First report of the novel atypical porcine pestivirus in piglets with congenital
tremor in Canada
Authors : Fanny G1. Dessureault1’*, Martin Choinière2, Carl A. Gagnon3,4,
1Complexe de diagnostic et d’épidémiosurveillance vétérinaires du Québec (CDEVQ); 3Swine
and poultry infectious diseases research center (CRIPA) and 4Service de diagnostic (SD), Faculté
de médecine vétérinaire (FMV), Université de Montréal (UdeM); 3200 rue Sicotte, Saint-Hyacinthe, Québec, Canada, J2S 2M2. 2Bureau vétérinaire Martin Choinière DMV Inc., 2694
chemin Bellerive, Carignan, Québec, Canada, J3L 4J9.
*Address all correspondence to Dr. Fanny Dessureault; e-mail: Fanny.dessureault@mapaq.gouv.qc.ca
Source of support: This work was funded by Ministère de l’Agriculture, des Pêcheries et de l’Alimentation du Québec (MAPAQ) and FMVof UdeM. CAG was financially supported by a Natural Sciences and Engineering Research Council of Canada (NSERC) discovery grants.
Abstract – Two 2-year-old nursing piglets with congenital tremor were submitted for
postmortem examination. Histopathology revealed severe hypomyelination in the spinal cord and quantitative real-time PCR detected the presence of atypical porcine pestivirus (APPV) in several tissues. This is the first reported case of APPV infection in Canada.
Résumé – Premier cas rapporté du nouveau pestivirus atypique porcin au Canada chez des porcelets avec tremblements congénitaux. Deux porcelets à la mamelle âgés de deux jours et présentant des tremblements congénitaux ont été soumis pour examen post-mortem.
L’histopathologie a révélé une hypomyélinisation sévère dans la moelle épinière et des analyses PCR en temps réel ont détecté la présence du pestivirus atypique porcin (APPV) dans plusieurs tissus. Il s’agit apparemment du premier cas signalé d’infection par le virus APPV au Canada.
(Introduction)
Syndrome of congenital tremor (CT), also referred to as dancing pigs (1), shivers (2) or myoclonia congenita (3, 4), is a well-recognized, usually sporadic but common phenomenon described in neonatal piglets (5, 6). The condition, first described in the United States by Kinsley (1922), is characterized by local or systemic muscle spasms appearing within hours of birth, ranging from mild tremors of the ears, flank and hind leg, to severe systemic tremors resulting in difficulties to stand and walk (5). The rhythmic tremors worsen with excitement but cease with sleep (6) and increased morbidity and mortality may result from difficulties or an inability to nurse.
CT is a non-specific clinical sign that has been classified according to the presence (Type A) or absence (Type B) of morphological lesions, mainly hypomyelination, in the central nervous system (7). Type A is subdivided into five subtypes; AI-V, with distinctive causes (6). Type AI is caused by transplacental infection with classical swine fever virus (CSFV), a pestivirus (7), while types AIII and AIV are inherited, and type AV is of toxic origin (6, 7, 8, 9). For a long time, CT type AII was thought to be of infectious origin (5, 7, 10), but it was not until recently that a link was established between CT type AII and a newly identified pestivirus tentatively named atypical porcine pestivirus (APPV) (11, 12, 13, 14).
There is no apparent breed predilection with CT type AII, but the sporadic disease is more frequent in litters of gilts (5). Also, the high prevalence of the disease, which is associated with increased mortality and morbidity in piglets, make it of great clinical interest. Moreover, the
clinical resemblance between CT type AI (caused by CSFV) and AII (caused by APPV), make it critical to be able to differentiate the two conditions.
APPV has been identified in several countries around the world (11, 12, 13, 14, 15, 16, 17, 18), but the infection has apparently never been identified in Canada. Here we present the first report of APPV infection in piglets with CT type AII in Canada.
Case description
Two 2-day old nursing piglets F1 Yorkshire-Landrace from a farrow-to-finish multiplier farm with a high health status were submitted for postmortem examination with a clinical history of congenital tremor (CT). The farm, located in western Canada, housed 400 Landrace breeding sows in sanitized facilities equipped with air filtration system. A complete vaccination program was in place and the sows were in very good health, with a negative status reported in the herd for several infectious agents including porcine reproductive and respiratory syndrome virus (PRRSV), influenza virus, Mycoplasma hyopneumoniae, Actinobacillus pleuropneumoniae and transmissible gastroenteritis coronavirus.
