Serpentes

Chapter 37

Serpentes Robert J. Ossiboff University of Illinois, Zoological Pathology Program, Brookfield, IL, United States

INTRODUCTION All snakes are members of the suborder Serpentes, a monophyletic clade deeply embedded within the phylogeny of the Squamata (lizards). There are more than 3500 species of snakes. They are distinguished from other legless lizards by morphologic features of the skull and dentition that first appear in the fossil record during the Middle Jurassic-Lower Cretaceous periods. It is hypothesized that changes to the cranium preceded the loss of limbs, and that the first snakes were likely short-bodied lizards with limbs (Caldwell et al., 2015). Ophidian taxonomy, like that for much of herpetofauna, is fraught with disagreement and is in a near constant state of flux. It is generally accepted that the basal division of extant species occurs between the Scolecophidia (blind and thread snakes) and the Alethinophidia (the “advanced” snakes). The Alethinophidia contains at least five superfamilies, including the Acrochordoidea (file snakes), the Uropeltoidea (pipe and shield-tailed snakes), the Pythonoidea (pythons and sunbeam snakes), the Booidea (boas), and the incredibly diverse Colubroidea (colubrids, elapids, viperids, and numerous others) (Pyron et al., 2011; Vidal et al., 2007). Approximately one in nine snake species are estimated to be threatened with extinction according to an assessment of the conservation status of reptiles globally (Böhm et al., 2013). Species inhabiting freshwater environments, tropical regions, and oceanic islands are considered at greatest risk. Habitat loss, climate change, over collection, and infectious diseases are all contributors to species declines. As reptilian macro- and microscopic anatomy can be quite intimidating, at least initially, a significant portion of this chapter is dedicated to highlighting some fundamental ophidian gross and histologic features. Many of these features are of particular importance in differentiating normal species variation from pathology associated with common Pathology of Wildlife and Zoo Animals. http://dx.doi.org/10.1016/B978-0-12-805306-5.00037-7 Copyright © 2018 Elsevier Inc. All rights reserved.

snake diseases. In this chapter, particular emphasis will also be placed on infectious diseases, particularly viruses and fungi, of snakes. Ophidian virology and mycology are rapidly evolving areas of reptile medicine and pathology that can have direct implications for conservation of free ranging species, ex situ conservation, and individuals within zoological and captive collections.

UNIQUE FEATURES Clinical Pathology Cellular components of snake blood include nucleated erythrocytes, nucleated thrombocytes, granulocytes (heterophils, eosinophils, and basophils) and mononuclear cells (lymphocytes, monocytes, azurophils, and plasma cells). Mature circulating erythrocytes are ellipsoid with oval, centrally located nuclei. While species differences exist, erythrocytes are on average 17.3 (± 2 ) µm in length and 9.8 (± 1.5) µm in width; erythrocyte nuclei are 7 (± 1.1) µm in length and 3.7 (± 0.6) µm in width with an average nuclear surface to cell surface ratio of 0.155 (± 0.063) (Saint Girons, 1970). Very low numbers of immature erythrocytes may be present in circulating blood of healthy animals. Increased circulating immature erythrocytes can be seen in young snakes or snakes undergoing ecdysis. Snake total red blood cell counts (TRBCs) are lower than those of mammals and birds but higher than those of chelonians, though there is significant variation in the TRBCs between snake species (378,000–1,900,000 erythrocytes/µL) (Saint Girons, 1970). Thrombocytes play a major role in coagulation and wound healing. Unlike mammalian platelets, thrombocytes are nucleated with clear cytoplasm and a single, central, hyperchromatic nucleus with dense chromatin. Circulating thrombocyte numbers are high, and ∼1.5–3 thrombocytes can be present per single leukocyte (Saint Girons, 1970). 897

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Thrombocytes may closely resemble small lymphocytes, which is important in the proper differentiation during blood film evaluation. Periodic acid-Schiff with diastase (PAS) staining can assist in differentiating positively stained thrombocytes from nonstaining lymphocytes (Alleman, Jacobson, and Raskin, 1999). On blood films, another differentiating feature is the tendency of thrombocytes to form aggregates not observed with small lymphocytes. Care must be taken when interpreting complete blood count results, as lymphocyte counts can be artifactually elevated due to the inclusion of thrombocytes in calculated values. Snake heterophils are the largest circulating granulocytes (17.7±3 µm). They are rounded cells with colorless cytoplasm that contain fusiform, eosinophilic granules (Saint Girons, 1970). The nuclei of snake heterophils are eccentrically placed and may be oval to lobulated depending on the species. Snake eosinophils can easily be confused for heterophils, and discrepancies in granulocyte differentiation between even closely related species are present in the literature (Alleman et al., 1999; Troiano et al., 1997). Distinct, spherical, cytoplasmic granules are a characteristic feature of ophidian eosinophils (Strik et al., 2007). Azurophils are large and round cells with basophilic to amphophilic, punctate to granular cytoplasm that may contain clear or pale basophilic, well demarcated vacuoles. They are commonly encountered leukocytes in snake blood, and in some studies, they are the most numerous circulating leukocyte (Troiano et al., 1997). Though reptilian azurophils are most commonly considered to be of monocytic origin, this may not be the case for snakes. Cytochemical staining profiles of azurophils in two viperids, the eastern diamondback rattlesnake and the jararaca, are much more similar to those of mammalian neutrophils than mammalian monocytes (Alleman et al., 1999; Egami and Sasso 1988). Furthermore, ophidian azurophils also produce an oxidative burst similar to mammalian neutrophils that is absent in other reptilian monocytes (Heard et al., 2004). As such, it is possible that snake azurophils are fundamentally unique to those of chelonians and other squamates. Basophils, the smallest of the snake granulocytes (11.5±3 µm), are circular cells with central, weakly staining, round nuclei that are often obscured by dense, basophilic, metachromatic granules (Saint Girons, 1970). The granules can variably dissolve in water-based stains, and alcohol fixation and Romanowsky type stains are preferred for optimal basophil visualization (Campbell, 2014). Though the exact function of snake basophils is unknown, based on findings in basophils of other reptiles, the cells are believed to function similar to their mammalian counterparts (Strik et al., 2007). Lymphocytes are round with small to moderate amounts of transparent, weakly basophilic cytoplasm and a large, round to ovoid nucleus with clumped chromatin. Nuclei are

typically located centrally in small lymphocytes, though eccentric nuclei can be observed in larger lymphocytes (Saint Girons, 1970). As in mammals, B- and T-lymphocytes are present. Large, reactive lymphocytes with abundant, variably vacuolated, basophilic cytoplasm can be observed under antigenic stimulation. Plasma cells, which are rarely observed in the blood of healthy reptiles, can increase in number under antigenic stimulation. Ophidian plasma cells are similar to their mammalian counterparts and are round to oval with abundant basophilic cytoplasm and a prominent, pale staining, perinuclear halo. Activated plasma cells with abundant well demarcated, round to ovoid, eosinophilic, cytoplasmic vacuoles (Mott-like cells) can also be seen with chronic stimulation. Monocytes are large, variable, round to amoeboid leukocytes with abundant pale basophilic cytoplasm and pleomorphic nuclei with finely clumped chromatin. While not abundant in the blood of healthy snakes, monocyte numbers can increase with chronic stimulation. Activated monocytes may contain cytoplasmic vacuoles (Strik et al., 2007).

Anatomic Features The epidermis of snakes is arranged in scales that are overlain by a vertical arrangement of two types of keratin: inner, elastic, helical, α-keratin, and outer sheets of mechanically resistant, β-keratin (Fig. e1). The two types of keratin contribute to the four cornified layers that exist above the stratum basale: the α-, mesos, β-, and Oberhäutchen layers. The innermost α- and mesos layers consist of α-keratin and contain distinct cellular outlines. The β- and outermost Oberhäutchen layers are composed of β-keratin and are fused into a compact stratum lacking cell boundaries. Keratinocytes producing each type of keratin originate from a uniform stratum basale prior to differentiation into either an α- or β-subtype (Landman, 1986). While the distribution of α-keratin is fairly uniform across the body, β-keratin is thickest on the scales and thinnest, down to a single cell layer, in the hinge region between scales. Snake skin lacks osteoderms and dermal glands, and a variety of pigment cells (chromatophores) can be present. Generally, a single layer of pteridine- and/or carotenoid-containing xanthophores and erythrophores are positioned subjacent to the epidermal basal lamina. These are first subtended by a variably thick layer of iridophores with their characteristic refractile, cytoplasmic crystals and then by variable numbers of large melanophores with prominent dendritic processes (Landman, 1986). Melanophores are also variably distributed within the stratum basale of the epidermis and melanosomes can be present in the β-layer of the corneum. The ophidian spinal column is divided into three segments: precloacal, cloacal/sacral and caudal. Vertebral numbers can be quite variable, ranging from 160 to greater

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than 400, with a majority of vertebral bodies being in the precloacal and cloacal/sacral segments (120 to greater than 320). In the precloacal and cloacal/sacral segments, ribs articulate with vertebral bodies by two opposing ligaments oriented dorsally and ventrally. The first pair of ribs attaches to vertebral bodies 2, 3, or 4 depending on the snake family; the atlas never bears ribs. In the cloacal/sacral region, the ribs are forked, and while the first pair of ribs are usually articulated, the rest are generally fused with the vertebral bodies. The vestigial pelvic girdle, which is present in some snakes (Scolephidia, Anilliidae, Boidae), occurs at the junction between the precaudal and cloacal/sacral regions. In the caudal segment, ribs are always fused to the vertebrae. While the length of the caudal segment is variable, the caudal segment is always shorter than the combined length of the precloacal and cloacal/sacral segments. Intervertebral disks are absent in snakes, and adjacent vertebrae are connected via modified ball-and-socket articular joints between the cartilaginous surfaces of the convex condyle and the concave cotyle (Fig. e2) (Hoffstetter and Gasc, 1969). The ophidian heart has four chambers: a single ventricle, a pair of atria, and the sinus venosus. The sinus venosus lies dorsally at the confluence of the three caval veins; the walls of the sinus venosus are the thinnest of the heart chambers and contain cardiac muscle with pacemaker activity. The right atrium in snakes is larger and more muscular than the left, and the junction between the atria and the ventricle is marked by the coronary sulcus. The ventricle is usually elongate, and is composed of a distinct compact layer of cardiac muscle surrounding a spongy, medullary myocardium organized into a series of ridges that serve to divide the ventricular lumen into “chambers.” Three twisting arterial trunks, the pulmonary, left, and right aortic arches, emerge from the heart base. The position of the heart within the coelomic cavity is more caudal in aquatic species than in terrestrial species, and a more cranially positioned heart is found in climbing, arboreal species and those that behaviorally raise their heads, such as elapids (Farrell et al., 1998). The ventricular myocardium is histologically organized into a compact (cortical) zone surrounding a loosely arranged, spongy, medullary region with sparse connective tissue. The muscle fibers are arranged in sheets that run lengthwise in the outermost cortical zone and form two spiral layers, forming an inner, circular layer that contributes to the inner portion of the cortical zone and the medullary zone. In the atria, a prominent cortical layer is absent (Farrell et al., 1998). The ophidian trachea is composed of hundreds of rings of hyaline cartilage. The majority of the tracheal rings are incomplete, although a short glottal segment composed of complete tracheal rings can be seen in certain species. The trachea most commonly enters the caudal margin of the most cranial portion of the lung. In some species, the trachea abruptly ends and the open, central lumen (­vorbronchus)

