Corallus hortulanus testes histology: morphological and reproductive aspects.

In general snakes show differentiate anatomical, biological and behavioral particularities compared to other species. Basic information about the snakes anatomy, physiology and reproductive biology is scarce in several species, making the reproduction a challenge. Thus, the present work aims to evaluate morphological aspects of the Corallus hortulanus testes, correlating these findings with environmental factors and reproductive aspects. The testes of three specimens of Corallus hortulanus were cut to a thickness of 3µm in microtome, stained with 1% toluidine blue, photo documented and described. Seasonality was observed in the sperm production of Corallus hortulanus, with the presence of mature spermatozoa in the wettest and hottest periods of the year, as well as the largest testicular volume in these periods.

Synergistic effects of musking and autohaemorrhaging on the duration of death feigning in dice snakes (Natrix tessellata).

Predation exerts a significant selection pressure on prey, shaping a multitude of traits that serve as antipredator defences. In turn, natural selection could favour combinations of antipredator defences with synergistic effects that enhance prey survival. An especially interesting antipredator defence is death feigning (DF), present in a wide variety of taxa and usually characterized by the prey lying motionless often along with defaecation, musking and autohaemorrhaging (AH). All these aspects of the DF display should work in conjunction with one another, intensifying the overall effect of the display and in turn facilitating quicker escape. To confirm this hypothesis, we tested 263 dice snakes (Natrix tessellata) directly in the field. We noted the occurrence of smearing faeces, musk and AH, and we measured the duration of DF, expecting to see a negative association between the occurrence of these behaviours and the duration of DF. Our results affirm our hypothesis: dice snakes that smeared themselves in musk and faeces prior to DF and had AH during DF spent significantly less time in DF. Our results highlight the functional integration of antipredator behaviours across different phases of predator-prey interactions, emphasizing the need for future research to prioritize studying the sequential display of behaviours.

Natrix natrix after dark: citizen science sheds light on the common grass snake's nightlife.

Activity patterns in animals are often species-specific, and can be generally categorized as diurnal, crepuscular, or nocturnal. Understanding these patterns provides insight into ecological adaptations and behaviors. The common grass snake (Natrix natrix), one of the most common and widespread European snake species, is traditionally considered diurnal, with scarce evidence of its crepuscular and nocturnal activity. We aimed to document the distribution, environmental conditions, and potential phenotype associations of nighttime activity in N. natrix. We used citizen science data from iNaturalist (1992-2022), Observation.org (2012-2022), together with personal field observations (2010-2023) to collect 127 crepuscular and nocturnal activity records. Most observations occurred between May and August, coinciding with the peak activity period of grass snakes across their distribution range. Statistical analyses revealed no significant difference in mean daily temperatures between crepuscular and nocturnal observations. However, striped individuals displayed nocturnal activity at higher temperatures, consistent with their distribution in warmer regions, but failed to register any difference when tested on a geographic subsample, that accounted for sympatry of the phenotypes. Surprisingly, we found no significant impact of moon presence or moonlight on nighttime activity or age class, contrary to expectations based on other snake species' responses. While our study reveals that nocturnal activity in the common grass snake is geographically widespread, further research is warranted to understand its drivers and ecological implications. This study highlights the value of citizen science platforms for biological and ecological research, offering unparalleled spatial and temporal coverage by their users. In conclusion, our work extends the knowledge of nocturnal behavior in N. natrix and underlines the critical role of citizen science in discovering behavioral aspects of common and widespread species.

One-Step Affinity Purification of Leucine-Rich α2-Glycoproteins from Snake Sera and Characterization of Their Phospholipase A2-Inhibitory Activities as ß-Type Phospholipase A2 Inhibitors.