A sudden outbreak of CT was observed at birth in piglets of gilts, while piglets of sows showed no clinical signs. Prior to this outbreak, CT had never been reported in the herd, which produced 24.56 weaned piglets per sow over the 12 previous months. Clinician also reported normal morbidity and mortality rates in piglets and sows before the outbreak.
About six months earlier, five new gilts were introduced into the herd. No clinical sign was reported in those gilts and other sows prior or after farrowing. However, 100% (5/5) of the new gilt litters were affected by CT, with 100% of piglets within a same litter displaying systemic tremors and difficulties standing or walking since birth (total of 61 piglets affected). In the same period of time, a total of 19 gilts had farrowed but CT was observed only in piglets from the newly introduced gilts (5/19). Clinical signs were severe enough to impair nursing and cause mortality. At 3 weeks of age (time of weaning) mortality in affected litter had reached between 13.3% (2/15) and 41.2% (7/17) piglets per litter (total of 24.6% piglets with CT; 15/61), in comparison to an average mortality rate of 12.7% piglets per litter in other gilt litters. It was also
noted that the average litter size was slightly smaller in CT affected litters (12.2 piglets per litter) in comparison to other gilt litters (13.3 piglets per litter). No stillborn was present in the affected litters. In surviving piglets, clinical signs of CT had completely vanished by three weeks of age.
At postmortem examination, a few gross changes were noted and body condition was within normal limits. The stomach of the first subject, a 1.5 kg female, was full of partly digested milk, while the stomach of the second subject, a 1.2 kg male, was empty. Meconium was still present in the colon of this subject. No obvious macroscopic lesions were noted in the central nervous system (CNS).
Most of the brain, the entire spinal cord and sections of lungs, heart, liver, spleen, kidneys, skeletal muscles, small intestine, large intestine, tonsils and thymus were fixed in 10% buffered formalin for 24 hours, processed routinely for histological examination and stained with hematoxylin phloxine saffron (HPS) stain. The same process was applied on the spinal cord of a control subject, a healthy 2-day old piglet from another farm free of CT. Luxol fast blue (LFB) stain was used on sections of thoracic spinal cord of the control and affected piglets. Fresh specimens of lungs, ileum and spiral colon from piglets with CT were submitted for aerobic culture and various pool of tissues from the same subjects were frozen at -20oC and later tested
by a reverse transcription quantitative real-time polymerase chain reaction (RT-qPCR) for PRRSV (single pool of lungs, tonsils, thymus and inguinal lymph nodes from both subjects), porcine circovirus type 2 (PCV2; single pool of lungs, tonsils, thymus and inguinal lymph nodes from both subjects) and APPV (pool A with lungs, tonsils, thymus and inguinal lymph node from both subjects; pool B, one per subject, with thymus, spleen, kidney, ileum, cortex, cerebellum and brainstem). Sections of formalin fixed and paraffin embedded thoracic spinal cord from the control piglet and the piglets with CT were also tested for APPV by qPCR. The APPV RT-qPCR assay used was adapted from the report of Arruda et al (2016). Briefly, this RT-RT-qPCR
assay targets the NS3 gene of the virus and produce an amplicon of 123 nucleotides (nt) in length.
Microscopically, significant findings were noted only in affected animals. In sections of cervical, thoracic and lumbar spinal cord, HPS stain revealed a few vacuoles multifocally in lateral and ventral funiculi, and oligodendrocytes appeared more prominent in comparison to the control piglet. LFS stain revealed evident hypomyelination in the white matter of the spinal cord of piglets with CT compared to the healthy control. In both piglets, an overall decrease in myelin was noted in the white matter of spinal cord, but the changes were most severe in bilateral and ventral funiculi (Figure 1). In comparison, myelinisation in the adjacent nerve roots (internal control) and in the spinal cord of the healthy subject (external control) appeared normal. No changes were noted in the cerebral cortex, cerebellum and brainstem of affected piglets. No bacterial pathogens were isolated from culture.
Molecular diagnostic results for PRRSV and PCV2 were negative. All frozen samples tested for APPV by RT-qPCR were positive, with Ct values of 26.09 for pool A, and 31.34 (subject 1) and 28.39 (subject 2) for pool B. Sections of formalin fixed and paraffin embedded thoracic spinal cord from control piglet were negative for APPV by RT-qPCR, but sections from diseases piglets were positive, with Ct values of 33.68 (subject 1) and 34.90 (subject 2). To confirm the APPV RT-qPCR results, PCR amplicons were sequenced. Interestingly, PCR
amplicons were entirely related to a United-States (US) APPV reference strain (isolate 20130103; GenBank accession number KU194230) with X % identity.
