We have developed and characterised a mouse model of Japanese encephalitis virus (JEV) infection via
footpad inoculation in order to better mimic viral transmission by mosquito bites. Two-week-old and
5-week-old mice consistently developed signs of infection such as ruffled fur, weight loss, hunchback
posture, tremors, mask-like facies and occasionally, hindlimb paralysis at 4 days post infection (dpi)
and 11-13 dpi, respectively. Most of the animals died within 24 to 48 hours following the onset of signs
of infection, with mortalities of 100% and 33.3% in 2-week-old and 5-week-old mice, respectively.
Mild meningitis and variable parenchymal inflammation with formation of microglial nodules, focal
necrosis and neuronophagia, and perivascular cuffing by inflammatory cells were observed in the
caudate nucleus, putamen, thalamus, cerebral cortex, brainstem, and spinal cord. Viral antigens/RNA
were demonstrated by immunohistochemisty and in situ hybridization, respectively, in most of these
areas as well as in the hippocampus and cerebellum, albeit more focally. The pathological findings in
this mouse model were generally similar to human Japanese encephalitis (JE) and other established JE
models but perhaps, compared to other JEV mouse models, it demonstrates lethal encephalitic infection
more consistently. We believe that our mouse model should be useful to study the pathogenesis of JE,
and for testing anti-viral drugs and vaccines
In the last few decades, there is an increasing emergence and re-emergence of viruses, such as West Nile virus, Enterovirus 71 and henipaviruses that cause epidemic viral encephalitis and other central nervous system (CNS) manifestations. The mortality and morbidity associated with these outbreaks are significant and frequently severe. While aspects of epidemiology, basic virology, etc., may be known, the pathology and pathogenesis are often less so, partly due to a lack of interest among pathologists or because many of these infections are considered "third world" diseases. In the study of epidemic viral encephalitis, the pathologist's role in unravelling the pathology and pathogenesis is critical. The novel henipavirus infection is a good example. The newly created genus Henipavirus within the family Paramyxoviridae consists of two viruses, viz., Hendra virus and Nipah virus. These two viruses emerged in Australia and Asia, respectively, to cause severe encephalitides in humans and animals. Studies show that the pathological features of the acute encephalitis caused by henipaviruses are similar and a unique dual pathogenetic mechanism of vasculitis-induced microinfarction and parenchymal cell infection in the CNS (mainly neurons) and other organs causes severe tissue damage. Both viruses can cause relapsing encephalitis months and years after the acute infection due to a true recurrent infection as evidenced by the presence of virus in infected cells. Future emerging viral encephalitides will no doubt continue to pose considerable challenges to the neuropathologist, and as the West Nile virus outbreak demonstrates, even economically advanced nations are not spared.
Coxsackievirus A16 (CV-A16) is the leading cause of hand-foot-mouth disease (HFMD), which usually
presents as mild and self-limiting symptoms in young children. Rarely, CV-A16 has been reported
to cause severe and fatal neurological complications but little is known about these complications.
In the present study, 1-day and 7-day old mouse models of CV-A16 were developed using a clinical
strain via subcutaneous inoculation. All infected mice exhibited clinical signs of infection, including
reduced mobility, limb weakness and paralysis between 3 to 6 days post-infection. Pathologically,
the main organs involved were the central nervous system (CNS), skeletal muscles and brown fat. In
the CNS, viral antigens as demonstrated by immunohistochemistry, were localized mainly to neurons
in the brain stem and spinal cord, suggesting that CV-A16 is neurotropic although inflammation is
very mild. The skeletal muscles showed necrosis and myositis due to viral infection as evidenced by
the dense viral antigens. Focal viral antigens were also detected in the brown fat. These preliminary
pathological findings indicate that our mouse models can be further developed to be useful models
for pathogenesis studies, and vaccine and anti-viral drug evaluation.
Mitochondrial DNA (mtDNA) deletions are a major cause of chronic progressive external ophthalmoplegia (CPEO) and Kearns-Sayre syndrome (KSS). We analyzed single mtDNA deletions in 11 CPEO and one KSS patients by means of Southern blot and long polymerase chain reaction (PCR) assays. The deletion sizes ranged from 3.4 kb to 6.9 kb whereas the heteroplasmy level varied from 18.8% to 85.5%. Two unique deletions sized 4320 bp and 4717 bp were found. This study represents the first genetic screen of mtDNA disorders in Malaysia, and it follows the data seen in other published reports on CPEO and KSS genetic aetiology.
