Spina bifida, diplomyelia and multiple malformations in a Texel lamb

This report described the clinical and pathological aspects of open spina bifida and diplomyelia along with multiple congenital malformations in a Texel lamb. Clinically, paresis of the thoracic limbs, paralysis of the pelvic limbs and a cutaneous opening in the lumbosacral region were observed. At necropsy, there was a focally extensive disruption of the skin associated with an absence of the dorsal portions of the lumbosacral vertebrae. Additionally, diplomyelia of the lumbar segment, mild hydromyelia of thoracic segment, and moderate communicating hydrocephalus of the lateral and third ventricles were noted. Possible viral etiologies (bovine viral diarrhea virus, bluetongue virus, and Schmallemberg virus) were not detected by RT-PCR, and toxic plants were not identified. Therefore, a possible genetic cause may

Spina bifida is a congenital defect characterized by the absence of a dorsal portion of the vertebrae as a consequence of closure failure of the neural tube, which may result in meningocele or meningomyelocele, and it is usually accompanied by amyelia, hydromyelia, and dysraphism (POTTER et al., 2010;CANTILE & YOUSSEF, 2016). Such defects have no sex predisposal and are commonly detected in the lumbosacral region (POTTER et al., 2010;STEELE et al., 2019). Malformations of this nature are mainly related to hereditary, nutritional, viral, and/or environmental factors (HUT et al., 2017) (REZAZADEH & DEZFOULI, 2008). Open spina bifida is characterized by protrusion of the meninges and spinal cord exposure due to lack of skin coverage, while in cystic spina bifida the protrusion is into the subarachnoid space. Both may cause displacement of nerve roots, leading to neurological signs (CANTILE & YOUSSEF, 2016) which vary according to the affected region (i.e., difficulty standing, fecal and urinary incontinence, paraparesis of pelvic limbs) with no signs of brain involvement (GÓRRIZ-Silveira et al. MARTÍN et al., 2019;STEELE et al., 2019). The present study aimed to describe the clinical and pathological aspects of open spina bifida and diplomyelia along with other central nervous system malformations in a Texel lamb, as well as to discuss the pathogenesis of these conditions in this lamb. To the authors` knowledge, this is the first case report of open spina bifida in a lamb in Rio Grande do Sul State, Brazil.
A seven-day-old, male Texel lamb had a history of paresis of the thoracic limbs, paralysis of the pelvic limbs and dysuria, with urine release only through bladder compression since birth. The animal was alert, with suckling reflexes, regular body condition, and with moderate atrophy of the pelvic limb skeletal muscles. In the lumbar region, a dorsal skin defect measuring 7×2 cm was observed ( Figure 1A). Due to the poor prognosis, the lamb was euthanized. The lamb was born in a property located in Encruzilhada do Sul, Rio Grande do Sul State, with an area of 174.81 hectares and a herd composed of 320 adult female sheep for the purpose of producing lambs for meat production. These females were raised extensively on ryegrass pastures (Lolium multiflorum) and improved native pasture. In the lambing period of the year of birth of the lamb here reported (2018), the property had an abortion rate of 5.1%, but no abnormal alterations were observed in the fetuses. Moreover, there was no history of detection of congenital malformations in the property, and toxic plants able to cause such lesions, as Conium maculatum or Mimosa tenuiflora, were not observed in the property. At necropsy, there was a focally extensive disruption of the skin in the lumbosacral region associated with absence of the dorsal portions of the 1 st to 6 th lumbar vertebrae and sacral vertebrae, with exposure of the vertebral canal and spinal cord (open spina bifida). The spinal cord of this segment was grayish and soft near to the lumbar failure, and within this area the spinal cord had two lateral portions outlined by different meninges (diplomyelia; Figure  1B). Moreover, there was mild cerebellar coning and the brain was moderately swollen with flattening of the sulci and narrowing of the gyri, which were caused by moderate communicating hydrocephalus involving the mesencephalic aqueduct, third and lateral ventricles ( Figure 1C). The urinary bladder, both ureters, and renal pelvis were markedly distended by urine (hydroureter and hydronephrosis). Samples representative of all organs were collected, routinely processed for histology and stained with hematoxylin and eosin (HE). The vertebral segment of the lumbosacral region with bone malformation was macerated, boiled in water and cleared by immersion in 10% hydrogen peroxide. With this analysis, a complete absence of the spinous and dorsal processes of the lumbosacral vertebrae was observed ( Figure 1D).
Microscopically, the malformed segment of the vertebrae contained a well-differentiated spinal cord divided into two lateral portions which were outlined by independent leptomeninges (diplomyelia). This tissue was mostly contained within the subcutaneous tissue, but often extended into a diffusely ulcerated epidermis ( Figure 1E). In addition, severe hyperemia, marked perivascular multifocal neutrophilic infiltrate, moderate white matter vacuolation intermixed by axonal spheroids, as well as countless necrotic neurons within the grey matter were noted in the spinal cord ( Figure 1F). The ependymal canal of these segments was moderately dilated and filled with fibrin. The ulcerated epidermis was covered by large amounts of fibrin and degenerated neutrophils, as well as it presented multifocal areas of neovascularization at the vicinities of the nervous tissue. The thoracic spinal cord exhibited moderate hydromyelia and a focal area of vacuolation of the grey matter, with rare axonal spheroids. The other organs were unremarkable.
The diagnosis in the present case of spina bifida and diplomyelia associated with multiple malformations in a Texel lamb was based on the clinical and pathological findings. Clinically, it presented neurological signs of compression and displacement of lumbosacral nerve roots responsible for the sensory and motor innervation of the pelvic limbs, bladder, and anus (DE LAHUNTA & GLASS, 2009;CANTILE & YOUSSEF, 2016). In addition, microscopic lesions of severe infiltrate, necrosis and vacuolation were compatible with the clinical signs. Clinical signs of dysuria and urinary incontinence can be explained by a lesion in the medullary region responsible for emptying the bladder, affecting the pudendal nerve and the innervation of the detrusor muscle and urethral muscles, inhibiting their contraction (DE LAHUNTA & GLASS, 2009). At necropsy, hydronephrosis, hydroureter, and marked bladder distention were observed, possibly due to a prolonged inability to urinate.
Hydromyelia in the thoracic and lumbar segments of the spinal cord is an unusual change in domestic animals, but often associated with cases of spina bifida (POTTER et al., 2010;HUT et al., 2017;GÓRRIZ-MARTÍN et al., 2019). The compression of the lumbar region of the spinal cord most likely prevented the cerebrospinal fluid (CSF) flow into the ependymal canal, generating hydromyelia at the proximal segments of the spinal cord. Additionally, a reduced resorption of the CSF at the subarachnoid spaces, which characterizes a communicating hydrocephalus, probably increased the pressure within the ventricular system, resulting in dilation of the mesencephalic aqueduct, third and lateral ventricles, as well as promoting cerebellar coning (DE LAHUNTA & GLASS, 2009). However, there were no clinical signs related to these changes.

