loggerhead turtles. Recognition of this novel organism in an endangered species is concerning; additional work is underway to further characterize the potential of this organism as a pathogen of sea turtles., (© 2022 Wiley-VCH GmbH.)

This report documents cases of fatal pulmonary mycosis caused by entomopathogenic fungi in the genera Metarhizium and Beauveria (Order Hypocreales) in a loggerhead sea turtle ( Caretta caretta ), a Chinese alligator ( Alligator sinensis ), two gopher tortoises ( Gopherus polyphemus ), a Cuvier's dwarf caiman ( Paleosuchus palpebrosus ), a false gharial ( Tomistoma schlegelii ), a green sea turtle ( Chelonia mydas ), and a Kemp's ridley sea turtle ( Lepidochelys kempii ), and a case of granulomatous coelomitis in a hawksbill sea turtle ( Eretmochelys imbricata ). Fungi identified in these cases included Beauveria bassiana , Beauveria brongniartii , Metarhizium anisopliae , Metarhizium robertsii , and one case of infection by a novel Metarhizium species. The animals were either housed at zoos or brought into rehabilitation from the wild. Although the majority of animals had comorbidities, the fungal infections were believed to be the primary cause of death. Fungal susceptibility testing was performed on two Beauveria spp. isolates, and revealed lower minimum inhibitory concentrations for itraconazole and voriconazole when compared to terbinafine and fluconazole. This case series demonstrates that a variety of reptile species from different orders are vulnerable to infection with Metarhizium, and multiple species of sea turtle are susceptible to infection with Beauveria .

Few aquatic animal negative-sense RNA viruses have been characterized, and their role in disease is poorly understood. Here, we describe a virus isolated from diseased freshwater turtles from a Florida farm in 2007 and from an ongoing epizootic among free-ranging populations of Florida softshell turtles (Apalone ferox), Florida red-bellied cooters (Pseudemys nelsoni), and peninsula cooters (Pseudemys peninsularis). Affected turtles presented with similar neurological signs, oral and genital ulceration, and secondary microbial infections. Microscopic lesions were most severe in the softshell turtles and included heterophilic/histiocytic meningoencephalitis, multi-organ vasculitis, and cytologic observation of leukocytic intracytoplasmic inclusions. The virus was isolated using Terrapene heart (TH-1) cells. Ultrastructurally, viral particles were round to pleomorphic and acquired an envelope with prominent surface projections by budding from the cell membrane. Viral genomes were sequenced from cDNA libraries of two nearly identical isolates and determined to be bi-segmented, with an ambisense coding arrangement. The larger segment encodes a predicted RNA-directed RNA polymerase (RdRP) and a putative zinc-binding matrix protein. The smaller segment encodes a putative nucleoprotein and an envelope glycoprotein precursor (GPC). Thus, the genome organization of this turtle virus resembles that of arenaviruses. Phylogenetic analysis shows that the RdRP of the turtle virus is highly diverged from the RdRPs of all known negative-sense RNA viruses and forms a deep branch within the phylum Negarnaviricota, that is not affiliated with any known group of viruses, even at the class level. In contrast, the GPC protein of the turtle virus is confidently affiliated with homologs from a distinct group of fish hantaviruses. Thus, the turtle virus is expected to become the founder of a new taxon of negative-sense RNA viruses, at least with a family rank, but likely, an order or even a class. These viruses probably evolved either by reassortment or by intrasegment recombination between a virus from a distinct branch of negarnaviruses distant from all known groups and a hanta-like aquatic virus. We suggest the provisional name Tosoviridae for the putative new family, with Turtle fraservirus 1 (TFV1) as the type species within the genus Fraservirus. A conventional RT-PCR assay, targeting the TFV1 RdRP, confirmed the presence of viral RNA in multiple tissues and exudates from diseased turtles. The systemic nature of the TFV1 infection was further supported by labeling of cells within lesions using in situ hybridization targeting the RNA of the TFV1 RdRP., Competing Interests: The authors have declared that no competing interests exist.

Objective: To characterize osteolytic lesions in cold-stunned Kemp's ridley sea turtles (Lepidochelys kempii) hospitalized for rehabilitation and describe methods used for the management of such lesions., Animals: 25 stranded, cold-stunned Kemp's ridley sea turtles hospitalized between 2008 and 2018., Procedures: Medical records of sea turtles with a diagnosis of osteolytic lesions were reviewed retrospectively to obtain the date of diagnosis, clinical signs, radiographic findings, microbial culture results, hematologic and plasma biochemical data, cytologic and histologic findings, antimicrobial history, time to first negative culture result, treatment duration, and outcome., Results: Lesions were identified radiographically a median of 50 days after admission and were located within epiphyses or metaphyses of various appendicular joints. Lesions were associated with periarticular swelling (n = 24), lameness (16), lethargy (2), and hyporexia (2). Bacterial culture yielded growth of single organisms (n = 16), multiple organisms (2), or no growth (6). Significant differences in hematologic and biochemical data were detected between the times of diagnosis and convalescence. Cytologic and histologic findings characterized the lesions as osteomyelitis leading to septic arthritis. Sixteen sea turtles were managed medically, and 8 were managed medically and surgically. Surgery resulted in rapid improvement in joint mobility and overall clinical status. Most (22/25 [88%]) sea turtles survived and were released after long-term management., Conclusions and Clinical Relevance: During rehabilitation, cold-stunned Kemp's ridley sea turtles may be affected by osteomyelitis. Medical management based on antimicrobial susceptibility testing was effective for most turtles. Long term management efforts in turtles are justified by high survival rate.