Snakes contain three types of phospholipase A2 (PLA2)-inhibitory proteins in their blood, PLIα, ß, and γ, which protect them from their own venom, PLA2. PLIß is the snake ortholog of leucine-rich α2 glycoprotein (LRG). Since autologous cytochrome c (Cyt c) serves as an endogenous ligand for LRG, in this study, we purified snake LRGs from various snake serum samples using Cyt c affinity chromatography. All purified snake LRGs were found to be dimers linked by disulfide bonds. Laticauda semifasciata and Naja kaouthia LRGs showed no inhibitory activity against L. semifasciata PLA2 and weak inhibitory activity against Gloydius brevicauda basic PLA2. Elaphe climacophora PLIß had weaker inhibitory activity against G. brevicauda basic PLA2 than G. brevicauda and Elaphe quadrivirgata PLIs, which are abundant in blood and known to neutralize G. brevicauda basic PLA2. Protobothrops flavoviridis LRG showed no inhibitory activity against basic venom PLA2, PL-X, or G. brevicauda basic PLA2. Binding analysis of P. flavoviridis LRG using surface plasmon resonance showed very strong binding to snake Cyt c, followed by that to horse Cyt c, weak binding to yeast Cyt c, and no binding to P. flavoviridis PL-X or BPI/II. We also deduced the amino acid sequences of L. semifasciata and P. flavoviridis LRG by means of cDNA sequencing and compared them with those of other known sequences of PLIs and LRGs. This study concluded that snake LRG can potentially inhibit basic PLA2, but, whether it actually functions as a PLA2-inhibitory protein, PLIß, depends on the snake.

Envenomation by a common green racer (Chlorosoma viridissimum) in the Brazilian Amazon: A report of three cases.

Envenomations by the common green racer (Chlorosoma viridissimum) are seldom reported in the literature. Herein, we report three cases caused by the same specimen of C. viridissimum in three different victims in the Brazilian Amazon. In all cases, the victims were either a biologist or biology students that were handling the animal and were bitten in their upper limbs. The victims showed only local symptoms, such as edema, tooth marks, pain, erythema, ecchymoses and bleeding. One of the patients presented extensive ecchymosis. Two patients sought medical care, but were only treated for local manifestations and evolved without complications. Chlorosoma viridissimum is capable of provoking mild to moderate signs and symptoms.

First case of envenomation in humans caused by the banded cat-eyed snake Leptodeira annulata (Linnaeus, 1758) (Squamata: Dipsadidae) in Brazil.

A 22-year-old female researcher was bitten by a Leptodeira annulata on the index finger of the left hand during a contention activity. After removing the snake, a little bleeding and redness was observed in the bite region, accompanied by fang marks. Thirty minutes later, edema had progressed to the dorsum of the hand. After four hours, edema persisted, but the bitten area was slightly whitened. Treatment consisted of antibiotics and anti-inflammatory drugs. The edema resolved completely and disappeared after 48 hours. Overall, this report presents the first case of envenomation in humans caused by Leptodeira annulata in Brazil.

Characterisation, prevalence and severity of skin lesions caused by ophidiomycosis in a population of wild snakes.

Ophidiomycosis is an emerging infectious disease affecting wild snakes in the Northern Hemisphere. Recently confirmed in Great Britain, the prevalence, severity and significance of ophidiomycosis has yet to be characterised in free-living snakes at a population level in Europe. Therefore, a population of barred grass snakes (Natrix helvetica) in eastern England was monitored for three seasons (May 2019 to October 2021), to investigate the prevalence (25.5%; 191/750 snakes) and severity of skin lesions and their aetiology. The most frequently observed skin lesion characteristics were changes in scale colour, crusting, and scale margin erosion. The majority of such lesions (96.9%; 185/191 snakes) was observed on the ventral surface along the length of the body. The severity of skin lesions was considered mild in more than half of the cases (53.1%; 98/191 snakes). Predominantly, skin lesions were observed in adult snakes (72.8%; 139/191 snakes). Combined histological examinations and qPCR tests of skin lesions from N. helvetica sloughs and/or carcasses confirmed a diagnosis of ophidiomycosis. Further targeted surveillance, supported by molecular and histological examinations to confirm skin lesion aetiology, is required to determine the extent to which our findings reflect the occurrence of ophidiomycosis in populations within wider landscapes.

Spatial and temporal assessment of snake encounters in urban Delhi, India.