Silveira et al.
At necropsy, there was complete absence of the spinous processes, dorsal part of the vertebral arch, caudal articular processes and articular nipple process, similarly to that described in other studies (POTTER et al., 2010;GUTZWILLER et al., 2014;STEELE et al., 2019). The cause of these malformations, as in the present case, is unknown, with sporadic case reports (GUTZWILLER et al., 2014). Nonetheless, a genetic component has been indicated in investigations with calves and lambs, suggesting a possible inherited recessive anomaly, with an influence of a dominant gene with low penetrance and variable expressiveness (GÓRRIZ-MARTÍN et al., 2019;STEELE et al., 2019).
Possible etiologies considered in this case were Pestivirus (BVDV), Orbivirus (BTV), and SV. The first is associated with birth defects, abortions, and stillbirths predominantly in cattle (CANTILE & YOUSSEF, 2016), while infection in sheep is associated with stillbirths and neonatal death (SCHLAFER & FOSTER, 2016), which are distinct from the present case. SV infection may cause vertebral malformations in sheep, but these are predominantly characterized by changes in position, torticollis, scoliosis, kyphosis, and/or lordosis (PEPPERKAMP et al., 2015;SCHLAFER & FOSTER, 2016). Moreover, changes in the central nervous system related to SV and BTV infections result mostly from tissue loss with microencephaly, hydranencephaly, and anencephaly, as well as dysplasia within the cerebellum, medulla oblongata, and spinal cord (PEPPERKAMP et al., 2015), differing from the present report, in which duplication of the spinal cord was evidenced (diplomyelia). Poisoning by plants with toxic teratogenic principles that may cause congenital anomalies in the nervous and musculoskeletal systems (RIET-CORREA et al., 2012) were other possible causes considered in the present case. Mimosa tenuiflora and M. ophthalmocentra, widely found in Brazil, are examples of these causes (DANTAS et al., 2010;RIET-CORREA et al., 2012), but none of these were observed on the property. This research highlighted the clinical presentation and the associated pathological findings of spina bifida in a lamb. Possible viral or toxic etiologies have not been identified and, therefore, a possible genetic cause could be considered.