AUTOLOGOUS OR ALLOGENIC BLOOD PLEURODESIS AS TREATMENT FOR PNEUMOCOELOM IN FOUR SEA TURTLES (LEPIDOCHELYS KEMPII, CARETTA CARETTA).

Pneumocoelom secondary to primary pulmonary disease was diagnosed via CT (three cases) or radiographs and coelioscopy (one case) in one Kemp's ridley (Lepidochelys kempii) and three loggerhead (Caretta caretta) sea turtles. All cases were suspected to be caused by trauma to the lung, and all exhibited positive buoyancy disorders. Coelomocentesis alone was ineffective at resolving each pneumocoelom, suggesting a large and persistent tear in the pulmonary parenchyma. An autologous (in three cases) or allogenic (in one case) blood patch pleurodesis was performed. The pneumocoelom was resolved in all four cases with no noted postoperative complications. Follow-up diagnostic imaging confirmed resolution of the pneumocoelom in three cases, and all four patients were successfully rehabilitated and had normal buoyancy at the time of release. One rehabilitated loggerhead was satellite tagged, and the tracking data demonstrated that the animal was still alive and moving normally 126 d postrelease, when the tag stopped transmitting. In sea turtles, an autologous or allogenic blood patch pleurodesis can be safely performed in cases of persistent pneumocoelom secondary to traumatic primary pulmonary disease.

RETROSPECTIVE HEMATOLOGY AND SERUM BIOCHEMISTRY OF RING-TAILED LEMURS (LEMUR CATTA) ON ST. CATHERINES ISLAND, GEORGIA, USA.

Annual health records were retrospectively analyzed for a colony of ring-tailed lemurs (Lemur catta) inhabiting St. Catherines Island, Georgia, USA to establish baseline hematological and serum biochemical parameters and determine sex- and age-related differences. Summarized complete blood count and serum biochemistry panel results are presented for 85 blood samples collected from 54 lemurs at annual health exams during 1998-2003. Within each of four age classes (infant, <1 yr; juvenile, 1-5 yr; adult, ≥ 6 yr), data were stratified and summarized based on sex. Lemur age was a significant positive predictor of mean corpuscular hemoglobin; absolute concentrations of neutrophils, monocytes, and band cells; serum concentrations of blood urea nitrogen (BUN), creatinine, globulins, lipase, and total protein; and gamma-glutamyl transferase (GGT) activity. Lemur age was a significant negative predictor of albumin:globulins ratio; alkaline phosphatase (ALP) activity; and serum concentrations of calcium, cholesterol, glucose, magnesium, phosphorus, potassium, and triglycerides. Neutrophil proportions increased with aging and lymphocyte proportions decreased with aging, particularly in females. Recent steep population declines of wild ring-tailed lemurs make their successful husbandry and medical care an increasingly pressing concern. These biomedical data will aid in clinical diagnosis and treatment of lemurs in human care, and support conservation efforts for this species.

PULMONARY AND COELOMIC MYCOSES DUE TO METARHIZIUM AND BEAUVERIA SPECIES IN REPTILES.

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.

PREVALENCE OF MULTIPLE REPTILIAN PATHOGENS IN THE OROPHARYNGEAL MUCOSA, CLOACAL MUCOSA, AND BLOOD OF DIAMONDBACK TERRAPIN (MALACLEMYS TERRAPIN) POPULATIONS FROM MARYLAND AND GEORGIA, USA.

The diamondback terrapin (Malaclemys terrapin) is a coastal turtle with a range from Massachusetts to Texas and is the only exclusively brackish water turtle in North America. Two populations of wild terrapins from Maryland (n=55) and Georgia (n=7) were examined and tested for potential reptile pathogens. Whole blood and a mucosal (combined oropharyngeal and cloacal) swab from each animal were evaluated by quantitative PCR for 15 potential pathogens including frog virus 3, box turtle Mycoplasmopsis, Mycoplasma agassizii, Mycoplasma testudineum, Salmonella Enteritidis, Salmonella Typhimurium, Borrelia burgdorferi, Anaplasma phagocytophilum, tortoise intranuclear coccidia, testudinid alphaherpesvirus 2, terrapene herpesvirus 1, and terrapene adenovirus. Swabs were positive for a DNA segment 100% homologous to M. testudineum in both populations, with Maryland animals 87% (48 of 55) positive and Georgia animals 86% (6 of 7) positive. Although Mycoplasmopsis spp. are important respiratory pathogens for members of the order Testudines, none of the animals in the study showed any sign of upper respiratory disease. Our data suggest that M. testudineum may survive in non-Testudinidae turtles without causing clinical sigs of disease and suggesting appropriate precautions should be taken in facilities that house multiple species of turtles simultaneously.