Delhi, the capital city of India is, highly urbanized and surrounded by remnant forest, farms, ridges, and other green areas experience regular snake encounters in and around residential, institutional, and industrial areas. A total of 41 months of sampling from January 2019 to May 2022 was conducted wherein we, studied the snake assemblage in Delhi to determine the species composition, encounter frequency, seasonal activity patterns, and probable encounter sites in an urban setup. We documented 372 individuals belonging to 15 species from seven families out of 23 species found in Delhi. Snakes were found inside forests, public parks, homes, drain networks, streets, office buildings, and even in school-college buildings. The most recorded species being Ptyas mucosa (37.37%, n = 139), Naja naja (19.62%, n = 73), and Lycodon aulicus (13.44%, n = 50). The highest numbers of incidents were reported in the month of July (22.04%, n = 82) and August (19.89%, n = 74) during the peak monsoon season, for identifying high encounter sites, we used a geostatistical modeling tool, Ordinary kriging to identify places having more snake occurrences. We further used a statistical spatial method called average nearest neighbor distance to detect the pattern distribution of snake species. Spatial interpolation done through Ordinary kriging highlighted two areas having concentrated snake encounters. The results of the average nearest neighbor distance analysis showed three species having clustered and two species having dispersed distribution. The incidence of snake encounters was found to be highly seasonal and appeared to be associated mainly with monthly rainfall, temperature, and humidity. The findings of this study on snakes' distribution patterns provide valuable insights into the conservation of these species. Understanding their habitat preferences and spatial distribution is crucial for the implementation of effective conservation strategies.

Developmental energetics in the oviparous corn snake, Pantherophis guttatus, confirms a conservative evolutionary pattern in snakes.

Eggs of oviparous reptiles are ideal models for studying evolutionary patterns of embryonic metabolism since they allow tracking of energy allocation during development. Analyzing oxygen consumption of whole eggs throughout development indicates three patterns among reptiles. Embryos initially grow and consume oxygen exponentially, but oxygen consumption slows, or drops before hatching in some species. Turtles, crocodilians, and most lizards follow curves with initial exponential increases followed by declines, whereas embryonic snakes that have been studied exhibit a consistently exponential pattern. This study measured oxygen consumption of corn snake, Pantherophis guttatus, embryos to determine if this species also exhibits an exponential increase in oxygen consumption. Individual eggs, sampled weekly from oviposition to hatching, were placed in respirometry chambers for 24-h during which oxygen consumption was recorded. Embryos were staged and carcasses and yolk were weighed separately. Results indicate steady inclines in oxygen consumption during early stages of development, with a rapid increase prior to hatching. The findings support the hypothesis that embryonic oxygen consumption of snakes differs from most other non-avian reptiles. Total energy required for development was determined based on calorimetry of initial yolk compared to hatchlings and residual yolk and by integration of the area under the curve plotting oxygen consumption versus age of embryos. The cost of development estimates based on these two methods were 6.4 and 10.0 kJ, respectively. Our results emphasize the unique physiological aspects of snake embryogenesis and illustrate how the study of physiological characteristics can contribute to the broader understanding of reptilian evolution.

Patterns of recombination in snakes reveal a tug-of-war between PRDM9 and promoter-like features.

In some mammals, notably humans, recombination occurs almost exclusively where the protein PRDM9 binds, whereas in vertebrates lacking an intact PRDM9, such as birds and canids, recombination rates are elevated near promoter-like features. To determine whether PRDM9 directs recombination in nonmammalian vertebrates, we focused on an exemplar species with a single, intact PRDM9 ortholog, the corn snake (Pantherophis guttatus). Analyzing historical recombination rates along the genome and crossovers in pedigrees, we found evidence that PRDM9 specifies the location of recombination events, but we also detected a separable effect of promoter-like features. These findings reveal that the uses of PRDM9 and promoter-like features need not be mutually exclusive and instead reflect a tug-of-war that is more even in some species than others.

Association between anthropization and rodent reservoirs of zoonotic pathogens in Northwestern Mexico.

The world is facing a major pulse of ecological and social changes that may favor the risk of zoonotic outbreaks. Such risk facilitation may occur through the modification of the host's community diversity and structure, leading to an increase in pathogen reservoirs and the contact rate between these reservoirs and humans. Here, we examined whether anthropization alters the relative abundance and richness of zoonotic reservoir and non-reservoir rodents in three Socio-Ecological Systems. We hypothesized that anthropization increases the relative abundance and richness of rodent reservoirs while decreasing non-reservoir species. We first developed an Anthropization index based on 15 quantitative socio-ecological variables classified into five groups: 1) Vegetation type, 2) Urbanization degree, 3) Water quality, 4) Potential contaminant sources, and 5) Others. We then monitored rodent communities in three regions of Northwestern Mexico (Baja California, Chihuahua, and Sonora). A total of 683 rodents of 14 genera and 27 species were captured, nine of which have been identified as reservoirs of zoonotic pathogens (359 individuals, 53%). In all regions, we found that as anthropization increased, the relative abundance of reservoir rodents increased; in contrast, the relative abundance of non-reservoir rodents decreased. In Sonora, reservoir richness increased with increasing anthropization, while in Baja California and Chihuahua non-reservoir richness decreased as anthropization increased. We also found a significant positive relationship between the anthropization degree and the abundance of house mice (Mus musculus) and deer mice (Peromyscus maniculatus), the most abundant reservoir species in the study. These findings support the hypothesis that reservoir species of zoonotic pathogens increase their abundance in disturbed environments, which may increase the risk of pathogen exposure to humans, while anthropization creates an environmental filtering that promotes the local extinction of non-reservoir species.