Discussion
The results are consistent with an outbreak of CT type AII caused by APPV infection. APPV is a recently-discovered porcine pestivirus, first described by Hause & al. (2015) following high throughput sequencing. Pestiviruses are enveloped, highly variable RNA viruses belonging to the family Flaviviridae (19). Several pestiviruses are of great socioeconomic relevance, notably CSFV in pigs, bovine viral diarrhea virus type 1 (BVDV-1) and type 2 (BVDV-2) in cattle, and border disease virus (BDV) in sheep and goats (20). Fetopathogenicity is a common feature of intrauterine pestivirus infections and in pigs, CSFV highly virulent strains cause severe clinical signs and high mortality resulting in significant decrease in productivity (19, 20). For a long time, it was believed that pestiviruses were affecting only ungulate species, but recently several novel pestiviruses have been identified in various domestic and wild species including in non-ungulate species such as the rat (21) and bat (22).
APPV is one of the novel emerging pestiviruses identified in pigs along with
Bungowannah virus (23) and Lindavirus (24). Bungowannah virus has been associated with stillbirths or myocardiopathy and sudden death in piglets in Australia (25), while Lindavirus has been identified in Austria from piglets with CT and severe hypomyelination (24). However, despite the fact that clinical signs and lesions observed in piglets infected with Lindavirus are
quite similar to changes associated with APPV infection, phylogenic analysis revealed that Lindavirus has closer identity with Bungowannah than APPV.
Phylogenic analysis have revealed only distant genetic relation between APPV and ruminant pestiviruses and CSFV, with less than 50% nt identity with CSFV (11). A distant genetic relation was also noted between APPV and Bungowannah virus and the rat pestivirus, while the bat pestivirus appears to be more closely related (Hause, 2015).
Like other pestiviruses, APPV display a large genetic variation, with a large number of strains (13, 14, 15, 16, 17, 18, 26). The reports of such various strains in commercial pig populations in several countries including Austria, China, Germany, Great Britain, Italy,
Netherlands, Serbia, Spain, Switzerland and the US, indicate it is also widely distributed (12, 13, 14, 15, 16, 17, 18, 26). The prevalence reported in herds is variable but can be high (13, 15, 18). A strong association between CT type AII and APPV have also been observed by several authors (12, 13, 14, 16, 17) and reproduction of CT in piglets has been achieved experimentally following inoculation of APPV to pregnant gilts at day 32 of gestation (14), and following inoculation of APPV to pregnant gilts and fetal amniotic vesicle at day 45 or 62 of gestation (12). In both studies, the inoculation times were established with the knowledge of foetal development of immunocompetence (around 70 days of gestation; 5, 27, 28, 29) and CNS (at 62 days of gestation; 30, 31) in pigs.
Congenital infection with APPV is typically associated with outbreaks of CT in litters of gilts (13, 14, 18) although piglets born from higher parity sows can also be affected. While the infection does not affect the number of piglets born per litter, the prevalence of CT within a litter, which is highly variable, can reach 100% (14). In the present case, the cause for a slightly lower number of piglets per litter within CT-affected litters in comparison to other litters remains
unknown. Morbidity is typically increased with CT, resulting frequently in growth retardation, but mortality can be normal or increased (13, 16, 17).
Intensity of CT can be quite variable (12, 14, 17) and mild tremor could be unnoticed or mistaken for shivering from cold. While piglets will recover completely typically within 2-3 weeks (13), some authors (16) report the persistence of mild tremor of ear tips and flank for up to 14 weeks before complete disappearance. Like with other types of CT and as mentioned earlier, no shaking is noted in piglets during relaxation or sleep, but tremors are increased or induced by stress.
In the present case, CT was the only clinical sign reported. However, other changes such as splay leg (12, 14, 16), which reflect muscle weakness, or an abnormal posture with xyphosis and ears on the neck (14) have also been associated with APPV infection.