The complex pathophysiology of traumatic brain injury, its cascading effects and a varied outcome suggest that factors such as genetics may permeate and modulate the neurocognitive outcomes in patients with mild traumatic brain injury (mTBI). This study was conducted to determine the relationship between genetic polymorphism of apolipoprotein E, and neurocognitive and functional outcomes in mTBI. Twenty-one patients with mTBI were recruited prospectively. The severity of the injury was established with the Glasgow Coma Score (GCS). Other assessments included the CT Scan of the head on admission, Disability Rating Scale, Chessington Occupational Therapy Neurological Assessment (COTNAB) and Glasgow Outcome Scale (GOS). The Spearmen correlation analysis of ApoE allele status and the cognitive and functional assessments saw some association with the Sensory Motor Ability - Coordination (-0.526, p
Enterovirus A71 (EV-A71) belongs to the species group A in the Enterovirus genus within the Picornaviridae family. EV-A71 usually causes self-limiting hand, foot and mouth disease or herpangina but rarely causes severe neurological complications such as acute flaccid paralysis and encephalomyelitis. The pathology and neuropathogenesis of these neurological syndromes is beginning to be understood. EV-A71 neurotropism for motor neurons in the spinal cord and brainstem, and other neurons, is mainly responsible for central nervous system damage. This review on the general aspects, recent developments and advances of EV-A71 infection will focus on neuropathogenesis and its implications on other neurotropic enteroviruses, such as poliovirus and the newly emergent Enterovirus D68. With the imminent eradication of poliovirus, EV-A71 is likely to replace it as an important neurotropic enterovirus of worldwide importance.
The genus Henipavirus within the family Paramyxoviridae includes the Hendra virus (HeV) and Nipah virus (NiV) which were discovered in the 1990s in Australia and Malaysia, respectively, after emerging to cause severe and often fatal outbreaks in humans and animals. While HeV is confined to Australia, more recent NiV outbreaks have been reported in Bangladesh, India and the Philippines. The clinical manifestations of both henipaviruses in humans appear similar, with a predominance of an acute encephalitic syndrome. Likewise, the pathological features are similar and characterized by disseminated, multi-organ vasculopathy comprising endothelial infection/ulceration, vasculitis, vasculitis-induced thrombosis/occlusion, parenchymal ischemia/microinfarction, and parenchymal cell infection in the central nervous system (CNS), lung, kidney and other major organs. This unique dual pathogenetic mechanism of vasculitis-induced microinfarction and neuronal infection causes severe tissue damage in the CNS. Both viruses can also cause relapsing encephalitis months and years after the acute infection. Many animal models studied to date have largely confirmed the pathology of henipavirus infection, and provided the means to test new therapeutic agents and vaccines. As the bat is the natural host of henipaviruses and has worldwide distribution, spillover events into human populations are expected to occur in the future.
Enterovirus A71 (EV-A71) causes hand-foot-and-mouth disease (HFMD), which may be complicated by fatal encephalomyelitis. Although fecal-oral or oral-oral routes are important in person-to-person transmission, how viral shedding and exposure may predispose individuals to infection remains unknown. We investigated person-to-person transmission by using a model of HFMD and encephalomyelitis based on EV-A71 oral infection of 2-week-old hamsters. Animals (index animals) infected with 104 50% cell culture infective doses of virus uniformly developed severe disease four days post-infection (dpi), whereas littermate contacts developed severe disease after six to seven days of exposure to index animals. Virus was detected in oral washes and feces at 3-4 dpi in index animals and at three to eight days after exposure to index animals in littermate contact animals. In a second experiment, non-littermate contact animals exposed for 8 or 12 h to index animals developed the disease six and four days post-exposure, respectively. Tissues from killed index and contact animals, studied by light microscopy, immunohistochemistry and in situ hybridization, exhibited mild inflammatory lesions and/or viral antigens/RNA in the squamous epithelia of the oral cavity, tongue, paws, skin, esophagus, gastric epithelium, salivary glands, lacrimal glands, central nervous system neurons, muscles (skeletal, cardiac and smooth muscles) and liver. Orally shed viruses were probably derived from infected oral mucosa and salivary glands, whereas fecal viruses may have derived from these sites as well as from esophageal and gastric epithelia. Asymptomatic seroconversion in exposed mother hamsters was demonstrated. Our hamster model should be useful in studying person-to-person EV-A71 transmission and how drugs and vaccines may interrupt transmission.
Background and Objective: There is a great challenge to establish a level 4 epilepsy care offering
complete evaluation for epilepsy surgery including invasive monitoring in a resource-limited country.