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continues caudally. In other species, the trachea continues along the dorsal surface of the lungs before transitioning into a bronchus. The glottis, larynx, and trachea are lined by ciliated, columnar epithelial cells, nonciliated epithelial cells, and admixed goblet cells. Snake lungs are elongate and asymmetrical, with the left lung typically being smaller than the right. In certain species, the left lung is very rudimentary. The pulmonary parenchyma can be divided into three regions: a cranial, thick, vascular, respiratory portion; a transitional, thinwalled, semisaccular, variably respiratory portion; and a caudal, membranous, avascular, saccular, nonrespiratory portion. Lesions are most commonly observed in the respiratory portions of the lung, though extension into the nonrespiratory portion can be seen in severe pneumonia. Trabeculae along the luminal surface of the vascular and semisaccular portions of the lung form a network of faveolar spaces (Wallach, 1998). Histologically, trabeculae consist of fibrovascular septa ending in cords of smooth muscle cells lined by cuboidal to columnar, ciliated epithelial cells; secretory epithelial cells surround faveolar spaces. The faveoli are primarily lined by flattened, Type I pneumocytes and fewer cuboidal, Type II pneumocytes (Fig. 37.1). Pneumocyte hyperplasia is a prominent feature of certain viral pneumonias. Mucosal associated lymphoid tissue (MALT) aggregates are present multifocally along the bronchus and within the respiratory portions of the lung. The aggregates are composed of lymphocytes, plasma cells, macrophages, and interstitial cells. As the lung transitions into the nonrespiratory, saccular portion, the pulmonary lining changes to a columnar to squamous epithelium (Wallach, 1998). The snake thyroid gland is unpaired and ovoid to spherical, lies cranial to the heart and ventral to the trachea, and it is in close relation to the parathyroid glands, ­ultimobranchial

FIGURE 37.1  Normal lung from a boa constrictor. This section of the vascular, respiratory portion of the lung highlights faveolar septa lined by flattened pneumocytes.

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body and thymus (Lynn, 1970). The thyroid gland plays a crucial role in the process of ecdysis (shedding), and while histologic features can vary depending on the stage of epidermal regeneration, there can be substantial variation within individuals and species. Variable numbers of pairs (2–5) of parathyroid glands have been described in snakes; the glands are small (0.5–1 mm in diameter) and are not intimately associated with vessels (Clark, 1970). The thymus is composed predominantly of lymphoid cells (thymocytes) arranged in lobules with a medulla and a cortex. The number of thymocytes decreases with age. Myoid cells are also present, at highest concentration within the medullary areas. Hassall’s corpuscles, as seen in mammals, are absent in the reptilian thymus. Instead, thymic epithelial cells form cyst-like structures that surround granulocytes, myoid cells, and variable amounts of degenerative cellular debris (Bockman, 1970). The oral mucosa can show considerable variation between species and is composed of squamous, nonkeratinized epithelial cells, columnar epithelial cells, ciliated epithelial cells, and goblet cells. The palatine glands present in most other reptiles are absent in snakes, but other oral glands including sublingual, lingual, labial, and (in some species) venom glands are present. Oral glands can be serous, mucinous, or mixed. Snake teeth are ankylosed to the rims of shallow depressions in the dentary (mandible), maxilla, palatine, and pterygoid regions, and there is rapid, regular resorption, and replacement of teeth. One to four replacement teeth develop along the bases of their predecessors, lying with their tips angled posteriorly. In certain snakes, particularly the elapids and viperids, there is considerable modification of a subset of conical teeth to form fangs, wherein one wall, most often the mesial, is pushed inward to either create a partial crescential or complete tubular cavity to channel venom gland secretions (Edmund, 1969). In certain species, such as egg-eating snakes, there is a dramatic reduction to absence of dentition. Breakage or slitting of ingested eggs is achieved by lengthened projections of the ventral portion of the vertebral bodies immediately caudal to the head (hypapophyses) (Hoffstetter and Gasc, 1969). Swallowing of large food items in all snakes is permitted by an incredible degree of cranial kinesis afforded by loose attachment of the skull and quadrate bones. The esophagus is thin-walled and incredibly distensible with prominent mucosal folds when empty. It is lined by intermixed cuboidal to columnar ciliated epithelial cells and goblet cells interspersed with MALT aggregates. In some species, particularly boids, the aggregates can form well-defined structures (esophageal “tonsils”) (Jacobson, 2007b). The stomach is thick-walled and muscular with prominent rugae that clearly delineate the junction between the esophagus and stomach (Luppa, 1977). The luminal diameter of the stomach is typically not much greater than

that of either the esophagus or the duodenum. The stomach is composed of two segments, the fundus (corpus) and pars pylorica. The fundic portion of the stomach is composed of densely packed glands. Three types of glands are described in the fundus of reptile stomachs, and while there are species differences, all have been described in snakes. The glands are variably composed of neck cells (found in Type 2 glands), densely eosinophilic, granular, serous cells (found in Type 1, 2, and 3 glands), and lightly basophilic, “clear” mucus cells (found in Type 1 glands). Gastric glands terminate individually or in pairs in gastric pits. The gastric pits are lined by a basal layer of cuboidal cells with basal nuclei and a luminal layer of tall columnar cells with apical nuclei and apical cap-like expansions (Luppa, 1977). Additional populations of spindle-shaped and enterochromaffin-like cells are scattered throughout the fundic, and to a lesser extent, the pyloric glands. These have been shown in other reptiles to have probable neuroendocrine functions (D’Este et al., 1993). Pyloric glands are shorter and have fewer branches than the fundic glands. They are lined exclusively by mucus cells; gastric pits in the pyloric region resemble those in the fundus. No pyloric sphincter is present (Luppa, 1977). The snake intestinal tract is the shortest of the reptiles. The small intestine is partially coiled and adjacent, transverse folds are linked by connective tissue. The small intestinal mucosa is composed of densely packed villi arranged in ribbon-like folds. Villi are lined by a simple epithelium composed primarily of enterocytes and goblet cells with fewer granular, Paneth-like cells and enterochromaffin cells. Cryptlike folds described in other reptiles are absent in snakes. MALT aggregates can be found within the lamina propria, and a distinct muscularis mucosae is not always appreciable. The lamina muscularis is composed of internal circular and outer longitudinal smooth muscle layers. The large intestine is thin-walled, and while folds similar to those noted for the small intestine are present, the density of the folds greatly decreases caudally (Parsons and Cameron, 1977). The large intestine and cloaca are histologically similar to the small intestine, though the mucosal enterocytes are subjectively taller and MALT aggregates are in greater concentration (Luppa, 1977). Cloacal glands (musk glands) are also present in some species, particularly of the Colubridae. Ophidian livers are elongate, dorsoventrally flattened, and brown to dark brown in appearance. The dorsally positioned portal vein and the ventrally positioned hepatic vein divide the liver into left and right lobes. In many species, the lobes are of similar length, while in others the left lobe can extend beyond the right lobe both cranially and dorsally (Schaffner, 1998). Reptile hepatocellular organization is intermediate between that of fish and mammals. Snake hepatocytes are arranged primarily in two-cell thick plates and occasional tubules that converge toward terminal hepatic venules. Sinusoids are lined by fenestrated endothelial cells, pigmented

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macrophages, Kupffer cells, and stellate (Ito) cells (Schaffner, 1998). Both vacuolated and empty Ito cells have been demonstrated in snake livers (Taira and Mutoh, 1981). Bile canaliculi form a reticular network throughout the parenchyma that drain into ductules located primarily near portal tracts. Pigmented macrophages are present individually as well as in aggregates (pigmented macrophage centers). They contain iron and pigment and can exhibit significant variability in their light microscopic appearance among fish, amphibians, and reptiles. Because of this, the term “pigmented macrophage” is preferred over the more commonly used term, “melanomacrophage.” Pigmented macrophage centers can increase in size and number with age and chronic antigenic stimulation (absolute hypertrophy and hyperplasia). Relative hepatic pigmented macrophage center hypertrophy and hyperplasia can also be observed with chronic inanition and concomitant hepatocellular atrophy. The gall bladder lies a considerable distance caudal to the liver in close association with the pancreas and spleen. The close spatial relationship of the spleen, pancreas, and gall bladder can complicate gross and histologic evaluation of these organs, as bile imbibition can intensify regional postmortem autolysis. Snake spleens are generally oval or spherical and located cranial to the pancreas. However, in many species the two organs are in intimate contact with one another and are referred to as the splenopancreas. The spleen is composed of nodules of lymphoid tissue that encircle terminal splenic arteries and are separated by bands of fibrous tissue (Tanaka, 1998). Classic red pulp, as observed in mammalian spleens, is practically absent in snakes (Jacobson 2007b; Tanaka 1998). Histologically, splenic parenchyma consists of three tissue zones: fibrous capsule/trabeculae, lymphoid tissue, and a perilymphoid fibrous zone (PLFZ) (Tanaka, 1998). Lymphoid tissue surrounds terminal arteries and can appear nodular due to the presence of the fibrous trabeculae and PLFZs. Lymphoid aggregates are composed of lymphocytes, reticular cells, macrophages, and dendritic cells (Fig. e3). Periarteriolar lymphoid sheaths surrounding septal arterioles are absent. The PLFZ contains numerous small venous vessels and many infiltrating lymphocytes, macrophages, and other interstitial cells. While with routine staining, the distinction between the PLFZ and white pulp can be unclear, reticulin histochemical staining of connective tissue fibers in the PLFZ can clearly differentiate the two zones (Tanaka, 1998). Pigmented macrophage centers are also present (Fig. e3), that like their hepatic counterparts can increase in size and number with age and chronic antigenic stimulation. The pancreatic parenchyma is composed of branching exocrine tubules and irregularly branching endocrine cords. Parallel rows of polarized zymogen cells surround a central lumen. Alternating alpha and beta endocrine cells lie in close proximity to exocrine ducts and tubules and can be found