The histological changes observed in this case report are consistent with lesions reported by previous authors (13, 16). Congenital tremor associated with APPV infection is typically characterized histologically by hypomyelination in the lateral white matter of the spinal cord only (13) or in the white matter of the cerebellum and spinal cord (16). By electron microscopy, hypomyelination but also myelin disruption and myelin breakdown have been described, while no detectable loss of oligodendrocytes have been noted by immunoperoxidase (16). Such findings suggest that APPV has deleterious effects on foetal oligodendrocytes, likely impacting on myelin development and function, or leading to degeneration of present myelin.
By fluorescent in situ hybridization, the presence of the virus has been noted in the central nervous system (cerebellum inner granular layer, spinal ganglia neurons) and lymphoid tissues (glandular epithelium of tonsils and follicular centers of mandibular lymph node, but absent in the thymus) of piglets with CT type AII (13). The pathogenesis is still unclear and the presence
of APPV in central nervous system may explain neurological symptoms, but its presence in a wide variety of other tissues suggests it replicates systemically.
Apart from the central nervous system (brain, cerebellum, brainstem, spinal cord and trigeminal ganglia) and the lymphoid organs (thymus, tonsils, spleen, and mandibular, tracheal, inguinal or mesenteric lymph nodes), the virus has also been identified by PCR from peripheral nerves, heart, lungs, liver, kidney, bladder, pancreas, small intestine, colon, salivary glands, skeletal muscle, umbilical blood, serum, cerebrospinal fluid, saliva, nasal swabs, rectal swabs and even semen (12, 13, 14, 16, 18). The frequency of identification and the viral genome load vary between samples and between studies, however lymphoid organs seem to be the most frequently positive samples and usually with a higher viral genome load (13, 14, 17, 18).
No relationship has been established between the virus concentration in blood and
severity of the disease (14), but the presence of APPV in samples such as rectal and nasal swabs, salivary glands, saliva, intestines and semen represent potential sources for viral shedding into the environment.
One important observation in the literature is the high prevalence of APPV infection in herds or individuals with no obvious association with clinical disease (13, 14, 15, 26) and studies suggest that CT type AII only occurs when naïve sows are infected in a certain gestation period, prior to the development of immunocompetence of the foetuses (12, 14, 16), while the infection may not induce clinical signs in an immune herd. Within clinical outbreaks of CT, asymptomatic viremic piglets have been reported (17) and persistent shedding of the virus after the resolution of clinical disease has also been frequently observed (14, 16, 17). Moreover, the high prevalence of APPV in tonsils or serum from apparently healthy pigs support the suspicion for the presence of persistently infected subjects and may explain that CT can be recurrent at farm level (14). In
such cases, CT will usually reappears after the introduction of new gilts, allowing clinically healthy but chronically or persistently infected pigs to infect naïve subjects (12, 14, 15, 16, 17).
Detection of the virus, sometimes with prolonged shedding (14, 16), from feces, saliva, intestines, pancreas, salivary glands (14, 16, 18) and even semen of pigs suggest that the transmission of the virus in herd is orofecal, but also possibly venereal. At the same time, the higher viral loads in lymphoid tissues suggest that replication occur in those, and like other pestiviruses, an immunosuppressive capacity is possible and should be investigated. In piglets, transplacental transmission of the virus is the most likely transmission route and sows serum are usually negative for APPV at time of delivery (13, 14).
Cell culture isolation was first achieved by Beer et al. (2016), but several authors have failed to replicate the virus (11, 12, 13, 14, 18) suggesting that in vitro infectivity is limited. Recently, isolation and passage on different porcine cells was achieved in culture (16, 32), but infection of subjects with isolated virus remain to be done.
The present case appears to be the first report of APPV in piglets with congenital tremor in Canada, however the virus was probably already present in herds for several years and just unrecognized. Some of the mains characteristics of the virus appear to be a wide prevalence worldwide, with great variation in strains and the presence of prolonged or persistent carriers and shedders that help to maintain the infection within herds. While presence of the virus correlates with CT in piglets, no clinical disease has been associated with the infection in adults. Thus, many questions remain to be answered, notably regarding the exact prevalence of the virus in Canadian herds, the route of transmission of the infection and its pathogenesis. A better
understanding of the APPV pathogenicity, epidemiology and immunobiology would be important to control the infection and to promote development of vaccines and other prevention strategies.