This study aimed to report the setup of a level 4 comprehensive epilepsy program in Malaysia and the
outcome of epilepsy surgery over the past 4 years.
Methods: This is a retrospective study analyzing
cases with intractable epilepsy in a comprehensive epilepsy program in University Malaya Medical
Center (UMMC), Kuala Lumpur, from January 2012 to August 2016.
Results: A total of 92 cases
had comprehensive epilepsy evaluation from January 2012 till August 2016. The mean age was 35.57
years old (range 15-59) and 54 (58.7%) were male. There were 17 cases having epilepsy surgery
after stage-1 evaluation. Eleven cases had mesial temporal sclerosis and 81% achieved Engel class
I surgical outcome. Six cases had lesionectomy and 60% had Engel class I outcome. A total of 16
surgeries were performed after stage-2 evaluation, including invasive EEG monitoring in 9 cases.
Among those with surgery performed more than 12 months from the time of data collection, 5/10
(50%) achieved Engel I outcome, whereas 2 (20%) had worthwhile improvement (Engel class III)
with 75% and 90% seizure reduction.
Conclusion: Level 4 epilepsy care has an important role and is possible with joint multidisciplinary
effort in a middle-income country like Malaysia despite resource limitation.
Primary angiitis of the central nervous system (PACNS) is a rare vasculitis restricted to the central nervous system without systemic involvement. Delay in diagnosis and treatment is common due to its non-specific symptoms and lack of non-invasive diagnostic tests. Myelopathy can occur in PACNS, during the clinical course of the illness, with or without cerebral symptoms. We describe here a 51 year-old ethnic Chinese woman who presented initially with paraparesis without cerebral symptoms. The diagnosis of PACNS was eventually made from brain biopsy when she subsequently developed cerebral involvement. Despite aggressive treatment, the patient developed progressive neurological deterioration and died. This patient demonstrates the rare occurrence of myelopathy as the sole initial presentation of PACNS.
Myopathies, although presenting more commonly in the younger age group, can occur and contribute significantly to disability in the elderly. To describe the spectrum of elderly myopathies, we reviewed 52 elderly patients (> 65 years) from the University of Malaya Medical Centre muscle biopsy databank, constituting 6.8% of 759 adult patients (> 18 years) who underwent muscle biopsy between 1992 and 2012. Commonest were the inflammatory myopathies (41/52, 78.8%), of which 43.9% had dermatomyositis; 23.9% polymyositis; 14.6% sporadic inclusion body myositis; 9.8% undifferentiated myositis and 2.4% overlap myositis. Seven patients (13.4%) had genetic myopathy; 2 muscular dystrophy and 5 chronic progressive external ophthalmoplegia, while 4 patients (7.7%) had drug-associated myopathy, 3 with statins. Malignancies were seen in 9.8% of inflammatory myopathies at diagnosis. Both acquired and genetic myopathies are seen in elderly Malaysians of all ethnicities and should not be misdiagnosed as some are potentially treatable and/or associated with malignancy.
Dengue virus is a flavivirus transmitted by the mosquitoes, Aedes aegypti and Aedes albopictus. Dengue infection by all four serotypes (DEN 1 to 4) is endemic globally in regions with tropical and subtropical climates, with an estimated 100-400 million infections annually. Among those hospitalized, the mortality is about 1%. Neurological involvement has been reported to be about 5%. The spectrum of neurological manifestations spans both the peripheral and central nervous systems. These manifestations could possibly be categorized into those directly related to dengue infection, i.e. acute and chronic encephalitis, indirect complications leading to dengue encephalopathy, and post-infectious syndrome due to immune-mediated reactions, and manifestations with uncertain mechanisms, such as acute transverse myelitis, acute cerebellitis and myositis. The rising trend in global dengue incidence calls for attention to a more explicit definition of each neurological manifestation for more accurate epidemiological data. The actual global burden of dengue infection with neurological manifestation is essential for future planning and execution of strategies, especially in the development of effective antivirals and vaccines against the dengue virus. In this article, we discuss the recent findings of different spectrums of neurological manifestations in dengue infection and provide an update on antiviral and vaccine development and their challenges.