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beneath the pancreatic capsule, adjacent to the spleen, and occasionally within the spleen (Miller and Lagios, 1970). Snake kidneys are elongate and composed of offset renal lobules that confer a “stacked-coin” gross appearance. The number of lobules is variable by species, as is the relative position of the left and right kidneys. While the right kidney is most often situated cranially to the left, the two can be found at about the same position in some species. Snake glomeruli are round to ovoid and small (60– 120 µm) with a connective tissue center and capillaries that are located primarily at the glomerular surface. Bowman’s capsule leads to a short, nonsecretory neck segment lined by variably ciliated, cuboidal epithelial cells. The neck leads into the proximal convoluted tubule (PCT). In some snakes, short blind aglomerular diverticulae may originate from the PCT (Fox, 1977). Tubular epithelial cells of the PCT may contain abundant secretory granules and lipid droplets. The nephron proceeds into the nonsecretory, distal convoluted tubule composed of an initial ciliated region followed by a section containing abundant mucus cells. In males only, a terminal segment of the distal convoluted tubule termed as the sexual segment of the nephron, is significantly hypertrophied and lined by tubular epithelial cells containing large, prominent, brightly eosinophilic, and cytoplasmic granules (Fig. 37.2) (Gist, 2011). The sexual segment of the nephron can be seen grossly as linear, pale tan or white, radiating parenchymal streaks. Secretory activity of the segment is seasonally variable, and regression can occur. Ureters travel from the kidneys to papillae on the dorsal surface of the cloaca; snakes lack a true urinary bladder though mild dilatation of the ureters prior to their cloacal insertion can be observed in many snakes (Fox, 1977).

FIGURE 37.2  Normal kidney from a male emerald tree boa. Large, prominent, eosinophilic cytoplasmic granules are characteristic of epithelial cells lining the terminal segment of the distal convoluted tubule in males (sexual segment of the nephron) but not females.

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In both sexes, the right gonad lies cranial to the left. Testicles are cylindrical to flattened, elongated, and covered by a thin, elastic tunica. Seasonal variation in testicular volume is common and can be dramatic in some species. Interstitial cells can be a prominent feature of snake testicles. Both the size and appearance of interstitial cells may exhibit seasonal variability, and they can account for up to 20% of the total testicular volume (Fox, 1977). Efferent ducts connect seminiferous tubules to a narrow epididymis along the full length of the testicle. A convoluted vas deferens travels caudally along the lateral surface of the kidney and follows the ureter to a common insertion at the cloaca near the paired hemipenes (Fox, 1977). Ovaries are sac-like and elongated with a central lymph-filled cavity surrounded by loose stroma with embedded follicles, oocytes, and/or corpora lutea; the two ovaries are often of unequal length. Oviducts receive ovulated eggs in the thin, funnel shaped, and highly convoluted infundibula that connect to the uterus and end in a highly muscular vagina that opens to the cloaca (Gist, 2011). Some sources identify an additional oviductal segment (uterine tube) between the infundibulum and uterus in at least some snake species (Girling, 2002), and the left oviduct is absent in some species (Fox, 1977). The histologic organization of the oviduct varies based on whether the species is oviparous, ovoviviparous, or viviparous. The adrenal glands are paired and filiform, lying within the gonadal mesentery dorsal to the gonads. Like the closely associated gonads, the right gland is located cranial to the left. Glands are composed of anastomosing cords of interrenal cells interposed with islets of chromaffin cells and no gross corticomedullary distinction is present (Gabe, 1970). Like other reptiles, snake adipose stores are concentrated in coelomic fat bodies. However, unlike lacertids and crocodilians, numerous discrete adipose lobules can extend ventrally along the full length of the coelomic cavity in well- and overconditioned animals. Snake eyes are approximately spherical. The lens is also nearly spherical, and unlike other reptiles, has sutures. The sclera is completely fibrous and lacks cartilage or bone. The cornea is very large, and may be more strongly curved than the rest of the eye. Bowman’s membrane is absent, and Descemet’s membrane is very thin (Underwood, 1970). The spectacle, a fixed, transparent, scale, overlies the cornea. The snake vomeronasal (Jacobson’s) organ is a paired, highly developed, auxiliary, olfactory sense organ. Each organ is a nearly spherical structure that is positioned caudal to the external naris and cranial to the globe and sandwiched between the ventral oral cavity and dorsal nasal cavity. The organs are separated from the rest of the nasal cavity but communicate with the oral cavity/choana by very narrow ducts that transmit olfactory cues from the tongue to the glands (Jacobson, 2007b). The lumen of the vomeronasal organ contains three different regions (Fig. 37.3).

FIGURE 37.3  A longitudinal section of a normal vomeronasal (Jacobson’s) organ in a smooth green snake. Dorsally, the vomeronasal organ is lined by pseudostratified epithelium containing bipolar neurons with dendritic terminations at the luminal surface. These detect olfactory cues transmitted to the lumen of the vomeronasal organ by narrow ducts connected to the oral cavity.

The d­ orsal (concave) region is lined by a pseudostratified epithelium containing bipolar neurons whose dendrites terminate at the luminal surface. The ventral (convex) region is lined by a layer of simple ciliated columnar cells that overlie the mushroom body, a projection of hyaline cartilage and fibrovascular connective tissue. The aforementioned regions are connected by a shallow, stratified, layer of cuboidal epithelial cells (Gharzi et al., 2013). Pit organs are radiant heat receptors that are highly developed in some snakes, which include the Boidae, Pythonidae, and Crotalinae subfamily of the Viperidae. In the Crotalinae, facial pits are prominent, anterior-facing, deep depressions on either side of the face between the eye and the nostril. A sensory membrane divides the pit into two chambers and contains densely packed free nerve endings from the ophthalmic and maxillary divisions of the trigeminal nerve. In the Boidae and Pythonidae, labial pits are shallow depressions found in labial, supralabial, infralabial, and/or rostral scales. Free nerve endings from all three main branches of the trigeminal nerve extend up into the epidermal scales at the base of the labial pits (Barrett et al., 1970). Specialized oral glands that secrete proteins and polypeptides and assist in prey immobilization, digestion, and defense have evolved in several families. The Viperidae (night adders, pit, and pit-less vipers) and Elapidae (adders, cobras, coral snakes, kraits, mambas, and sea snakes) are front-fanged snakes, and have large venom glands located laterally on the head and ventrocaudal to the orbit. Secretions from the venom gland (and in many species an additional accessory gland) pass through a variable number of ducts to the fangs. The glands consist of simple to compound, branching tubules surrounding a ­central

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lumen (Kochva, 1978). In contrast, many snakes in the family Colubridae are rear-fanged and venom is produced by Duvernoy’s gland, a modified parotid gland (Hill and Mackessy, 2000). Duvernoy’s glands are located immediately dorsal to the maxilla, lack a central reservoir, and are composed of predominantly serous cells that produce proteolytic venom. Venom is transmitted via ducts to the base of solid, maxillary teeth (Weinstein et al., 2009). Many snakes of the genera Crotalus and Sistrurus accumulate shed keratin from modified tail tip scales to form a rattle. The interlocked, shed scales accumulate at the tip of the tail, and with high-frequency contraction of modified tail shaker muscles, vibrate against each other to create the distinctive and characteristic rattling sound (Savitzky and Moon, 2008).

NON-INFECTIOUS DISEASES Nutritional As in other reptiles, gout, the deposition of uric acid in tissues, is commonly encountered in the kidney and viscera of snakes. The kidneys can be swollen and contain multifocal to miliary, white-tan, pinpoint nodules (Fig. 37.4A). On gross examination, care must be taken to differentiate tubular urate stasis or the sexual segment of the nephron in males, seen as radiating white or tan streaking, from true renal gout. Impression smears or squash preps of the kidney can be used to differentiate urate stasis and renal gout. In true renal gout, characteristic radiating, urate tophi will be present (Fig. e4), while in cases of urate stasis, round to amorphous urates will be seen. Histologically, radiating, acicular clefts (gout tophi) that often appear as empty spaces (the crystals dissolve during routine histologic processing) are surrounded by epithelioid macrophages, multinucleated giant cells, and heterophils (Fig. 37.4B). In snakes, secondary gout associated with dehydration is most common, and uric acid depo-

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sition in the kidneys typically precedes that of other tissues (commonly the pericardial sac, pleura, and the serosa of the liver and other coelomic organs). Articular (intervertebral) accumulations of uric acid are not as commonly seen in snakes, but can occur.

Age-Related/Degenerative Chronic vertebral osteopathy, chronic proliferative, degenerative, and/or inflammatory lesions in the vertebral bodies, intervertebral joints, and ribs, is a common finding in snakes. Lesions can include a spectrum of changes, which include bony remodeling, proliferative new bone and fibrocartilage formation, sclerosis, vertebral and costal spondylosis and ankylosis, degeneration of the articular cartilage, osteomyelitis, osteoarthritis, osteonecrosis, and pathologic fractures (Fitzgerald and Vera 2006; Jacobson 2007a). Chronic, noninflammatory lesions can include sclerotic changes in both cortical and trabecular bone that are characterized by widening and confluence of trabeculae by disorganized, irregular patches of woven and lamellar bone with mosaic reversal lines, which can be quite dramatic. Fibrocartilaginous proliferation can be a prominent feature. An increase in osteoclastic activity is variably reported. In other cases, chronic noninflammatory, degenerative changes may be admixed with small foci of chronic inflammation. This suggests that many cases may fall on a spectrum of disease progression from initial inflammatory lesions to chronic, aseptic, degenerative, and proliferative foci. Some comparisons have been made between chronic vertebral osteopathy in snakes and osteitis deformans (Paget’s disease) in humans (Preziosi et al., 2007). Features of vertebral osteopathy in snakes do not follow the same hypothesized pathologic progression as the human disease, and the designation of reptile lesions as osteitis deformans should be avoided (Jacobson, 2007a).