Correction: Turner et al. Steatitis in Cold-Stunned Kemp's Ridley Sea Turtles (Lepidochelys kempii). Animals 2021, 11, 898.

In the original publication [...].

Health and nutrition of loggerhead sea turtles (Caretta caretta) in the southeastern United States.

Loggerhead sea turtles (Caretta caretta) are opportunistic carnivores that feed primarily on benthic invertebrates and fish. Sea turtle rehabilitation requires provision of a species-specific, balanced diet that supplies nutrition similar to that of a wild diet; this can be challenging because free-ranging loggerheads' diets vary depending on their life stage and geographic location, with predominant prey species dictated by local availability. The goal of this study was to better understand the nutritional needs of subadult and adult loggerheads in rehabilitation. This was accomplished by conducting a retrospective survey of stomach contents identified during gross necropsy of 153 deceased loggerheads that stranded in coastal Georgia, USA. A total of 288 different forage items were identified; the most frequently observed prey items belong to the subphylum Crustacea (N = 131), followed by bony fish (Osteichthyes; N = 45), gastropod mollusks (N = 40), bivalve mollusks (N = 23), and Atlantic horseshoe crabs (Limulus polyphemus; N = 15). The proportions of certain prey items differed significantly with turtle size; adult turtles ate proportionately more gastropods (p = 0.001), and subadults ate proportionately more fish (p = 0.01). Stomach contents information was used to determine common local prey items (blue crab, cannonball jellyfish, horseshoe crab, whelk), which were evaluated for nutritional content. Additionally, we compared hematology and plasma biochemistry profiles (including proteins, trace minerals, and vitamins) between four cohorts of loggerhead turtles, including free-ranging subadults and adults, nesting females, and loggerheads undergoing rehabilitation. This information was applied to inform a regionally specific, formulated diet for tube feeding, and a supplement containing vitamins and minerals for captive loggerheads, to more closely approximate the nutritional content of their natural diet. Assessing the regional and temporal variability in loggerhead diets is an important component in their effective conservation because resultant data can be used to help understand the impacts of environmental perturbations on benthic food webs.

Multi-Injection Pharmacokinetics of Meloxicam in Kemp's Ridley (Lepidochelys kempii) and Green (Chelonia mydas) Sea Turtles after Subcutaneous Administration.

The objective of this study was to determine the pharmacokinetics and safety of multiple injections of meloxicam (MLX) administered subcutaneously (SQ) in Kemp's ridley (Lepidochelys kempii) and green (Chelonia mydas) sea turtles. Based on results from a previously published single-injection study, a multiple-injection regimen was derived for the Kemp's ridleys, which consisted of administering MLX at a dose of 1 mg/kg SQ every 12 h for 5 days, and for green turtles at a dose of 1 mg/kg SQ every 48 h for three treatments. Six turtles of each species were used for the study, and blood samples were taken at multiple time intervals. The terminal half-life after the last dose for the Kemp's ridley sea turtles was calculated at 7.18 h, and for the green sea turtles at 23.71 h. Throughout the multiple injections, MLX concentrations remained above 0.57 µg/mL, a concentration targeted in humans for the analgesic and anti-inflammatory effects. No negative side effects or changes to blood parameters evaluated were observed during the study in either species. The results of this study suggest MLX should be administered SQ to Kemp's ridley sea turtles at a dosage of 1 mg/kg every 12 h and in green sea turtles at a dose of 1 mg/kg every 48 h. The novelty of this work is that it is a multiple-injection study. Multiple injections were administered and produced concentrations that were considered therapeutic in humans, and the turtles did not have any adverse side effects. Furthermore, there were large differences in the pharmacokinetic values between green and Kemp's ridley sea turtles.

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 degradation.

BIOMEDICAL EVALUATION OF THE GOLDEN-CROWNED SIFAKA (PROPITHECUS TATTERSALLI) IN DARAINA, MADAGASCAR.

Complete medical examinations were performed on 25 wild golden-crowned sifaka (Propithecus tattersalli) from northeastern Madagascar. Each animal received a complete physical examination and weight, body temperature, heart rate, and respiratory rate were documented. Blood samples were collected for packed cell volume, estimated total white blood cell count, serum biochemical profile, fat-soluble vitamin analysis, trace mineral analysis, and Toxoplasma gondii serology. All animals examined were adults and determined to be in good health and body condition. No ectoparasites were observed. Fecal samples were collected for endoparasite examination and bacterial culture; while no endoparasites were observed, fecal samples from two females cultured positive for Bacillus cereus. One male lemur had a positive antibody titer to Toxoplasma gondii immunoglobulin G. These baseline health data provide an important foundation for continued monitoring of this critically endangered species.