Finally, the diagnosis of APPV is particularly important since CSFV, a federally
reportable disease in Canada, is a clinical differential diagnosis for CT in piglets. As mentioned earlier, CT type AI and type AII cannot be clinically distinguished and that’s why adequate diagnostic tools are essentials. Fortunately, there is no evidence so far of cross reactivity with diagnostic tests routinely used for the diagnosis of classical swine fever (32) and it appears unlikely that APPV could negatively impact the diagnosis and surveillance programs in place for CSFV.
Acknowledgments
We would also like to thank Dr Lalitha Peddireddi for the confirmation of our atypical porcine pestivirus positive samples by her diagnostic laboratory at Kansas State veterinary Diagnostic Laboratory.
References
1. Kinsley AT. Dancing pigs. Vet Med 1922; 17:123.
2. Nissley SM. Shivers in pigs. J Am Vet Med Assoc 1932; 81: 551.
3. Kernkamp HCH. Myoclonia congenita: A disease of newborn pigs. Vet Med 1950; 45:189. 4. Maplesden DC, Brown GCT. Myoclonia congenital in a litter of young pigs. Can J Vet Res
1957; 21: 170-172.
5. Done S, Williamson SM, Strugnell BW. Nervous and locomotor systems. In: Zimmerman JJ, Karriker LA, Ramirez A, Schwarts KJ, Stevenson GW, ed. Diseases of Swine. 10th ed. Ames,
Iowa: Wiley-Blackwell. 2012: 303-305.
6. Cantile C, Youssef S. Nervous system. In: Maxie MG, ed. Jubb, Kennedy and Palmer’s Pathology of Domestic Animals. 6th ed. Vol 1. London, UK: Saunders Elsevier, 2016:336-338.
7. Done JT. Congenital nervous diseases of pigs: A review. Lab Animal 1968; 2: 207-217. 8. Blakemore WF, Harding JD, Done JT. Ultrastructural observations on the spinal cord of a
9. Blakemore WF, Harding JD. Ultrastructural observations on the spinal cords piglets with congenital tremor type AIV. Res Vet Sci 1974; 17:248-255.
10. Blomström A-L, Ley Cecilia, Jacobson M. Astrovirus as a possible cause of congenital tremor type AII in piglets? Acta Vet Scand 2014; 56:82.
11. Hause BM, Collin EA, Peddireddi L, Yuan F, Chen Z, Hesse RA, Gauger PC, Clement T, Fang Y, Anderson G. Discovery of a novel putative atypical porcine pestivirus in pigs in the USA. J Gen Virol 2015; 96:2994-2998.
12. Arruda BL, Arruda PH, Magstadt DR, Schwartz KJ, Dohlman T, Schleining JA, Patterson AR, Visek CA, Victoria JG. Identification of a divergent lineage porcine pestivirus in nursing piglets with congenital tremors and reproduction of disease following experimental inoculation. PLOS ONE 2016; doi:10.1371/journal.pone.0150104.
13. Postel A, Hansmann F, Baechlein C, Fischer N, Alawi M, Grundhoff A, Derking S,
Tenhündfeld J, Pfankuche VM, Herder V, Baumgärtner W, Wendt M, Becher P. Presence of atypical porcine pestivirus (APPV) genomes in newborn piglets correlated with congenital tremor. Sci Rep 2016; doi 10.1038/srep27735.
14. de Groof A, Deij M, Guelen L, van Grinsven L, van Os-Galdos L, Vogels W, Derks C, Cruijsen T, Geurts V, Vrijenhoek M, Suijskens J, van Doorn P, van Leengoed L, Schrier C, van der Hoek L. Atypical Porcine Pestivirus: A possible cause of congenital tremor type A-II in newborn piglets. Viruses 2016; 8:271; doi:10.3390/v8100271.
15. Beer M, Wernike K, Dräger C, Höper D, Pohlmann A, Bergermann C, Schröder C,
Klinkhammer S, Blome S, Hoffmann B. High prevalence of highly variable atypical porcine pestiviruses found in Germany. Transbound Emerg Dis 2016; 64:e22-e26.
16. Schwarz L, Riedel C, Högler S, Sinn LJ, Voglmayr T, Wöchtl B, Dinhopl N, Rebel-Bauder B, Weissenböck, Ladinig A, Rümenapf T, Lamp B. Congenital infection with atypical
porcine pestivirus (APPV) is associated with disease and viral persistence. Vet Res 2017; 48:1; doi 10.1186/s13567-016-0406-1.