First report of clinically significant bites due to Platyceps ventromaculatus (GRAY, 1834) from Thar Desert region, Rajasthan, India.

BACKGROUND: Platyceps ventromaculatus is a non-front-fanged colubrid snake of unknown medical significance. In this study, we highlight the clinical manifestations and outcomes of P. ventromaculatus bites. We also emphasise the need to create awareness among clinicians and the public for its potential to be confused with serious venomous bites such as Echis carinatus sochureki. METHODS: This series is part of an ongoing observational clinical study from our tertiary care hospital in Jodhpur, India on the profile and outcomes of snakebite envenoming. Data was collected after approval from Institute Ethics Committee. The date and time of the bite, geographical location, type of human-snake conflict, time-to-reach a healthcare facility, antivenom used (dose), and outcomes were recorded. We retrospectively examined our clinical data for images suggestive of P. ventromaculatus and present the clinical details of these patients. The photographs were identified utilising taxonomic keys for species identification. RESULTS: A total of four images and three patients with bites due to P. ventromaculatus were identified. The clinical effects included mild local erythema, pain, transient local bleeding, and edema. All bites occurred during daylight hours, 2 during agricultural activities, and one at home. Twenty-minute Whole Blood Clotting Test was persistently prolonged for 12 h after the bite in one patient. All patients were treated symptomatically, observed at the emergency department, and discharged within 24 h. None of the patients received antivenom. CONCLUSIONS: To our knowledge, P. ventromaculatus has so far not been reported to result in envenoming or medically significant bites. This study highlights that Platyceps bite can present with clinically significant local and possibly systemic findings that may lead to confusion with saw-scaled viper (Echis) envenoming. Clinicians must receive appropriate training so as to be aware and recognize regional snake species that do not require antivenom so as to avoid unnecessary antivenom administration.

Through the Looking Glass: Revisiting Smoking and Health.

January 11, 2024, marked the 60th anniversary of the initial U.S. Surgeon General report "Smoking and Health," which definitively linked cigarette smoking and lung cancer.1 Similar to the old Virginia Slims slogan "you've come a long way, baby," smoking rates have diminished greatly since the release of that report.2 However, smoking still represents the leading cause of preventable deaths.3 Numerous countries have enacted policies aimed at decreasing conventional cigarette use, such as including warning labels on tobacco products, limiting advertising, and imposing bans on particular products. Such measures have contributed to significant reductions in cigarette use.

Effects of ophidiomycosis on movement, survival, and reproduction of eastern foxsnakes (Pantherophis vulpinus).

Ophidiomycosis (snake fungal disease) is caused by the fungal pathogen Ophidiomyces ophidiicola, which causes dermal lesions, occasional systemic infections, and in some cases, mortality. To better understand potential conservation implications of ophidiomycosis (i.e., population-level effects), we investigated its impacts on individual fitness in a population of endangered eastern foxsnakes (Pantherophis vulpinus). We tracked 38 foxsnakes over 6 years and quantified body condition, movement patterns, oviposition rates, and survival. Body condition, distance travelled, and oviposition rates were similar between snakes with and without ophidiomycosis. Interestingly, snakes that tested positive for the pathogen travelled farther, suggesting that movement through a greater diversity of habitats increases risk of exposure. Ophidiomycosis did not negatively affect survival, and most apparently infected snakes persisted in a manner comparable to snakes without ophidiomycosis. Only one mortality was directly attributed to ophidiomycosis, although infected snakes were overrepresented in a sample of snakes killed by predators. Overall, our results suggest that ophidiomycosis may have sublethal effects on eastern foxsnakes, but do not suggest direct effects on survival, ovipositioning, or viability of the study population.

Bolder together: conformity drives behavioral plasticity in eastern gartersnakes.