PHARMACOKINETIC BEHAVIOR OF MELOXICAM IN LOGGERHEAD (CARETTA CARETTA), KEMP'S RIDLEY (LEPIDOCHELYS KEMPII), AND GREEN (CHELONIA MYDAS) SEA TURTLES AFTER SUBCUTANEOUS ADMINISTRATION.

The objective of this study was to determine the pharmacokinetics of a single dose of meloxicam administered subcutaneously (SQ) to three species of sea turtles: loggerheads (Caretta caretta), Kemp's ridley (Lepidochelys kempii), and greens (Chelonia mydas). A dose of 1 mg/kg was given to the Kemp's ridleys and greens, whereas the loggerheads received 2 mg/kg. After SQ administration, the half-life (t1/2) of meloxicam administered at 1 mg/kg in the Kemp's ridleys was 5.51 hr but could not be determined in the greens. The half-life of meloxicam administered at 2 mg/kg in the loggerheads was 2.99 hr. The maximum concentration (Cmax) for meloxicam after SQ administration at 1 mg/kg in the Kemp's ridleys was 6.76 µg/ml and in the greens was 9.35 µg/ml. The Cmax in loggerheads for meloxicam after SQ administration at 2 mg/kg was 3.63 µg/mL. Meloxicam administered SQ at a dose of 1 mg/kg to the Kemp's ridley and greens provided measurable plasma concentrations of meloxicam for 48 and 120 hr, respectively, with no adverse side effects. In loggerheads, meloxicam administered SQ at a dose of 2 mg/kg provided measurable plasma levels of meloxicam for only 24 hr. Plasma levels of meloxicam of greater than 0.5 µg/ml are considered to be therapeutic in humans. Results suggested that administration of meloxicam SQ at 1 mg/kg in Kemp's ridleys and greens would result in plasma concentrations greater than 0.5 µg/ml for 12 and 120 hr, respectively. The administration of 2 mg/kg meloxicam to loggerhead turtles resulted in plasma concentrations greater than 0.5 µg/ml for only 4 hr.

Steatitis in Cold-Stunned Kemp's Ridley Sea Turtles (Lepidochelys kempii).

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 33­469). 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.

Epidemiological Investigation of a Mortality Event in a Translocated Gopher Tortoise (Gopherus polyphemus) Population in Northwest Florida.

Nokuse Plantation, a 22,055 ha private conservation preserve in northwest Florida, is a recipient site for gopher tortoises translocated from development sites in Florida. Since 2006, Nokuse has received over 5,000 tortoises from multiple development sites. During 2013-2015, 52 tortoises were found sick (n = 14) or dead (n = 38) in multiple soft-release enclosures in which tortoises consistently exhibited clinical signs, with additional sick (n = 5) and dead (n = 5) tortoises presenting similarly during 2016-2017. When found alive, tortoises behaved abnormally (e.g., frequently out of burrows during cold weather, pacing along enclosure fencing), appeared emaciated, were lethargic, and had developed redness under plastron scutes. Similar numbers of male (n = 28) and female (n = 32) tortoises were recovered along with two of unidentified sex, including mainly adults (n = 59) and three subadults. Physical examination, blood analysis, and other diagnostics were indicative of starvation and dehydration. Most sick tortoises provided with supportive care recovered. Necropsy findings generally confirmed starvation, with no evidence of infectious pathogens or contaminants. There were no apparent differences in quality of habitat, plant community, or soil or water among affected and unaffected enclosures. Botanical surveys indicated adequate forage quality and quantity, with no poisonous exotic or native plants detected. No land management practices changed prior to this event. Analysis of epidemiological data and demographic factors from before and during this mortality event identified initial density of tortoises in the enclosures as exerting the strongest influence on detection of tortoise morbidity and mortality. We believe that the stress associated with mixing tortoises from different populations and at higher densities during translocation impacted an individual tortoise's ability to obtain or absorb adequate nutrients from foraging, ultimately leading to a wasting condition consistent with starvation. Based on our findings, we recommend a maximum of 3 gopher tortoises per ha in soft-release enclosures for translocation, but further research is warranted to investigate the complexity of stress and social pressures associated with translocation.

Diagnosis and Surgical Management of a Paratesticular Cyst in a Rehabilitating Juvenile Male Green Turtle (Chelonia mydas).