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 degradation.

Sea turtles, like other air-breathing diving vertebrates, commonly experience significant gas embolism (GE) when incidentally caught at depth in fishing gear and brought to the surface. To better understand why sea turtles develop GE, we built a mathematical model to estimate partial pressures of N 2 (PN 2 ), O 2 (PO 2 ), and CO 2 (PCO 2 ) in the major body-compartments of diving loggerheads ( Caretta caretta ), leatherbacks ( Dermochelys coriacea ), and green turtles ( Chelonia mydas ). This model was adapted from a published model for estimating gas dynamics in marine mammals and penguins. To parameterize the sea turtle model, we used values gleaned from previously published literature and 22 necropsies. Next, we applied this model to data collected from free-roaming individuals of the three study species. Finally, we varied body-condition and cardiac output within the model to see how these factors affected the risk of GE. Our model suggests that cardiac output likely plays a significant role in the modulation of GE, especially in the deeper diving leatherback turtles. This baseline model also indicates that even during routine diving behavior, sea turtles are at high risk of GE. This likely means that turtles have additional behavioral, anatomical, and/or physiologic adaptions that serve to reduce the probability of GE but were not incorporated in this model. Identifying these adaptations and incorporating them into future iterations of this model will further reveal the factors driving GE in sea turtles., Competing Interests: AF was employed without salary by the company Global Diving Research Inc. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Robinson, García-Párraga, Stacy, Costidis, Blanco, Clyde-Brockway, Haas, Harms, Patel, Stacy and Fahlman.)

The pathogenesis of steatitis that infrequently occurs in cold-stunned Kemp’s ridley sea turtles (KRT; Lepidochelys kempii ) has been undetermined. The objectives of this study were to investigate the clinical ( n = 23) and histologic findings ( n = 11) in cold-stunned KRT, and to compare plasma concentrations of α-tocopherol (vitamin E), thiobarbituric acid reactive substances (TBARS), and the TBARS to vitamin E (T/E) ratio (an assessment of oxidative stress) between cold-stunned KRT with clinically and/or histologically confirmed steatitis ( n = 10) and free-ranging KRT ( n = 9). None of the cold-stunned turtles had clinically detectable steatitis at admission, and the median number of days to diagnosis of steatitis was 71 (range 33–469). Histologic findings of affected adipose tissue included heterophilic ( n = 9) and/or histiocytic ( n = 5) steatitis, fat necrosis ( n = 7), myonecrosis ( n = 2), and intralesional bacteria ( n = 6). Cold-stunned KRT had significantly lower plasma vitamin E concentrations (median = 3.5 nmol/g), lower plasma TBARS concentrations (median = 1.6 nmol/g), and higher T/E ratios (median = 0.50), than controls (62.3 nmol/g; 2.1 nmol/g; 0.03, respectively). These results suggest a multifactorial etiology for the development of steatitis in KRT during rehabilitation, including tissue injury, septicemia, and various factors resulting in imbalances of anti-/oxidative status. By highlighting the need to provide more effective vitamin E supplementation, and the need to re-assess specific components of the diet, this study may lead to reduced incidence and improved medical management of steatitis in cold-stunned sea turtles.

Sea turtle fibropapillomatosis (FP) is a tumor promoting disease that is one of several threats globally to endangered sea turtle populations. The prevalence of FP is highest in green sea turtle ( Chelonia mydas ) populations, and historically has shown considerable temporal growth. FP tumors can significantly affect the ability of turtles to forage for food and avoid predation and can grow to debilitating sizes. In the current study, based in South Texas, we have applied transcriptome sequencing to FP tumors and healthy control tissue to study the gene expression profiles of FP. By identifying differentially expressed turtle genes in FP, and matching these genes to their closest human ortholog we draw on the wealth of human based knowledge, specifically human cancer, to identify new insights into the biology of sea turtle FP. We show that several genes aberrantly expressed in FP tumors have known tumor promoting biology in humans, including CTHRC1 and NLRC5 , and provide support that disruption of the Wnt signaling pathway is a feature of FP. Further, we profiled the expression of current targets of immune checkpoint inhibitors from human oncology in FP tumors and identified potential candidates for future studies., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Blackburn, Leandro, Nahvi, Devlin, Leandro, Martinez Escobedo, Peralta, George, Stacy, deMaar, Blangero, Keniry and Curran.)