FIGURE 37.4  Renal gout in a corn snake. (A) Tophi composed of crystallized uric acid and associated inflammation are grossly visible as white, pinpoint foci that disrupt the normal parenchyma. (B) Histologically, granulomatous inflammation surrounds empty, radiating, acicular clefts. The precipitated uric acid dissolves during routine processing.

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rodents). Secondary bacterial infections are a common complication of husbandry-related traumatic injuries.

Miscellaneous

FIGURE 37.5  Necrotizing vertebral osteomyelitis in a corn snake due to Salmonella enterica arizonae. Well-defined, solid aggregates of heterophilic and granulomatous inflammation surround necrotic eosinophilic tissue and cellular debris and disrupt the normal architecture of the vertebral body.

Several differential etiologies have been proposed for chronic vertebral osteopathy in snakes including trauma, viral infection, nutritional deficiencies (hypovitaminosis A and hypervitaminosis D), chronic inactivity secondary to cage confinement, and chronic or resolved bacterial infections (Fitzgerald and Vera, 2006). A number of bacteria, notably Salmonella spp., Pseudomonas spp., and Staphylococcus spp., have been associated with spinal infection and active granulomatous and heterophilic inflammation (Jacobson, 2007a). Additionally, a specific serotype of Salmonella enterica arizonae (56:Z4, Z23) has been associated with osteotropism in a captive colony of ridgenose rattlesnakes. Infection produced heterophilic and granulomatous osteomyelitis, osteonecrosis, sequestra, and pathologic fractures (Ramsay et al., 2002). Salmonella enterica arizonae can be commonly isolated from other cases of vertebral osteomyelitis in snakes where culture is productive (Fig. 37.5). Spinal bacterial osteomyelitis can be a primary source of septicemia, and close examination of the spinal column for any irregularities or foci of bony proliferation is a necessary component of a snake necropsy.

Trauma Husbandry-related trauma can be an important cause of morbidity and mortality in captive snakes. Thermal burns can occur on the dorsum (due to heat lamps and/or radiant heat panels) or ventrum (due to heating pads, cable, and/or tape) and can be associated with extensive epidermal, dermal, and muscular necrosis. Localized facial trauma secondary to repetitive trauma (face-rubbing) with either the habitat furniture or the enclosure itself is also common. Bite wounds can occur if the snake is being fed live prey (typically larger

Ecdysis (shedding) in snakes is periodic and complete in most species. In the largest snake species (such as Burmese pythons and the anacondas), partial shedding can be observed and is considered normal. Ecdysis is preceded by the synchronous division and differentiation of the epidermis to create a new set of cornified epidermal layers. A cleavage zone forms between the Oberhäutchen of the new, inner cornified layer and the α-layer of the old, outer cornified layer, and is filled with lymph (Landman, 1986). In captivity, dysecdysis (incomplete ecdysis and shed retention) is common (Fig. e5). Dysecdysis can be a manifestation of chronic dehydration, malnutrition, underlying disease, and/ or improper husbandry conditions, such as low humidity and inappropriate temperatures. Chronic dysecdysis can be associated with dermatitis as either a sequela to the dermatitis or the cause of it; this is particularly true for scale retention around the margins of the oral cavity. Retention of the spectacle, the modified scale overlying the cornea, can result in secondary bacterial infections and inflammation. Edema, hyperkeratosis, and granulocyte infiltration of the spectacle (spectaculitis) with or without associated bacterial and/or fungal infections are reported (Da Silva et al., 2015). Ovoretention and dystocia are common in oviparous snakes and, in severe cases, can be associated with oviductal prolapse (Stahl, 2002). Salpingitis can either be a cause or sequela of ovoretention/dystocia. Ovoretention of nonviable ova (slugs) can also occur (Fig. 37.6). Chronic ovoretention is most often seen as a sterile condition, though bacterial salpingitis may result if there is rupture and release of yolk material. While yolk coelomitis can occur in snakes, it is far less common than in other squamates. Given the close anatomic proximity of the ovary and the adrenal gland, close examination of any antemortem biopsy submissions of the squamate female reproductive tract for the presence of adrenal tissue is recommended/warranted. The male hemipenes are sac-like extensions inverted within the base of the tail. Paraphimosis with prolapse and inflammation of the hemipenes is not uncommon. Inflammation of the hemipenes and the hemipenal sac are often associated with bacterial infections in the presence or absence of concurrent cloacitis; squamous metaplasia of the hemipenis and hemipenal sac mucosa is typical with chronic hemipenal inflammation.

Neoplastic Neoplasia is a common finding in captive snakes, and in retrospective studies of reptile neoplasia, snakes are more often diagnosed with neoplasia than other reptile ­species

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FIGURE 37.6  Chronic sterile ovoretention (oviductal slugs) in an Aruba Island rattlesnake. Yellow to green aggregates of inspissated yolk material (slugs) within the oviduct are variably associated with congestion and hemorrhage and/or surrounded by abundant translucent, yellow fluid. (Photo Courtesy of R. Ossiboff, Wildlife Conservation Society).

(Steeil et al., 2013). While certain neoplasms are more frequently encountered than others, a great diversity of ophidian tumors have been reported (Catao-Dias and ­Nichols 1999; Garner et al., 2004; Mauldin and Done 2006; Sykes and Trupkiewicz, 2006). Hematopoietic neoplasms, most notably lymphoma (Figs. 37.7–37.9), are common in snakes. Neoplastic aggregate distribution is most often multicentric, though solitary tumors, for example, in esophageal MALT (Fig. 37.8) are reported and leukemic extension and vascular tropisms can all be observed. Leukemia in the absence of solitary or multicentric lymphoma has also been reported in a number of species (Fig. 37.9) (Mauldin and Done, 2006). Neoplastic lymphocytes can have variable morphology, but are most commonly large and blast-like (Garner et al., 2004). Immunohistochemical characterization of neoplastic lymphocytes can be difficult due to a lack of appropriate reagents. CD3+ T-cell lymphomas seem to be most common in snakes (Figs. 37.7C, and 37.8), but as commercially available CD3 antibodies can be successfully used in many snake species while B-cell markers (BLA36, CD79a, and CD20) are of limited diagnostic utility, this observation may be biased. Mast cell tumors and histiocytic origin round cell tumors have also been reported in snakes (Garner et al., 2004; Mauldin and Done, 2006; Schumacher et al., 1998). IBA1 is a highly evolutionarily conserved protein expressed in the cytoplasm of activated macrophages and microglial cells, and shows promise as a marker of cells of histiocytic lineage in snakes as well as other nonmammalian vertebrates. Neoplasms are common throughout the ophidian gastrointestinal tract. Tumors of the oral cavity are observed frequently, with one of the most common being solitary or multifocal squamous cell carcinoma (Mauldin and

FIGURE 37.7  Thymic lymphoma in a ball python. (A) A large, mottled, light tan/red, multinodular mass effaces the thymus and expands the soft tissues cranial to and surrounding the heart. (B) The tumor is composed of sheets of neoplastic round cells with moderate anisocytosis and anisokaryosis. (C) Neoplastic cells are characterized by strong, membranepositive immunolabeling for CD3, consistent with T-lymphocyte origin.

Done, 2006). Neoplastic cells can be quite invasive and locally destructive, and metastasis, while reported, is uncommon (Steeil et al., 2013). Squamous cell carcinoma can arise

906 Pathology of Wildlife and Zoo Animals

FIGURE 37.8  T-cell lymphoma in esophageal mucosal associated lymphoid tissue (MALT) aggregates in a boa constrictor. MALT aggregates expanded by neoplastic lymphocytes are markedly congested and project from the mucosal surface. There are also multifocal areas of mucosal petechiation and ecchymosis. (Photo Courtesy of William R. Pritchard Veterinary Medical Teaching Hospital).

FIGURE 37.9  Leukemia in a western hognose. Sheets of neoplastic round cells expand and fill the lumina of faveolar capillaries and interstitial vessels.

de novo or in the face of chronic stomatitis. An association between chronic inflammation and neoplasia is possible but has not been fully examined. A ­variety of other epithelial and mesenchymal tumors, ­particularly ­fibrosarcoma, have been reported in the oral cavity (Mauldin and Done, 2006). Commercially available pan-cytokeratin (AE1/AE3) antibodies are often useful in immunostaining snake epithelial cells, while vimentin immunoreactivity is inconsistent and therefore of limited diagnostic utility. In a report of an oral amelanotic melanophoroma in a boa constrictor, neoplastic cells were positive for melan-A immunohistochemical staining (Thompson et al., 2015). Oral tumors of odontogenic origin are sporadically reported in snakes (Mauldin