A juvenile green turtle (Chelonia mydas) undergoing rehabilitation for cold stunning exhibited an asymmetric bulging of the left caudal plastron and was diagnosed with a large intra-coelomic mass based on radiographical findings. Ultrasonography further identified a fluid-filled structure within the caudal coelom. Cytological evaluation of fluid obtained from the structure was consistent with a transudate, and thus, a cyst of unknown origin was suspected. Computed tomography imaging was pursued to further characterize the extent and location of the mass, which occupied ~50% of the total coelomic cavity volume. Conservative management with monitoring and occasional drainage of the mass did not result in improvements; thus, an exploratory laparoscopy for further investigation and surgical planning was elected. Intra-coelomic surgery was performed to remove a thick-walled cystic mass associated with the left gonad. Histopathology confirmed a paratesticular cyst continuous with, and possibly originating from, the epididymis. Post-surgical recurrence of the cyst was not appreciated, and the animal was successfully released 1 year after admission. Unrelated to the cyst, the turtle developed acute severe anemia on two occasions throughout rehabilitation that responded to modification of antimicrobial treatment and subsequent steroid administration. To the authors' knowledge, this is the first report of a paratesticular cyst in a reptile.

Pathogen Surveillance and Detection of Ranavirus (Frog virus 3) in Translocated Gopher Tortoises (Gopherus polyphemus).

Emerging pathogens may pose additional threats to already vulnerable populations of chelonians, such as gopher tortoises (Gopherus polyphemus). In response to a mortality event on a translocation site in northwest Florida, US during 2013-15, 13 gopher tortoises were necropsied and their tissues were screened for 12 pathogens, including Mycoplasma agassizii, Mycoplasma testudineum, and Frog virus 3-like ranavirus (FV3). The DNA of FV3 was detected via quantitative PCR in the gastrointestinal tract of three tortoises. Subsequently, pathogen surveillance was performed on whole blood and oral-cloacal swab samples of live translocated tortoises from two different enclosures within the site (n=68), rehabilitated tortoises from the site (n=18), and tortoises prior to release on site (n=35) during 2015-17. Mycoplasma spp. were present in all groups and years of live tortoises tested. The DNA of FV3 was detected in 15 individuals both with and without clinical signs of disease in 2016. We recaptured 20 tortoises and captured an additional 20 tortoises in 2017 for surveillance, yet FV3 DNA was no longer detected, even in those that had previously tested positive (n=7). The results of this study contribute to the epidemiology of ranavirus in chelonians and suggests that gopher tortoises could be reservoirs for FV3. We recommend that the status of Ranavirus infection should be included for health screens for gopher tortoises in translocation programs.

The effect of diet on the gastrointestinal microbiome of juvenile rehabilitating green turtles (Chelonia mydas).

Threatened and endangered green turtles (Chelonia mydas) are unique because as juveniles they recruit from pelagic to near-shore waters and shift from an omnivorous to primarily herbivorous diet (i.e. seagrass and algae). Nevertheless, when injured and ill animals are admitted to rehabilitation, animal protein (e.g. seafood) is often offered to combat poor appetite and emaciation. We examined how the fecal microbiome of juvenile green turtles changed in response to a dietary shift during rehabilitation. We collected fecal samples from January 2014 -January 2016 from turtles (N = 17) in rehabilitation at the Georgia Sea Turtle Center and used next generation sequencing to analyze bacterial community composition. Samples were collected at admission, mid-rehabilitation, and recovery, which entailed a shift from a mixed seafood-vegetable diet at admission to a primarily herbivorous diet at recovery. The dominant phyla changed over time, from primarily Firmicutes (55.0%) with less Bacteroidetes (11.4%) at admission, to primarily Bacteroidetes (38.4%) and less Firmicutes (31.8%) at recovery. While the microbiome likely shifts with the changing health status of individuals, this consistent inversion of Bacteroidetes and Firmicutes among individuals likely reflects the increased need for protein digestion, for which Bacteroidetes are important. Firmicutes are significant in metabolizing plant polysaccharides; thus, fewer Firmicutes may result in underutilization of wild diet items in released individuals. This study demonstrates the importance of transitioning rehabilitating green turtles to an herbivorous diet as soon as possible to afford them the best probability of survival.

Tick paralysis in a free-ranging bobcat (Lynx rufus).