Injuries inflicted by sharks are a frequent observation in stranded sea turtles. Sharks prey on live turtles and scavenge carcasses, which can create uncertainty as to the cause of stranding when sea turtles are found dead with shark-bite wounds. Consequently, attributing the cause of stranding to a shark attack based purely on the presence of the characteristic wounds can overestimate predation by sharks as a cause of mortality. To better characterize the timing of shark-bite wounds relative to death of sea turtles in the southeastern USA, we performed necropsies on 70 stranded turtles that were found dead in which the predominant observation was bite wounds without any grossly evident vital responses (inflammation or healing). Postmortem examination included assessment for evidence of exsanguination and histopathological evaluation of skeletal muscle comprising wound margins. We characterized wounds as antemortem, perimortem, or postmortem based on specific criteria related to the presence or absence of supravital and intravital responses. Most (80%) shark-bite wounds were postmortem, 10% were antemortem, and 10% were perimortem. We found that antemortem and postmortem wounds were similar in extent and location except for wounds that primarily involved the shell, which were never found in cases of scavenging. For sea turtles found dead in the southeastern USA, our findings suggest that most shark-bite wounds without externally evident vital responses are due to scavenging. Additionally, this scavenging can significantly damage a carcass, potentially obscuring the detection of other causes of mortality. These findings should be considered when using data derived from stranded sea turtles to conduct mortality assessments.

Decompression sickness (DCS) has been described mainly in loggerhead turtles Caretta caretta bycaught in trawls and gillnets. Here we present cases of gas emboli (GE) in 8 green turtles Chelonia mydas and 2 Kemp's ridleys Lepidochelys kempii entrained in hopper dredges that were working at 8.8-15.2 m depths during shipping channel maintenance or beach renourishment activities. Turtle weights ranged from 2.2 to 6.7 kg. All were found alive with blunt force injuries from passage through the dredge and were taken to rehabilitation facilities. Four green turtles died or were euthanized within 24 h. Six turtles survived. Radiographic or ultrasonographic evidence of GE was detected in 4 turtles, including 3 mortalities. Computed tomography (CT) revealed perirenal and cervical GE in 4 turtles, including 1 mortality. No GE were detected in 2 of the survivors. Upon necropsy, GE were found in mesenteric vessels, the right atrium, and kidneys. Histopathology confirmed that tissues were in a good state of preservation without evidence of bacterial overgrowth or putrefactive gas formation. Death likely resulted primarily from massive tissue trauma from the dredge, but moderate GE could have led to DCS and complicated recovery. The surviving turtles weighed less than those that did not survive. Besides hypothesized stress/exercise-induced circulatory changes of blood through the lungs and pressure reduction of forced surfacing from depth, drastic pressure change within the dredge pipes before and after the pump could contribute to GE. Hopper dredge entrainment is an additional cause of GE and potential DCS in sea turtles.

Fish within the family Tetraodontidae are potential sources of both endogenous tetrodotoxins (TTXs) and dietary derived saxitoxins (STXs). Ingestion of fish tissues containing these toxins by other vertebrates can lead to severe illness and death. The Caribbean sharpnose puffer ( Canthigaster rostrata ) is a widespread tetraodontid species within the western Atlantic. Mass settlement of juveniles into foraging habitats have been associated with large-scale puffer fish mortality events. In 2013, 2014, and 2017, puffer mortality events on the southern Caribbean coast of Costa Rica were also associated with strandings of green turtles ( Chelonia mydas ) found to have fed on C. rostrata . Stranded sea turtles were found dead without apparent cause or alive with severe neurological signs that resolved during short periods of captivity. Puffer fish and turtle organ samples were analyzed for both TTXs and STXs. Concentrations of TTXs were extremely low in the fish (0.5-0.7 μg/g) and undetectable in turtle stomach contents. However, concentrations of STXs in whole fish (16.6-47.5 μg STX-eq/g) exceeded the 0.8 μg STX-eq/g human seafood safety threshold for STXs by orders of magnitude. Saxitoxins were also detected in samples of stomach contents (ingested fish), brain, lung, kidney, and serum from three affected turtles. Study results indicate that saxitoxicosis resulting from opportunistic foraging on C. rostrata during fish mortality events may be a significant factor in episodic stranding of green sea turtles in this region., (Copyright © 2019 Barrientos, Hernández-Mora, Alegre, Field, Flewelling, McGrath, Deeds, Chacón, Rojas Arrieta, Vargas, Artavia and Stacy.)