and Done, 2006). An ameloblastoma was documented in a free ranging, black rat snake (Comolli et al., 2015). Gastric adenocarcinomas are locally invasive and expansile mural masses composed of tubulopapillary arrangements of columnar to cuboidal neoplastic epithelial cells with a prominent scirrhous response (Baron et al., 2014; Martin et al., 1994). Intestinal adenocarcinomas are common, particularly in colubrids. Both the distal small intestine and colon are common sites, though cloacal adenocarcinomas are also reported. In some tumors, neoplastic epithelial cells may contain a single, large, cytoplasmic vacuole (signet ring morphology); in others, cells may contain abundant, PAS-positive cytoplasm with distension of glandular lumina by wispy, basophilic material (mucinous morphology) (Latimer and Rich, 1998). Intestinal adenocarcinomas can be exophytic or mural, and, like their gastric counterparts, locally invasive with a prominent scirrhous reaction (Figs. 37.10A,B) (Jessup 1980; ­Latimer and Rich 1998; Mauldin and Done, 2006). Visceral ­metastasis of intestinal adenocarcinomas is uncommon but reported (Orós et al., 2004). Benign and malignant smooth ­muscle, endothelial, and unclassified mesenchymal origin neoplasms are also reported in the gastrointestinal tract (Mauldin and Done, 2006). Benign and malignant hepatic and biliary tumors are also a common diagnosis (Fig. 37.11); colubrids appear to be at higher risk for these neoplasms than other snakes (Garner et al., 2004; Sykes and Trupkiewicz, 2006). Both can be solitary or multifocal, and biliary neoplasms are often cystic. Carcinomas of the biliary duct and/or gall bladder have been reported in several viperids (Mauldin and Done, 2006). Adenocarcinomas of both the exocrine pancreas and of pancreatic ducts have been reported in multiple species of colubrids and viperids; the tumors, while well differentiated, can be extremely aggressive and metastasize (Garner et al., 2004). Exocrine pancreas adenomas and nodular exocrine hyperplasia are a frequent, generally incidental, finding. Renal adenocarcinomas are frequently diagnosed in snakes; colubrids are overrepresented (Fig. 37.12). The tumors are most often large and solitary, and composed of variably sized tubules lined by well-differentiated epithelial cells surrounded by a scirrhous stroma that expand and disrupt the normal parenchyma. Urate tophi are often present within the neoplastic tubules and the adjacent desmoplastic stroma (Garner et al., 2004). Large, cystic fluid-filled cavities containing urine and urates can also be seen. Renal tubular adenomas can be an incidental finding during postmortem evaluation and consist of solid, nodular foci of tubular proliferations that may compress the adjacent tissue but lack an associated desmoplastic response. Tumors of the female reproductive tract are more common than testicular tumors. Ovarian carcinoma, sarcoma, and benign granulosa cell tumors have all been reported

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FIGURE 37.10  Colonic adenocarcinoma in a gopher snake. (A) There is abrupt neoplastic transformation of the colonic mucosa with the formation of papilliferous projections. Neoplastic infiltrate of the colonic wall. The submucosa is segmentally expanded by colorless space (edema). (B) Islands and tubuloglandular arrangements of neoplastic epithelial cells invade the colon transmurally and are associated with a prominent scirrhous response and heterophilic inflammation.

FIGURE 37.11  Hepatocellular carcinoma in a Maximilian’s viper. A moderately demarcated, nodular, hepatocellular neoplasm expands the hepatic parenchyma regionally. Large areas of necrosis within the mass impart a mottled dark brown/tan appearance. The hepatic capsule overlying the mass is thickened. (Photo Courtesy of R. Ossiboff, Wildlife Conservation Society).

FIGURE 37.12  Renal adenocarcinoma in a Mexican black kingsnake. The caudal pole of the kidney is expanded by a cystic, multilobulated neoplasm. The neoplasm is composed of nodules of light brown neoplastic renal tissue admixed with large, fluid and urate-filled cavities. (The colon has been reflected ventrally to expose the kidney). (Photo Courtesy of J. Rodriguez-Ramos Fernandez, Wildlife Conservation Society).

(Mauldin and Done, 2006). Oviductal adenocarcinomas are exophytic and papillary and composed of acini or cords of neoplastic cells that may have areas of hemorrhage, necrosis, and inflammation. The biologic behavior of these tumors appears to be variable; in one retrospective study, no metastases were noted, while in another case series, visceral metastases were noted in all cases (Garner et al., 2004; Pereira and Viner, 2008). Tumors of the integument are commonly encountered in snakes. Squamous cell carcinomas are most commonly associated with the cloacal region and can originate from the hemipenes, cloacal glands, or cloacal skin (Garner et al., 2004). Chromatophoromas are neoplasms arising from the pigmented cells in either the epidermis (melanophores) or dermis (xanthophores, iridophores, and melanophores). Melanophoromas are most common, and are generally grossly gray or black and composed of neoplastic spindled cells with variably prominent black cytoplasmic granules (melanosomes) (Heckers et al., 2012). Both melan-A and S100 positive immunohistochemical staining has been reported in melanophoromas. These tumors can exhibit variable biological activity, with some being invasive with intravascular and visceral metastases. Iridophoromas are grossly white to light gray, dermal masses composed of neoplastic spindle cells with aggregates of golden brown to yellow-green cytoplasmic pigment, that is, refractile under polarized light. Iridophoromas in reptiles are most often benign, though malignant forms have been reported (Heckers et al., 2012). Definitive diagnosis of xanthophoromas is more difficult due to the absence of clear, distinguishing light microscopic features between neoplastic xanthophores and dedifferentiated melanophores and

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INFECTIOUS DISEASES DNA Viruses

FIGURE 37.13  Intratracheal chondroma in a ball python. A nodular mass composed of well-differentiated chondrocytes amid chondroid matrix projects into the lumen of the trachea. There is squamous metaplasia and lymphocytic and heterophilic inflammation of the overlying mucosa.

i­ridophores, and additional ­diagnostics including electron microscopy may be necessary. The cytoplasmic pigmentcontaining pterinosomes of xanthophores may exhibit a positive histochemical reaction with Fontana-Masson stains (Gregory et al., 1997). Cutaneous soft tissue sarcomas are also commonly found in snakes (Dietz et al., 2016). Locally invasive sarcomas composed of interlacing bundles and streams of neoplastic spindle cells, interpreted as fibrosarcomas are most commonly reported, and colubrids and viperids are overrepresented (Garner et al., 2004). Due to a lack of appropriate reagents and diagnostic criteria, further characterization of most soft tissue sarcomas in reptiles is limited. Endocrine tumors in snakes are rare. Thyroid and parathyroid adenomas are reported. Both pheochromocytomas and interrenal (adrenocortical) adenocarcinomas are also documented. Neoplastic interrenal cells from a woma python were positive for melan A, an immunohistochemical marker also used for neoplastic adrenocortical cells in mammals (Kaye et al., 2016). Neoplasms of the respiratory, cardiovascular, musculoskeletal, and central nervous systems are rare, though sporadically reported (Fig. e6), and intratracheal chondromas resulting in airway obstruction have been described in ball pythons (Fig. 37.13) (Diethelm et al., 1996; Drew et al., 1999). Hemangiosarcomas of the heart and spleen, respectively, have been described in colubrids (Shoemaker et al., 2016; Tuttle et al., 2006). Neoplastic cells in a case of cardiac hemangiosarcoma in a Madagascar giant hognose snake were positive for factor VIII-related antigen (Shoemaker et al., 2016).

Reports of herpesvirus infections in snakes are uncommon. Herpesviruses of the venom gland have been reported in cobras and kraits. In cobras with herpesvirus infection, inflammation, degeneration, and necrosis of the venom gland epithelium is associated with decreased venom production (Simpson et al., 1979). Hepatocellular necrosis with amphophilic intranuclear hepatocellular inclusions were observed in a clutch of captive boa constrictors, with concurrent pancreatic, renal, and adrenal intranuclear inclusions. While a herpesviral infection was not proven, the findings were strongly suggestive of a herpesvirus infection (Hauser et al., 1983). Herpesviruses have also been found as part of mixed infections in snakes with gastrointestinal disease (Marschang, 2014). A novel ranavirus was isolated and characterized from a group of green tree pythons with systemic disease. Lesions in affected snakes included ulcerative and necrotizing rhinitis, stomatitis, pharyngitis, and hepatic and splenic necrosis; viral inclusion bodies were not seen (Hyatt et al., 2002). Erythrocytic iridoviruses are seen in many species of squamates and chelonians. Erythrocyte inclusions in free-ranging northern water, plains garter, and eastern ribbon snakes attributed to assembly sites of snake erythrocytic iridoviruses have been reported in North America (Jacobson, 2007; Smith et al., 1994). Historical reports of snake erythrocytic inclusions were often attributed to protozoal organisms (Toddia or Pirhemocyton) in the absence of ultrastructural confirmation; as such, the prevalence of snake erythrocytic iridoviruses may be quite high. Iridoviral erythrocytic inclusions can be seen in both healthy and anemic snakes (Jacobson 2007; Johnsrude et al., 1997). Scattered reports of other viral infections of snakes are present in the literature. These include several adenoviruses that are associated with sporadic gastrointestinal disease. In boa constrictors, adenoviruses can cause hepatocellular necrosis with large, characteristic basophilic intranuclear hepatocellular viral inclusions (Jacobson, 2007). Adenoviral-like inclusion bodies can also be seen in enterocytes of a variety of colubrids with enteritis (Jacobson, 2007; ­Wozniak et al., 2000b).

RNA Viruses Inclusion body disease (IBD) is a global, transmissible, and progressively fatal disease of almost exclusively captive boids (boas and pythons). IBD was initially suspected to be caused by a retrovirus. However, subsequent research has identified divergent arenaviruses (family ­Arenaviridae;

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genus Reptarenavirus) in snakes with IBD. Arenaviruses are enveloped, segmented, single stranded, negative sense RNA viruses that were previously believed to infect only rodents and humans. All known snake arenaviruses are more closely related to each other than to the mammalian arenaviruses, and form the recently created genus Reptarenavirus. Though Koch’s postulates have not been yet entirely fulfilled and arenaviruses can be identified in apparently clinically healthy snakes, there is ample epidemiologic and in vitro evidence implicating highly divergent arenaviruses as the cause of snake IBD in North America and Europe (Bodewes et al., 2013; Hepojoki et al., 2015; Hetzel et al., 2013; Stenglein et al., 2012). One recent study identified arenaviral coinfections to be common in snakes clinical for IBD (Hepojoki et al., 2015), while another study identified the potential for high arenaviral prevalence in overtly healthy boids (Chang et al., 2016). Additional studies to determine whether certain arenaviral strains or species are associated with certain tissue tropisms and the development of clinical disease are needed. IBD was first identified in the 1970s in North America, but has since been reported in captive snakes in Europe, Australia, and Africa (Chang and Jacobson, 2010). Clinical signs of snakes with IBD include central nervous system abnormalities, such as flaccid paralysis, stargazing, and torticollis as well as chronic regurgitation and loss of body condition. Some individuals may succumb to the disease in weeks while others may survive for months; as such, it is a challenging disease to manage in captive collections. The most striking diagnostic feature of inclusion body disease is the presence of variably sized, eosinophilic to amphophilic, intracytoplasmic inclusions (Fig. 37.14). Inclusion bodies are composed primarily of the ∼68 kDa reptarenaviral nucleoprotein (also referred to as inclusion body disease protein) and cells with inclusions can frequently be observed in the absence of associated ­ inflammation

FIGURE 37.14  Inclusion body disease in a boa constrictor. Prominent, eosinophilic, intracytoplasmic, viral inclusion bodies are present in epithelial cells throughout the esophageal mucosa.