CASE DESCRIPTION: A free-ranging male bobcat (Lynx rufus) was evaluated because of signs of pelvic limb paralysis. CLINICAL FINDINGS: Physical examination of the anesthetized animal revealed tick infestation, normal mentation, and a lack of evidence of traumatic injuries. Radiography revealed no clinically relevant abnormalities. Hematologic analysis results were generally unremarkable, and serologic tests for exposure to feline coronavirus, FeLV, FIV, and Toxoplasma gondii were negative. Results of PCR assays for flea- and common tick-borne organisms other than Bartonella clarridgeiae were negative. TREATMENT AND OUTCOME: Ticks were manually removed, and the patient received supportive care and fipronil treatment. The bobcat made a full recovery within 72 hours after treatment for ticks, and a presumptive diagnosis of tick paralysis was made. Identified tick species included Dermacenter variabilis, Amblyomma americanum, and Ixodes scapularis. CLINICAL RELEVANCE: To the authors' knowledge, tick paralysis has not previously been reported in felids outside Australia. This disease should be considered a differential diagnosis in felids, including exotic cats, with signs of neuromuscular disease of unknown etiopathogenesis.

Tumor re-growth, case outcome, and tumor scoring systems in rehabilitated green turtles with fibropapillomatosis.

Fibropapillomatosis (FP) is an infectious, neoplastic disease of major concern in sea turtle rehabilitation facilities. Rehabilitating sea turtles that undergo tumor removal surgery often have tumor regrowth and may experience mortality. We evaluated tumor score, removal, and regrowth in rehabilitating green sea turtles with FP in 4 rehabilitation facilities in the southeastern USA during 2009-2017. Of 756 cases, 312 (41%) underwent tumor removal surgery, 155 (50%) of those had tumor regrowth within an average of 46 ± 45 d, and 85 (27%) had multiple (>1) regrowth events. Of 756 turtles with FP, 563 (75%) did not survive after admission into a rehabilitation facility, including 283 (37%) that were euthanized and 280 that died without euthanasia (37%), and 193 survived, including 186 (25%) released and 7 (1%) placed in permanent captive care. Tumor removal surgery increased the odds of tumor regrowth but also enhanced survivorship, whereas tumor regrowth was not a significant predictor of case outcome. Three FP tumor scoring systems were used to assign tumor scores to 449 cases, and differing results emphasize that tumor scoring systems should be applied to the situations and/or location(s) for which they were intended. FP tumor score was not a significant predictor for the event or extent of FP tumor regrowth after surgical excision. Under current rehabilitation regimes, outcomes of rehabilitation for tumored turtles have a low probability of success. The results of this study may be used to help guide clinical decision-making and determine prognoses for rehabilitating sea turtles with FP.

Caryospora-Like Coccidia Infecting Green Turtles (Chelonia mydas): An Emerging Disease With Evidence of Interoceanic Dissemination.

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.

ADVANCING TRANSFUSION MEDICINE IN SEA TURTLES: OPTIMIZATION OF A CROSS-MATCHING PROTOCOL.

Whole blood transfusions are an essential treatment modality during rehabilitation of stranded sea turtles, however, standardized protocols for transfusions are not available in reptile medicine. The objective of this study was to optimize a cross-matching protocol for sea turtle blood transfusions. Fresh venous blood samples from 15 turtles (n = 14 green turtles, Chelonia mydas [Cm]; n = 1 loggerhead sea turtle, Caretta caretta [Cc]) were tested using a temperature-appropriate (i.e., reflecting body temperature), time-sensitive protocol in 26 reactions using two procedures for cross-match evaluation at 30 and 60 min at ambient air and water bath temperature. There were no significant differences between both protocols at 30- and 60-min incubation times or between microscopic evaluations at 2 or 5 min. The major cross-match identified 7/22 incompatible Cm-Cm reactions as observed by microscopic erythrocyte agglutination. Minor cross-matches resulted in 6/22 incompatible Cm-Cm reactions. About half of all Cm-Cm reactions (12/22) were compatible by major and minor cross-match. All Cc-Cm reactions (4/4) were incompatible. A higher than expected proportion of incompatible Cm cross-matching reactions suggests preexisting antibodies to nonself erythrocyte antigens in this species, or other factors promoting erythrocyte aggregation or agglutination. Preliminary data across Cm and Cc suggests cross-species incompatibility. These results indicate that sea turtles may react to donor erythrocytes even at the first transfusion. Concurrent major and minor cross-matching using the proposed protocol at 30-min incubation at room temperature should be considered a necessary and effective way to test for patient and donor incompatibilities in sea turtles.

A novel host-adapted strain of Salmonella Typhimurium causes renal disease in olive ridley turtles (Lepidochelys olivacea) in the Pacific.

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.