Protozoa morphologically consistent with Caryospora sp. are one of the few pathogens associated with episodic mass mortality events involving free-ranging sea turtles. Parasitism of green turtles ( Chelonia mydas ) by these coccidia and associated mortality was first reported in maricultured turtles in the Caribbean during the 1970s. Years later, epizootics affecting wild green turtles in Australia occurred in 1991 and 2014. The first clinical cases of Caryospora -like infections reported elsewhere in free-ranging turtles were from the southeastern US in 2012. Following these initial individual cases in this region, we documented an epizootic and mass mortality of green turtles along the Atlantic coast of southern Florida from November 2014 through April 2015 and continued to detect additional, sporadic cases in the southeastern US in subsequent years. No cases of coccidial disease were recorded in the southeastern US prior to 2012 despite clinical evaluation and necropsy of stranded sea turtles in this region since the 1980s, suggesting that the frequency of clinical coccidiosis has increased here. Moreover, we also recorded the first stranding associated with infection by a Caryospora -like organism in Hawai'i in 2018. To further characterize the coccidia, we sequenced part of the 18S ribosomal and mitochondrial cytochrome oxidase I genes of coccidia collected from 62 green turtles found in the southeastern US and from one green turtle found in Hawai'i. We also sequenced the ribosomal internal transcribed spacer regions from selected cases and compared all results with those obtained from Caryospora -like coccidia collected from green turtles found in Australia. Eight distinct genotypes were represented in green turtles from the southeastern US. One genotype predominated and was identical to that of coccidia collected from the green turtle found in Hawai'i. We also found a coccidian genotype in green turtles from Florida and Australia with identical 18S and mitochondrial sequences, and only slight inter-regional differences in the internal transcribed spacer 2. We found no evidence of geographical structuring based on phylogenetic analysis. Low genetic variability among the coccidia found in green turtle populations with minimal natural connectivity suggests recent interoceanic dissemination of these parasites, which could pose a risk to sea turtle populations., (Copyright © 2019 Stacy, Chapman, Stockdale-Walden, Work, Dagenais, Foley, Wideroff, Wellehan, Childress, Manire, Rodriguez, Zachariah, Staggs, Zirkelbach, Nahvi, Crowder, Boylan, Marquardt, Pelton and Norton.)

Salmonella spp. are frequently shed by wildlife including turtles, but S. enterica subsp. enterica serovar Typhimurium or lesions associated with Salmonella are rare in turtles. Between 1996 and 2016, we necropsied 127 apparently healthy pelagic olive ridley turtles (Lepidochelys olivacea) that died from drowning bycatch in fisheries and 44 live or freshly dead stranded turtles from the west coast of North and Central America and Hawaii. Seven percent (9/127) of pelagic and 47% (21/44) of stranded turtles had renal granulomas associated with S. Typhimurium. Stranded animals were 12 times more likely than pelagic animals to have Salmonella-induced nephritis suggesting that Salmonella may have been a contributing cause of stranding. S. Typhimurium was the only Salmonella serovar detected in L. olivacea, and phylogenetic analysis from whole genome sequencing showed that the isolates from L. olivacea formed a single clade distinct from other S. Typhimurium. Molecular clock analysis revealed that this novel clade may have originated as recently as a few decades ago. The phylogenetic lineage leading to this group is enriched for non-synonymous changes within the genomic area of Salmonella pathogenicity island 1 suggesting that these genes are important for host adaptation.

Data on Karenia brevis red tides (≥105 cells l-1) and on dead or debilitated (i.e. stranded) Kemp's ridleys Lepidochelys kempii, loggerheads Caretta caretta, green turtles Chelonia mydas, hawksbills Eretmochelys imbricata, and leatherbacks Dermochelys coriacea documented in Florida during 1986-2013 were evaluated to assess red tides as a sea turtle mortality factor. Unusually large numbers of stranded sea turtles were found coincident with red tides primarily along Florida's Gulf coast but also along a portion of Florida's Atlantic coast. These strandings were mainly adult and large immature loggerheads and Kemp's ridleys, and small immature green turtles and hawksbills. Unusually large numbers of stranded leatherbacks never coincided with red tide. For the 3 most common species, results of stranding data modeling, and of investigations that included determining brevetoxin concentrations in samples collected from stranded turtles, all indicated that red tides were associated with greater and more frequent increases in the numbers of stranded loggerheads and Kemp's ridleys than in the number of stranded green turtles. The mean annual number of stranded sea turtles attributed to K. brevis red tide was 80 (SE = 21.6, range = 2-338). Considering typical stranding probabilities, the overall mortality was probably 5-10 times greater. Red tide accounted for a substantial portion of all stranded loggerheads (7.1%) and Kemp's ridleys (17.7%), and a smaller portion of all stranded green turtles (1.6%). Even though K. brevis red tides occur naturally, the mortality they cause needs to be considered when managing these threatened and endangered species.

Five green (Chelonia mydas) and 11 Kemp's ridley (Lepidochelys kempii) sea turtles found dead, or that died soon after stranding, on the southern Texas (USA) coast during 2 Karenia brevis blooms (October 2015, September-October 2016) were tested for exposure to brevetoxins (PbTx). Tissues (liver, kidney) and digesta (stomach and intestinal contents) were analyzed by ELISA. Three green turtles found alive during the 2015 event and 2 Kemp's ridley turtles found alive during the 2016 event exhibited signs of PbTx exposure, including lethargy and/or convulsions of the head and neck. PbTx were detected in 1 or more tissues or digesta in all 16 stranded turtles. Detected PbTx concentrations ranged from 2 to >2000 ng g-1. Necropsy examination and results of PbTx analysis indicated that 10 of the Kemp's ridleys and 2 of the green turtles died from brevetoxicosis via ingestion. This is the first documentation of sea turtle mortality in Texas attributed to brevetoxicosis.