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(Chang and Jacobson, 2010; Hetzel et al., 2013; Wozniak et al., 2000a). Within the central nervous system, inclusions may be observed in neurons and glial cells. In snakes (particularly pythons) with a more rapid progression of neurologic disease, inclusions are often limited to the central nervous system. In snakes with a more chronic manifestation of ­disease, such as has been reported in boa constrictors, inclusions can be seen throughout the body, including enteric, respiratory and renal epithelial cells, hepatocytes, pancreatic acinar cells, and mononuclear cells (Jacobson, 2007). Inclusions in lymphocytes are prominent within lymphoid aggregates in the esophagus (esophageal tonsils), though they can be observed s­ ystemically and in circulating leukocytes in blood smears (Chang et al., 2016). Secondary infections (bacterial, fungal, and protozoal) are common and a significant factor in IBD mortalities; concurrent lymphoid neoplasia has also been reported (Jacobson, 2007). The nidoviruses (family Coronaviridae; subfamily Torovirinae) are an important emerging group of viruses in reptiles, and particularly in boid snakes. Nidoviruses are large, enveloped, negative sense, single-stranded RNA viruses in the same subfamily as fish bafiniviruses and mammalian toroviruses, though they are at present not further classified within the Torovirinae. Fatal respiratory disease in ball pythons and an Indian python have been reported (Bodewes et al., 2014; Stenglein et al., 2014; Uccellini et al., 2014). Additional nidoviruses from a number of other python and boa species are pending ­characterization. Pythons with nidoviral infections most often present with signs of respiratory disease, including open mouth, labored, and audible breathing and copious mucoid to exudative, oral and glottal/tracheal discharge. On gross examination, catarrhal and necroexudative stomatitis, tracheitis, and pneumonia are present (Fig. 37.15A). ­Aggregates of mucus and exudate frequently expand faveoli and occlude the vorbronchus; similar material swallowed from the oral cavity can also be present along the length of the gastrointestinal tract. Histologically, ­significant ­epithelial hyperplasia and necrosis accompanied by lymphoplasmacytic and heterophilic inflammation can be seen in the oral and nasal cavities, glottis, trachea, proximal esophagus, and the lungs; pneumocyte hyperplasia can be profound (Fig. 37.15B). Abundant viral RNA can be observed within oral, tracheal, esophageal, and pulmonary epithelial cells (Fig. 37.15C). Secondary Gram-negative bacterial stomatitis and bronchopneumonia are common. Paramyxoviruses are another important group of viruses responsible for respiratory disease in captive snakes. Paramyxoviruses are enveloped, negative sense, and single stranded RNA viruses. All identified ophidian paramyxoviruses (OPMVs) to date are members of the genus Ferlavirus, though substantial viral diversity has been reported (Wellehan and Johnson, 2005). OPMVs are primarily reported in captive viperids and crotalids, though

910 Pathology of Wildlife and Zoo Animals

FIGURE 37.16  Lung from a king snake with paramyxoviral pneumonia. Faveolar pneumocytes are hyperplastic and small numbers of syncytial cells are present. Small numbers of pneumocytes also have pale eosinophilic, intracytoplasmic viral inclusion bodies. Intact and degenerate heterophils and cellular debris accumulate within the faveolar space.

FIGURE 37.15  Nidoviral pneumonia in a ball python. (A) The vorbronchus and faveolar lumina are expanded and obscured by accumulations of mucus, inflammatory exudate and necrotic debris; the pulmonary parenchyma is edematous and congested. (B) Severely hyperplastic faveolar pneumocytes surround sloughed and exudative cellular debris; numerous lymphocytes, plasma cells, and fewer granulocytes are present within the interstitium. (C) Positive in situ hybridization to Ball Python Nidovirus RNA colocalizes viral RNA with pulmonary epithelial hyperplasia and inflammation.

colubrids, elapids, and boids are also susceptible (Hyndman et al., 2013). Gross lesions are primarily noted in the lungs, which are often thickened and edematous. Hemorrhage and necroexudative material can accumulate in airways along the length of the lung, including the caudal, saccular nonrespiratory portion. Hemorrhage and exudate in the oral and nasal cavities, glottis, and esophagus as well as coelomic cavity hemorrhage may also be present. Histologic pulmonary changes are similar to those in other viral pneumonias of snakes, with prominent pneumocyte hyperplasia and variable lymphocytic, interstitial inflammation. Concurrent heterophilic bronchopneumonia associated with secondary Gram-negative bacterial infections is common. The presence of both pulmonary epithelial syncytia and eosinophilic, intracytoplasmic inclusion bodies within pneumocytes are diagnostic features of OPMVs (Fig. 37.16). The presence of these features in the concurrent absence of a segmental esophagitis that is typical in nidovirus infections can help differentiate between the two etiologies. Pancreatic inflammation and necrosis may be present in OPMV infection, and in some snakes, may be striking (Hyndman et al., 2013; Jacobson, 2007). Snakes with OPMV may also have concurrent neurologic disease with meningoencephalitis, lymphocytic perivascular cuffing, demyelination, axonal and neuronal degeneration, and/or intracytoplasmic glial and intranuclear neuronal viral inclusion bodies (Hyndman et al., 2013). A number of reoviruses (family Reoviridae; genus Orthoreovirus) have been reported in snakes. Reovirus infections are a diagnostic challenge due to the overlap of respiratory and neurorespiratory lesions with those of nidovirus and paramyxovirus infections, respectively. Reports of reoviral disease in snakes are characterized by mild proliferative pneumonia and tracheitis (­Jacobson,  2007;

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Bacteria

FIGURE 37.17  Cranial medulla of a carpet python with neurologic disease and Sunshine virus infection. Severe white matter spongiosis, which appears as variably sized clear spaces, and mild to moderate gliosis are characteristic lesions. (Photo Courtesy of C. Shilton and K. Dyrting, Berrimah Veterinary Laboratories).

­ amirande et al., 1999). Inclusion bodies, epithelial syncyL tia, and hemorrhage, which can be diagnostic features of OPMV infection, are not reported, nor is segmental esophagitis, a common diagnostic finding in nidovirus infections. In cases of suspected viral respiratory disease, virus isolation on ophidian cell lines, Vero, and CV-1 cells, is recommended, as is targeted PCR testing for commonly reported etiologies. Sunshine coast virus (family Sunviridae; genus ­Sunshinevirus) has been associated with neurorespiratory disease in a number of Australian python species. The virus was originally believed to be a paramyxovirus, but was recently reclassified to its own family in the order Mononegavirales (Afonso et al., 2016). Gross lesions are generally unremarkable. Histologically, central nervous system changes are characterized by spongiosis and gliosis of the hindbrain white matter (Fig. 37.17); extension into the gray matter and neuronal necrosis can be seen in severe cases. There can also be mild bronchointerstitial pneumonia (Hyndman et al., 2012). Picornaviruses, have been reported in cases of gastrointestinal disease in snakes and retrovirus particles have been identified in neoplasms from snakes (Jacobson, 2007). As diagnosticians and scientists, pathologists should be particularly cautious and conservative when interpreting findings and associations between lesions and proposed pathogens. This is particularly true for advanced molecular techniques. It is not difficult to find viruses when you go looking for them. The challenge is to ensure the link between the virus and the lesion is strong. Virus isolation, immunohistochemistry, in situ hybridization, and/or electron microscopy should all be utilized when available to implicate a novel pathogen in a disease process.

Many Gram-negative organisms, including Pseudomonas, Aeromonas, Serratia, Morganella, and Providencia spp., while considered to be part of the normal oral flora of healthy snakes, can also be opportunistic pathogens (Rosenthal and Mader, 2006). Bacterial stomatitis is a common disease in captive snakes that is often associated other problems (e.g., poor husbandry, concurrent disease, dysecdysis). Ulcerative and heterophilic inflammation of the oral mucosa, particularly at the junction between the labial scales, is a common finding early in disease; inflammation, exudatation, and necrotic debris accumulation can progress to involve large areas of the oral and pharyngeal mucosa, surround the teeth, and extend into the choana. Heterophilic exudate can accumulate within the lingual sheath, and with chronicity can manifest as severe necrotizing glossitis. Chronic oral inflammation may extend to the bones of the dentary and skull causing extensive osteomyelitis and bone loss. Septicemia and bronchopneumonia are commonly encountered sequelae. Salmonella is a component of the normal gastrointestinal flora of reptiles and a common pathogen in snakes. In a retrospective study of reptile submissions from a zoological collection, Salmonella was cultured from one-third of all submitted snake fecal samples and approximately one-third of all Salmonella positive samples were from clinically ill snakes (Clancy et al., 2016). Salmonella enterica subspecies enterica (I), arizonae (IIIa), and diarizonae (IIIb) are all commonly reported in snakes (Clancy et al., 2016; Jacobson, 2007a). Septicemia, heterophilic dermatitis, necrotizing inflammation of the gastrointestinal tract (stomach, small intestine, large intestine, and stomach), and osteomyelitis (as previously discussed in Chronic Vertebral Osteopathy; Fig. 37.5) can all be observed in cases of ophidian salmonellosis. Mycobacterial infections are encountered in captive snakes, albeit not as commonly as in some other reptiles and amphibians. Lesions may be localized (e.g., oral, pulmonary, oviductal, or dermal) or disseminated. Pulmonary mycobacteriosis is particularly common (Figs. 37.18, e7), and boids seem to be overrepresented. Typical reptilian ­granulomatous inflammation, with central cores of necrotic debris surrounded by epithelioid macrophages, multinucleated giant cells, and variable numbers of heterophils with an outer rim of lymphocytes and plasma cells, is characteristic for the disease (Fig. 37.18A). Bacteria are often readily identified with acid-fast histochemical stains (Fig. 37.18B). In reptiles and amphibians, Fite-Faraco staining may be more sensitive at detecting mycobacterial organisms than Ziehl-Neelsen staining. Mycobacterium marinum, M. haemophilum, M. chelonae, M. kansasii, and M. leprae are all reported in snakes (Hernandez-Divers and Shearer, 2002; Mitchell 2012; Reavill and Schmidt, 2012). The obligate intracellular bacterium Chlamydophila pneumonia is a cause of granulomatous disease in