Enterovirus 71 (EV71)-associated hand, foot, and mouth disease may be complicated by encephalomyelitis. We investigated EV71 brainstem infection and whether this infection could be ameliorated by passive immunization in a mouse model. Enterovirus 71 was injected into unilateral jaw/facial muscles of 2-week-old mice, and hyperimmune sera were given before or after infection. Harvested tissues were studied by light microscopy, immunohistochemistry, in situ hybridization, and viral titration. In unimmunized mice, viral antigen and RNA were detected within 24 hours after infection only in ipsilateral cranial nerves, motor trigeminal nucleus, reticular formation, and facial nucleus; viral titers were significantly higher in the brainstem than in the spinal cord samples. Mice given preinfection hyperimmune serum showed a marked reduction of ipsilateral viral antigen/RNA and viral titers in the brainstem in a dose-dependent manner. With optimum hyperimmune serum given after infection, brainstem infection was significantly reduced in a time-dependent manner. A delay in disease onset and a reduction of disease severity and mortality were also observed. Thus, EV71 can directly infect the brainstem, including the medulla, via cranial nerves, most likely by retrograde axonal transport. This may explain the sudden cardiorespiratory collapse in human patients with fatal encephalomyelitis. Moreover, our results suggest that passive immunization may still benefit EV71-infected patients who have neurologic complications.
Enterovirus A71 (EV-A71) causes self-limiting, hand-foot-and-mouth disease (HFMD) that may rarely be complicated by encephalomyelitis. Person-to-person transmission is usually by fecal-oral or oral-oral routes. To study viral replication sites in the oral cavity and other tissues, and to gain further insights into virus shedding and neuropathogenesis, we developed a consistent, orally-infected, 2-week-old hamster model of HFMD and EV-A71 encephalomyelitis. Tissues from orally-infected, 2-week-old hamsters were studied by light microscopy, immunohistochemistry and in situ hybridization to detect viral antigens and RNA, respectively, and by virus titration. Hamsters developed the disease and died after 4-8 days post infection; LD50 was 25 CCID50. Macroscopic cutaneous lesions around the oral cavity and paws were observed. Squamous epithelium in the lip, oral cavity, paw, skin, and esophagus, showed multiple small inflammatory foci around squamous cells that demonstrated viral antigens/RNA. Neurons (brainstem, spinal cord, sensory ganglia), acinar cells (salivary gland, lacrimal gland), lymphoid cells (lymph node, spleen), and muscle fibres (skeletal, cardiac and smooth muscles), liver and gastric epithelium also showed varying amounts of viral antigens/RNA. Intestinal epithelium, Peyer's patches, thymus, pancreas, lung and kidney were negative. Virus was isolated from oral washes, feces, brain, spinal cord, skeletal muscle, serum, and other tissues. Our animal model should be useful to study squamous epitheliotropism, neuropathogenesis, oral/fecal shedding in EV-A71 infection, person-to-person transmission, and to test anti-viral drugs and vaccines.
Radiology-pathology correlation is essential for multidisciplinary collaboration in diagnosis and understanding the mechanism of CNS damage in infectious processes. The microscopic acute inflammatory processes are well established and are supplemented by a variety of less-invasive microbial and immunohistochemical investigations. Understanding the pathogenesis of pathogen spread and neuroinvasion, vascular and immune-mediated brain, and spinal cord damage are essential for interpreting radiological images.
Encephalomyelitis is a well-known complication of hand, foot, and mouth disease (HFMD) due to Enterovirus 71 (EV71) infection. Viral RNA/antigens could be detected in the central nervous system (CNS) neurons in fatal encephalomyelitis but the mechanisms of neuronal cell death is not clearly understood. We investigated the role of absent in melanoma 2 (AIM2) inflammasome in neuronal cell death, and its relationship to viral replication. Our transcriptomic analysis, RT-qPCR, Western blot, immunofluorescence and flow cytometry studies consistently showed AIM2 gene up-regulation and protein expression in EV-A71-infected SK-N-SH cells. Downstream AIM2-induced genes, CARD16, caspase-1 and IL-1β were also up-regulated and caspase-1 was activated to form cleaved caspase-1 p20 subunits. As evidenced by 7-AAD positivity, pyroptosis was confirmed in infected cells. Overall, these findings have a strong correlation with decreases in viral titers, copy numbers and proteins, and reduced proportions of infected cells. AIM2 and viral antigens were detected by immunohistochemistry in infected neurons in inflamed areas of the CNS in EV-A71 encephalomyelitis. In infected AIM2-knockdown cells, AIM2 and related downstream gene expressions, and pyroptosis were suppressed, resulting in significantly increased virus infection. These results support the notion that AIM2 inflammasome-mediated pyroptosis is an important mechanism of neuronal cell death and it could play an important role in limiting EV-A71 replication.