Neospirorchis (Digenea: "Spirorchiidae") are blood flukes of sea turtles. Trematodes tentatively identified as Neospirorchis sp. infect various sites within sea turtles inhabiting waters of the southeastern United States, but efforts to obtain specimens adequate for morphologic study has proven difficult. Two genetic targets, the internal transcribed spacer region of the ribosomal RNA gene and the partial mitochondrial cytochrome c oxidase subunit I gene, were used to investigate potential diversity among parasite specimens collected from stranded sea turtles. Sequence data were obtained from 215 trematode and egg specimens collected from 92 individual free-ranging cheloniid sea turtles comprising 4 host species. Molecular analysis yielded more than 20 different genotypes. We were able to assign 1 genotype to 1 of the 2 recognized species, Neospirorchis pricei Manter and Larson, 1950 . In many examples, genotypes exhibited host and site specificity. Our findings indicate considerable diversity of parasites resembling Neospirorchis with evidence of a number of uncharacterized blood flukes that require additional study.

Incidental capture, or 'bycatch' in fishing gear is a major global threat to sea turtle populations. A recent study showed that underwater entrapment in fishing gear followed by rapid decompression may cause gas bubble formation within the blood stream (embolism) and tissues leading to organ injury, impairment, and even mortality in some bycaught individuals. We analyzed data from 128 capture events using logistic and ordinal regression to examine risk factors associated with gas embolism in sea turtles captured in trawls and gillnets. Likelihood of fatal decompression increases with increasing depth of gear deployment. A direct relationship was found between depth, risk and severity of embolism, which has not been previously demonstrated in any breath-hold diving species. For the trawl fishery in this study, an average trawl depth of 65 m was estimated to result in 50% mortality in by-caught turtles throughout the year. This finding is critical for a more accurate estimation of sea turtle mortality rates resulting from different fisheries and for devising efforts to avoid or minimize the harmful effects of capture.

Blood flukes of the family Spirorchiidae are significant pathogens of both free-ranging and captive marine turtles. Despite a significant proportion of marine turtle mortality being attributable to spirorchiid infections, details of their life cycles remain almost entirely unknown. Here we report on the molecular elucidation of the complete life cycle of a marine spirorchiid, identified as Amphiorchis sp., infecting vermetid gastropods and captive hatched neonate Caretta caretta in the Oceanogràfic Aquarium, in Valencia, Spain. Specimens of a vermetid gastropod, Thylaeodus cf. rugulosus (Monterosato, 1878), collected from the aquarium filtration system housing diseased C. caretta, were infected with sporocysts and cercariae consistent with the family Spirorchiidae. We generated rDNA sequence data [internal transcribed spacer 2 (ITS2) and partial 28S rDNA] from infections from the vermetid which were identical to sequences generated from eggs from the serosa of the intestine of neonate C. caretta, and an adult spirorchiid from the liver of a C. caretta from Florida, USA. Given the reliability of these markers in the delineation of trematode species, we consider all three stages to represent the same species and tentatively identify it as a species of Amphiorchis Price, 1934. The source of infection at the Oceanogràfic Foundation Rehabilitation Centre, Valencia, Spain, is inferred to be an adult C. caretta from the western Mediterranean being rehabilitated in the same facility. Phylogenetic analysis suggests that this Amphiorchis sp. is closely related to other spirorchiids of marine turtles (species of Carettacola Manter & Larson, 1950, Hapalotrema Looss, 1899 and Learedius Price, 1934). We discuss implications of the present findings for the control of spirorchiidiasis in captivity, for the better understanding of epidemiology in wild individuals, and the elucidation of further life cycles., (Copyright © 2016 Australian Society for Parasitology. Published by Elsevier Ltd. All rights reserved.)

Mycobacteriosis is infrequently reported in free-ranging sea turtles. Nontuberculous Mycobacterium haemophilum was identified as the causative agent of disseminated mycobacteriosis in a juvenile leatherback turtle (Dermochelys coriacea) that was found stranded on the Atlantic coast of Florida. Disseminated granulomatous inflammation was identified histologically, most notably affecting the nervous system. Identification of mycobacterial infection was based on cytologic, molecular, histologic, and microbiologic methods. Among stranded sea turtles received for diagnostic evaluation from the Atlantic and Gulf of Mexico coasts of the United States between 2004 and 2015, the diagnosis of mycobacteriosis was overrepresented in stranded oceanic-phase juveniles compared with larger size classes, which suggests potential differences in susceptibility or exposure among different life phases in this region. We describe M. haemophilum in a sea turtle, which contributes to the knowledge of diseases of small juvenile sea turtles, an especially cryptic life phase of the leatherback turtle., (© 2016 The Author(s).)

Histologic lesions incidental to the cause of death were observed in the adrenal glands of 17 subadult and adult leatherback sea turtles ( Dermochelys coriacea ) found dead or moribund on or near shore in North America. Round bodies, 250-300 μm in diameter composed of an outer capsule and large multinucleated cells surrounding a central mass of acellular material were distributed throughout the affected glands. Protozoal etiology was suspected based on some resemblance to coccidia; however, features diagnostic for coccidial infection were lacking in all but one case, which had a focal area of adrenalitis containing zoites. A novel eucoccidian partial 18S rRNA genetic sequence was consistently detected in adrenal glands with lesions. With the use of quantitative PCR, a specific area of the V4 region of the coccidian 18S gene was quantified in affected adrenal glands and correlated significantly with density of the histologic lesions. A second distinct, but closely related, 18S sequence was also amplified from the adrenal gland of one turtle and from a fecal sample containing unsporulated coccidian oocysts. The two 18S sequences identified from leatherback sea turtles form a clade within the family Eimeriidae. Further investigation is required to understand better the morphology of the life stages, life cycle, and potential effects of this coccidian parasite on adrenal function.