912 Pathology of Wildlife and Zoo Animals

FIGURE 37.18  Pulmonary mycobacteriosis in an emerald tree boa. (A) Granulomas with prominent multinucleated giant cells often surround cores of hypereosinophilic debris along with edema and lymphoplasmacytic inflammation expand faveolar septa. (B) Abundant acid fast positive bacilli are present within granulomatous foci. Fite-Faraco.

captive puff adders and emerald tree boas (Jacobson et al., 1989, 2002). In both species, there is variable and widespread, visceral granulomatous inflammation with intragranulomatous, small, basophilic, punctate inclusions. Ultrastructural, immunohistochemical, and molecular testing support the diagnosis (Bodetti et al., 2002; Jacobson et al., 2004). While reports of snake chlamydophilosis are limited, it should be included as a differential etiology in cases of granulomatous inflammation with no clear evidence of a mycobacterial pathogen. There is limited literature on the significance of mycoplasma infections in snakes. A Mycoplasma sp. isolated from a Burmese python with proliferative tracheitis and pneumonia was associated with the surface of the tracheal and pulmonary airways by ultrastructural examination (Penner et al., 1997). Concurrent infections of snakes with nidoviruses and Mycoplasma spp. have also been seen by the author. Dermatophilus-like bacteria have been associated with subcutaneous masses in a boa constrictor and a king cobra (Jacobson 2007a; Wellehan et al., 2004). Numerous mixed aerobic and anaerobic bacteria have been isolated from foci of dermal inflammation in snakes, including, but not limited to Morganella, Fusobacterium, and Serratia (Jacobson, 2007a).

Fungi Snake fungal disease (SFD) (Fig. 37.19A–D) is an emerging disease of wild and captive snakes caused by the ascomycete Ophidiomyces ophiodiicola. First reported in free-ranging North American snakes in 2008, SFD has since

been documented in wild crotalids (timber, eastern Massasauga, and pygmy rattlesnakes) and sympatric colubrids (including but not limited to garter, water, and milk snakes) throughout the eastern and midwestern United States of America (Allender et al., 2011; Paré 2014). Ophidiomycosis has also been documented globally in captive snakes (Sigler et al., 2013). The most consistent and striking feature of SFD cases is a fungal dermatitis. In mild ­infections or early in the disease course, typical lesions consist of superficial foci of dermatitis and scale loss (Fig. 37.19A). In some species, particularly crotalids, individuals with severe and chronic infections can have substantial facial disfiguration (Fig. 37.19B). Histologically, the lesions are characterized by necroulcerative, heterophilic, and granulomatous dermatitis (Fig. 37.19C). Overlying serocellular crusts are composed of abundant necrotic debris and proteinaceous material that subtend the stratum corneum and frequently contain myriad profiles of up to 5 µm diameter, parallel walled, branching, septate hyphae. Along the surface of the crusts, cylindrical fission arthroconidia may be present (Fig. 37.19D). The arthroconidia are a diagnostic feature of Ophidiomyces and other closely related fungi previously grouped into Chrysosporium anamorph of Nannizziopsis vriesii (CANV). Histochemical stains [Grocott-Gomori ­Methenamine Silver (GMS); Periodic acid-Schiff (PAS)] may assist with arthroconidial identification. In some cases, the superficial dermatitis can resolve with successive rounds of ecdysis. In severe cases that result in facial distortion, deep, chronic granulomatous, and heterophilic inflammation can be associated with severe osteolysis and osteomyelitis (Fig. 37.19C). Disseminated infections

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FIGURE 37.19  Snake fungal disease (Ophidiomyces ophiodiicola infection). (A) Early or mild O. ophiodiicola infection often presents as one or multiple area/s of mild, superficial dermatitis with localized scale loss (B) Severe, chronic infections can cause multinodular inflammation and severe facial and oral disfiguration. (C) Infection is associated with foci of granulomatous and heterophilic inflammation that in this case abut and replace bony trabeculae of the face. (D) Periodic acid-Schiff staining of the superficial dermis highlights the presence of cylindrical arthroconidia at the air-lesion interface and invasive, septate, branching hyphae that are invasive throughout the inflammatory foci. Small numbers of cocci are also present along the surface of the lesion. (Part A: timber rattlesnake; Photo Courtesy of R. Ossiboff, Cornell University Wildlife Health Program; Part B: eastern massasauga rattlesnake; Photo Courtesy of W. Edwards, University of Illinois Zoological Pathology Program; Parts C and D: timber rattlesnake; Photo Courtesy of R. Ossiboff, Cornell University Wildlife Health Program).

c­haracterized by widespread visceral granulomas with intralesional fungal elements can also be seen. While the identification of arthroconidia is consistent with SFD, additional ancillary testing is required for a definitive diagnosis of ophidiomycosis. As fungal culture alone may result in the isolation of saprophytic organisms or concurrent, opportunistic copathogens, both fungal culture and quantitative PCR specific for O. ophiodiicola are recommended. For field surveillance, the examination of shed skins can be quite useful in identifying the presence of SFD. Snake sheds can be examined for the presence of focal or localized thickenings of the corneum or crusts, particularly along the ventral scales. Histology, fungal culture, and

nucleic acid extraction for molecular testing can all successfully and reliably be performed on shed skins. Fungal dermatitides histologically similar to SFD but caused by other fungi previously classified as CANV can also be seen, particularly in aquatic snakes. In captive tentacled snakes, fungal dermatitis is a common finding (often due to suboptimal water quality and parameters) and two species of Paranannizopsis (P. californiensis and P. ­crustacea) have been identified (Sigler et al., 2013). Likewise, P. australasiensis has been reported as a cause of necrotizing dermatitis in aquatic file snakes (Sigler et al., 2013). In general, dermatomycoses are common in snakes and opportunistic hyalohyphomycotic fungi, such as

914 Pathology of Wildlife and Zoo Animals

­ aecilomyces, Fusarium, Aspergillus, and Penicillium P are often causative. For this reason, accurate description of fungal morphology and additional ancillary diagnostics are very important for proper diagnosis. Reports of infections by yeast and dimorphic fungi in snakes are rare. Granulomatous cryptococcal pneumonia and meningoencephalitis have been documented in a green anaconda (McNamara et al., 1994). Coccidioides immitisassociated pneumonia with mixed inflammation surrounding round, thick-walled spherules was reported in a Sonoran gopher snake (Timm et al., 1988).

Metazoa Metazoan parasitism in snakes is primarily an issue in free ranging or wild-caught animals in captive collections, though there are some exceptions. Rhabditoid nematodes are particularly significant clinically in snakes. Rhabdias and Strongyloides are direct developing and free-living parasites that can also enter a parasitic, parthenogenetic phase. The larvae of both genera initially target pulmonary tissues. Rhabdias spp. remain in the lung for maturation into adults while Strongyloides spp. complete their maturation in the intestines. In snakes with severe infections or extensive larval migration there can be proliferative pneumonia and secondary bacterial infections. Strongyloides infections can result in proliferative enteritis (Jacobson 2007c). Rhabditoid nematodes are a cause of ocular disease in captive ball pythons (Fig. 37.20A–C). Snakes infected with ­Serpentirhabdias dubielzigi can present with opaque spectacles and swelling of the oral, facial, and periocular tissues (Fig. 37.20A). Histologically, the subspectacular space, the potential space between the cornea and the overlying spectacle, is expanded by inflammatory and proteinaceous debris (Fig. 37.20B) and abundant nematode parasites at multiple life stages, including ova, larvae, and adults (Fig. 37.20C). Adult Serpentirhabdias have characteristic double lateral alae, and ova are commonly larvated or ­embryonated. Parasites and mixed granulocytic and granulomatous inflammation and edema can extend into the connective tissues of the head (Hausmann et al., 2015; Lucio-Forster et al., 2015). Ascarids are common in free-ranging snakes, and are often associated with gastric lesions. Adults of Ophidascaris spp. may embed in the caudal esophageal and gastric mucosa and wall, resulting in ulceration, inflammation, and fibrosis. Visceral larval migration can cause multifocal granulocytic and granulomatous inflammation. ­Kalicephalus, the snake hookworm, has a direct life cycle, and as such can be maintained in wild caught snakes in captivity. In severe infections, gastritis, ulcerative enteritis and hemorrhage, or impaction may be observed (Jacobson 2007c; Klaphake et al., 2005).

Dracunculoidea larvae can cause raised, pustular-like lesions in a wide variety of snakes. Macdonaldius, a filarid of ophidians, is most often an incidental finding. Intravascular adults and microfilariae are seen histologically, and can be associated with vascular obstruction, localized necrosis, and inflammation; microfilariae can also be observed in blood smears (Jacobson 2007c). The majority of cestode infections in snakes are subclinical. Bothridium and Bothriocephalus are parasites of boid snakes that are rarely associated with disease, even in heavy infections (Fig. e8). Tetrahydria, larval cyclophyllidean cestodes of the genus Mesocestoides, are commonly found in snakes and develop following the ingestion of a cysticercoid-containing arthropod. Tetrahydria can be found within white nodules in the intestine, liver, or serosal surfaces of the coelomic viscera (Jacobson, 2007c). Renifers (digenetic trematodes of the families ­Ochetosomatidae and P ­ lagiorchiidae) are common inhabitants of the oral cavity, proximal esophagus, and lungs of free-ranging snakes. Adults migrate through the oral cavity into the glottis, t­rachea, and lungs where they attach to the mucosal epithelium and can produce focal lesions and predispose to secondary bacterial infections. Plagiorchiid trematodes of the genus Styphlodora reside in renal collecting ducts and ureters and can cause tubular dilatation and chronic interstitial nephritis (Jacobson, 2007c). Pentastomids are superficially segmented p­ arasitic crustaceans that are commonly found in reptiles (Fig. 37.21). In snakes, Armillifer, Kiricephalus, P ­orocephalus and ­Raillietiella are reported. The ­ parasites are ingested as larva and migrate extensively before attaining maturity in the lung (Armillifer, Porocephalus, and Raillietiella) or subcutis (Kiricephalus) (Jacobson, 2007c). While adults residing within the lungs are not often associated with ­significant lesions other than obstruction, there can be significant inflammation associated with parasite migration and molting.