Coxsackievirus A16 (CV-A16) and enterovirus A71 (EV-A71) are closely related enteroviruses that cause the same hand, foot, and mouth disease (HFMD), but neurological complications occur only very rarely in CV-A16 compared to EV-A71 infections. To elucidate host responses that may be able to explain these differences, we performed transcriptomic analysis and real-time quantitative PCR (RT-qPCR) in CV-A16-infected neuroblastoma cells (SK-N-SH), and the results showed that the radical S-adenosylmethionine domain containing 2 (RSAD2) was the highest upregulated gene in the antimicrobial pathway. Increased RSAD2 expression was correlated with reduced viral replication, while RSAD2 knockdown cells were correlated with increased replication. EV-A71 replication showed no apparent correlation to RSAD2 expressions. Absent in melanoma 2 (AIM2), which is associated with pyroptotic cell death, was upregulated in EV-A71-infected neurons but not in CV-A16 infection, suggesting that the AIM2 inflammasome played a significant role in suppressing EV-A71 replication. Chimeric viruses derived from CV-A16 and EV-A71 but containing swapped 5' nontranslated regions (5' NTRs) showed that RSAD2 expression/viral replication and AIM2 expression/viral replication patterns may be linked to the 5' NTRs of parental viruses. Differences in secondary structure of internal ribosomal entry sites within the 5' NTR may be responsible for these findings. Overall, our results suggest that CV-A16 and EV-A71 elicit different host responses to infection, which may help explain the apparent lower incidence of CV-A16-associated neurovirulence in HFMD outbreaks compared to EV-A71 infection.IMPORTANCE Although coxsackievirus A16 (CV-A16) and enterovirus A17 (EV-A71) both cause hand, foot, and mouth disease, EV-A71 has emerged as a leading cause of nonpolio, enteroviral fatal encephalomyelitis among young children. The significance of our research is in the identification of the possible differing and novel mechanisms of CV-A16 and EV-A71 inhibition in neuronal cells that may impact viral neuropathogenesis. We further showed that viral 5' NTRs may play significant roles in eliciting different host response mechanisms.
Japanese encephalitis virus (JEV) causes central nervous system neuronal injury and inflammation. A clear understanding of neuronal responses to JEV infection remains elusive. Using the Affymetrix array to investigate the transcriptome of infected SK-N-MC cells, 1316 and 2737 dysregulated genes (≥ 2/-2 fold change, P
Background: Rosette-forming glioneuronal tumor (World Health Organization grade I) is considered as a benign tumor with very low potential for progression. The potential for malignant transformation of this tumor is not known and has never been reported before in literature.
Case Description: We report a 42-year-old man, diagnosed with rosette-forming glioneuronal tumor of the fourth ventricle with a positive isocitrate dehydrogenase 1 mutation, progressed to glioblastoma after 6 years from diagnosis. We discuss the clinical history, radiological findings, and histopathological characteristic with immunohistochemistry findings observed in this unique case.
Conclusions: Despite being acceptable as benign, based on our observations in this case, there is a potential for malignant transformation of rosette-forming glioneuronal tumor. The role of isocitrate dehydrogenase 1 mutation leading to malignant transformation could not be established as our finding is novel and further prospective studies are required to prove this association.
Japanese encephalitis (JE) is a known CNS viral infection that often involves the thalamus early. To investigate the possible role of sensory peripheral nervous system (PNS) in early neuroinvasion, we developed a left hindlimb footpad-inoculation mouse model to recapitulate human infection by a mosquito bite. A 1-5 days postinfection (dpi) study, demonstrated focal viral antigens/RNA in contralateral thalamic neurons at 3 dpi in 50% of the animals. From 4 to 5 dpi, gradual increase in viral antigens/RNA was observed in bilateral thalami, somatosensory, and piriform cortices, and then the entire CNS. Infection of neuronal bodies and adjacent nerves in dorsal root ganglia (DRGs), trigeminal ganglia, and autonomic ganglia (intestine, etc.) was also observed from 5 dpi. Infection of explant organotypic whole brain slice cultures demonstrated no viral predilection for the thalamus, while DRG and intestinal ganglia organotypic cultures confirmed sensory and autonomic ganglia susceptibility to infection, respectively. Early thalamus and sensory-associated cortex involvement suggest an important role for sensory pathways in neuroinvasion. Our results suggest that JE virus neuronotropism is much more extensive than previously known, and that the sensory PNS and autonomic system are susceptible to infection.