Leatherback turtles (Dermochelys coriacea) undergo substantial cyclical changes in body condition between foraging and nesting. Ultrasonography has been used to measure subcutaneous fat as an indicator of body condition in many species but has not been applied in sea turtles. To validate this technique in leatherback turtles, ultrasound images were obtained from 36 live-captured and dead-stranded immature and adult turtles from foraging and nesting areas in the Pacific and Atlantic oceans. Ultrasound measurements were compared with direct measurements from surgical biopsy or necropsy. Tissue architecture was confirmed histologically in a subset of turtles. The dorsal shoulder region provided the best site for differentiation of tissues. Maximum fat depth values with the front flipper in a neutral (45-90°) position demonstrated good correlation with direct measurements. Ultrasound-derived fat measurements may be used in the future for quantitative assessment of body condition as an index of health in this critically endangered species.

Leatherback sea turtles are globally distributed and endangered throughout their range. There are limited data available on disease in this species. Initial observations of solitary large intestinal diverticulitis in multiple leatherbacks led to a multi-institutional review of cases. Of 31 subadult and adult turtles for which complete records were available, all had a single exudate-filled diverticulum, as large as 9.0 cm in diameter, arising from the large intestine immediately distal to the ileocecal junction. All lesions were chronic and characterized by ongoing inflammation, numerous intralesional bacteria, marked attenuation of the muscularis, ulceration, and secondary mucosal changes. In three cases, Morganella morganii was isolated from lesions. Diverticulitis was unrelated to the cause of death in all cases, although risk of perforation and other complications are possible., (© The Author(s) 2014.)

Three wild immature green sea turtles Chelonia mydas were found alive but lethargic on the shores of the Indian River Lagoon and Gulf of Mexico in Florida, USA, and subsequently died. Necropsy findings in all 3 turtles included partial occlusion of the trachea by a mass comprised of granulomatous inflammation. Pigmented fungal hyphae were observed within the lesion by histology and were characterized by culture and sequencing of the internal transcribed spacer 2 domain of the rRNA gene and D1/D2 region of the fungal 28s gene. The dematiaceous fungus species Veronaea botryosa was isolated from the tracheal mass in 2 cases, and genetic sequence of V. botryosa was detected by polymerase chain reaction in all 3 cases. Genetic sequencing and fungal cultures also detected other dematiaceous fungi, including a Cladosporium sp., an Ochroconis sp., and a Cochliobolus sp. These cases are the first report of phaeohyphomycosis caused by V. botryosa in wild marine animals.

Wounds were created by incision in skeletal muscle of 2 mixed-breed canine cadavers at multiple time points from 0.5 to 74.5 hours postmortem and were exposed to artificial seawater (35 parts per thousand), 0.9% saline (8 parts per thousand), or freshwater for 24 hours before fixation for histology. Discoid and segmental disintegration of myofibers deep to the severed edges was observed in injuries inflicted within 6.5 hours of death and exposed to 0.9% saline and seawater and was not observed in injuries made at later time points or in other treatments. Exposure to artificial seawater had pronounced effects on histomorphology that markedly diminished with increasing postmortem wounding interval. In a third cadaver, these changes were shown to be detectable with confidence following up to 10 days of submergence in seawater at 22.2°C despite decomposition. These findings are important for evaluation of skeletal muscle injuries that are exposed to seawater, such as those occurring in marine animals, and may assist in recognizing wounds inflicted either antemortem or within the supravital period., (© The Author(s) 2014.)

Abstract We observed predation by river otters (Lontra canadensis) on large numbers of Florida cooter (Pseudemys floridana) and Florida softshell turtles (Apalone ferox) in two small lakes in North Central Florida, USA during a period of unusually low water levels. Carcasses were strewn on the shoreline and accumulated around floating boat docks, where some residents observed turtles being killed. We found 76 carcasses, including predominantly skeletons, and two live, severely injured turtles from one lake; however, numerous remains undoubtedly were unrecovered. The otters frequently eviscerated the turtles and removed the head and one or more appendages, including the phallus of mature males. In skeletal remains, injuries inflicted by otters were nonspecific, indistinguishable from damage caused by scavengers, or easily missed in incomplete carcasses. This report of large-scale mortality of freshwater turtles in Florida suggests that otters could have a significant impact on local turtle populations.