Personality traits drive individual differences in behaviour that are consistent across time and context. Personality limits behavioural plasticity, which could lead to maladaptive choices if animals cannot adapt their behavior to changing conditions. Here, we assessed consistency and flexibility in one personality trait, boldness, across non-social and social contexts in eastern gartersnakes (Thamnophis sirtalis sirtalis). Snakes explored a novel open arena either alone or in a pair. Pairs were assigned based on the data from the solo trials, such that each snake was paired once with a bolder and once with a less bold partner. We predicted that snakes would conform when in a social context, displaying plasticity in their personality, and causing boldness scores to converge. We found that snakes were consistent within contexts (solo or paired), but changed their behavior across contexts (from solo to paired). Plasticity in boldness resulted from an interaction between conformity and repeatable individual differences in plasticity. In line with some data on other species, snakes conformed more when they were the less bold partner. Personality reflects a consistent bias in decision-making, but our results highlight that the cognitive processes that drive the expression of personality traits in behavior are flexible and sensitive to social context. We show that both consistency and plasticity combine to shape snake social behavior in ways that are responsive to competition. This pattern of behavior may be particularly beneficial for species in which group-living is seasonal.

Comparative morphology of oral glands in snakes of the family Homalopsidae reveals substantial variation and additional independent origins of salt glands within Serpentes.

Using diffusible iodine-based contrast-enhanced computed tomography (diceCT), we examined the morphology of the oral glands of 12 species of the family Homalopsidae. Snakes of this family exhibit substantial interspecific morphological variation in their oral glands. Particular variables are the venom glands, ranging from large (e.g., Subsessor bocourti) to small (e.g., Erpeton tentaculatum). The supra- and infralabial glands are more uniform in morphology, being the second most developed in almost all the sampled species. Premaxillary glands distinct from the supralabial glands were observed in five species (Myron richardsonii, Bitia hydroides, Cantoria violacea, Fordonia leucobalia, and Gerarda prevostiana), in addition to Cerberus rynchops, the only species in which this condition was previously documented associated with the excretion of salt. In the three species of the saltwater group of homalopsids (C. violacea, F. leucobalia, and G. prevostiana), the premaxillary glands also extend posteriorly, occupying a large area above the supralabial gland, a condition not observed in any other species of snake studied thus far. Character evolution analyses indicate that premaxillary glands differentiated from the supralabial gland and evolved independently three or four times in the family, always in lineages that invaded marine habitats. Our results suggest that the differentiated premaxillary glands are likely salt glands, as is the case in C. rynchops. If corroborated, this increases to six or seven the number of independent evolutionary origins of salt glands in snakes that have undergone an evolutionary transition to marine life.

Coinfection by Mycobacterium marinum and Mycolicibacterium fortuitum in a captive adult diamondback water snake causing disseminated mycobacteriosis with acute cutaneous ulceration.

An adult male captive diamondback water snake (Nerodia rhombifer) was found dead after a 1-d history of lethargy and cutaneous ulcers. The snake had eaten 2 sunfish (Mola spp.) 5 d before death. Gross examination revealed white-to-tan nodules in the lung and liver and segmental intestinal impactions with digested fish. Histopathology confirmed disseminated granulomas with numerous intrahistiocytic acid-fast bacteria in the skin, skeletal muscle, lung, liver, and intestines. Mycobacterium marinum and Mycolicibacterium fortuitum were identified by culture of the hepatic granuloma, followed by PCR and rpoB gene sequencing. To our knowledge, this is the first description of M. marinum and M. fortuitum coinfection in this species. Although M. fortuitum has been isolated from reptiles, lesions associated with its presence in tissues have not been described previously. Interestingly, the mineralization within granulomas that we observed in our case is not reported in mycobacterial infection in reptiles, whereas this finding is common in mammals.

CONTROLLED CLINICAL TRIAL USING TERBINAFINE NEBULIZATION TO TREAT WILD LAKE ERIE WATERSNAKES (NERODIA SIPEDON INSULARUM) WITH OPHIDIOMYCOSIS.