17. Muñoz-González S, Canturri A, Pérez-Simó M, Bohórquez JA, Rosell R, Cabezón O, Segalés J, Domingo M, Ganges L. First report of the novel atypical porcine pestivirus in Spain and a retrospective study. Transbound Emerg Dis 2017; 1-5.
18. Yuan J, Han Z, Li J, Huang Y, Yang J, Ding H, Z J, Zhu M, Zhang Y, Liao J, Zhao M, Chen J. Atypical Porcine Pestivirus as a novel type of pestivirus in pigs in China. Front Microbiol 2017; doi 10.3389/fmicb.2017.00862.
19. Flaviviridae. In: Murphy FA, Gibbs EPJ, Horzinek MC, Studdert MJ, ed. Veterinary Virology. 3rd ed. San Diego, California: Academic Press. 1999: 555-569.
20. Kirkland PD, Le Potier M-F, Vannier P, Finlaison D. Pestiviruses. In: Zimmerman JJ, Karriker LA, Ramirez A, Schwarts KJ, Stevenson GW, ed. Diseases of Swine. 10th ed. Ames,
IA: Wiley-Blackwell. 2012: 538-553.
21. Firth C, Bhat M, Firth MA, Williams SH, Frye MJ, Simmonds P, Conte JM, Ng J, Garcia J, Bhuva NP, Lee B, Che X, Quan P-L, Lipkin WI. Detection of zoonotic pathogens and characterization of novel viruses carried by commensal Rattus norvegicus in New York City. mBio 2014; 5:e01933-14.
22. Wu Z, Ren X, Yang L, Hu Y, Yang J, He G, Zhang J, Dong J, Sun L, Du J, Liu L, Xue Y, Wang J, Yang F, Zhang S, Jin Q. Virome analysis for identification of novel mammalian viruses in bat species from Chinese provinces. J Virol 2012; 86:10999-11012.
23. Kirkland PD, Frost MJ, Finlaison DS, King KR, Ridpath JF, Gu X. Identification of a novel virus in pigs - Bungowannah virus: a possible new species of pestivirus. Virus Res 2007; 129:26-34.
24. Lamp B, Schwarz L, Högler S, Riedel C, Sinn Leonie, Rebel-Bauder B., Weissenböck H, Ladinig A, Rümenapf T. Novel Pestivirus Species in pigs, Austria, 2015. Emerg Infect Dis 2017; 23:1176-1179f.
25. Kirkland PD, Read AJ, Frost J, Finlaison DS. Bungowannah virus – a probable new species of pestivirus – what have we found in the last 10 years?
Anim Health Res Rev 2015; 16:60-63.
26. Postel A, Meyer D, Cagatay G, Feliziani F, De Mia G, Fischer N, et al. High Abundance and Genetic Variability of Atypical Porcine Pestivirus in Pigs from Europe and Asia. Emerg Infect Dis 2017;23(12):2104-2107.
27. Salmon H. Immunité chez le fœtus et le nouveau-né : modèle porcin. Reprod Nutr Dévelop 1984; 24:197-206.
28. Nielsen J, Rønsholt L, Sørensen KJ. Experimental in utero infection of pig foetuses with porcine parvovirus (PPV). Vet Microbiol 1991; 28: 1-11.
29. Sinkora M, Butler JE. The ontogeny of the porcine immune system. Dev Comp Immunol 2009; 33: 273-283.
30. Pond WG, Boleman SL, Fiorotto ML, Ho H, Knabe DA, Mersmann HJ, Savell JW, Su DR. Perinatal ontogeny of brain growth in the domestic pig. Proc Soc Exp Biol Med 2000; 223:102-108.
31. Sweasey D, Patterson DSP, Glancy EM. Biphasic myelination and the fatty acid composition of cerebrosides and cholesterol esters in the developing central nervous system of the
domestic pig. J Neurochem 1976; 27:375-380.
32. Postel A, Meyer D, Petrov A, Becher P. Recent emergence of a novel porcine pestivirus: interference with classical swine fever diagnosis? Emerg Microbes Infect 2017; 6; doi 10.1038/emi.2017.5
Figure legend(s)
Figure 1. Luxol Fast Blue staining of the thoracic spinal cord of two days old piglets. A.
Unaffected piglet. There is regular myelination. B. Congenital tremor affected piglets. There is an important loss of myelin from the periphery of the cord, more severe in the lateral and ventral funiculi. Notice the normal myelination in the nerve roots (internal control). Scale bars = 1mm.
Figures Figure 1.