Case information and postmortem examination findings are presented for 11 adult female sea turtles in reproductive form that died in Florida, USA. All had abundant, large vitellogenic follicles, and most were either gravid or had recently nested. Species included six loggerheads (Caretta caretta) and five green turtles (Chelonia mydas). Identified proximate causes of death included falls or entrapment by obstructions on nesting beaches, burial under collapsed dunes, and other traumatic injuries of different causes. Evidence of yolk embolization was found in 10 cases and suspected in an 11th turtle. Ten turtles also had various amounts of free intracoelomic yolk. Although the effects of yolk embolization are uncertain at this time, precedence of pathologic importance in other species suggests that embolism may complicate traumatic injuries, including seemingly minor events.

In 2005 and 2006, the central west Florida coast experienced two intense Karenia brevis red tide events lasting from February 2005 through December 2005 and August 2006 through December 2006. Strandings of sea turtles were increased in the study area with 318 turtles (n = 174, 2005; n = 144, 2006) stranding between 1 January 2005 and 31 December 2006 compared to the 12-yr average of 43 +/- 23 turtles. Live turtles (n = 61) admitted for rehabilitation showed clinical signs including unresponsiveness, paresis, and circling. Testing of biological fluids and tissues for the presence of brevetoxin activity by enzyme-linked immunosorbent assay found toxin present in 93% (52 of 56) of live stranded sea turtles, and 98% (42 of 43) of dead stranded sea turtles tested. Serial plasma samples were taken from several live sea turtles during rehabilitation and toxin was cleared from the blood within 5-80 days postadmit depending upon the species tested. Among dead animals the highest brevetoxin levels were found in feces, stomach contents, and liver. The lack of significant pathological findings in the majority of animals necropsied supports toxin-related mortality.

A seven-year old California sea lion (Zalophus californianus) presented with focally extensive, bilaterally symmetric, proliferative axillary skin lesions and preputial lesions. A second California sea lion in the same population presented with similar proliferative lesions on the underside of the tail. Histopathology revealed epidermal hyperplasia with severe hyperkeratosis, with proliferating keratinocytes forming broad, branching pegs that extended into the dermis. Pan-papillomaviral consensus PCR was used to obtain initial E1 sequence template and the complete genome was determined using a combination of rolling circle amplification and specific-primer PCR. Analysis revealed a novel papillomavirus, Zalophus californianus papillomavirus 1 (ZcPV1), with seven open reading frames encoding five early proteins (E6, E7, E1, E2 and E4) and two late proteins (L1 and L2). Phylogenetic analysis revealed that (ZcPV1) is most closely related to Equine papillomavirus 1 (EcPV1) in the genus Zetapapillomavirus, and Canine papillomaviruses 3 and 4 (CPV3, CPV4) in the genus Chipapillomavirus. The lesions regressed without intervention over a period of several months., (Copyright © 2011 Elsevier B.V. All rights reserved.)

Marine spirorchiid trematodes are associated with morbidity and mortality in sea turtles worldwide. The intermediate hosts remain unknown, and discovery efforts are hindered by the large number and great diversity of potential hosts within sea turtle habitats, as well the potential for low prevalence and overdispersion. A high-throughput DNA extraction and polymerase chain reaction-based method was developed to detect the internal transcribed spacer 2 (ITS2) region of the ribosomal gene of 2 spirorchiid genera, Learedius and Hapalotrema , within pooled samples of gastropod tissues. A model system consisting of freshwater snail ( Pomacea bridgesii ) tissues and DNA extracts spiked with adult Learedius learedi and known quantities of spirorchiid DNA was used to develop and test the technique. Threshold of detection was found to be equivalent to an early prepatent infection within 1.5 g of gastropod tissue. This technique was used to screen approximately 25 species of marine gastropods at a captive facility where green turtles ( Chelonia mydas ) become infected by L. learedi . The parasite was detected in a sample of knobby keyhole limpet ( Fissurella nodosa ), thus providing the first evidence of an intermediate host for a marine spirorchiid trematode. This technique has many potential applications in trematode life cycle discovery studies.

Bacteria of the genus Helicobacter are associated with disease in humans and animals. Reports of infection in reptiles are very limited. In the present study, pathological findings and molecular characterization are reported for a Helicobacter species associated with septicemia in a pancake tortoise (Malacochersus tornieri). Disseminated infection resulted in regional cellulitis and edema of the head and neck, and pericarditis. Spiral bacteria were identified in cytological preparations and Warthin-Starry-stained sections of pathological lesions. Amplification of partial sequence of the 16S ribosomal gene using polymerase chain reaction identified the organism as Helicobacter and suggest that it is a novel species.

Cryptosporidiosis in squamates is well documented, but there is very limited information available on cryptosporidiosis in testudines. We describe three cases of cryptosporidiosis in tortoises with associated pathology. Two Russian tortoises (Agrionemys [Testudo] horsfieldii) and a pancake tortoise (Malacochersus tornieri), all from separate collections, were found dead. At necropsy, two had histological evidence of intestinal cryptosporidiosis and one had gastric cryptosporidiosis. Consensus Cryptosporidium sp. PCR and sequencing was used to identify the Cryptosporidium sp. present in these three tortoises. In the juvenile Russian tortoise with gastric cryptosporidiosis, the organism had 98% homology with a previously reported sequence from an Indian star tortoise isolate. A second chelonian Cryptosporidium sp. was identified in the pancake tortoise and the second Russian tortoise. This sequence was 100% identical to a shorter gene sequence previously reported in a marginated tortoise. This is the first report coordinating pathology with Cryptosporidium characterization in chelonians. The two Cryptosporidium sp. found in tortoises segregate according to site of infection, and there may be further differences in pathology, host range, and transmission. These Cryptosporidium sp. appear to be able to infect diverse tortoise host species. This may be an under-recognized problem in tortoises., ((c) 2010 Elsevier B.V. All rights reserved.)