Ophidiomycosis (snake fungal disease) is an important infectious disease caused by the fungus Ophidiomyces ophidiicola. To mitigate the disease's impact on individual snakes, a controlled clinical trial was conducted using terbinafine nebulization to treat snakes with ophidiomycosis. Fifty-three wild-caught Lake Erie watersnakes (Nerodia sipedon insularum) with apparent ophidiomycosis (skin lesions present, qPCR positive for O. ophidiicola) were divided into treatment and control groups: treatment snakes were nebulized with a 2 mg/ml terbinafine solution for 30 min daily for 30 d; control snakes received nebulization with 0.9% saline or no nebulization. Weekly physical exams were conducted to assign disease severity scores based on the number, type, location, and size of lesions, and qPCR was repeated after each 30-d course of treatment. Persistently qPCR-positive snakes received multiple nebulization courses. Terbinafine nebulization showed mixed results as a treatment for ophidiomycosis: 29.2% of animals treated with terbinafine showed molecular resolution of external disease, based on antemortem swabbing, following 3-6 mon of daily nebulization; this was significantly more than with saline nebulization (5%), but molecular resolution also occurred in 11.1% of snakes that received no treatment. Terbinafine nebulization did not significantly decrease clinical disease, as measured by disease severity scores. Evaluating molecular response to treatment using fungal quantities, terbinafine nebulization significantly reduced fungal quantity after three or more courses of treatment. These results indicate that, although terbinafine nebulization is a promising treatment for ophidiomycosis, snakes may require multiple nebulization courses and disease may not always resolve completely, despite treatment. This treatment may be most useful in snakes from managed populations that can be treated for several months, rather than wild snakes who are not releasable after multiple months in captivity.

SERUM FENTANYL CONCENTRATIONS AND BEHAVIOR ASSOCIATED WITH TRANSDERMAL FENTANYL APPLICATION ON HEALTHY CORN SNAKES (PANTHEROPHIS GUTTATUS).

Snakes are common household pets and frequently managed in zoos. Geriatric snakes commonly develop osteoarthritis, leading to a declining quality of life that often results in euthanasia. Anecdotally, the application of transdermal fentanyl patches (TFP) appears to contribute to clinical improvement, including increased activity level, in osteoarthritic snakes presumed to be in pain. This study evaluated serum fentanyl concentrations over time and the effects of TFP on the normal behavior of healthy, captive, adult corn snakes (Pantherophis guttatus) using constant video monitoring. Serum fentanyl concentrations were evaluated over 4 wk during 12.5 µg/h TFP application, and the results demonstrated long-lasting (>4 wk) serum concentrations that were consistent with analgesic efficacy in mammalian species during TFP application. At 4 wk of TFP application, mean serum fentanyl concentrations were 11.5 ± 5.5 ng/ml. Snakes were videotaped for 1 wk prior to and 2 wk after 12.5 µg/h TFP application, and behavior was evaluated by an ethogram. Behavioral changes associated with TFP application included decreased mean time spent active, decreased mean number of climbs, and decreased mean number of water visits; feeding behavior was unchanged. Overall, these results suggest that TFP application may provide safe, clinically effective analgesia in healthy corn snakes for at least 4 wk without inducing deleterious side effects, and may therefore be appropriate analgesia for management of osteoarthritic snakes.

COMPARING THE EFFECTS OF LITHIUM HEPARIN AND DIPOTASSIUM ETHYLENEDIAMINETETRAACETIC ACID ON HEMATOLOGIC VALUES IN PRAIRIE RATTLESNAKES (CROTALUS VIRIDIS) AND LAKE ERIE WATER SNAKES (NERODIA SIPEDON INSULARUM).

Anticoagulants prevent clotting of blood samples and preserve cellular morphology for hematologic evaluations, but studies comparing anticoagulants are limited in snakes. The objective of this study was to evaluate the effects of lithium heparin (LH) and dipotassium ethylenediaminetetraacetic acid (EDTA) on hematologic values in prairie rattlesnakes (PR; Crotalus viridis, n = 16) and Lake Erie water snakes (LEWS; Nerodia sipedon insularum, n = 21). Venipuncture was performed and blood samples were immediately aliquoted into LH and EDTA microtainers. Packed cell volume (PCV), total solids (TS), 100-cell differential counts, and Avian Leukopet white blood cell counts (WBC) were determined for each anticoagulant. Passing-Bablok regression and Bland-Altman plots revealed that anticoagulant choice did not constantly or proportionally bias the values of any WBC parameter. Mixed models demonstrated that blood anticoagulated with EDTA had higher PCV in PR (P = 0.04) and TS in both species (P < 0.05). However, the magnitude of the differences attributable to anticoagulant choice was relatively small and likely not clinically important. Hemolysis was not appreciated in any samples. Our findings demonstrate that LH and EDTA are equally appropriate for use in PR and LEWS, but may require separate reference values.