Protozoa Cryptosporidial infections are an important cause of gastric, and, to a lesser degree, enteric disease in snakes. Cryptosporidium serpentis infections can present as enlarged and palpably firm stomachs that may cause bulging of the body wall. Grossly, the gastric mucosa is thickened with prominent rugae and mucus accumulation (Fig. 37.22A). Histologically, the gastric mucosa is hyperplastic with increased amounts of connective tissue in the lamina propria and submucosa (Fig. 37.22B). Gastric glands are dilated and there can be significant atrophy of granular, serous cells with hyperplasia of mucus cells. Variable numbers of intracellular, extracytoplasmic cryptosporidial oocysts (3–6 µm) are attached to the m ­ icrovillar surface (Fig. 37.22C) and can

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FIGURE 37.21  Armillifer armillatus in the lung of a free-ranging African rock python. Armillifer, a parasitic crustacean, attaches to the luminal surface of the lung by way of compound hooks that surround its mouth. (Photo Courtesy of B. Cossic, Cornell University).

be highlighted with Giemsa histochemical staining (Brownstein et al., 1977). Though more common in lizards, chronic enteritis associated with Cryptosporidium spp. at the apical margin of enterocytes, can also be seen in snakes. Definitive identification and speciation of Cryptosporidium can be difficult. Frozen feces or tissues (stomach and small intestine) are the most likely to be successful for molecular characterization; formalin-fixed paraffin embedded tissues are of limited diagnostic utility. Entamoeba invadens is a common cause of necrotizing hepatitis and enterocolitis in snakes. Infections in captive snakes are often seen in mixed reptile collections that include subclinical reservoir animals/species, such as herbivorous lizards and turtles. While many snake species are susceptible, some snake species, including garter snakes and northern black racers, seem to be more resistant to infection though individuals can still succumb to amoebiasis. Ingested quadrinucleate cysts develop into ­ FIGURE 37.20  Ocular rhabdiasis in a ball python. (A) The spectacle bulges out from the globe due to the accumulation of fluid, exudate and hemorrhage within the subspectacular space. There is also periorbital and subcutaneous swelling of the right lateral side of the head. (B) The ­subspectacular space, the potential space between the cornea and the overlying spectacle, is expanded by inflammatory and proteinaceous debris. (C) Numerous transverse sections of larval and mature nematode parasites (the type host of Serpentirhabdias dubielzigi) expand the subspectacular space. Double lateral alae are a characteristic feature of adults, and the presence of abundant larvated and/or embryonated ova is a common finding. (Part A: Photo Courtesy of C. Mans, University of Wisconsin School of Veterinary Medicine and A. Lucio-Forster and D. Bowman, Cornell University; Part C: Photo Courtesy of University of Wisconsin School of Veterinary Medicine Pathology Service and A. Miller, A. Lucio-Forster and D. Bowman, Cornell University).

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FIGURE 37.23  Hepatic amoebiasis in a garter snake. Within foci of hepatocellular degeneration and necrosis are small numbers of extracellular, eosinophilic, amoebic trophozoites with a round to ovoid ­amphophilic nucleus (Entamoeba invadens, presumptive). Amoeba are accompanied by moderate numbers intact and degenerate heterophils and fewer lymphocytes.

invasive ­trophozoites within the alimentary tract and cause segmental to diffuse mucosal necrosis of primarily the large intestine though the small intestine and stomach can also be affected. Widespread mucosal necrosis permits amoebae access to the portal circulation and the liver. Necrotizing hepatitis due to ascension of both amoebic trophozoites and alimentary bacteria is common (Fig. 37.23) (Jacobson, 2007c). Periodic acid-Schiff staining is useful to highlight trophozoites. Monocercomonas-like trichomonads are commonly seen in the gastrointestinal tract of snakes and are most often an incidental finding, though sporadic individual reports associate the organisms with pathology of a variety of tissues (gall bladder, oviducts, stomach, and intestine). Numerous Eimeria and Isospora have also been identified in snakes. Their presence is most often ­incidental. Hemogregarine apicomplexans are the most common intraerythrocytic parasite of snakes and even severe infections are typically clinically incidental (­Jacobson,  2007c).

Ectoparasites

FIGURE 37.22  Gastric cryptosporidiosis in a corn snake. (A) The gastric mucosa is severely hyperplastic with abundant mucus. (B) Gastric glands are branching and tortuous and there are increased amounts of fibrous connective tissue in the lamina propria with admixed inflammation. (C) Numerous cryptosporidial oocysts stud the surface of the gastric epithelial cells. (Photo Courtesy of W. Yau, University of Georgia Zoo and Exotic Animal Pathology Service).

Mites are a common finding in free-ranging snakes. They are also common in captive snakes with suboptimal husbandry and are frequently associated with dysecdysis. Severe ­Ophionyssus natricis acariasis can result in anemia and debilitation. O. natricis is also suspected as having a role in the transmission of disease in captive snakes, ­including divergent arenaviruses associated with IBD and certain bacterial diseases due to Aeromonas hydrophila. Entonyssus and Hamertonia spp. are snake lung mites that are reported as incidental findings in free ranging and recently wild-caught snakes (Jacobson, 2007c).

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E-SLIDES 37.e1

37.e2 37.e3

37.e4

37.e5 37.e6

37.e7

37.e8

Renal gout, corn snake, kidney. The renal parenchyma is disrupted by crystallized aggregates of uric acid (urate tophi) surrounded by granulomatous inflammation. As uric acid dissolves during routine processing, tophi are most often characterized by empty, radiating, acicular clefts histologically. (see Figs. 37.4 and Fig. 37.e4). eSlide: VM05037 Salmonella vertebral osteomyelitis, corn snake, vertebral column. Heterophilic and granulomatous inflammation are associated with osteonecrosis, bone lysis, and osteomyelitis and disrupt the normal architecture of the vertebral body. New bone proliferation is also present multifocally. (see Fig. 37.5). eSlide: VM05039 Colonic adenocarcinoma, gopher snake, large intestine. There is abrupt neoplastic transformation of the colonic mucosa. Neoplastic cells form papilliferous projections into the lumen of the colon. Islands and tubuloglandular arrangements of neoplastic epithelial cells invade the colon transmurally and are associated with a prominent scirrhous response and heterophilic inflammation. (see Fig. 37.10). eSlide: VM05032 Renal adenocarcinoma with gout, African egg-eating snake, kidney. The renal parenchyma is expanded by a cystic, multilobulated neoplasm that expands and replaces normal tissue. The neoplasm is composed of numerous, variably sized tubular structures lined by well-differentiated epithelial cells surrounded by a scirrhous stroma. Urate tophi (gout) are often present within the neoplastic tubules and adjacent desmoplastic stroma. (see Fig. 37.12). eSlide: VM05116 Inclusion body disease, red-tailed boa, kidney. Prominent, round to ovoid, eosinophilic, intracytoplasmic, viral inclusions are present in epithelial cells of renal tubules and collecting ducts. (see Fig. 37.14). eSlide: VM05034 Nidoviral pneumonia, ball python, lung. Concurrent nidoviral interstitial pneumonia and secondary Gram negative bacterial bronchopneumonia in the lung of a ball python. Faveolar pneumocytes are moderately and multifocally hyperplastic; lymphocytes, plasma cells, and granulocytes are present within the interstitium along with moderate amounts of colorless space (edema). The faveolar lumina contain aggregates of intact and degenerate heterophils, macrophages, erythrocytes, cellular debris, and coccobacilli. (see Fig. 37.15). eSlide: VM05035 Snake fungal disease (Ophidiomyces ophiodiicola), timber rattlesnake, skin. Necrotizing heterophilic and granulomatous dermatitis in a timber rattlesnake with Ophidiomyces ophiodiicola. A prominent serocellular crust composed of abundant necrotic debris, intact and degenerate heterophils, proteinaceous material, keratin, and profiles of parallel-walled septa with transverse walls and acute-angle branching overlies the ulcerated epidermis. Along the surface of the crust, low numbers of cylindrical arthroconidia are present. (see Fig. 37.19). eSlide: VM05036 Gastric cryptosporidiosis, indigo snake, stomach. The gastric mucosa is severely hyperplastic. Gastric glands are branching and tortuous and surrounded by increased amounts of fibrous connective tissue with admixed inflammation. Numerous cryptosporidial oocysts stud the surface of the gastric epithelial cells. (see Fig. 37.22). eSlide: VM05033

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E-ONLY CONTENTS

FIGURE e1  Normal epidermis from a red Thai bamboo ratsnake. The hinge region between adjacent scales is composed primarily of alpha keratin, while beta keratin is thickest on the scales. Snakes lack osteoderms and dermal glands. FIGURE e4  Renal gout in a corn snake. Radiating crystallized aggregates of uric acid in renal cytologic squash preps.

FIGURE e2  Normal vertebrae from a ball python. Adjacent vertebrae are connected via modified ball-and-socket articular joints between the cartilaginous surfaces of the convex condyle and the concave cotyle.

FIGURE e5  Dysecdysis and retained spectacle in a corn snake. Dull, slightly opaque, retained keratin (shed) is present on the surface of the skin. On close inspection, a retained, slightly opaque spectacle is also present.

FIGURE e3  Normal spleen from an Amazon tree boa. Moderate numbers of pigmented macrophages are present both within the perilymphoid fibrous zone and interspersed among lymphocytes, reticular cells, macrophages, and dendritic cells of the lymphoid tissue zone.

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FIGURE e8  Cestodiasis in a green tree python. Moderate numbers of cestodes (Bothridium sp.) are adhered to the mucosa of the small intestine (Photo courtesy of R. Ossiboff, Wildlife Conservation Society)

FIGURE e6  Cardiac sarcoma in an ashy pit viper. A tan, multinodular mass composed of neoplastic mesenchymal cells expands and infiltrates the cardiac atria. (Photo Courtesy of D. McAloose, Wildlife Conservation Society)

FIGURE e7  Pulmonary mycobacteriosis in a pacific gopher snake. Multifocal, well demarcated foci of granulomatous pneumonia due to Mycobacterium chelonae expand the pulmonary parenchyma and are visible through the pleura. (Photo Courtesy of William R Pritchard Veterinary Medical Teaching Hospital)

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