Spirorchiid trematodes are implicated as an important cause of stranding and mortality in sea turtles worldwide. However, the impact of these parasites on sea turtle health is poorly understood due to biases in study populations and limited or missing data for some host species and regions, including the southeastern United States. We examined necropsy findings and parasitological data from 89 loggerhead Caretta caretta and 59 green turtles Chelonia mydas that were found dead or moribund (i.e. stranded) in Florida (USA) and evaluated the role of spirorchiidiasis in the cause of death. High prevalence of infection in the stranding population was observed, and most infections were regarded as incidental to the cause of death. Spirorchiidiasis was causal or contributory to death in some cases; however, notable host injury and/or large numbers of parasites were observed in some animals, including nutritionally robust turtles, with no apparent relationship to cause of death. New spirorchiid species records for the region were documented and identified genera included Neospirorchis, Hapalotrema, Carettacola, and Learedius. Parasites inhabited and were associated with injury and inflammation in a variety of anatomic locations, including large arteries, the central nervous system, endocrine organs, and the gastrointestinal tract. These findings provide essential information on the diversity of spirorchiids found in Florida sea turtles, as well as prevalence of infection and the spectrum of associated pathological lesions. Several areas of needed study are identified with regard to potential health implications in the turtle host, and findings caution against over-interpretation in individual cases.

In this paper we describe the characterization of the genomes of two sea turtle papillomaviruses, Chelonia mydas PV (CmPV-1) and Caretta caretta PV (CcPV-1). The isolation and sequencing of the first non-avian reptilian PVs extend the evolutionary history of PVs to include all amniotes. PVs have now been described in mammals, birds and non-avian reptiles. The chelonian PVs form a distinct clade most closely related to the avian PVs. Unlike the avian PVs, both chelonian PVs have canonical E6 and E7 ORFs, indicating that these genes were present in the common ancestor to mammalian and non-mammalian amniote PVs. Rates of evolution among the non-mammalian PVs were generally slower than those estimated for mammalian PVs, perhaps due to lower metabolic rates among the ectothermic reptiles.

A wild peninsula ribbon snake (Thamnophis sauritus sackenii) in Florida was found to have hypochromic erythrocytes containing two different types of inclusions: purple granular inclusions, and pale orange or pink crystalloid inclusions that were round, oval, rectangular, or hexagonal in shape. Transmission electron microscopy revealed hexagonal or pleomorphic, homogenous inclusions and enveloped particles morphologically consistent with a member of the Iridoviridae. Histopathology of the animal revealed necrotizing hepatitis consistent with sepsis. Consensus PCR was used to amplify a 628-bp region of iridoviral DNA-dependent DNA polymerase. Bayesian and maximum likelihood phylogenetic analysis found that this virus was distinct from other known iridoviral genera and species, and may represent a novel genus and species.

A subadult loggerhead turtle, Caretta caretta, presented with generalized small, white, raised lesions over its neck, shoulders, and all four flippers. A juvenile green turtle, Chelonia mydas, recently treated for fibropapillomatosis, presented with four similar localized lesions on one flipper. To diagnose the conditions, biopsies of the lesions were taken for histopathology, electron microscopy, and molecular diagnostics. Histopathologic findings were similar in the two turtles and skin lesions were characterized by multifocal areas of epidermal hyperplasia accompanied by variation and abnormalities in the nuclear morphology of keratinocytes and a few intranuclear inclusions in some cells. Transmission electron microscopy revealed multiple epithelial cells with large intranuclear aggregates of virions consistent in morphology with papillomavirus. Papillomavirus was detected in samples from both turtles by polymerase chain reaction (PCR). Sequence analysis of the partial sequence of the papillomavirus E1 gene revealed two viruses (CcPV and CmPV) that were distinct from each other and from other species in Papillomaviridae, and likely represent two novel species and perhaps a new genus.

Eighteen green turtles Chelonia mydas recovered from the Atlantic and Gulf coasts of Florida and Tortuguero National Park, Costa Rica, were diagnosed with renal oxalosis by histopathological examination. Affected sea turtles included 14 adults and 4 immature animals, which comprised 26% (18/69) of green turtle necropsy cases available for review. Calcium oxalate deposition ranged from small to moderate amounts and was associated with granuloma formation and destruction of renal tubules. All affected turtles died from traumatic events or health problems unrelated to renal oxalosis; however, 1 immature turtle had notable associated renal injury. Crystal composition was confirmed by infrared and scanning electron microscopy and energy dispersive X-ray analysis. The source of calcium oxalate is unknown and is presumed to be of dietary origin.